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Sample records for micropatterned hepatocellular co-cultures

  1. Scanning electron microscopical observation of an osteoblast/osteoclast co-culture on micropatterned orthopaedic ceramics.

    Science.gov (United States)

    Halai, Mansur; Ker, Andrew; Meek, Rm Dominic; Nadeem, Danish; Sjostrom, Terje; Su, Bo; McNamara, Laura E; Dalby, Matthew J; Young, Peter S

    2014-01-01

    In biomaterial engineering, the surface of an implant can influence cell differentiation, adhesion and affinity towards the implant. On contact with an implant, bone marrow-derived mesenchymal stromal cells demonstrate differentiation towards bone forming osteoblasts, which can improve osteointegration. The process of micropatterning has been shown to improve osteointegration in polymers, but there are few reports surrounding ceramics. The purpose of this study was to establish a co-culture of bone marrow-derived mesenchymal stromal cells with osteoclast progenitor cells and to observe the response to micropatterned zirconia toughened alumina ceramics with 30 µm diameter pits. The aim was to establish whether the pits were specifically bioactive towards osteogenesis or were generally bioactive and would also stimulate osteoclastogenesis that could potentially lead to osteolysis. We demonstrate specific bioactivity of micropatterns towards osteogenesis, with more nodule formation and less osteoclastogenesis compared to planar controls. In addition, we found that that macrophage and osteoclast-like cells did not interact with the pits and formed fewer full-size osteoclast-like cells on the pitted surfaces. This may have a role when designing ceramic orthopaedic implants.

  2. Micropatterned co-culture of hepatocyte spheroids layered on non-parenchymal cells to understand heterotypic cellular interactions

    International Nuclear Information System (INIS)

    Otsuka, Hidenori; Sasaki, Kohei; Okimura, Saya; Nagamura, Masako; Nakasone, Yuichi

    2013-01-01

    Microfabrication and micropatterning techniques in tissue engineering offer great potential for creating and controlling cellular microenvironments including cell–matrix interactions, soluble stimuli and cell–cell interactions. Here, we present a novel approach to generate layered patterning of hepatocyte spheroids on micropatterned non-parenchymal feeder cells using microfabricated poly(ethylene glycol) (PEG) hydrogels. Micropatterned PEG-hydrogel-treated substrates with two-dimensional arrays of gelatin circular domains (ϕ = 100 μm) were prepared by photolithographic method. Only on the critical structure of PEG hydrogel with perfect protein rejection, hepatocytes were co-cultured with non-parenchymal cells to be led to enhanced hepatocyte functions. Then, we investigated the mechanism of the functional enhancement in co-culture with respect to the contributions of soluble factors and direct cell–cell interactions. In particular, to elucidate the influence of soluble factors on hepatocyte function, hepatocyte spheroids underlaid with fibroblasts (NIH/3T3 mouse fibroblasts) or endothelial cells (BAECs: bovine aortic endothelial cells) were compared with physically separated co-culture of hepatocyte monospheroids with NIH3T3 or BAEC using trans-well culture systems. Our results suggested that direct heterotypic cell-to-cell contact and soluble factors, both of these between hepatocytes and fibroblasts, significantly enhanced hepatocyte functions. In contrast, direct heterotypic cell-to-cell contact between hepatocytes and endothelial cells only contributed to enhance hepatocyte functions. This patterning technique can be a useful experimental tool for applications in basic science, drug screening and tissue engineering, as well as in the design of artificial liver devices. (paper)

  3. Engineering blood vessels through micropatterned co-culture of vascular endothelial and smooth muscle cells on bilayered electrospun fibrous mats with pDNA inoculation.

    Science.gov (United States)

    Liu, Yaowen; Lu, Jinfu; Li, Huinan; Wei, Jiaojun; Li, Xiaohong

    2015-01-01

    Although engineered blood vessels have seen important advances during recent years, proper mechanical strength and vasoactivity remain unsolved problems. In the current study, micropatterned fibrous mats were created to load smooth muscle cells (SMC), and a co-culture with endothelial cells (EC) was established through overlaying on an EC-loaded flat fibrous mat to mimic the layered structure of a blood vessel. A preferential distribution of SMC was determined in the patterned regions throughout the fibrous scaffolds, and aligned fibers in the patterned regions provided topological cues to guide the orientation of SMC with intense actin filaments and extracellular matrix (ECM) production in a circumferential direction. Plasmid DNA encoding basic fibroblast growth factors and vascular endothelial growth factor were integrated into electrospun fibers as biological cues to promote SMC infiltration into fibrous mats, and the viability and ECM production of both EC and SMC. The layered fibrous mats with loaded EC and SMC were wrapped into a cylinder, and engineered vessels were obtained with compact EC and SMC layers after co-culture for 3 months. Randomly oriented ECM productions of EC formed a continuous endothelium covering the entire lumenal surface, and a high alignment of ECM was shown in the circumferential direction of SMC layers. The tensile strength, strain at failure and suture retention strength were higher than those of the human femoral artery, and the burst pressure and radial compliance were in the same range as the human saphenous vein, indicating potential as blood vessel substitutes for transplantation in vivo. Thus, the establishment of topographical cues and biochemical signals in fibrous scaffolds demonstrates advantages in modulating cellular behavior and organization found in complex multicellular tissues. Copyright © 2014 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  4. Metformin affects the features of a human hepatocellular cell line (HepG2) by regulating macrophage polarization in a co-culture microenviroment.

    Science.gov (United States)

    Chen, Miaojiao; Zhang, Jingjing; Hu, Fang; Liu, Shiping; Zhou, Zhiguang

    2015-11-01

    Accumulating evidence suggests an association between diabetes and cancer. Inflammation is a key event that underlies the pathological processes of the two diseases. Metformin displays anti-cancer effects, but the mechanism is not completely clear. This study investigated whether metformin regulated the microenvironment of macrophage polarization to affect the characteristics of HepG2 cells and the possible role of the Notch-signalling pathway. RAW264.7 macrophages were cultured alone or co-cultured with HepG2 cells and treated with metformin. We analysed classical (M1) and alternative (M2) gene expression in RAW264.7 cells using quantitative real-time polymerase chain reaction. Changes in mRNA and protein expressions of Notch signalling in both cell types were also detected using quantitative real-time polymerase chain reaction and Western-blotting analyses. The proliferation, apoptosis and migration of HepG2 cells were detected using Cell Titer 96 AQueous One Solution Cell Proliferation Assay (MTS) (Promega Corporation, Fitchburg, WI, USA), Annexin V-FITC/PI (7SeaPharmTech, Shanghai, China) and the cell scratch assay, respectively. Metformin induced single-cultured RAW264.7 macrophages with an M2 phenotype but attenuated the M2 macrophage differentiation and inhibited monocyte chemoattractant protein-1 (MCP-1) secretion in a co-culture system. The co-cultured group of metformin pretreatment activated Notch signalling in macrophages but repressed it inHepG2 cells. Co-culture also promoted the proliferation and migration of HepG2 cells. However, along with the enhanced apoptosis, the proliferation and the migration of HepG2 cells were remarkably inhibited in another co-culture system with metformin pretreatment. Metformin can skew RAW264.7 macrophages toward different phenotypes according to changes in the microenvironment, which may affect the inflammatory conditions mediated by macrophages, induce apoptosis and inhibit the proliferation and migration of HepG2

  5. SU-8 micropatterning for microfluidic droplet and microparticle focusing

    International Nuclear Information System (INIS)

    Debuisson, Damien; Senez, Vincent; Arscott, Steve

    2011-01-01

    We demonstrate micropatterned surfaces consisting of concentric circles and spirals which can focus an evaporating sessile droplet to a specific location on a surface. We also study the micropattern geometry to focus microparticles contained within the droplet. The micropatterned surfaces are fabricated using the photoresist SU-8. Our process enables the modification of the surface wetting via the formation of smooth trench-like defects in the SU-8 which define the micropatterns; the geometry of these micropatterns determines the droplet/microparticle focusing. It is clearly shown that the introduction of small gaps into the micropatterns promotes microparticle centring due to the modification of the depinning angle of the droplet. We also show that the use of spiral micropatterns promotes microparticle centring. Finally, microparticle focusing can be enhanced by modification of surface wetting via the addition of a thin fluorocarbon hydrophobic layer onto the SU-8

  6. R&D on a new type of micropattern gaseous detector: The Fast Timing Micropattern detector

    Energy Technology Data Exchange (ETDEWEB)

    Abbaneo, D.; Abbas, M. [CERN, Geneva (Switzerland); Abbrescia, M. [INFN Bari and University of Bari, Bari (Italy); Akl, M. Abi [Texas A& M University at Qatar, Doha (Qatar); Aboamer, O. [Academy of Scientific Research and Technology, Egyptian Network of High Energy Physics, ASRT-ENHEP, Cairo (Egypt); Acosta, D. [University of Florida, Gainesville (United States); Ahmad, A. [National Center for Physics, Quaid-i-Azam University Campus, Islamabad (Pakistan); Ahmed, W. [INFN Bari and University of Bari, Bari (Italy); Aleksandrov, A. [Institute for Nuclear Research and Nuclear Energy, Sofia (Bulgaria); Altieri, P. [INFN Bari and University of Bari, Bari (Italy); Asawatangtrakuldee, C. [Peking University, Beijing (China); Aspell, P. [CERN, Geneva (Switzerland); Assran, Y. [Academy of Scientific Research and Technology, Egyptian Network of High Energy Physics, ASRT-ENHEP, Cairo (Egypt); Awan, I. [National Center for Physics, Quaid-i-Azam University Campus, Islamabad (Pakistan); Bally, S. [CERN, Geneva (Switzerland); Ban, Y. [Peking University, Beijing (China); Banerjee, S. [Saha Institute of Nuclear Physics, Kolkata (India); Barashko, V. [University of Florida, Gainesville (United States); Barria, P. [Universite Libre de Bruxelles, Brussels (Belgium); Bencze, G. [Institute for Particle and Nuclear Physics, Wigner Research Centre for Physics, Hungarian Academy of Sciences, Budapest (Hungary); and others

    2017-02-11

    This contribution introduces a new type of Micropattern Gaseous Detector, the Fast Timing Micropattern (FTM) detector, utilizing fully Resistive WELL structures. The structure of the prototype will be described in detail and the results of the characterization study performed with an X-ray gun will be presented, together with the first results on time resolution based on data collected with muon/pion test beams.

  7. A rapid co-culture stamping device for studying intercellular communication

    Science.gov (United States)

    Hassanzadeh-Barforoushi, Amin; Shemesh, Jonathan; Farbehi, Nona; Asadnia, Mohsen; Yeoh, Guan Heng; Harvey, Richard P.; Nordon, Robert E.; Warkiani, Majid Ebrahimi

    2016-10-01

    Regulation of tissue development and repair depends on communication between neighbouring cells. Recent advances in cell micro-contact printing and microfluidics have facilitated the in-vitro study of homotypic and heterotypic cell-cell interaction. Nonetheless, these techniques are still complicated to perform and as a result, are seldom used by biologists. We report here development of a temporarily sealed microfluidic stamping device which utilizes a novel valve design for patterning two adherent cell lines with well-defined interlacing configurations to study cell-cell interactions. We demonstrate post-stamping cell viability of >95%, the stamping of multiple adherent cell types, and the ability to control the seeded cell density. We also show viability, proliferation and migration of cultured cells, enabling analysis of co-culture boundary conditions on cell fate. We also developed an in-vitro model of endothelial and cardiac stem cell interactions, which are thought to regulate coronary repair after myocardial injury. The stamp is fabricated using microfabrication techniques, is operated with a lab pipettor and uses very low reagent volumes of 20 μl with cell injection efficiency of >70%. This easy-to-use device provides a general strategy for micro-patterning of multiple cell types and will be important for studying cell-cell interactions in a multitude of applications.

  8. Micropatterning on glass with deep UV

    OpenAIRE

    sprotocols

    2015-01-01

    Authors: Nicolas CARPI, Matthieu PIEL, Ammar Azioune & Jenny Fink ### Abstract This protocol describes a method to print micropatterns on glass with extra-cellular matrix proteins to promote cell adhesion. The non-adhesive part is made with polylysine grafted polyethyleneglycol (PLL-g-PEG). This technique is reproducible, cheap, fast and can achieve high resolution (~1 µm). ### Introduction This protocol explains how to make high resolution adhesive micropattens of protein...

  9. Progress in micro-pattern gas detectors

    International Nuclear Information System (INIS)

    Bellazzini, Ronaldo

    2001-01-01

    Micro-Pattern Gas Detectors are position-sensitive proportional counters whose sense electrodes are constructed using micro-electronics , thin-film or advanced PCB techniques.The feature size attainable using these methods is of the order of a few microns and the detectors demonstrate excellent spatial resolution and fast charge collection. I will review recent progress on Micro patterned Gas Detectors for tracking and other cross-disciplinary applications.I will focus on the design principles,performance capability and limitations. A short list of interesting applications will be discussed

  10. Cell Migration According to Shape of Graphene Oxide Micropatterns

    Directory of Open Access Journals (Sweden)

    Sung Eun Kim

    2016-10-01

    Full Text Available Photolithography is a unique process that can effectively manufacture micro/nano-sized patterns on various substrates. On the other hand, the meniscus-dragging deposition (MDD process can produce a uniform surface of the substrate. Graphene oxide (GO is the oxidized form of graphene that has high hydrophilicity and protein absorption. It is widely used in biomedical fields such as drug delivery, regenerative medicine, and tissue engineering. Herein, we fabricated uniform GO micropatterns via MDD and photolithography. The physicochemical properties of the GO micropatterns were characterized by atomic force microscopy (AFM, scanning electron microscopy (SEM, and Raman spectroscopy. Furthermore, cell migration on the GO micropatterns was investigated, and the difference in cell migration on triangle and square GO micropatterns was examined for their effects on cell migration. Our results demonstrated that the GO micropatterns with a desired shape can be finely fabricated via MDD and photolithography. Moreover, it was revealed that the shape of GO micropatterns plays a crucial role in cell migration distance, speed, and directionality. Therefore, our findings suggest that the GO micropatterns can serve as a promising biofunctional platform and cell-guiding substrate for applications to bioelectric devices, cell-on-a-chip, and tissue engineering scaffolds.

  11. Developing and evaluating new micropattern gas detectors

    International Nuclear Information System (INIS)

    Villa, Marco

    2014-02-01

    Micropattern gas detectors (MPGDs) were introduced in the late 1980s in order to overcome the limited rate capability of traditional proportional counters. Thanks to their microscopic electrode structures, MPGDs are faster and more precise than the previous gas detectors and soon gained popularity. Two of the most successful MPGDs are the gas electron multiplier (GEM) and the micro-mesh gaseous structure (Micromegas). In this thesis I present the features of GEMs and Micromegas, some of their current applications and the research and development that I have done on these technologies. My activity covered two main topics: the test and enhancement of single-mask GEMs for large-area applications and the study of spark-tolerant Micromegas for the upgrade of the ATLAS Small Wheels.

  12. Subsurface oxidation for micropatterning silicon (SOMS).

    Science.gov (United States)

    Zhang, Feng; Sautter, Ken; Davis, Robert C; Linford, Matthew R

    2009-02-03

    Here we present a straightforward patterning technique for silicon: subsurface oxidation for micropatterning silicon (SOMS). In this method, a stencil mask is placed above a silicon surface. Radio-frequency plasma oxidation of the substrate creates a pattern of thicker oxide in the exposed regions. Etching with HF or KOH produces very shallow or much higher aspect ratio features on silicon, respectively, where patterning is confirmed by atomic force microscopy, scanning electron microscopy, and optical microscopy. The oxidation process itself is studied under a variety of reaction conditions, including higher and lower oxygen pressures (2 and 0.5 Torr), a variety of powers (50-400 W), different times and as a function of reagent purity (99.5 or 99.994% oxygen). SOMS can be easily executed in any normal chemistry laboratory with a plasma generator. Because of its simplicity, it may have industrial viability.

  13. Micropatterned substrates for studying astrocytes in culture

    Directory of Open Access Journals (Sweden)

    William Lee

    2009-12-01

    Full Text Available Recent studies of the physiological roles of astrocytes have ignited renewed interest in the functional significance of these glial cells in the central nervous system. Many of the newly discovered astrocytic functions were initially demonstrated and characterized in cell culture systems. We discuss the use of microculture techniques and micropatterning of cell-adhesive substrates in studies of astrocytic Ca2+ excitability and bidirectional neuron-astrocyte signaling. This culturing approach aims to reduce the level of complexity of the system by limiting the interacting partners and by controlling the localization of cells. It provides tight control over experimental conditions allowing detailed characterization of cellular functions and intercellular communication. Although such a reductionist approach yields some difference in observations between astrocytic properties in culture and in situ, general phenomena discovered in cell culture systems, however, have also been found in vivo.

  14. Hepatocellular calcification

    DEFF Research Database (Denmark)

    Ladefoged, Claus; Frifelt, J J

    1987-01-01

    Autopsy of a twenty year old girl dying from complications of renal and cardiac failure demonstrated severe hepatocellular calcification, a rare finding. The pathogenesis is thought to be a combination of dystrophic calcification caused by severe centrilobular necrosis and metastatic calcificatio...

  15. Opposite cytokine synthesis by fibroblasts in contact co-culture with osteosarcoma cells compared with transwell co-cultures.

    Science.gov (United States)

    David, Manu S; Kelly, Elizabeth; Zoellner, Hans

    2013-04-01

    We recently reported exchange of membrane and cytoplasm during contact co-culture between human Gingival Fibroblasts (h-GF) and SAOS-2 osteosarcoma cells, a process we termed 'cellular sipping' to reflect the manner in which cells become morphologically diverse through uptake of material from the opposing cell type, independent of genetic change. Cellular sipping is increased by Tumor Necrosis Factor-α (TNF-α), and we here show for the first time altered cytokine synthesis in contact co-culture supporting cellular sipping compared with co-culture where h-GF and SAOS-2 were separated in transwells. SAOS-2 had often undetectably low cytokine levels, while Interleukin-6 (IL-6), Granulocyte Colony Stimulating Factor (G-CSF) and Granulocyte-Macrophage Colony Stimulating Factor (GM-CSF) were secreted primarily by TNF-α stimulated h-GF and basic Fibroblast Growth Factor (FGF) was prominent in h-GF lysates (p cultures permitting cellular sipping had lower IL-6, G-CSF and GM-CSF levels, as well as higher lysate FGF levels compared with TNF-α treated h-GF alone (p cultures in transwells, with increased IL-6, G-CSF and GM-CSF levels (p cultures where cellular sipping occurs, potentially contributing to tumor inflammatory responses. Crown Copyright © 2013. Published by Elsevier Ltd. All rights reserved.

  16. The study and fabrication of DLC micropattern on roll mold

    Science.gov (United States)

    Kwon, Young Woo; Lee, Tae Dong; Park, Yeong Min; Cho, Hyun; Kim, Jin Kon; Kim, Tae Gyu

    2015-03-01

    Diamond-like carbon (DLC) coating is becoming a promising protective coating layers due to its superior properties. In this study, instead of protective coating, DLC film was applied as the only component for micropattern then etched with lithography and lift-off process selectively. Furthermore, DLC film has been fabricated on aluminum roll mold. Then UV curing resin was applied to form the pattern on the polyethylene terephthalate (PET) film. The dimension and formation of the DLC micropattern on roll mold were analyzed. Moreover, the Raman spectroscopic of nitrogen-doped DLC film was analyzed.

  17. Hepatocellular carcinoma

    International Nuclear Information System (INIS)

    Farooqi, J.I.; Farooqi, R.J.

    2001-01-01

    Hepatocellular carcinoma (HCC) is a common cause of cancer mortality. Hepatitis B and C viruses, aflatoxin and alga toxin in the contaminated drinking water are the major etiological factors. Rapidly progressing medical imaging has resulted in the improved treatment results. Surgical resection has a major role for influencing prognosis of HCC. Local cancer therapies based on the advances in early diagnosis are progressing rapidly. Multimodality combination and sequential treatment has proved effective, unfortunately systemic chemotherapy for HCC remains disappointed. All of these have resulted in the improved prognosis of HCC. (author)

  18. Adhesive Micropatterns for Cells: A Microcontact Printing Protocol

    OpenAIRE

    sprotocols

    2014-01-01

    Authors: Manuel Théry and Matthieu Piel Corresponding authors ([](); []()) ### INTRODUCTION This protocol describes a simple, fast, and efficient method for making adhesive micropatterns that can be used to control individual cell shape and adhesion patterns. It is based on the use of an elastomeric stamp containing microfeatures to print proteins on the substrate of choice. The process can be subdiv...

  19. Micropatterning on cylindrical surfaces via electrochemical etching using laser masking

    International Nuclear Information System (INIS)

    Cho, Chull Hee; Shin, Hong Shik; Chu, Chong Nam

    2014-01-01

    Highlights: • Various micropatterns were fabricated on the cylindrical surface of a stainless steel shaft. • Selective electrochemical dissolution was achieved via a series process of laser masking and electrochemical etching. • Laser masking characteristics on the non-planar surface were investigated. • A uniform mask layer was formed on the cylindrical surface via synchronized laser line scanning with a rotary system. • The characteristics of electrochemical etching on the non-planar surface were investigated. - Abstract: This paper proposes a method of selective electrochemical dissolution on the cylindrical surfaces of stainless steel shafts. Selective electrochemical dissolution was achieved via electrochemical etching using laser masking. A micropatterned recast layer was formed on the surface via ytterbium-doped pulsed fiber laser irradiation. The micropatterned recast layer could be used as a mask layer during the electrochemical etching process. Laser masking condition to form adequate mask layer on the planar surface for etching cannot be used directly on the non-planar surface. Laser masking condition changes depending on the morphological surface. The laser masking characteristics were investigated in order to form a uniform mask layer on the cylindrical surface. To minimize factors causing non-uniformity in the mask layer on the cylindrical surface, synchronized laser line scanning with a rotary system was applied during the laser masking process. Electrochemical etching characteristics were also investigated to achieve deeper etched depth, without collapsing the recast layer. Consequently, through a series process of laser masking and electrochemical etching, various micropatternings were successfully performed on the cylindrical surfaces

  20. Effects of Co-Culture Media on Hepatic Differentiation of hiPSC with or without HUVEC Co-Culture.

    Science.gov (United States)

    Freyer, Nora; Greuel, Selina; Knöspel, Fanny; Strahl, Nadja; Amini, Leila; Jacobs, Frank; Monshouwer, Mario; Zeilinger, Katrin

    2017-08-07

    The derivation of hepatocytes from human induced pluripotent stem cells (hiPSC) is of great interest for applications in pharmacological research. However, full maturation of hiPSC-derived hepatocytes has not yet been achieved in vitro. To improve hepatic differentiation, co-cultivation of hiPSC with human umbilical vein endothelial cells (HUVEC) during hepatic differentiation was investigated in this study. In the first step, different culture media variations based on hepatocyte culture medium (HCM) were tested in HUVEC mono-cultures to establish a suitable culture medium for co-culture experiments. Based on the results, two media variants were selected to differentiate hiPSC-derived definitive endodermal (DE) cells into mature hepatocytes with or without HUVEC addition. DE cells differentiated in mono-cultures in the presence of those media variants showed a significant increase ( p < 0.05) in secretion of α-fetoprotein and in activities of cytochrome P450 (CYP) isoenzymes CYP2B6 and CYP3A4 as compared with cells differentiated in unmodified HCM used as control. Co-cultivation with HUVEC did not further improve the differentiation outcome. Thus, it can be concluded that the effect of the used medium outweighed the effect of HUVEC co-culture, emphasizing the importance of the culture medium composition for hiPSC differentiation.

  1. Micropatterning of a nanoporous alumina membrane with poly(ethylene glycol) hydrogel to create cellular micropatterns on nanotopographic substrates.

    Science.gov (United States)

    Lee, Hyun Jong; Kim, Dae Nyun; Park, Saemi; Lee, Yeol; Koh, Won-Gun

    2011-03-01

    In this paper, we describe a simple method for fabricating micropatterned nanoporous substrates that are capable of controlling the spatial positioning of mammalian cells. Micropatterned substrates were prepared by fabricating poly(ethylene glycol) (PEG) hydrogel microstructures on alumina membranes with 200 nm nanopores using photolithography. Because hydrogel precursor solution could infiltrate and become crosslinked within the nanopores, the resultant hydrogel micropatterns were firmly anchored on the substrate without the use of adhesion-promoting monolayers, thereby allow tailoring of the surface properties of unpatterned nanoporous areas. For mammalian cell patterning, arrays of microwells of different dimensions were fabricated. These microwells were composed of hydrophilic PEG hydrogel walls surrounding nanoporous bottoms that were modified with cell-adhesive Arg-Gly-Asp (RGD) peptides. Because the PEG hydrogel was non-adhesive towards proteins and cells, cells adhered selectively and remained viable within the RGD-modified nanoporous regions, thereby creating cellular micropatterns. Although the morphology of cell clusters and the number of cells inside one microwell were dependent on the lateral dimension of the microwells, adhered cells that were in direct contact with nanopores were able to penetrate into the nanopores by small extensions (filopodia) for all the different sizes of microwells evaluated. Copyright © 2010 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  2. Liver (Hepatocellular) Cancer Screening

    Science.gov (United States)

    ... Treatment Liver Cancer Prevention Liver Cancer Screening Research Liver (Hepatocellular) Cancer Screening (PDQ®)–Patient Version What is ... These are called diagnostic tests . General Information About Liver (Hepatocellular) Cancer Key Points Liver cancer is a ...

  3. Cell micro-patterning by atom beam exposure

    International Nuclear Information System (INIS)

    Adachi, Taiji; Kajita, Fumiaki; Sato, Katsuya; Matsumoto, Koshi; Tagawa, Masahiro

    2003-01-01

    This study aimed to develop a new cell micro-patterning method by controlling material surface affinity of the cell using atomic oxygen beam exposure. Surfaces of low-density polyethylene (LDPE) and tetrafluoroethylene-hexafluoropropylene (FEP) were exposed to the atomic oxygen beam. On the LDPE surface, the roughness measured by atomic force microscopy (AFM) did not change much, however, the oxygen concentration on the surface measured by X-ray photoelectron spectroscopy (XPS) significantly increased that resulted in increase in wettability. Contrary to this, on the FEP surface, the oxygen concentration showed no significant change, but roughness of the surface remarkably increased and the wettability decreased. As a result of the surface modification, affinity of the osteoblastic cells on the FEP surface increased, which was also confirmed by increase in the cell area. Finally, cell micro-patterning on the FEP surface was carried out based on difference in the affinity between modified and unmodified surfaces patterned by masking method. (author)

  4. Biomimicking micropatterned surfaces and their effect on marine biofouling.

    Science.gov (United States)

    Brzozowska, Agata M; Parra-Velandia, Fernando J; Quintana, Robert; Xiaoying, Zhu; Lee, Serina S C; Chin-Sing, Lim; Jańczewski, Dominik; Teo, Serena L-M; Vancso, Julius G

    2014-08-05

    When synthetic materials are submerged in marine environments, dissolved matter and marine organisms attach to their surfaces by a process known as marine fouling. This phenomenon may lead to diminished material performance with detrimental consequences. Bioinspired surface patterning and chemical surface modifications present promising approaches to the design of novel functional surfaces that can prevent biofouling phenomena. In this study, we report the synergistic effects of surface patterns, inspired by the marine decapod crab Myomenippe hardwickii in combination with chemical surface modifications toward suppressing marine fouling. M. hardwickii is known to maintain a relatively clean carapace although the species occurs in biofouling communities of tropical shallow subtidal coastal waters. Following the surface analysis of selected specimens, we designed hierarchical surface microtopographies that replicate the critical features observed on the crustacean surface. The micropatterned surfaces were modified with zwitterionic polymer brushes or with layer-by-layer deposited polyelectrolyte multilayers to enhance their antifouling and/or fouling-release potential. Chemically modified and unmodified micropatterned surfaces were subjected to extensive fouling tests, including laboratory assays against barnacle settlement and algae adhesion, and field static immersion tests. The results show a statistically significant reduction in settlement on the micropatterned surfaces as well as a synergistic effect when the microtopographies are combined with grafted polymer chains.

  5. Cell behaviors on micro-patterned porous thin films

    International Nuclear Information System (INIS)

    Phong, Ho Quoc; Wang Shuling; Wang, Meng-Jiy

    2010-01-01

    Thin polymer films with patterned surfaces have drawn tremendous attention in manufacturing advanced electronic, mechanical devices and in biomaterials due to the advantageous properties such as mechanical strength, chemical resistance and optic transparency. The applications can be extended to the fields such as catalysts, antireflection coatings, template for inorganic growth masks, and substrates for cell culturing providing the patterned surface containing micron-sized features. Various methods have been used to fabricate polymers with micro-patterned surfaces such as photolithographic, ink-jet printing, nonsolvent, spin coating in a dry environment, self-organization, and the condensation of monodisperse water droplet on the polymer solution. The physiological functions of mature cells depend on the microenvironment/niche surrounding which can provide proper factors to regulate cell proliferation and differentiation. While designing appropriate scaffolds for tissue engineering, the microstructure is one of the most important factors to be considered. In this work, a facile single-step phase separation method was used to create micro-patterned polymer thin films with concaves or convexes with sizes ranged from 7 to 70 μm. The effects of water content, casting volume and the addition of surfactant on the distribution of pores and substrate morphology were examined. Moreover, detailed observations of fibroblast cells on the micro-patterned thin films were presented to compare and elucidate the roles of surface micro-features and chemical functionalities.

  6. R and D on a New Technology of Micro-pattern Gaseous Detectors Fast Timing Micro-pattern Detector

    CERN Document Server

    Salva Diblen, Sinem

    2016-01-01

    After the upgrades of the Large Hadron Collider (LHC) planned for the second and the third Long Shutdown (LS), the LHC luminosity will approach very high values. Such conditions will affect the performance of the CMS muon system, especially in the very forward region, due to the harsh expected background environment and high pile-up conditions. The CMS collaboration considers upgrading the muon forward region to take advantage of the pixel tracking coverage extension a new detector, ME0 station, possibly behind the new forward calorimeter. New resistive micro-pattern gaseous detectors that are able to handle the very demanding spatial, time resolution and rate capability, are being considered. In this contribution we introduce a new type of MPGD technology the Fast Timing Micro-pattern (FTM) detector, utilizing a fully resistive WELL structure. It consists of a stack of several coupled layers where drift and WELL multiplication stages alternate in the structure, yielding a significant improvement in timing p...

  7. Elsevier R&D on a new type of micropattern gaseous detector: The Fast Timing Micropattern detector

    CERN Document Server

    Abbaneo, D; Abbrescia, M; Abi Akl, M; Aboamer, O; Acosta, D; Ahmad, A; Ahmed, W; Aleksandrov, A; Altieri, P; Asawatangtrakuldee, C; Aspell, P; Assran, Y; Awan, I; Bally, S; Ban, Y; Banerjee, S; Barashko, V; Barria, P; Bencze, G; Beni, N; Benussi, L; Bhopatkar, V; Bianco, S; Bos, J; Bouhali, O; Braghieri, A; Braibant, S; Buontempo, S; Calabria, C; Caponero, M; Caputo, C; Cassese, F; Castaneda, A; Cauwenbergh, S; Cavallo, F R; Celik, A; Choi, M; Choi, S; Christiansen, J; Cimmino, A; Colafranceschi, S; Colaleo, A; Conde Garcia, A; Czellar, S; Dabrowski, M M; De Lentdecker, G; Oliveira, R De; De Robertis, G; Dildick, S; Dorney, B; Endroczi, G; Errico, F; Fallavollita, F; Fenyvesi, A; Ferry, S; Furic, I; Giacomelli, P; Gilmore, J; Golovtsov, V; Guiducci, L; Guilloux, F; Gutierrez, A; Hadjiiska, R M; Hauser, J; Hoepfner, K; Hohlmann, M; Hoorani, H; Iaydjiev, P; Jeng, Y G; Kamon, T; Karchin, P; Korytov, A; Krutelyov, S; Kumar, A; Kim, H; Lee, J; Lenzi, T; Litov, L; Loddo, F; Madorsky, A; Maerschalk, T; Maggi, M; Magnani, A; Mal, P K; Mandal, K; Marchioro, A; Marinov, A; Majumdar, N; Merlin, J A; Mitselmakher, G; Mohanty, A K; Mohapatra, A; Molnar, J; Muhammad, S; Mukhopadhyay, S; Naimuddin, M; Nuzzo, S; Oliveri, E; Pant, L M; Paolucci, P; Park, I; Passeggio, G; Pavlov, B; Philipps, B; Piccolo, D; Postema, H; Puig Baranac, A; Radi, A; Radogna, R; Raffone, G; Ranieri, A; Rashevski, G; Ressegotti, M; Riccardi, C; Rodozov, M; Rodrigues, A; Ropelewski, L; RoyChowdhury, S; Ryu, G; Ryu, M S; Safonov, A; Salva, S; Saviano, G; Sharma, A; Sharma, A; Sharma, R; Shah, A H; Shopova, M; Sturdy, J; Sultanov, G; Swain, S K; Szillasi, Z; Talvitie, J; Tatarinov, A; Tuuva, T; Tytgat, M; Vai, I; Van Stenis, M; Venditti, R; Verhagen, E; Verwilligen, P; Vitulo, P; Volkov, S; Vorobyev, A; Wang, D; Wang, M; Yang, U; Yang, Y; Yonamine, R; Zaganidis, N; Zenoni, F; Zhang, A

    2017-01-01

    Micropattern gaseous detectors (MPGD) underwent significant upgrades in recent years, introducing resistive materials to build compact spark-protected devices. Exploiting this technology further, various features such as space and time resolution, rate capability, sensitive area, operational stability and radiation hardness can be improved. This contribution introduces a new type of MPGD, namely the Fast Timing Micropattern (FTM) detector, utilizing a fully resistive WELL structure. It consists of a stack of several coupled layers where drift and WELL multiplication stages alternate in the structure, yielding a significant improvement in timing properties due to competing ionization processes in the different drift regions. Two FTM prototypes have been developed so far. The first one is uWELL-like, where multiplication takes place in the holes of a kapton foil covered on both sides with resistive material. The second one has a resistive Micromegas-like structure, with multiplication developing in a region del...

  8. Staging Scenes of Co-Cultural Communication: Acting out Aspects of Marginalized and Dominant Identities

    Science.gov (United States)

    Root, Elizabeth

    2018-01-01

    Courses: Intercultural Communication, Interracial Communication, or an Interpersonal Communication class that covers co-cultural theory. Objectives: Students will be able to demonstrate a practical application of co-cultural theory by creating scenes that illustrate different communicative approaches and desired outcomes based on communication…

  9. Micropatterning of Functional Conductive Polymers with Multiple Surface Chemistries in Register

    DEFF Research Database (Denmark)

    Lind, Johan Ulrik; Acikgöz, Canet; Daugaard, Anders Egede

    2012-01-01

    A versatile procedure is presented for fast and efficient micropatterning of multiple types of covalently bound surface chemistry in perfect register on and between conductive polymer microcircuits. The micropatterning principle is applied to several types of native and functionalized PEDOT (poly(3...... functionalized conjugated polymer systems....

  10. A long-term three dimensional liver co-culture system for improved prediction of clinically relevant drug-induced hepatotoxicity

    International Nuclear Information System (INIS)

    Kostadinova, Radina; Boess, Franziska; Applegate, Dawn; Suter, Laura; Weiser, Thomas; Singer, Thomas; Naughton, Brian; Roth, Adrian

    2013-01-01

    Drug-induced liver injury (DILI) is the major cause for liver failure and post-marketing drug withdrawals. Due to species-specific differences in hepatocellular function, animal experiments to assess potential liabilities of drug candidates can predict hepatotoxicity in humans only to a certain extent. In addition to animal experimentation, primary hepatocytes from rat or human are widely used for pre-clinical safety assessment. However, as many toxic responses in vivo are mediated by a complex interplay among different cell types and often require chronic drug exposures, the predictive performance of hepatocytes is very limited. Here, we established and characterized human and rat in vitro three-dimensional (3D) liver co-culture systems containing primary parenchymal and non-parenchymal hepatic cells. Our data demonstrate that cells cultured on a 3D scaffold have a preserved composition of hepatocytes, stellate, Kupffer and endothelial cells and maintain liver function for up to 3 months, as measured by the production of albumin, fibrinogen, transferrin and urea. Additionally, 3D liver co-cultures maintain cytochrome P450 inducibility, form bile canaliculi-like structures and respond to inflammatory stimuli. Upon incubation with selected hepatotoxicants including drugs which have been shown to induce idiosyncratic toxicity, we demonstrated that this model better detected in vivo drug-induced toxicity, including species-specific drug effects, when compared to monolayer hepatocyte cultures. In conclusion, our results underline the importance of more complex and long lasting in vitro cell culture models that contain all liver cell types and allow repeated drug-treatments for detection of in vivo-relevant adverse drug effects. - Highlights: ► 3D liver co-cultures maintain liver specific functions for up to three months. ► Activities of Cytochrome P450s remain drug- inducible accross three months. ► 3D liver co-cultures recapitulate drug-induced liver toxicity

  11. A long-term three dimensional liver co-culture system for improved prediction of clinically relevant drug-induced hepatotoxicity

    Energy Technology Data Exchange (ETDEWEB)

    Kostadinova, Radina; Boess, Franziska [Non-Clinical Safety, Hoffmann-La Roche Ltd, Grenzacherstrasse 124, Building 73 / Room 117b, 4070 Basel (Switzerland); Applegate, Dawn [RegeneMed, 9855 Towne Centre Drive Suite 200, San Diego, CA 92121 (United States); Suter, Laura; Weiser, Thomas; Singer, Thomas [Non-Clinical Safety, Hoffmann-La Roche Ltd, Grenzacherstrasse 124, Building 73 / Room 117b, 4070 Basel (Switzerland); Naughton, Brian [RegeneMed, 9855 Towne Centre Drive Suite 200, San Diego, CA 92121 (United States); Roth, Adrian, E-mail: adrian_b.roth@roche.com [Non-Clinical Safety, Hoffmann-La Roche Ltd, Grenzacherstrasse 124, Building 73 / Room 117b, 4070 Basel (Switzerland)

    2013-04-01

    Drug-induced liver injury (DILI) is the major cause for liver failure and post-marketing drug withdrawals. Due to species-specific differences in hepatocellular function, animal experiments to assess potential liabilities of drug candidates can predict hepatotoxicity in humans only to a certain extent. In addition to animal experimentation, primary hepatocytes from rat or human are widely used for pre-clinical safety assessment. However, as many toxic responses in vivo are mediated by a complex interplay among different cell types and often require chronic drug exposures, the predictive performance of hepatocytes is very limited. Here, we established and characterized human and rat in vitro three-dimensional (3D) liver co-culture systems containing primary parenchymal and non-parenchymal hepatic cells. Our data demonstrate that cells cultured on a 3D scaffold have a preserved composition of hepatocytes, stellate, Kupffer and endothelial cells and maintain liver function for up to 3 months, as measured by the production of albumin, fibrinogen, transferrin and urea. Additionally, 3D liver co-cultures maintain cytochrome P450 inducibility, form bile canaliculi-like structures and respond to inflammatory stimuli. Upon incubation with selected hepatotoxicants including drugs which have been shown to induce idiosyncratic toxicity, we demonstrated that this model better detected in vivo drug-induced toxicity, including species-specific drug effects, when compared to monolayer hepatocyte cultures. In conclusion, our results underline the importance of more complex and long lasting in vitro cell culture models that contain all liver cell types and allow repeated drug-treatments for detection of in vivo-relevant adverse drug effects. - Highlights: ► 3D liver co-cultures maintain liver specific functions for up to three months. ► Activities of Cytochrome P450s remain drug- inducible accross three months. ► 3D liver co-cultures recapitulate drug-induced liver toxicity

  12. Biomaterials in co-culture systems: towards optimizing tissue integration and cell signaling within scaffolds.

    Science.gov (United States)

    Battiston, Kyle G; Cheung, Jane W C; Jain, Devika; Santerre, J Paul

    2014-05-01

    Most natural tissues consist of multi-cellular systems made up of two or more cell types. However, some of these tissues may not regenerate themselves following tissue injury or disease without some form of intervention, such as from the use of tissue engineered constructs. Recent studies have increasingly used co-cultures in tissue engineering applications as these systems better model the natural tissues, both physically and biologically. This review aims to identify the challenges of using co-culture systems and to highlight different approaches with respect to the use of biomaterials in the use of such systems. The application of co-culture systems to stimulate a desired biological response and examples of studies within particular tissue engineering disciplines are summarized. A description of different analytical co-culture systems is also discussed and the role of biomaterials in the future of co-culture research are elaborated on. Understanding the complex cell-cell and cell-biomaterial interactions involved in co-culture systems will ultimately lead the field towards biomaterial concepts and designs with specific biochemical, electrical, and mechanical characteristics that are tailored towards the needs of distinct co-culture systems. Copyright © 2014 Elsevier Ltd. All rights reserved.

  13. Precise micropatterning of silver nanoparticles on plastic substrates

    Energy Technology Data Exchange (ETDEWEB)

    Ammosova, Lena; Jiang, Yu; Suvanto, Mika; Pakkanen, Tapani A., E-mail: tapani.pakkanen@uef.fi

    2017-04-15

    Highlights: • Silver ink has been deposited on plastic substrate and silver nanoparticles have been produced. • 3D control allows both ink superimposing and deposition on complicated surfaces. • Polyol method ensures the formation of metallic mircopatterns with high uniformity. • Substrate wettability, ink volume, and sintering temperature influences deposited patterns. - Abstract: Conventional fabrication methods to obtain metal patterns on polymer substrates are restricted by high operating temperature and complex preparation steps. The present study demonstrates a simple yet versatile method for preparation of silver nanoparticle micropatterns on polymer substrates with various surface geometry. With the microworking robot technique, we were able not only to directly structure the surface, but also precisely deposit silver nanoparticle ink on the desired surface location with the minimum usage of ink material. The prepared silver nanoparticle ink, containing silver cations and polyethylene glycol (PEG) as a reducing agent, yields silver nanoparticle micropatterns on plastic substrates at low sintering temperature without any contamination. The influence of the ink behaviour was studied, such as substrate wettability, ink volume, and sintering temperature. The ultraviolet visible (UV–vis), scanning electron microscopy (SEM), and energy dispersive spectroscopy (EDS) measurements revealed the formation of micropatterns with uniformly distributed silver nanoparticles. The prepared patterns are expected to have a broad range of applications in optics, medicine, and sensor devices owing to the unique properties of silver. Furthermore, the deposition of a chemical compound, which is different from the substrate material, not only adds a fourth dimension to the prestructured three-dimensional (3D) surfaces, but also opens new application areas to the conventional surface structures.

  14. Precise micropatterning of silver nanoparticles on plastic substrates

    International Nuclear Information System (INIS)

    Ammosova, Lena; Jiang, Yu; Suvanto, Mika; Pakkanen, Tapani A.

    2017-01-01

    Highlights: • Silver ink has been deposited on plastic substrate and silver nanoparticles have been produced. • 3D control allows both ink superimposing and deposition on complicated surfaces. • Polyol method ensures the formation of metallic mircopatterns with high uniformity. • Substrate wettability, ink volume, and sintering temperature influences deposited patterns. - Abstract: Conventional fabrication methods to obtain metal patterns on polymer substrates are restricted by high operating temperature and complex preparation steps. The present study demonstrates a simple yet versatile method for preparation of silver nanoparticle micropatterns on polymer substrates with various surface geometry. With the microworking robot technique, we were able not only to directly structure the surface, but also precisely deposit silver nanoparticle ink on the desired surface location with the minimum usage of ink material. The prepared silver nanoparticle ink, containing silver cations and polyethylene glycol (PEG) as a reducing agent, yields silver nanoparticle micropatterns on plastic substrates at low sintering temperature without any contamination. The influence of the ink behaviour was studied, such as substrate wettability, ink volume, and sintering temperature. The ultraviolet visible (UV–vis), scanning electron microscopy (SEM), and energy dispersive spectroscopy (EDS) measurements revealed the formation of micropatterns with uniformly distributed silver nanoparticles. The prepared patterns are expected to have a broad range of applications in optics, medicine, and sensor devices owing to the unique properties of silver. Furthermore, the deposition of a chemical compound, which is different from the substrate material, not only adds a fourth dimension to the prestructured three-dimensional (3D) surfaces, but also opens new application areas to the conventional surface structures.

  15. Innovative applications and developments of micro-pattern gaseous detectors

    CERN Document Server

    Francke, Tom

    2014-01-01

    Study of nature and the world around us has been a primary motivation for scientists and researchers for centuries. Advanced methods in the study of elementary particles have led to even greater discoveries in recent years. "Innovative Applications and Developments of Micro-Pattern Gaseous Detectors" focuses on the analysis and use of various gas detection systems, providing a detailed description of some of the most commonly used gas detectors and the science behind them. From early detectors to modern tools and techniques, this book will be of particular use to practitioners and researchers in chemical engineering and materials science, in addition to students and academicians concentrating in the field.

  16. Semiconductor micropattern pixel detectors a review of the beginnings

    CERN Document Server

    Heijne, Erik H M

    2001-01-01

    The innovation in monolithic and hybrid semiconductor 'micropattern' or 'reactive' pixel detectors for tracking in particle physics was actually to fit logic and pulse processing electronics with µW power on a pixel area of less than 0.04 mm2, retaining the characteristics of a traditional nuclear amplifier chain. The ns timing precision in conjunction with local memory and logic operations allowed event selection at > 10 MHz rates with unambiguous track reconstruction even at particle multiplicities > 10 cm-2. The noise in a channel was ~100 e- r.m.s. and enabled binary operation with random noise 'hits' at a level 30 Mrad, respectively.

  17. Type conversion of secretomes in a 3D TAM2 and HCC cell co-culture system and functional importance of CXCL2 in HCC.

    Science.gov (United States)

    Lu, Yu; Li, Shan; Ma, Liping; Li, Yan; Zhang, Xiaolian; Peng, Qiliu; Mo, Cuiju; Huang, Li; Qin, Xue; Liu, Yinkun

    2016-04-27

    Macrophages play important roles in the tumor microenvironment, driving cancer progression and metastasis, particularly in hepatocellular carcinoma (HCC). However, few studies have assessed the exact secretome composition in HCC. In the present study, the impact of different phenotype of macrophages on HCC cells was investigated. Alternatively activated macrophages (M2) were found to significantly increase the proliferation, migration, and invasion abilities of SMMC7721 cells (all P cultured with SMMC7721 cells to reconstruct the tumor microenvironment. Conditioned medium from 3D single cultures of M2, SMMC7721 cells, and their co-culture system were analyzed using quantitative proteomics via iTRAQ labeling combined with mass spectrometric analysis. Secretome analysis revealed a total of 159 differential secreted proteins in the co-culture system compared to the single culture systems, with 63 being up-regulated (>1.3-fold) and 96 down-regulated (culture system and HCC tissues, and was selected for further investigation. Functional effects data suggested that recombinant human CXCL2 significantly enhanced the migration, invasion ability of SMMC7721 cells, and weakened adhesion ability. While CXCL2 neutralization and CXCR2 blockage significantly inhibited the effects of CXCL2 on SMMC7721 cells, indicating that CXCL2 may play pivotal role in HCC metastasis.

  18. Expression of HSP27, HSP72 and MRP proteins in in vitro co-culture ...

    Indian Academy of Sciences (India)

    Prakash

    2009-12-09

    Dec 9, 2009 ... in co-cultures of human colon carcinoma cell spheroids obtained from different grades of ...... of adherens junction in prostate epithelial cells; Exp. Cell. Res. ... tumour-associated antigens is related to estrogen receptor status;.

  19. A yeast co-culture-based biosensor for determination of waste water contamination levels.

    Science.gov (United States)

    Yudina, N Yu; Arlyapov, V A; Chepurnova, M A; Alferov, S V; Reshetilov, A N

    2015-10-01

    Artificial microbial co-cultures were formed to develop the receptor element of a biosensor for assessment of biological oxygen demand (BOD). The co-cultures possessed broad substrate specificities and enabled assays of water and fermentation products within a broad BOD range (2.4-80 mg/dm(3)) with a high correlation to the standard method (R = 0.9988). The use of the co-cultures of the yeasts Pichia angusta, Arxula adeninivorans and Debaryomyces hansenii immobilized in N-vinylpyrrolidone-modified poly(vinyl alcohol) enabled developing a BOD biosensor possessing the characteristics not inferior to those in the known biosensors. The results are indicative of a potential of using these co-cultures as the receptor element base in prototype models of instruments for broad application. Copyright © 2015 Elsevier Inc. All rights reserved.

  20. Neural differentiation of adipose-derived stem cells by indirect co-culture with Schwann cells

    Directory of Open Access Journals (Sweden)

    Li Xiaojie

    2009-01-01

    Full Text Available To investigate whether adipose-derived stem cells (ADSCs could be subject to neural differentiation induced only by Schwann cell (SC factors, we co-cultured ADSCs and SCs in transwell culture dishes. Immunoassaying, Western blot analysis, and RT-PCR were performed (1, 3, 7, 14 d and the co-cultured ADSCs showed gene and protein expression of S-100, Nestin, and GFAP. Further, qRT-PCR disclosed relative quantitative differences in the above three gene expressions. We think ADSCs can undergo induced neural differentiation by being co-cultured with SCs, and such differentia­tions begin 1 day after co-culture, become apparent after 7 days, and thereafter remain stable till the 14th day.

  1. Self-organization of human embryonic stem cells on micropatterns

    Science.gov (United States)

    Deglincerti, Alessia; Etoc, Fred; Guerra, M. Cecilia; Martyn, Iain; Metzger, Jakob; Ruzo, Albert; Simunovic, Mijo; Yoney, Anna; Brivanlou, Ali H.; Siggia, Eric; Warmflash, Aryeh

    2018-01-01

    Fate allocation in the gastrulating embryo is spatially organized as cells differentiate to specialized cell types depending on their positions with respect to the body axes. There is a need for in vitro protocols that allow the study of spatial organization associated with this developmental transition. While embryoid bodies and organoids can exhibit some spatial organization of differentiated cells, these methods do not yield consistent and fully reproducible results. Here, we describe a micropatterning approach where human embryonic stem cells are confined to disk-shaped, sub-millimeter colonies. After 42 hours of BMP4 stimulation, cells form self-organized differentiation patterns in concentric radial domains, which express specific markers associated with the embryonic germ layers, reminiscent of gastrulating embryos. Our protocol takes 3 days; it uses commercial microfabricated slides (CYTOO), human laminin-521 (LN-521) as extra-cellular matrix coating, and either conditioned or chemically-defined medium (mTeSR). Differentiation patterns within individual colonies can be determined by immunofluorescence and analyzed with cellular resolution. Both the size of the micropattern and the type of medium affect the patterning outcome. The protocol is appropriate for personnel with basic stem cell culture training. This protocol describes a robust platform for quantitative analysis of the mechanisms associated with pattern formation at the onset of gastrulation. PMID:27735934

  2. Decolorization of synthetic brilliant green carpet industry dye through fungal co-culture technology.

    Science.gov (United States)

    Kumari, Simpal; Naraian, Ram

    2016-09-15

    Aim of the present study was to evaluate the efficiency of fungal co-culture for the decolorization of synthetic brilliant green carpet industry dye. For this purpose two lignocellulolytic fungi Pleurotus florida (PF) and Rhizoctonia solani (RS) were employed. The study includes determination of enzyme profiles (laccase and peroxidase), dye decolorization efficiency of co-culture and crude enzyme extracts. Both fungi produced laccase and Mn peroxidase and successfully decolorized solutions of different concentrations (2.0, 4.0, 6.0, & 8.0(w/v) of dye. The co-culture resulted highest 98.54% dye decolorization at 2% (w/v) of dye as compared to monocultures (82.12% with PF and 68.89% with RS) during 12 days of submerged fermentation. The lower levels of dyes were rapidly decolorized, while higher levels in slow order as 87.67% decolorization of 8% dye. The promising achievement of the study was remarkable decolorizing efficiency of co-culture over monocultures. The direct treatment of the mono and co-culture enzyme extracts to dye also influenced remarkable. The highest enzymatic decolorization was through combined (PF and RS) extracts, while lesser by monoculture extracts. Based on the observations and potentiality of co-culture technology; further it can be exploited for the bioremediation of areas contaminated with hazardous environmental pollutants including textile and other industry effluents. Copyright © 2016 Elsevier Ltd. All rights reserved.

  3. Detection limits of Legionella pneumophila in environmental samples after co-culture with Acanthamoeba polyphaga

    Science.gov (United States)

    2013-01-01

    Background The efficiency of recovery and the detection limit of Legionella after co-culture with Acanthamoeba polyphaga are not known and so far no investigations have been carried out to determine the efficiency of the recovery of Legionella spp. by co-culture and compare it with that of conventional culturing methods. This study aimed to assess the detection limits of co-culture compared to culture for Legionella pneumophila in compost and air samples. Compost and air samples were spiked with known concentrations of L. pneumophila. Direct culturing and co-culture with amoebae were used in parallel to isolate L. pneumophila and recovery standard curves for both methods were produced for each sample. Results The co-culture proved to be more sensitive than the reference method, detecting 102-103 L. pneumophila cells in 1 g of spiked compost or 1 m3 of spiked air, as compared to 105-106 cells in 1 g of spiked compost and 1 m3 of spiked air. Conclusions Co-culture with amoebae is a useful, sensitive and reliable technique to enrich L. pneumophila in environmental samples that contain only low amounts of bacterial cells. PMID:23442526

  4. Advances in tissue engineering through stem cell-based co-culture.

    Science.gov (United States)

    Paschos, Nikolaos K; Brown, Wendy E; Eswaramoorthy, Rajalakshmanan; Hu, Jerry C; Athanasiou, Kyriacos A

    2015-05-01

    Stem cells are the future in tissue engineering and regeneration. In a co-culture, stem cells not only provide a target cell source with multipotent differentiation capacity, but can also act as assisting cells that promote tissue homeostasis, metabolism, growth and repair. Their incorporation into co-culture systems seems to be important in the creation of complex tissues or organs. In this review, critical aspects of stem cell use in co-culture systems are discussed. Direct and indirect co-culture methodologies used in tissue engineering are described, along with various characteristics of cellular interactions in these systems. Direct cell-cell contact, cell-extracellular matrix interaction and signalling via soluble factors are presented. The advantages of stem cell co-culture strategies and their applications in tissue engineering and regenerative medicine are portrayed through specific examples for several tissues, including orthopaedic soft tissues, bone, heart, vasculature, lung, kidney, liver and nerve. A concise review of the progress and the lessons learned are provided, with a focus on recent developments and their implications. It is hoped that knowledge developed from one tissue can be translated to other tissues. Finally, we address challenges in tissue engineering and regenerative medicine that can potentially be overcome via employing strategies for stem cell co-culture use. Copyright © 2014 John Wiley & Sons, Ltd.

  5. Testicular Sertoli cells influence the proliferation and immunogenicity of co-cultured endothelial cells

    International Nuclear Information System (INIS)

    Fan, Ping; He, Lan; Pu, Dan; Lv, Xiaohong; Zhou, Wenxu; Sun, Yining; Hu, Nan

    2011-01-01

    Research highlights: → The proliferation of dramatic increased by co-cultured with Sertoli cells. → VEGF receptor-2 expression of ECs was up-regulated by co-cultured with Sertoli cells. → The MHC expression of ECs induced by INF-γ and IL-6, IL-8 and sICAM induced by TNF-α decreased respectively after co-cultured with Sertoli cells. → ECs co-cultured with Sertoli cells also didn't increase the stimulation index of spleen lymphocytes. -- Abstract: The major problem of the application of endothelial cells (ECs) in transplantation is the lack of proliferation and their immunogenicity. In this study, we co-cultured ECs with Sertoli cells to monitor whether Sertoli cells can influence the proliferation and immunogenicity of co-cultured ECs. Sertoli cells were isolated from adult testicular tissue. ECs were divided into the control group and the experimental group, which included three sub-groups co-cultured with 1 x 10 3 , 1 x 10 4 or 1 x 10 5 cell/ml of Sertoli cells. The growth and proliferation of ECs were observed microscopically, and the expression of vascular endothelial growth factor (VEGF) receptor-2 (KDR) was examined by Western blotting. In another experiment, ECs were divided into the control group, the single culture group and the co-culture group with the optimal concentration of Sertoli cells. After INF-γ and TNF-α were added to the culture medium, MHC II antigen expression was detected by immunofluorescence staining and western blotting; interleukin (IL)-6, IL-8 and soluble intercellular adhesion molecule (sICAM) were measured in the culture medium by ELISA. We demonstrated that 1 x 10 4 cell/ml Sertoli cells promoted the proliferation of co-cultured ECs more dramatically than that in other groups (P 4 cell/ml of the Sertoli cells was most effective in the up-regulation of KDR expression in the co-cultured ECs (P < 0.05). Sertoli cells can effectively suppress INF-γ-induced MHC II antigen expression in co-cultured ECs compared with single

  6. Formation of controllable polymer micropatterns through liquid film electro-dewetting

    Science.gov (United States)

    Zhou, Shangru; Zheng, Huai; Li, Guoliang; Liu, Jie; Liu, Sheng

    2018-04-01

    Controllable polymer micropatterns, served as indispensable function structures, are extensively required in many micro/nano scientific areas and engineering applications. Exploring advanced methods of fabricating micropatterns is always a research hotspot. In this article, we introduce a novel method of patterning polymer by the electro-dewetting induced by corona discharge. For the first time, it is observed experimentally that liquid polymer on conductive/non-conductive patterned substrates, spontaneously converges from non-conductive areas to conductive areas under the action of ion wind. Taking advantage of such a flow phenomenon, controllable polymer micropatterns including microbump arrays and microwell arrays are fabricated successfully. Their sizes range from hundreds of microns to millimeters. Micropattern surfaces present an ultra-smooth characteristic, with roughness in the nanometer range.

  7. Radiosensitivity of hepatocellular carcinoma

    International Nuclear Information System (INIS)

    Hennequin, C.; Quero, L.; Rivera, S.

    2011-01-01

    The frequency of hepatocellular carcinoma (HCC) is increasing in the western world and the role of radiotherapy is more and more discussed. Classically, hepatocellular carcinoma was considered as a radioresistant tumour: in fact, modern radio-biologic studies, performed on cell lines directly established from patients, showed that hepatocellular carcinoma has the same radiosensitivity than the other epithelial tumours. From clinical studies, its α/β ratio has been estimated to be around 15 Gy. Radiosensitivity of normal hepatic parenchyma is now well evaluated and some accurate NTCP models are available to guide hepatic irradiation. The biology of hepatocellular carcinoma is also better described: the combination of radiotherapy and targeted therapies will be a promising approach in the near future. (authors)

  8. Cryotherapy for hepatocellular carcinoma

    DEFF Research Database (Denmark)

    Awad, Tahany; Thorlund, Kristian; Gluud, Christian

    2009-01-01

    BACKGROUND: Hepatocellular carcinoma is the most common primary malignant cancer of the liver. Evidence for the role of cryotherapy in the treatment of hepatocellular carcinoma is controversial. OBJECTIVES: The aim of this review is to evaluate the potential benefits and harms of cryotherapy...... for the treatment of hepatocellular carcinoma. SEARCH STRATEGY: We searched The Cochrane Hepato-Biliary Group Controlled Trials Register, the Cochrane Central Register of Controlled Trials (CENTRAL) in The Cochrane Library, MEDLINE, EMBASE, and LILACS until June 2009. We identified further studies by searching...... of benefit but included for the assessment of harm. Both severe and non-severe adverse events were reported, but the true nature and extent of harm was difficult to asses. AUTHORS' CONCLUSIONS: At present, there is no evidence to recommend or refute cryotherapy for patients with hepatocellular carcinoma...

  9. Preparation and surface characterization of plasma-treated and biomolecular-micropatterned polymer substrates

    Science.gov (United States)

    Langowski, Bryan Alfred

    A micropatterning process creates distinct microscale domains on substrate surfaces that differ from the surfaces' original chemical/physical properties. Numerous micropatterning methods exist, each having relative advantages and disadvantages in terms of cost, ease, reproducibility, and versatility. Polymeric surfaces micropatterned with biomolecules have many applications, but are specifically utilized in tissue engineering as cell scaffolds that attempt to controlled tissue generation in vivo and ex vivo. As the physical and chemical cues presented by micropatterned substrates control resulting cellular behavior, characterization of these cues via surface-sensitive analytical techniques is essential in developing cell scaffolds that mimic complex in vivo physicochemical environments. The initial focus of this thesis is the chemical and physical characterization of plasma-treated, microcontact-printed (muCP) polymeric substrates used to direct nerve cell behavior. Unmodified and oxygen plasma-treated poly(methyl methacrylate) (PMMA) substrates were analyzed by surface sensitive techniques to monitor plasma-induced chemical and physical modifications. Additionally, protein-micropattern homogeneity and size were microscopically evaluated. Lastly, poly(dimethylsiloxane) (PDMS) stamps and contaminated PMMA substrates were characterized by spectroscopic and microscopic methods to identify a contamination source during microcontact printing. The final focus of this thesis is the development of microscale plasma-initiated patterning (muPIP) as a versatile, reproducible micropatterning method. Using muPIP, polymeric substrates were micropatterned with several biologically relevant inks. Polymeric substrates were characterized following muPIP by surface-sensitive techniques to identify the technique's underlying physical and chemical bases. In addition, neural stem cell response to muPIP-generated laminin micropatterns was microscopically and biologically evaluated

  10. Methods of Micropatterning and Manipulation of Cells for Biomedical Applications

    Directory of Open Access Journals (Sweden)

    Adrian Martinez-Rivas

    2017-11-01

    Full Text Available Micropatterning and manipulation of mammalian and bacterial cells are important in biomedical studies to perform in vitro assays and to evaluate biochemical processes accurately, establishing the basis for implementing biomedical microelectromechanical systems (bioMEMS, point-of-care (POC devices, or organs-on-chips (OOC, which impact on neurological, oncological, dermatologic, or tissue engineering issues as part of personalized medicine. Cell patterning represents a crucial step in fundamental and applied biological studies in vitro, hence today there are a myriad of materials and techniques that allow one to immobilize and manipulate cells, imitating the 3D in vivo milieu. This review focuses on current physical cell patterning, plus chemical and a combination of them both that utilizes different materials and cutting-edge micro-nanofabrication methodologies.

  11. GEMs with Double Layred Micropattern Electrodes and their Applications

    CERN Document Server

    Di Mauro, A.; Nappi, E.; Oliveira, R.; Peskov, V.; Pietropaolo, F.; Picchi, P.

    We have developed and tested several new designs of GEM detectors with micropattern electrodes manufactured by microelectronic technology. In one design, the inner layer of the detector electrode consists of thin metallic strips and the outer layer is made of a resistive grid manufactured by a screen printing technology. In other designs, the electrodes were made of metallic strips fed by HV via micro-resistors manufactured by a screen printing technology. Due to these features, the new detectors have several important advantages over conventional GEMs or ordinary thick GEMs. For example, the resistive grid (in the first design) and the screen printed resistors (in other designs) limited the current in case of discharges, making these detectors intrinsically spark-protected. We will here describe our tests with the photosensitive versions of these detectors (coated with CsI layers) and the efforts of implementing them in several applications. In particular, we will focus on our activity towards the ALICE RICH...

  12. A novel fast timing micropattern gaseous detector: FTM

    CERN Document Server

    De Oliveira, Rui; Sharma, Archana

    2015-01-01

    In recent years important progress in micropattern gaseous detectors has been achieved in the use of resistive material to build compact spark-protected devices. The novel idea presented here consists of the polarisation of WELL structures using only resistive coating. This allows a new device to be built with an architecture based on a stack of several coupled layers where drift and WELL multiplication stages alternate in the structure. The signals from each multiplication stage can be read out from any external readout boards through the capacitive couplings. Each layer provides a signal with a gain of 10^4-10^5. The main advantage of this new device is the dramatic improvement of the timing provided by the competition of the ionisation processes in the different drift regions, which can be exploited for fast timing at the high luminosity accelerators (e.g. HL-LHC upgrade) as well as far applications like medical imaging.

  13. A novel fast timing micropattern gaseous detector: FTM

    CERN Document Server

    De Oliveira, Rui; Maggi, Marcello

    2015-01-01

    In recent years important progress in micropattern gaseous detectors has been achieved in the use of resistive material to build compact spark-protected devices. The novel idea presented here consists of the polarisation of WELL structures using only resistive electrodes. This allows a new device to be built with an architecture based on a stack of several coupled layers where drift and WELL multiplication stages alternate in the structure. The signals from each multiplication stage can be read out from any external readout boards through the capacitive couplings. Each layer provides a signal with a gain of 10^4 - 10^5. The main advantage of this new device is the dramatic improvement of the timing provided by the competition of the ionisation processes in the different drift regions, which can be exploited for fast timing at the high luminosity accelerators (e.g. HL-LHC upgrade) as well as applications outside particle physics.

  14. A co-cultured skin model based on cell support membranes

    International Nuclear Information System (INIS)

    Dai, N.-T.; Yeh, M.-K.; Liu, Demeral David; Adams, E.F.; Chiang, C.-H.; Yen, C.-Y.; Shih, C.-M.; Sytwu, H.-K.; Chen, Tim-Mo; Wang, H.-J.; Williamson, M.R.; Coombes, A.G.A.

    2005-01-01

    Tissue engineering of skin based on collagen: PCL biocomposites using a designed co-culture system is reported. The collagen: PCL biocomposites having collagen: PCL (w/w) ratios of 1:4, 1:8, and 1:20 have been proven to be biocompatible materials to support both adult normal human epidermal Keratinocyte (NHEK) and mouse 3T3 fibroblast growth in cell culture, respectively, by Dai, Coombes, et al. in 2004. Films of collagen: PCL biocomposites were prepared using non-crosslinking method by impregnation of lyophilized collagen mats with PCL/dichloromethane solutions followed by solvent evaporation. To mimic the dermal/epidermal structure of skin, the 1:20 collagen: PCL biocomposites were selected for a feasibility study of a designed co-culture technique that would subsequently be used for preparing fibroblast/biocomposite/keratinocyte skin models. A 55.3% increase in cell number was measured in the designed co-culture system when fibroblasts were seeded on both sides of a biocomposite film compared with cell culture on one surface of the biocomposite in the feasibility study. The co-culture of human keratinocytes and 3T3 fibroblasts on each side of the membrane was therefore studied using the same co-culture system by growing keratinocytes on the top surface of membrane for 3 days and 3T3 fibroblasts underneath the membrane for 6 days. Scanning electron microscopy (SEM) and immunohistochemistry assay revealed good cell attachment and proliferation of both human keratinocytes and 3T3 fibroblasts with these two types of cells isolated well on each side of the membrane. Using a modified co-culture technique, a co-cultured skin model presenting a confluent epidermal sheet on one side of the biocomposite film and fibroblasts populated on the other side of the film was developed successfully in co-culture system for 28 days under investigations by SEM and immunohistochemistry assay. Thus, the design of a co-culture system based on 1:20 (w/w) collagen: PCL biocomposite

  15. [Biological characteristics of mesenchymal stem cell and hematopoietic stem cell in the co-culture system].

    Science.gov (United States)

    Wei, Wei; Xu, Chao; Ye, Zhi-Yong; Huang, Xiao-Jun; Yuan, Jia-En; Ma, Tian-Bao; Lin, Han-Biao; Chen, Xiu-Qiong

    2016-10-25

    The aim of the present study was to obtain the qualified hematopoietic stem/progenitor cells (HSC/HPC) and human umbilical cord-mesenchymal stem cells (MSC) in vitro in the co-culture system. Cord blood mononuclear cells were separated from umbilical cord blood by Ficoll lymphocyte separation medium, and then CD34 + HSC was collected by MACS immunomagnetic beads. The selected CD34 + HSC/HPC and MSC were transferred into culture flask. IMDM culture medium with 15% AB-type cord plasma supplemented with interleukin-3 (IL-3), IL-6, thrombopoietin (TPO), stem cell factor (SCF) and FMS-like tyrosine kinase 3 ligand (Flt-3L) factors were used as the co-culture system for the amplification of HSC/HPC and MSC. The cellular growth status and proliferation on day 6 and 10 after co-culture were observed by using inverted microscope. The percentage of positive expression of CD34 in HSC/HPC, as well as the percentages of positive expressions of CD105, CD90, CD73, CD45, CD34 and HLA-DR in the 4 th generation MSC, was tested by flow cytometry. Semisolid colony culture was used to test the HSC/HPC colony forming ability. The osteogenic, chondrogenesis and adipogenic ability of the 4 th generation MSC were assessed. The karyotype analysis of MSC was conducted by colchicines. The results demonstrated that the HSC/HPC of co-culture group showed higher ability of amplification, CFU-GM and higher CD34 + percentage compared with the control group. The co-cultured MSC maintained the ability to differentiate into bone cells, fat cells and chondrocytes. And the karyotype stability of MSC remained normal. These results reveal that the appropriate co-culture system for MSC and HSC is developed, and via this co-culture system we could gain both two kinds of these cells. The MSCs under the co-culture system maintain the biological characteristics. The CFU-GM ability, cell counting and the flow cytometry results of HSC/HPC under the co-culture system are conform to the criterion, showing that

  16. Garlic exerts allelopathic effects on pepper physiology in a hydroponic co-culture system

    Directory of Open Access Journals (Sweden)

    Haiyan Ding

    2016-05-01

    Full Text Available A hydroponic co-culture system was adopted to determine the allelopathic potential of garlic on the growth of pepper plants. Different numbers of garlic plants (0, 2, 4, 8 and 12 were hydroponically co-cultured with two pepper plants to investigate allelopathic effects on the growth attributes and antioxidative defense system of the test pepper plants. The responses of the pepper plants depended on the number of garlic plants included in the co-culture system, indicating an association of pepper growth with the garlic root exudate concentration. When grown at a pepper/garlic ratio of 1:1 or 1:2, the pepper plant height, chlorophyll content, and peroxidase (POD, catalase (CAT and phenylalanine ammonia-lyase (PAL activities were significantly increased after 30 days of co-culture; in contrast, reduction in methane dicarboxylic aldehyde (MDA content was observed. However, when the pepper/garlic ratio was 1:4 or higher, these morphological indices and protective enzyme activities were significantly inhibited, whereas MDA levels in the pepper leaves were significantly increased due to severe membrane lipid peroxidation. The results indicate that although low concentrations of garlic root exudates appear to induce protective enzyme systems and promote pepper growth, high concentrations have deleterious effects. These findings suggest that further investigations should optimize the co-culture pepper/garlic ratio to reduce continuous cropping obstacles in pepper production.

  17. Co-culture systems and technologies: taking synthetic biology to the next level.

    Science.gov (United States)

    Goers, Lisa; Freemont, Paul; Polizzi, Karen M

    2014-07-06

    Co-culture techniques find myriad applications in biology for studying natural or synthetic interactions between cell populations. Such techniques are of great importance in synthetic biology, as multi-species cell consortia and other natural or synthetic ecology systems are widely seen to hold enormous potential for foundational research as well as novel industrial, medical and environmental applications with many proof-of-principle studies in recent years. What is needed for co-cultures to fulfil their potential? Cell-cell interactions in co-cultures are strongly influenced by the extracellular environment, which is determined by the experimental set-up, which therefore needs to be given careful consideration. An overview of existing experimental and theoretical co-culture set-ups in synthetic biology and adjacent fields is given here, and challenges and opportunities involved in such experiments are discussed. Greater focus on foundational technology developments for co-cultures is needed for many synthetic biology systems to realize their potential in both applications and answering biological questions. © 2014 The Author(s) Published by the Royal Society. All rights reserved.

  18. Modular co-culture engineering, a new approach for metabolic engineering.

    Science.gov (United States)

    Zhang, Haoran; Wang, Xiaonan

    2016-09-01

    With the development of metabolic engineering, employment of a selected microbial host for accommodation of a designed biosynthetic pathway to produce a target compound has achieved tremendous success in the past several decades. Yet, increasing requirements for sophisticated microbial biosynthesis call for establishment and application of more advanced metabolic engineering methodologies. Recently, important progress has been made towards employing more than one engineered microbial strains to constitute synthetic co-cultures and modularizing the biosynthetic labor between the co-culture members in order to improve bioproduction performance. This emerging approach, referred to as modular co-culture engineering in this review, presents a valuable opportunity for expanding the scope of the broad field of metabolic engineering. We highlight representative research accomplishments using this approach, especially those utilizing metabolic engineering tools for microbial co-culture manipulation. Key benefits and major challenges associated with modular co-culture engineering are also presented and discussed. Copyright © 2016 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  19. [Research progress of co-culture system for constructing vascularized tissue engineered bone].

    Science.gov (United States)

    Fu, Weili; Xiang, Zhou

    2014-02-01

    To review the research progress of the co-culture system for constructing vascularized tissue engineered bone. The recent literature concerning the co-culture system for constructing vascularized tissue engineered bone was reviewed, including the selection of osteogenic and endothelial lineages, the design and surface modification of scaffolds, the models and dimensions of the co-culture system, the mechanism, the culture conditions, and their application progress. The construction of vascularized tissue engineered bone is the prerequisite for their survival and further clinical application in vivo. Mesenchymal stem cells (owning the excellent osteogenic potential) and endothelial progenitor cells (capable of directional differentiation into endothelial cell) are considered as attractive cell types for the co-culture system to construct vascularized tissue engineered bone. The culture conditions need to be further optimized. Furthermore, how to achieve the clinical goals of minimal invasion and autologous transplantation also need to be further studied. The strategy of the co-culture system for constructing vascularized tissue engineered bone would have a very broad prospects for clinical application in future.

  20. Application of cell co-culture system to study fat and muscle cells.

    Science.gov (United States)

    Pandurangan, Muthuraman; Hwang, Inho

    2014-09-01

    Animal cell culture is a highly complex process, in which cells are grown under specific conditions. The growth and development of these cells is a highly unnatural process in vitro condition. Cells are removed from animal tissues and artificially cultured in various culture vessels. Vitamins, minerals, and serum growth factors are supplied to maintain cell viability. Obtaining result homogeneity of in vitro and in vivo experiments is rare, because their structure and function are different. Living tissues have highly ordered complex architecture and are three-dimensional (3D) in structure. The interaction between adjacent cell types is quite distinct from the in vitro cell culture, which is usually two-dimensional (2D). Co-culture systems are studied to analyze the interactions between the two different cell types. The muscle and fat co-culture system is useful in addressing several questions related to muscle modeling, muscle degeneration, apoptosis, and muscle regeneration. Co-culture of C2C12 and 3T3-L1 cells could be a useful diagnostic tool to understand the muscle and fat formation in animals. Even though, co-culture systems have certain limitations, they provide a more realistic 3D view and information than the individual cell culture system. It is suggested that co-culture systems are useful in evaluating the intercellular communication and composition of two different cell types.

  1. Use of bacterial co-cultures for the efficient production of chemicals.

    Science.gov (United States)

    Jones, J Andrew; Wang, Xin

    2017-12-02

    The microbial production of chemicals has traditionally relied on a single engineered microbe to enable the complete bioconversion of substrate to final product. Recently, a growing fraction of research has transitioned towards employing a modular co-culture engineering strategy using multiple microbes growing together to facilitate a divide-and-conquer approach for chemical biosynthesis. Here, we review key success stories that leverage the unique advantages of co-culture engineering, while also addressing the critical concerns that will limit the wide-spread implementation of this technology. Future studies that address the need to monitor and control the population dynamics of each strain module, while maintaining robust flux routes towards a wide range of desired products will lead the efforts to realize the true potential of co-culture engineering. Copyright © 2017 Elsevier Ltd. All rights reserved.

  2. Microbial transformation of tannin-rich substrate to gallic acid through co-culture method.

    Science.gov (United States)

    Banerjee, Rintu; Mukherjee, Gargi; Patra, Krushna Chandra

    2005-05-01

    Modified solid-state fermentation (MSSF) of tannin-rich substrate yielding tannase and gallic acid was carried out using a co-culture of the filamentous fungi, Rhizopus oryzae (RO IIT RB-13, NRRL 21498) and Aspergillus foetidus (GMRB013 MTCC 3557). Powdered fruits of Terminalia chebula and powdered pod cover of Caesalpinia digyna was used in the process and the different process parameters for maximum production of tannase and gallic acid by co-culture method were optimized through media engineering. MSSF was carried out at the optimum conditions of 30 degrees C and 80% relative humidity. The optimal pH and incubation period was 5.0 and 48 h respectively. Through the co-culture technique the maximum yield of tannase and gallic acid was found to be 41.3 U/ml and 94.8% respectively.

  3. Chondrogenesis of synovium-derived mesenchymal stem cells in gene-transferred co-culture system.

    Science.gov (United States)

    Varshney, Rohan R; Zhou, Ruijie; Hao, Jinghua; Yeo, Suan Siong; Chooi, Wai Hon; Fan, Jiabing; Wang, Dong-An

    2010-09-01

    A co-culture strategy has been developed in this study wherein rabbit synovial mesenchymal stem cells (SMSCs) are co-cultured with growth factor (GF) transfected articular chondrocytes. Toward this end, both SMSCs and early passage rabbit articular chondrocytes that had been adenovirally transduced with transforming growth factor-beta 3 (TGF-beta3) gene were separately encapsulated in alginate beads and co-cultured in the same pool of chondrogenic medium. The chondrocytes act as transfected companion cells (TCCs) providing GF supply to induce chondrogenic differentiation of SMSCs that play the role of therapeutic progenitor cells (TPCs). Against the same TCC based TGF-beta3 release profile, the co-culture was started at different time points (Day 0, Day 10 and Day 20) but made to last for identical periods of exposure (30 days) so that the exposure conditions could be optimized in terms of initiation and duration. Transfection of TCCs prevents the stem cell based TPCs from undergoing the invasive procedure. It also prevents unpredictable complications in the TPCs caused by long-term constitutive over-expression of a GF. The adenovirally transfected TCCs exhibit a transient GF expression which results in a timely termination of GF supply to the TPCs. The TCC-sourced transgenic TGF-beta3 successfully induced chondrogenesis in the TPCs. Real-time PCR results show enhanced expression of cartilage markers and immuno/histochemical staining for Glycosaminoglycans (GAG) and Collagen II also shows abundant extracellular matrix (ECM) production and chondrogenic morphogenesis in the co-cultured TPCs. These results confirm the efficacy of directing stem cell differentiation towards chondrogenesis and cartilage tissue formation by co-culturing them with GF transfected chondrocytes.

  4. Three-dimensional co-culture facilitates the differentiation of embryonic stem cells into mature cardiomyocytes.

    Science.gov (United States)

    Ou, Dong-Bo; He, Yong; Chen, Rui; Teng, Ji-Wei; Wang, Hong-Tao; Zeng, Di; Liu, Xiong-Tao; Ding, Lu; Huang, Jin-Yan; Zheng, Qiang-Sun

    2011-12-01

    The cardiomyocyte (CM) differentiation of embryonic stem cells (ESCs) is routinely cultured as two-dimensional (2D) monolayer, which doesn't mimic in vivo physiological environment and may lead to low differentiated level of ESCs. Here, we develop a novel strategy that enhances CM differentiation of ESCs in collagen matrix three-dimensional (3D) culture combined with indirect cardiac fibroblasts co-culture. ESCs were cultured in hanging drops to form embryoid bodies (EBs) and then applied on collagen matrix. The EBs were indirectly co-cultured with cardiac fibroblasts by the hanging cell culture inserts (PET 1 µm). The molecular expressions and ultrastructural characteristics of ESC-derived CMs (ESCMs) were analyzed by real time RT-PCR, immunocytochemistry, and Transmission Electron Microscopy (TEM). We found that the percentage of beating EBs with cardiac fibroblasts co-culture was significantly higher than that without co-culture after differentiation period of 8 days. Type I collagen used as 3D substrates enhanced the late-stage CM differentiation of ESCs and had effect on ultrastructural mature of ESCMs in late-stage development. The combined effects of 3D and co-culture that mimic in vivo physiological environment further improved the efficiency of CM differentiation from ESCs, resulting in fiber-like structures of cardiac cells with organized sarcomeric structure in ESCMs. This novel 3D co-culture system emphasizes the fact that the ESC differentiation is actively responding to cues from their environment and those cues can drive phenotypic control, which provides a useful in vitro model to investigate CM differentiation of stem cells. Copyright © 2011 Wiley Periodicals, Inc.

  5. Developing a mesophilic co-culture for direct conversion of cellulose to butanol in consolidated bioprocess.

    Science.gov (United States)

    Wang, Zhenyu; Cao, Guangli; Zheng, Ju; Fu, Defeng; Song, Jinzhu; Zhang, Junzheng; Zhao, Lei; Yang, Qian

    2015-01-01

    Consolidated bioprocessing (CBP) of butanol production from cellulosic biomass is a promising strategy for cost saving compared to other processes featuring dedicated cellulase production. CBP requires microbial strains capable of hydrolyzing biomass with enzymes produced on its own with high rate and high conversion and simultaneously produce a desired product at high yield. However, current reported butanol-producing candidates are unable to utilize cellulose as a sole carbon source and energy source. Consequently, developing a co-culture system using different microorganisms by taking advantage of their specific metabolic capacities to produce butanol directly from cellulose in consolidated bioprocess is of great interest. This study was mainly undertaken to find complementary organisms to the butanol producer that allow simultaneous saccharification and fermentation of cellulose to butanol in their co-culture under mesophilic condition. Accordingly, a highly efficient and stable consortium N3 on cellulose degradation was first developed by multiple subcultures. Subsequently, the functional microorganisms with 16S rRNA sequences identical to the denaturing gradient gel electrophoresis (DGGE) profile were isolated from consortium N3. The isolate Clostridium celevecrescens N3-2 exhibited higher cellulose-degrading capability was thus chosen as the partner strain for butanol production with Clostridium acetobutylicum ATCC824. Meanwhile, the established stable consortium N3 was also investigated to produce butanol by co-culturing with C. acetobutylicum ATCC824. Butanol was produced from cellulose when C. acetobutylicum ATCC824 was co-cultured with either consortium N3 or C. celevecrescens N3-2. Co-culturing C. acetobutylicum ATCC824 with the stable consortium N3 resulted in a relatively higher butanol concentration, 3.73 g/L, and higher production yield, 0.145 g/g of glucose equivalent. The newly isolated microbial consortium N3 and strain C. celevecrescens N3

  6. Nanotopography enhanced mobility determines mesenchymal stem cell distribution on micropatterned semiconductors bearing nanorough areas.

    Science.gov (United States)

    Gallach Pérez, Darío; Punzón Quijorna, Esther; Sanz, Ruy; Torres-Costa, Vicente; García Ruiz, Josefa P; Manso Silván, Miguel

    2015-02-01

    Surface micropatterns are relevant instruments for the in vitro analysis of cell cultures in non-conventional planar conditions. In this work, two semiconductors (Si and TiO2) have been micropatterned by combined ion-beam/chemical-etching processes leading to selective areas bearing nanorough features. A preferential affinity of human mesenchymal stem cells (hMSCs) for planar areas versus nanotopographic ones is observed. Fluorescence microscopy after β-catenin staining suggests that hMSCs adhesion is inhibited on nanostructured porous silicon areas. This has a direct impact in the development of actin fibers and suggests different cell migration mechanisms on the materials of a micropattern. hMSCs organization on nanotopographic micropatterns has been modeled by using a simplified random walk approach. The model attributes preferential cell mobilities on the nanotopographic areas with respect to the planar and considers purely stochastic movement with no inertial term. Simulations of the cell distribution have been run on 1D and 2D micropatterns and compared with the real hMSC cultures. The simulations allow defining two regimes for cell organization as a function of cell density. hMSCs ordering on planar areas is diffusion-induced in most micropatterns but constriction forced disorder appears for high cell densities. The relative mobility on the planar versus nanotopographic areas can be used as a quality indicator of the nanotopography contrasts in the diffusion induced ordering regime. It is shown that the relative mobility is favorable for the TiO2 versus the Si based system, and allows envisaging its use for the calibrated design of nanotopography based micropatterned materials. Copyright © 2014 Elsevier B.V. All rights reserved.

  7. Solvent-Assisted Gel Printing for Micropatterning Thin Organic-Inorganic Hybrid Perovskite Films.

    Science.gov (United States)

    Jeong, Beomjin; Hwang, Ihn; Cho, Sung Hwan; Kim, Eui Hyuk; Cha, Soonyoung; Lee, Jinseong; Kang, Han Sol; Cho, Suk Man; Choi, Hyunyong; Park, Cheolmin

    2016-09-27

    While tremendous efforts have been made for developing thin perovskite films suitable for a variety of potential photoelectric applications such as solar cells, field-effect transistors, and photodetectors, only a few works focus on the micropatterning of a perovskite film which is one of the most critical issues for large area and uniform microarrays of perovskite-based devices. Here we demonstrate a simple but robust method of micropatterning a thin perovskite film with controlled crystalline structure which guarantees to preserve its intrinsic photoelectric properties. A variety of micropatterns of a perovskite film are fabricated by either microimprinting or transfer-printing a thin spin-coated precursor film in soft-gel state with a topographically prepatterned elastomeric poly(dimethylsiloxane) (PDMS) mold, followed by thermal treatment for complete conversion of the precursor film to a perovskite one. The key materials development of our solvent-assisted gel printing is to prepare a thin precursor film with a high-boiling temperature solvent, dimethyl sulfoxide. The residual solvent in the precursor gel film makes the film moldable upon microprinting with a patterned PDMS mold, leading to various perovskite micropatterns in resolution of a few micrometers over a large area. Our nondestructive micropatterning process does not harm the intrinsic photoelectric properties of a perovskite film, which allows for realizing arrays of parallel-type photodetectors containing micropatterns of a perovskite film with reliable photoconduction performance. The facile transfer of a micropatterned soft-gel precursor film on other substrates including mechanically flexible plastics can further broaden its applications to flexible photoelectric systems.

  8. Micropatterned Silica Films with Nanohydroxyapatite for Y-TZP Implants.

    Science.gov (United States)

    Miranda, R B P; Grenho, L; Carvalho, A; Fernandes, M H; Monteiro, F J; Cesar, P F

    2018-03-01

    This investigation aimed at developing micropatterned silica thin films (MSTFs) containing nanohydroxyapatite (nano-HA) microaggregates that were not completely covered by silica so that they could directly interact with the surrounding cells. The objectives were 1) to evaluate the effect of the presence of 2 films (MSTF with or without nano-HA addition) on the characteristic strength (σ 0 ) and Weibull modulus ( m) of a yttria-stabilized tetragonal zirconia polycrystal (Y-TZP) and 2) to evaluate the effect of these 2 films, as applied onto the Y-TZP surface, on the morphology, orientation, and proliferation of MG63 cells. Sol-gel process and soft lithography were used to apply the MSTF onto the Y-TZP specimens. Three experimental groups were produced: Y-TZP, Y-TZP + MSTF, and Y-TZP + MSTF + sprayed nano-HA. All surfaces were characterized by scanning electron microscopy and energy-dispersive X-ray spectroscopy and tested for 4-point flexural strength ( n = 30) in water at 37 °C. Weibull analysis was used to determine m and σ 0 (maximum likelihood method). In vitro biological behavior was performed with human osteoblast-like cells (MG63). Y-TZP was successfully coated with MSFT and MSFT + nano-HA. Scanning electron microscopy micrographs indicated that the microaggregates of nano-HA were not entirely covered by the silica. There was no statistically significant difference among the experimental groups for σ 0 and m. In the groups containing the films, the cells were elongated and aligned along the lines. The MSFT + nano-HA group showed significantly higher cell metabolic activity than that obtained for the Y-TZP group at day 7. This investigation was successful in producing an MSTF containing nano-HA microaggregates that remained exposed to the environment. The developed films did not jeopardize the structural reliability of a commercial Y-TZP, as confirmed by the Weibull statistics. The MG63 cells seeded over the films became elongated and aligned along the

  9. Ethanol production from corn cobs by co-culture of Saccharomyces ...

    African Journals Online (AJOL)

    Saccharomyces cerevisiae and Aspergillus niger were used in a co-culture for the simultaneous saccharification and fermentation (SSF) of 1% and 10% (w/v) dry pre-treated corn cobs to ethanol. Positive controls of glucose of same concentrations in a synthetic medium were also fermented. At 1% substrate concentration, ...

  10. Boosting dark fermentation with co-cultures of extreme thermophiles for biohythane production from garden waste.

    Science.gov (United States)

    Abreu, Angela A; Tavares, Fábio; Alves, Maria Madalena; Pereira, Maria Alcina

    2016-11-01

    Proof of principle of biohythane and potential energy production from garden waste (GW) is demonstrated in this study in a two-step process coupling dark fermentation and anaerobic digestion. The synergistic effect of using co-cultures of extreme thermophiles to intensify biohydrogen dark fermentation is demonstrated using xylose, cellobiose and GW. Co-culture of Caldicellulosiruptor saccharolyticus and Thermotoga maritima showed higher hydrogen production yields from xylose (2.7±0.1molmol(-1) total sugar) and cellobiose (4.8±0.3molmol(-1) total sugar) compared to individual cultures. Co-culture of extreme thermophiles C. saccharolyticus and Caldicellulosiruptor bescii increased synergistically the hydrogen production yield from GW (98.3±6.9Lkg(-1) (VS)) compared to individual cultures and co-culture of T. maritima and C. saccharolyticus. The biochemical methane potential of the fermentation end-products was 322±10Lkg(-1) (CODt). Biohythane, a biogas enriched with 15% hydrogen could be obtained from GW, yielding a potential energy generation of 22.2MJkg(-1) (VS). Copyright © 2016 Elsevier Ltd. All rights reserved.

  11. Triple co-culture cell model as an in vitro model for oral particulate vaccine systems

    DEFF Research Database (Denmark)

    Nielsen, Line Hagner; De Rossi, C.; Lehr, C-M.

    ; this was not observed with ovalbumin and blank solution. An example of the results is shown in Figure 2 for IL-17A. An established co-culture of Caco-2, THP-1 and MUTZ-3 cells showed promise as an in vitro model for testing of oral vaccine formulations. Mobility of co-culture immune cells as well as cytokine production......A triple co-culture cell model of Caco-2 cells, dendritic cells and macrophages (Figure 1) has previously been developed for studying intestinal permeability in a state of inflammation [1],[2]. The aim of this study was to investigate the applicability of this cell model for testing...... the model antigen ovalbumin was spray dried to obtain a particulate vaccine model system for testing in the cell model. The precursors were shown to form cubosomes when dispersed in aqueous medium, and was therefore used as the vaccine formulation for testing on the co-cultures. After 11 days, the TEER...

  12. Engineering systems for the generation of patterned co-cultures for controlling cell-cell interactions.

    Science.gov (United States)

    Kaji, Hirokazu; Camci-Unal, Gulden; Langer, Robert; Khademhosseini, Ali

    2011-03-01

    Inside the body, cells lie in direct contact or in close proximity to other cell types in a tightly controlled architecture that often regulates the resulting tissue function. Therefore, tissue engineering constructs that aim to reproduce the architecture and the geometry of tissues will benefit from methods of controlling cell-cell interactions with microscale resolution. We discuss the use of microfabrication technologies for generating patterned co-cultures. In addition, we categorize patterned co-culture systems by cell type and discuss the implications of regulating cell-cell interactions in the resulting biological function of the tissues. Patterned co-cultures are a useful tool for fabricating tissue engineered constructs and for studying cell-cell interactions in vitro, because they can be used to control the degree of homotypic and heterotypic cell-cell contact. In addition, this approach can be manipulated to elucidate important factors involved in cell-matrix interactions. Patterned co-culture strategies hold significant potential to develop biomimetic structures for tissue engineering. It is expected that they would create opportunities to develop artificial tissues in the future. This article is part of a Special Issue entitled Nanotechnologies - Emerging Applications in Biomedicine. 2010 Elsevier B.V. All rights reserved.

  13. Design of biomimetic cellular scaffolds for co-culture system and their application

    Science.gov (United States)

    Kook, Yun-Min; Jeong, Yoon; Lee, Kangwon; Koh, Won-Gun

    2017-01-01

    The extracellular matrix of most natural tissues comprises various types of cells, including fibroblasts, stem cells, and endothelial cells, which communicate with each other directly or indirectly to regulate matrix production and cell functionality. To engineer multicellular interactions in vitro, co-culture systems have achieved tremendous success achieving a more realistic microenvironment of in vivo metabolism than monoculture system in the past several decades. Recently, the fields of tissue engineering and regenerative medicine have primarily focused on three-dimensional co-culture systems using cellular scaffolds, because of their physical and biological relevance to the extracellular matrix of actual tissues. This review discusses several materials and methods to create co-culture systems, including hydrogels, electrospun fibers, microfluidic devices, and patterning for biomimetic co-culture system and their applications for specific tissue regeneration. Consequently, we believe that culture systems with appropriate physical and biochemical properties should be developed, and direct or indirect cell–cell interactions in the remodeled tissue must be considered to obtain an optimal tissue-specific microenvironment. PMID:29081966

  14. Co-culture systems-based strategies for articular cartilage tissue engineering.

    Science.gov (United States)

    Zhang, Yu; Guo, Weimin; Wang, Mingjie; Hao, Chunxiang; Lu, Liang; Gao, Shuang; Zhang, Xueliang; Li, Xu; Chen, Mingxue; Li, Penghao; Jiang, Peng; Lu, Shibi; Liu, Shuyun; Guo, Quanyi

    2018-03-01

    Cartilage engineering facilitates repair and regeneration of damaged cartilage using engineered tissue that restores the functional properties of the impaired joint. The seed cells used most frequently in tissue engineering, are chondrocytes and mesenchymal stem cells. Seed cells activity plays a key role in the regeneration of functional cartilage tissue. However, seed cells undergo undesirable changes after in vitro processing procedures, such as degeneration of cartilage cells and induced hypertrophy of mesenchymal stem cells, which hinder cartilage tissue engineering. Compared to monoculture, which does not mimic the in vivo cellular environment, co-culture technology provides a more realistic microenvironment in terms of various physical, chemical, and biological factors. Co-culture technology is used in cartilage tissue engineering to overcome obstacles related to the degeneration of seed cells, and shows promise for cartilage regeneration and repair. In this review, we focus first on existing co-culture systems for cartilage tissue engineering and related fields, and discuss the conditions and mechanisms thereof. This is followed by methods for optimizing seed cell co-culture conditions to generate functional neo-cartilage tissue, which will lead to a new era in cartilage tissue engineering. © 2017 Wiley Periodicals, Inc.

  15. Evaluation of Medicinal Plant Hepatotoxicity in Co-cultures of Hepatocytes and Monocytes

    Directory of Open Access Journals (Sweden)

    Bashar Saad

    2006-01-01

    Full Text Available Non-parenchymal cells might play an important role in the modulation of xenobiotic metabolism in liver and its pharmacological and toxicological consequences. Therefore, the role of cell-to-cell interactions in herbal induced liver toxicity was investigated in monocultures of cells from the human hepatocyte cell line (HepG2 and in co-cultures of cells from the HepG2 cell line and cells from the human monocyte cell line (THP1. Cells were treated with various concentrations (1–500 µg ml−1 of extracts of Pistacia palaestina, Juglans regia and Quercus ithaburensis for 24 h. Extracts from Cleome droserifolia, a known toxic plant, were taken as positive control. In the co-culture system, toxic effects were observed after exposure to extracts of Pistacia palestina and C. droserifolia. These two extracts significantly reduced by cell viability as measured the MTT test and the LDH assay. Whereas in hepatocyte cultures, only extracts of C. droserifolia were found to affect the cell viability. The production levels of albumin from hepatocytes were not affected by treatment with plant extracts in both culture systems. It seems that the observed reduction in cell viability after exposure to extracts of P. palestina in co-cultures but not in monocultures is a result of monocyte-derived factors. The use of liver cell co-cultures is therefore a useful approach to investigate the influence of intercellular communication on xenobiotic metabolism in liver.

  16. Immobilized anaerobic fermentation for bio-fuel production by Clostridium co-culture.

    Science.gov (United States)

    Xu, Lei; Tschirner, Ulrike

    2014-08-01

    Clostridium thermocellum/Clostridium thermolacticum co-culture fermentation has been shown to be a promising way of producing ethanol from several carbohydrates. In this research, immobilization techniques using sodium alginate and alkali pretreatment were successfully applied on this co-culture to improve the bio-ethanol fermentation performance during consolidated bio-processing (CBP). The ethanol yield obtained increased by over 60 % (as a percentage of the theoretical maximum) as compared to free cell fermentation. For cellobiose under optimized conditions, the ethanol yields were approaching about 85 % of the theoretical efficiency. To examine the feasibility of this immobilization co-culture on lignocellulosic biomass conversion, untreated and pretreated aspen biomasses were also used for fermentation experiments. The immobilized co-culture shows clear benefits in bio-ethanol production in the CBP process using pretreated aspen. With a 3-h, 9 % NaOH pretreatment, the aspen powder fermentation yields approached 78 % of the maximum theoretical efficiency, which is almost twice the yield of the untreated aspen fermentation.

  17. Effect of Fibroblast Co-culture on In Vitro Maturation and Fertilization of Mouse Preantral Follicles

    Directory of Open Access Journals (Sweden)

    Mahmoud Heidari

    2011-01-01

    Full Text Available Background: The aim of this study was to evaluate fibroblast co-culture on in vitro maturation andfertilization of prepubertal mouse preantral follicles.Materials and Methods: The ovaries of 12-14 day old mice were dissected and 120-150 μmintact preantral follicles with one or two layers of granulosa cells, and round oocytes were culturedindividually in α-minimal essential medium (α-MEM supplemented with 5% fetal bovine serum(FBS, 100 mIU/ml recombinant follicle stimulating hormone, 1% insulin, transferrin, seleniummix, 100 μg/ml penicillin and 50 μg/ml streptomycin as base medium for 12 days. A total number of226 follicules were cultured under two conditions: i base medium as control group (n=113; ii basemedium co-cultured with mouse embryonic fibroblast (MEF (n=113. Follicular diameters, alone,in addition to other factors were analyzed by student’s t-test and chi-square test, respectively.Results: The co-culture group showed significant differences (p<0.05 in growth rate (days 4, 6 and8 of the culture period and survival rate. However, there was no significant difference in antrumformation, ovulation rate and embryonic development of released oocytes. There were significantdifferences (p<0.05 in the estradiol and progesterone secretion at all days between the co-cultureand control groups.Conclusion: Fibroblast co-culture increased survival rate and steroid production of preantralfollicles by promoting granulosa cell proliferation.

  18. Boron nitride nanotube-mediated stimulation of cell co-culture on micro-engineered hydrogels.

    Directory of Open Access Journals (Sweden)

    Leonardo Ricotti

    Full Text Available In this paper, we describe the effects of the combination of topographical, mechanical, chemical and intracellular electrical stimuli on a co-culture of fibroblasts and skeletal muscle cells. The co-culture was anisotropically grown onto an engineered micro-grooved (10 µm-wide grooves polyacrylamide substrate, showing a precisely tuned Young's modulus (∼ 14 kPa and a small thickness (∼ 12 µm. We enhanced the co-culture properties through intracellular stimulation produced by piezoelectric nanostructures (i.e., boron nitride nanotubes activated by ultrasounds, thus exploiting the ability of boron nitride nanotubes to convert outer mechanical waves (such as ultrasounds in intracellular electrical stimuli, by exploiting the direct piezoelectric effect. We demonstrated that nanotubes were internalized by muscle cells and localized in both early and late endosomes, while they were not internalized by the underneath fibroblast layer. Muscle cell differentiation benefited from the synergic combination of topographical, mechanical, chemical and nanoparticle-based stimuli, showing good myotube development and alignment towards a preferential direction, as well as high expression of genes encoding key proteins for muscle contraction (i.e., actin and myosin. We also clarified the possible role of fibroblasts in this process, highlighting their response to the above mentioned physical stimuli in terms of gene expression and cytokine production. Finally, calcium imaging-based experiments demonstrated a higher functionality of the stimulated co-cultures.

  19. A New In Vitro Co-Culture Model Using Magnetic Force-Based Nanotechnology

    NARCIS (Netherlands)

    Takanari, Hiroki; Miwa, Keiko; Fu, Xian Ming; Nakai, Junichi; Ito, Akira; Ino, Kousuke; Honda, Hiroyuki; Tonomura, Wataru; Konishi, Satoshi; Opthof, Tobias; van der Heyden, Marcel A G; Kodama, Itsuo; Lee, Jong Kook

    2016-01-01

    Skeletal myoblast (SkMB) transplantation has been conducted as a therapeutic strategy for severe heart failure. However, arrhythmogenicity following transplantation remains unsolved. We developed an in vitro model of myoblast transplantation with “patterned” or “randomly-mixed” co-culture of SkMBs

  20. A New In Vitro Co-Culture Model Using Magnetic Force-Based Nanotechnology

    NARCIS (Netherlands)

    Takanari, Hiroki; Miwa, Keiko; Fu, Xianming; Nakai, Junichi; Ito, Akira; Ino, Kousuke; Honda, Hiroyuki; Tonomura, Wataru; Konishi, Satoshi; Opthof, Tobias; van der Heyden, Marcel Ag; Kodama, Itsuo; Lee, Jong-Kook

    2016-01-01

    Skeletal myoblast (SkMB) transplantation has been conducted as a therapeutic strategy for severe heart failure. However, arrhythmogenicity following transplantation remains unsolved. We developed an in vitro model of myoblast transplantation with "patterned" or "randomly-mixed" co-culture of SkMBs

  1. Engraftment Outcomes after HPC Co-Culture with Mesenchymal Stromal Cells and Osteoblasts

    Directory of Open Access Journals (Sweden)

    Matthew M. Cook

    2013-09-01

    Full Text Available Haematopoietic stem cell (HSC transplantation is an established cell-based therapy for a number of haematological diseases. To enhance this therapy, there is considerable interest in expanding HSCs in artificial niches prior to transplantation. This study compared murine HSC expansion supported through co-culture on monolayers of either undifferentiated mesenchymal stromal cells (MSCs or osteoblasts. Sorted Lineage− Sca-1+ c-kit+ (LSK haematopoietic stem/progenitor cells (HPC demonstrated proliferative capacity on both stromal monolayers with the greatest expansion of LSK shown in cultures supported by osteoblast monolayers. After transplantation, both types of bulk-expanded cultures were capable of engrafting and repopulating lethally irradiated primary and secondary murine recipients. LSKs co-cultured on MSCs showed comparable, but not superior, reconstitution ability to that of freshly isolated LSKs. Surprisingly, however, osteoblast co-cultured LSKs showed significantly poorer haematopoietic reconstitution compared to LSKs co-cultured on MSCs, likely due to a delay in short-term reconstitution. We demonstrated that stromal monolayers can be used to maintain, but not expand, functional HSCs without a need for additional haematopoietic growth factors. We also demonstrated that despite apparently superior in vitro performance, co-injection of bulk cultures of osteoblasts and LSKs in vivo was detrimental to recipient survival and should be avoided in translation to clinical practice.

  2. Expression of HSP27, HSP72 and MRP proteins in in vitro co-culture ...

    Indian Academy of Sciences (India)

    We studied the expression of inducible heat shock protein (HSP27, HSP72) and multidrug-resistance protein (MRP) in co-cultures of human colon carcinoma cell spheroids obtained from different grades of tumour with normal human colon epithelium, myofibroblast and endothelial cell monolayers. We also measured the ...

  3. Impact of sequential co-culture fermentations on flavour characters of Solaris wines

    DEFF Research Database (Denmark)

    Liu, Jing; Arneborg, Nils; Toldam-Andersen, Torben Bo

    2017-01-01

    -phenylethyl acetate. This co-culture fermentation also produced higher amounts of ethyl acetate and acetic acid, reducing the possibility of its use in winemaking. Three Metschnikowia strains, a M. chrysoperlae strain and two M. fructicola strains, gave a comparable production of volatile compounds...

  4. Functional 3-D cardiac co-culture model using bioactive chitosan nanofiber scaffolds.

    Science.gov (United States)

    Hussain, Ali; Collins, George; Yip, Derek; Cho, Cheul H

    2013-02-01

    The in vitro generation of a three-dimensional (3-D) myocardial tissue-like construct employing cells, biomaterials, and biomolecules is a promising strategy in cardiac tissue regeneration, drug testing, and tissue engineering applications. Despite significant progress in this field, current cardiac tissue models are not yet able to stably maintain functional characteristics of cardiomyocytes for long-term culture and therapeutic purposes. The objective of this study was to fabricate bioactive 3-D chitosan nanofiber scaffolds using an electrospinning technique and exploring its potential for long-term cardiac function in the 3-D co-culture model. Chitosan is a natural polysaccharide biomaterial that is biocompatible, biodegradable, non-toxic, and cost effective. Electrospun chitosan was utilized to provide structural scaffolding characterized by scale and architectural resemblance to the extracellular matrix (ECM) in vivo. The chitosan fibers were coated with fibronectin via adsorption in order to enhance cellular adhesion to the fibers and migration into the interfibrous milieu. Ventricular cardiomyocytes were harvested from neonatal rats and studied in various culture conditions (i.e., mono- and co-cultures) for their viability and function. Cellular morphology and functionality were examined using immunofluorescent staining for alpha-sarcomeric actin (SM-actin) and gap junction protein, Connexin-43 (Cx43). Scanning electron microscopy (SEM) and light microscopy were used to investigate cellular morphology, spatial organization, and contractions. Calcium indicator was used to monitor calcium ion flux of beating cardiomyocytes. The results demonstrate that the chitosan nanofibers retained their cylindrical morphology in long-term cell cultures and exhibited good cellular attachment and spreading in the presence of adhesion molecule, fibronectin. Cardiomyocyte mono-cultures resulted in loss of cardiomyocyte polarity and islands of non-coherent contractions. However

  5. Co-culture of 3D tumor spheroids with fibroblasts as a model for epithelial–mesenchymal transition in vitro

    International Nuclear Information System (INIS)

    Kim, Sun-Ah; Lee, Eun Kyung; Kuh, Hyo-Jeong

    2015-01-01

    Epithelial–mesenchymal transition (EMT) acts as a facilitator of metastatic dissemination in the invasive margin of malignant tumors where active tumor–stromal crosstalks take place. Co-cultures of cancer cells with cancer-associated fibroblasts (CAFs) are often used as in vitro models of EMT. We established a tumor–fibroblast proximity co-culture using HT-29 tumor spheroids (TSs) with CCD-18co fibroblasts. When co-cultured with TSs, CCD-18co appeared activated, and proliferative activity as well as cell migration increased. Expression of fibronectin increased whereas laminin and type I collagen decreased in TSs co-cultured with fibroblasts compared to TSs alone, closely resembling the margin of in vivo xenograft tissue. Active TGFβ1 in culture media significantly increased in TS co-cultures but not in 2D co-cultures of cancer cells–fibroblasts, indicating that 3D context-associated factors from TSs may be crucial to crosstalks between cancer cells and fibroblasts. We also observed in TSs co-cultured with fibroblasts increased expression of α-SMA, EGFR and CTGF; reduced expression of membranous β-catenin and E-cadherin, together suggesting an EMT-like changes similar to a marginal region of xenograft tissue in vivo. Overall, our in vitro TS–fibroblast proximity co-culture mimics the EMT-state of the invasive margin of in vivo tumors in early metastasis. - Highlights: • An adjacent co-culture of tumor spheroids and fibroblasts is presented as EMT model. • Activation of fibroblasts and increased cell migration were shown in co-culture. • Expression of EMT-related factors in co-culture was similar to that in tumor tissue. • Crosstalk between spheroids and fibroblasts was demonstrated by secretome analysis

  6. Co-culture of 3D tumor spheroids with fibroblasts as a model for epithelial–mesenchymal transition in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Sun-Ah, E-mail: j.sarah.k@gmail.com [Department of Biomedicine & Health Sciences, College of Medicine, The Catholic University of Korea, Seoul 137-701 (Korea, Republic of); Lee, Eun Kyung, E-mail: leeek@catholic.ac.kr [Department of Biochemistry, College of Medicine, The Catholic University of Korea, Seoul 137-701 (Korea, Republic of); Cancer Evolution Research Center, College of Medicine, The Catholic University of Korea, Seoul 137-701 (Korea, Republic of); Kuh, Hyo-Jeong, E-mail: hkuh@catholic.ac.kr [Department of Biomedicine & Health Sciences, College of Medicine, The Catholic University of Korea, Seoul 137-701 (Korea, Republic of); Cancer Evolution Research Center, College of Medicine, The Catholic University of Korea, Seoul 137-701 (Korea, Republic of)

    2015-07-15

    Epithelial–mesenchymal transition (EMT) acts as a facilitator of metastatic dissemination in the invasive margin of malignant tumors where active tumor–stromal crosstalks take place. Co-cultures of cancer cells with cancer-associated fibroblasts (CAFs) are often used as in vitro models of EMT. We established a tumor–fibroblast proximity co-culture using HT-29 tumor spheroids (TSs) with CCD-18co fibroblasts. When co-cultured with TSs, CCD-18co appeared activated, and proliferative activity as well as cell migration increased. Expression of fibronectin increased whereas laminin and type I collagen decreased in TSs co-cultured with fibroblasts compared to TSs alone, closely resembling the margin of in vivo xenograft tissue. Active TGFβ1 in culture media significantly increased in TS co-cultures but not in 2D co-cultures of cancer cells–fibroblasts, indicating that 3D context-associated factors from TSs may be crucial to crosstalks between cancer cells and fibroblasts. We also observed in TSs co-cultured with fibroblasts increased expression of α-SMA, EGFR and CTGF; reduced expression of membranous β-catenin and E-cadherin, together suggesting an EMT-like changes similar to a marginal region of xenograft tissue in vivo. Overall, our in vitro TS–fibroblast proximity co-culture mimics the EMT-state of the invasive margin of in vivo tumors in early metastasis. - Highlights: • An adjacent co-culture of tumor spheroids and fibroblasts is presented as EMT model. • Activation of fibroblasts and increased cell migration were shown in co-culture. • Expression of EMT-related factors in co-culture was similar to that in tumor tissue. • Crosstalk between spheroids and fibroblasts was demonstrated by secretome analysis.

  7. Optimization of the Static Human Osteoblast/Osteoclast Co-culture System

    Directory of Open Access Journals (Sweden)

    James Jam Jolly

    2018-03-01

    Full Text Available Osteoblasts (OBs and osteoclasts (OCs are 2 major groups of bone cells. Their cell-to-cell interactions are important to ensure the continuity of the bone-remodeling process. Therefore, the present study was carried out to optimize an OB/OC co-culture system utilizing the human OB cell line hFOB 1.19 and OCs extracted from peripheral blood mononuclear cells (PBMNCs. It was a 2-step procedure, involving the optimization of the OB culture and the co-culture of the OBs with PBMNCs at an optimum ratio. Firstly, pre-OBs were cultured to 90% confluency and the time required for differentiation was determined. OB differentiation was determined using the van Gieson staining to detect the presence of collagen and Alizarin Red for calcium. Secondly, OBs and OCs were co-cultured at the ratios of 1 OC: 1 OB, 1 OC: 4 OBs, 2 OCs: 1 OB, and 1 OC: 2 OBs. Tartrate-resistant acid phosphatase (TRAP staining was used to detect the differentiation of the OCs. The results showed that collagen was present on day 1, whereas calcium was detected as early as day 3. Based on the result of TRAP staining, 1 OC: 2 OBs was taken as the most appropriate ratio. No macrophage colony-stimulating factor and receptor activator of the nuclear factor-κB ligand were added because they were provided by the OBs. In conclusion, these optimization processes are vital as they ensure the exact time point and ratio of the OB/OC co-culture in order to produce a reliable and reproducible co-culture system.

  8. Treatment of real wastewater using co-culture of immobilized Chlorella vulgaris and suspended activated sludge.

    Science.gov (United States)

    Mujtaba, Ghulam; Lee, Kisay

    2017-09-01

    The use of algal-bacterial symbiotic association establishes a sustainable and cost-effective strategy in wastewater treatment. Using municipal wastewater, the removal performances of inorganic nutrients (nitrogen and phosphorus) and organic pollutants were investigated by the co-culture system having different inoculum ratios (R) of suspended activated sludge to alginate-immobilized microalgae Chlorella vulgaris. The co-culture reactors with lower R ratios obtained more removal of nitrogen than in pure culture of C. vulgaris. The reactor with R = 0.5 (sludge/microalgae) showed the highest performance representing 66% removal after 24 h and 95% removal after 84 h. Phosphorus was completely eliminated (100%) in the co-culture system with inoculum ratios of 0.5 and 1.0 after 24 h and in the pure C. vulgaris culture after 36 h. The COD level was greatly reduced in the activated sludge reactor, while, it was increasing in pure C. vulgaris culture after 24 h of incubation. However, COD was almost stabilized after 24 h in the reactors with high R ratios such as 2.0, 5.0, and 10 due to the higher concentration of activated sludge. The growth of C. vulgaris was promoted from 0.03 g/L/d to 0.05 g/L/d in the co-culture of low inoculum ratios such as R = 0.5, implying that there exist an optimum inoculum ratio in the co-culture system in order to achieve efficient removal of nutrients. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. Co-culture with Sertoli cells promotes proliferation and migration of umbilical cord mesenchymal stem cells

    International Nuclear Information System (INIS)

    Zhang, Fenxi; Hong, Yan; Liang, Wenmei; Ren, Tongming; Jing, Suhua; Lin, Juntang

    2012-01-01

    Highlights: ► Co-culture of Sertoli cells (SCs) with human umbilical cord mesenchymal stem cells (UCMSCs). ► Presence of SCs dramatically increased proliferation and migration of UCMSCs. ► Presence of SCs stimulated expression of Mdm2, Akt, CDC2, Cyclin D, CXCR4, MAPKs. -- Abstract: Human umbilical cord mesenchymal stem cells (hUCMSCs) have been recently used in transplant therapy. The proliferation and migration of MSCs are the determinants of the efficiency of MSC transplant therapy. Sertoli cells are a kind of “nurse” cells that support the development of sperm cells. Recent studies show that Sertoli cells promote proliferation of endothelial cells and neural stem cells in co-culture. We hypothesized that co-culture of UCMSCs with Sertoli cells may also promote proliferation and migration of UCMSCs. To examine this hypothesis, we isolated UCMSCs from human cords and Sertoli cells from mouse testes, and co-cultured them using a Transwell system. We found that UCMSCs exhibited strong proliferation ability and potential to differentiate to other cell lineages such as osteocytes and adipocytes. The presence of Sertoli cells in co-culture significantly enhanced the proliferation and migration potential of UCMSCs (P < 0.01). Moreover, these phenotypic changes were accompanied with upregulation of multiple genes involved in cell proliferation and migration including phospho-Akt, Mdm2, phospho-CDC2, Cyclin D1, Cyclin D3 as well as CXCR4, phospho-p44 MAPK and phospho-p38 MAPK. These findings indicate that Sertoli cells boost UCMSC proliferation and migration potential.

  10. Biorelevant media resistant co-culture model mimicking permeability of human intestine.

    Science.gov (United States)

    Antoine, Delphine; Pellequer, Yann; Tempesta, Camille; Lorscheidt, Stefan; Kettel, Bernadette; Tamaddon, Lana; Jannin, Vincent; Demarne, Frédéric; Lamprecht, Alf; Béduneau, Arnaud

    2015-03-15

    Cell culture models are currently used to predict absorption pattern of new compounds and formulations in the human gastro-intestinal tract (GIT). One major drawback is the lack of relevant apical incubation fluids allowing mimicking luminal conditions in the GIT. Here, we suggest a culture model compatible with biorelevant media, namely Fasted State Simulated Intestinal Fluid (FaSSIF) and Fed State Simulated Intestinal Fluid (FeSSIF). Co-culture was set up from Caco-2 and mucus-secreting HT29-MTX cells using an original seeding procedure. Viability and cytotoxicity assays were performed following incubation of FeSSIF and FaSSIF with co-culture. Influence of biorelevant fluids on paracellular permeability or transporter proteins were also evaluated. Results were compared with Caco-2 and HT29-MTX monocultures. While Caco-2 viability was strongly affected with FeSSIF, no toxic effect was detected for the co-cultures in terms of viability and lactate dehydrogenase release. The addition of FeSSIF to the basolateral compartment of the co-culture induced cytotoxic effects which suggested the apical mucus barrier being cell protective. In contrast to FeSSIF, FaSSIF induced a slight increase of the paracellular transport and both tested media inhibited partially the P-gp-mediated efflux in the co-culture. Additionally, the absorptive transport of propranolol hydrochloride, a lipophilic β-blocker, was strongly affected by biorelevant fluids. This study demonstrated the compatibility of the Caco-2/HT29-MTX model with some of the current biorelevant media. Combining biorelevant intestinal fluids with features such as mucus secretion, adjustable paracellular and P-gp mediated transports, is a step forward to more realistic in-vitro models of the human intestine. Copyright © 2015. Published by Elsevier B.V.

  11. Naringenin-responsive riboswitch-based fluorescent biosensor module for Escherichia coli co-cultures.

    Science.gov (United States)

    Xiu, Yu; Jang, Sungho; Jones, J Andrew; Zill, Nicholas A; Linhardt, Robert J; Yuan, Qipeng; Jung, Gyoo Yeol; Koffas, Mattheos A G

    2017-10-01

    The ability to design and construct combinatorial synthetic metabolic pathways has far exceeded our capacity for efficient screening and selection of the resulting microbial strains. The need for high-throughput rapid screening techniques is of upmost importance for the future of synthetic biology and metabolic engineering. Here we describe the development of an RNA riboswitch-based biosensor module with dual fluorescent reporters, and demonstrate a high-throughput flow cytometry-based screening method for identification of naringenin over producing Escherichia coli strains in co-culture. Our efforts helped identify a number of key operating parameters that affect biosensor performance, including the selection of promoter and linker elements within the sensor-actuator domain, and the effect of host strain, fermentation time, and growth medium on sensor dynamic range. The resulting biosensor demonstrates a high correlation between specific fluorescence of the biosensor strain and naringenin titer produced by the second member of the synthetic co-culture system. This technique represents a novel application for synthetic microbial co-cultures and can be expanded from naringenin to any metabolite if a suitable riboswitch is identified. The co-culture technique presented here can be applied to a variety of target metabolites in combination with the SELEX approach for aptamer design. Due to the compartmentalization of the two genetic constructs responsible for production and detection into separate cells and application as independent modules of a synthetic microbial co-culture we have subsequently reduced the need for re-optimization of the producer module when the biosensor is replaced or removed. Biotechnol. Bioeng. 2017;114: 2235-2244. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  12. Co-culture with Sertoli cells promotes proliferation and migration of umbilical cord mesenchymal stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Fenxi, E-mail: fxzhang0824@gmail.com [Department of Anatomy, Sanquan College, Xinxiang Medical University, Henan 453003, People' s Republic of China (China); Hong, Yan; Liang, Wenmei [Department of Histology and Embryology, Guiyang Medical University, Guizhou 550004, People' s Republic of China (China); Ren, Tongming [Department of Anatomy, Sanquan College, Xinxiang Medical University, Henan 453003, People' s Republic of China (China); Jing, Suhua [ICU Center, The Third Hospital of Xinxiang Medical University, Henan 453003, People' s Republic of China (China); Lin, Juntang [Stem Cell Center, Xinxiang Medical University, Henan 453003, People' s Republic of China (China)

    2012-10-12

    Highlights: Black-Right-Pointing-Pointer Co-culture of Sertoli cells (SCs) with human umbilical cord mesenchymal stem cells (UCMSCs). Black-Right-Pointing-Pointer Presence of SCs dramatically increased proliferation and migration of UCMSCs. Black-Right-Pointing-Pointer Presence of SCs stimulated expression of Mdm2, Akt, CDC2, Cyclin D, CXCR4, MAPKs. -- Abstract: Human umbilical cord mesenchymal stem cells (hUCMSCs) have been recently used in transplant therapy. The proliferation and migration of MSCs are the determinants of the efficiency of MSC transplant therapy. Sertoli cells are a kind of 'nurse' cells that support the development of sperm cells. Recent studies show that Sertoli cells promote proliferation of endothelial cells and neural stem cells in co-culture. We hypothesized that co-culture of UCMSCs with Sertoli cells may also promote proliferation and migration of UCMSCs. To examine this hypothesis, we isolated UCMSCs from human cords and Sertoli cells from mouse testes, and co-cultured them using a Transwell system. We found that UCMSCs exhibited strong proliferation ability and potential to differentiate to other cell lineages such as osteocytes and adipocytes. The presence of Sertoli cells in co-culture significantly enhanced the proliferation and migration potential of UCMSCs (P < 0.01). Moreover, these phenotypic changes were accompanied with upregulation of multiple genes involved in cell proliferation and migration including phospho-Akt, Mdm2, phospho-CDC2, Cyclin D1, Cyclin D3 as well as CXCR4, phospho-p44 MAPK and phospho-p38 MAPK. These findings indicate that Sertoli cells boost UCMSC proliferation and migration potential.

  13. Micropattern printing of adhesion, spreading, and migration peptides on poly(tetrafluoroethylene) films to promote endothelialization.

    Science.gov (United States)

    Gauvreau, Virginie; Laroche, Gaétan

    2005-01-01

    We report here the development of an original multistep micropatterning technique for printing peptides on surfaces, based on the ink-jet printer technology. Contrary to most micropatterning methods used nowadays, this technique is advantageous because it allows displaying 2D-arrays of multiple biomolecules. Moreover, this low cost procedure allies the advantages of computer-aided design with high flexibility and reproducibility. A Hewlett-Packard printer was modified to print peptide solutions, and Adobe Illustrator was used as the graphic-editing software to design high-resolution checkerboard-like micropatterns. In a first step, PTFE films were treated with ammonia plasma to introduce amino groups on the surface. These chemical functionalities were reacted with heterobifunctional cross-linker sulfo-succinimidyl 4-(N-maleimidomethyl)cycloexane-1-carboxylate (S-SMCC) to allow the subsequent surface covalent conjugation of various cysteine-modified peptides to the polymer substrate. These peptidic molecules containing RGD and WQPPRARI sequences were selected for their adhesive, spreading, and migrational properties toward endothelial cells. On one hand, our data demonstrated that the initial cell adhesion does not depend on the chemical structure and combination of the peptides covalently bonded either through conventional conjugation or micropatterning. On the other hand, spreading and migration of endothelial cells is clearly enhanced while coconjugating the GRGDS peptide in conjunction with WQPPRARI. This behavior is further improved by micropatterning these peptides on specific areas of the polymer surface.

  14. Morphology and intrinsic excitability of regenerating sensory and motor neurons grown on a line micropattern.

    Directory of Open Access Journals (Sweden)

    Ouafa Benzina

    Full Text Available Axonal regeneration is one of the greatest challenges in severe injuries of peripheral nerve. To provide the bridge needed for regeneration, biological or synthetic tubular nerve constructs with aligned architecture have been developed. A key point for improving axonal regeneration is assessing the effects of substrate geometry on neuronal behavior. In the present study, we used an extracellular matrix-micropatterned substrate comprising 3 µm wide lines aimed to physically mimic the in vivo longitudinal axonal growth of mice peripheral sensory and motor neurons. Adult sensory neurons or embryonic motoneurons were seeded and processed for morphological and electrical activity analyses after two days in vitro. We show that micropattern-guided sensory neurons grow one or two axons without secondary branching. Motoneurons polarity was kept on micropattern with a long axon and small dendrites. The micro-patterned substrate maintains the growth promoting effects of conditioning injury and demonstrates, for the first time, that neurite initiation and extension could be differentially regulated by conditioning injury among DRG sensory neuron subpopulations. The micro-patterned substrate impacts the excitability of sensory neurons and promotes the apparition of firing action potentials characteristic for a subclass of mechanosensitive neurons. The line pattern is quite relevant for assessing the regenerative and developmental growth of sensory and motoneurons and offers a unique model for the analysis of the impact of geometry on the expression and the activity of mechanosensitive channels in DRG sensory neurons.

  15. FGLD A novel and compact micro-pattern gas detector

    CERN Document Server

    Dick, Louis; Watts, David

    2004-01-01

    A new gas detector which combines in the same structure the gas amplification mechanism and the position sensitive readout, named the field gradient lattice detector (FGLD), is being developed at CERN. The detector, reminiscent in geometry of a multi-wire proportional chamber but with a different field configuration can be fabricated as two or more layers of micro-patterned parallel tracks on a variety of substrate materials. Two preliminary proof-of-concept designs without position sensitivity have been fabricated as copper tracks of 50 mum width and 150 mum pitch on polyimide in a 3D geometry and on epoxy in a 2D geometry. They have been shown to detect the 5.9 keV X-rays of an $^{55}Fe$ source with a stable gain ranging from 500 to 5000 in a 3 mm drift chamber containing an argon carbon-dioxide gas mixture. The elegance and compactness of the FGLD design make it a very attractive gas detector solution both economically and mechanically. Most interestingly, the 3D FGLD design on flexible polyimide should gr...

  16. Development of Hybrid and Monolithic Silicon Micropattern Detectors

    CERN Multimedia

    Beker, H; Snoeys, W; Campbell, M; Lemeilleur, F; Ropotar, I

    2002-01-01

    %RD-19 \\\\ \\\\ In a collaborative effort between particle physics institutes and microelectronics industry we are undertaking the development of true 2-dimensional semiconductor particle detectors with on-chip signal processing and information extraction: the so-called micropattern detector. This detector is able to cope in a robust way with high multiplicity events at high rates, while allowing for a longer detector lifetime under irradiation and a thinner sensitive depletion region. Therefore, it will be ideally suited for the complicated events in the LHC p-p collider experiments. Following a $^{\\prime}$stepping stone$^{\\prime}$ approach several telescopes of pixel planes, totalling now 600 cm$^{2}$ with \\(>\\)~1~M elements have been used in the WA97, NA50 and NA57 lead ion experiments. This new technology has facilitated the tracking considerably (see Fig.1). Not only Si but also GaAs and possibly diamond matrices can be connected to the readout matrix. Tests with GaAs pixel detectors with the RD-19 readout ...

  17. Engineering connectivity by multiscale micropatterning of individual populations of neurons.

    Science.gov (United States)

    Albers, Jonas; Toma, Koji; Offenhäusser, Andreas

    2015-02-01

    Functional networks are the basis of information processing in the central nervous system. Essential for their formation are guided neuronal growth as well as controlled connectivity and information flow. The basis of neuronal development is generated by guiding cues and geometric constraints. To investigate the neuronal growth and connectivity of adjacent neuronal networks, two-dimensional protein patterns were created. A mixture of poly-L-lysine and laminin was transferred onto a silanized glass surface by microcontact printing. The structures were populated with dissociated primary cortical embryonic rat neurons. Triangular structures with diverse opening angles, height, and design were chosen as two-dimensional structures to allow network formation with constricted gateways. Neuronal development was observed by immunohistochemistry to pursue the influence of the chosen structures on the neuronal outgrowth. Neurons were stained for MAP2, while poly-L-lysine was FITC labeled. With this study we present an easy-to-use technique to engineer two-dimensional networks in vitro with defined gateways. The presented micropatterning method is used to generate daisy-chained neuronal networks with predefined connectivity. Signal propagation among geometrically constrained networks can easily be monitored by calcium-sensitive dyes, providing insights into network communication in vitro. Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Reconstruction of Micropattern Detector Signals using Convolutional Neural Networks

    Science.gov (United States)

    Flekova, L.; Schott, M.

    2017-10-01

    Micropattern gaseous detector (MPGD) technologies, such as GEMs or MicroMegas, are particularly suitable for precision tracking and triggering in high rate environments. Given their relatively low production costs, MPGDs are an exemplary candidate for the next generation of particle detectors. Having acknowledged these advantages, both the ATLAS and CMS collaborations at the LHC are exploiting these new technologies for their detector upgrade programs in the coming years. When MPGDs are utilized for triggering purposes, the measured signals need to be precisely reconstructed within less than 200 ns, which can be achieved by the usage of FPGAs. In this work, we present a novel approach to identify reconstructed signals, their timing and the corresponding spatial position on the detector. In particular, we study the effect of noise and dead readout strips on the reconstruction performance. Our approach leverages the potential of convolutional neural network (CNNs), which have recently manifested an outstanding performance in a range of modeling tasks. The proposed neural network architecture of our CNN is designed simply enough, so that it can be modeled directly by an FPGA and thus provide precise information on reconstructed signals already in trigger level.

  19. Soft-Matter Printed Circuit Board with UV Laser Micropatterning.

    Science.gov (United States)

    Lu, Tong; Markvicka, Eric J; Jin, Yichu; Majidi, Carmel

    2017-07-05

    When encapsulated in elastomer, micropatterned traces of Ga-based liquid metal (LM) can function as elastically deformable circuit wiring that provides mechanically robust electrical connectivity between solid-state elements (e.g., transistors, processors, and sensor nodes). However, LM-microelectronics integration is currently limited by challenges in rapid fabrication of LM circuits and the creation of vias between circuit terminals and the I/O pins of packaged electronics. In this study, we address both with a unique layup for soft-matter electronics in which traces of liquid-phase Ga-In eutectic (EGaIn) are patterned with UV laser micromachining (UVLM). The terminals of the elastomer-sealed LM circuit connect to the surface mounted chips through vertically aligned columns of EGaIn-coated Ag-Fe 2 O 3 microparticles that are embedded within an interfacial elastomer layer. The processing technique is compatible with conventional UVLM printed circuit board (PCB) prototyping and exploits the photophysical ablation of EGaIn on an elastomer substrate. Potential applications to wearable computing and biosensing are demonstrated with functional implementations in which soft-matter PCBs are populated with surface-mounted microelectronics.

  20. Semiconductor micropattern pixel detectors: a review of the beginnings

    International Nuclear Information System (INIS)

    Heijne, E.H.M.

    2001-01-01

    The innovation in monolithic and hybrid semiconductor 'micropattern' or 'reactive' pixel detectors for tracking in particle physics was actually to fit logic and pulse processing electronics with μW power on a pixel area of less than 0.04 mm 2 , retaining the characteristics of a traditional nuclear amplifier chain. The ns timing precision in conjunction with local memory and logic operations allowed event selection at >10 MHz rates with unambiguous track reconstruction even at particle multiplicities >10 cm -2 . The noise in a channel was ∼100e - rms and enabled binary operation with random noise 'hits' at a level -8 . Rectangular pixels from 75 μmx500 μm down to 34 μmx125 μm have been used by different teams. In binary mode a tracking precision from 6 to 14 μm was obtained, and using analog interpolation one came close to 1 μm. Earlier work, still based on charge integrating imaging circuits, provided a starting point. Two systems each with more than 1 million sensor + readout channels have been built, for WA97-NA57 and for the Delphi very forward tracker. The use of 0.5 μm and 0.25 μm CMOS and enclosed geometry for the transistors in the pixel readout chips resulted in radiation hardness of ∼2 Mrad, respectively, >30 Mrad

  1. A defined co-culture of Geobacter sulfurreducens and Escherichia coli in a membrane-less microbial fuel cell.

    Science.gov (United States)

    Bourdakos, Nicholas; Marsili, Enrico; Mahadevan, Radhakrishnan

    2014-04-01

    Wastewater-fed microbial fuel cells (MFCs) are a promising technology to treat low-organic carbon wastewater and recover part of the chemical energy in wastewater as electrical power. However, the interactions between electrochemically active and fermentative microorganisms cannot be easily studied in wastewater-fed MFCs because of their complex microbial communities. Defined co-culture MFCs provide a detailed understanding of such interactions. In this study, we characterize the extracellular metabolites in laboratory-scale membrane-less MFCs inoculated with Geobacter sulfurreducens and Escherichia coli co-culture and compare them with pure culture MFCs. G. sulfurreducens MFCs are sparged to maintain anaerobic conditions, while co-culture MFCs rely on E. coli for oxygen removal. G. sulfurreducens MFCs have a power output of 128 mW m(-2) , compared to 63 mW m(-2) from the co-culture MFCs. Analysis of metabolites shows that succinate production in co-culture MFCs decreases current production by G. sulfurreducens and that the removal of succinate is responsible for the increased current density in the late co-culture MFCs. Interestingly, pH adjustment is not required for co-culture MFCs but a base addition is necessary for E. coli MFCs and cultures in vials. Our results show that defined co-culture MFCs provide clear insights into metabolic interactions among bacteria while maintaining a low operational complexity. © 2013 Wiley Periodicals, Inc.

  2. Co-culture of microalgae, cyanobacteria, and macromycetes for exopolysaccharides production: process preliminary optimization and partial characterization.

    Science.gov (United States)

    Angelis, S; Novak, A C; Sydney, E B; Soccol, V T; Carvalho, J C; Pandey, A; Noseda, M D; Tholozan, J L; Lorquin, J; Soccol, C R

    2012-07-01

    In this study, the biomass and exopolysaccharides (EPS) production in co-cultures of microalgae/cyanobacteria and macromycetes was evaluated as a technology for producing new polysaccharides for medical and/or industrial application. Based on biomass and EPS productivity of monocultures, two algae and two fungi were selected and cultured in different co-culture arrangements. The hydrosoluble EPS fractions from mono- and co-cultures were characterized by ¹³C NMR spectroscopy and gas chromatography coupled to mass spectrometry and compared. It was found that co-cultures resulted in the production of an EPS different from those produced by monocultures, showing fungal predominance with microalgal/cyanobacterial traces. Co-cultures conditions were screened (temperature, agitation speed, fungal and microalgae inoculation rate, initial pH, illumination rate, and glucose concentration) in order to achieve maximum biomass and EPS production, resulting in an increase of 33 and 61% in exopolysaccharides and biomass productions, respectively (patent pending).

  3. An experimental approach towards the development of an in vitro cortical-thalamic co-culture model.

    Science.gov (United States)

    Kanagasabapathi, Thirukumaran T; Massobrio, Paolo; Tedesco, Mariateresa; Martinoia, Sergio; Wadman, Wytse J; Decré, Michel M J

    2011-01-01

    In this paper, we propose an experimental approach to develop an in vitro dissociated cortical-thalamic co-culture model using a dual compartment neurofluidic device. The device has two compartments separated by 10 μm wide and 3 μm high microchannels. The microchannels provide a physical isolation of neurons allowing only neurites to grow between the compartments. Long-term viable co-culture was maintained in the compartmented device, neurite growth through the microchannels was verified using immunofluorescence staining, and electrophysiological recordings from the co-culture system was investigated. Preliminary analysis of spontaneous activities from the co-culture shows a distinctively different firing pattern associated with cultures of individual cell types and further analysis is proposed for a deeper understanding of the dynamics involved in the network connectivity in such a co-culture system.

  4. Micropatterning of nanocomposite polymer scaffolds using sacrificial phosphate glass fibers for tendon tissue engineering applications.

    Science.gov (United States)

    Alshomer, Feras; Chaves, Camilo; Serra, Tiziano; Ahmed, Ifty; Kalaskar, Deepak M

    2017-04-01

    This study presents a simple and reproducible method of micropatterning the novel nanocomposite polymer (POSS-PCU) using a sacrificial phosphate glass fiber template for tendon tissue engineering applications. The diameters of the patterned scaffolds produced were dependent on the diameter of the glass fibers (15 μm) used. Scaffolds were tested for their physical properties and reproducibility using various microscopy techniques. For the first time, we show that POSS-PCU supports growth of human tenocytes cells. Furthermore, we show that cellular alignment, their biological function and expression of various tendon related proteins such as scleraxis, collagen I and III, tenascin-C are significantly elevated on the micropatterned polymer surfaces compared to flat samples. This study demonstrated a simple, reproducible method of micropatterning POSS-PCU nanocomposite polymer for novel tendon repair applications, which when provided with physical cues could help mimic the microenvironment of tenocytes cells. Copyright © 2017 Elsevier Inc. All rights reserved.

  5. Preparation and regulating cell adhesion of anion-exchangeable layered double hydroxide micropatterned arrays.

    Science.gov (United States)

    Yao, Feng; Hu, Hao; Xu, Sailong; Huo, Ruijie; Zhao, Zhiping; Zhang, Fazhi; Xu, Fujian

    2015-02-25

    We describe a reliable preparation of MgAl-layered double hydroxide (MgAl-LDH) micropatterned arrays on gold substrate by combining SO3(-)-terminated self-assembly monolayer and photolithography. The synthesis route is readily extended to prepare LDH arrays on the SO3(-)-terminated polymer-bonded glass substrate amenable for cell imaging. The anion-exchangeable MgAl-LDH micropattern can act both as bioadhesive region for selective cell adhesion and as nanocarrier for drug molecules to regulate cell behaviors. Quantitative analysis of cell adhesion shows that selective HepG2 cell adhesion and spreading are promoted by the micropatterned MgAl-LDH, and also suppressed by methotrexate drug released from the LDH interlayer galleries.

  6. Modelling the effects of PSII inhibitor pulse exposure on two algae in co-culture.

    Science.gov (United States)

    Copin, Pierre-Jean; Chèvre, Nathalie

    2018-03-01

    A weakness of standard testing procedures is that they do not consider interactions between organisms, and they focus only on single species. Furthermore, these procedures do not take into account pulse exposure. However, pulse exposure is of particular importance because in streams, after crop application and during and after precipitation, herbicide concentrations fluctuate widely and can exceed the Annual Average Environmental Quality Standards (AA-EQS), which aim to protect the aquatic environment. The sensitivity of the algae Scenedesmus vacuolatus and Pseudokirchneriella subcapitata in a co-culture exposed to pulses is thus analysed in this study. As a first step, the growths of the algae in co-culture are investigated. For initial cell densities fixed, respectively, to 100,000 and 50,000 cells/mL, the growth of each alga is exponential over at least 48 h. S. vacuolatus seems to influence the growth of P. subcapitata negatively. Allelopathy is a possible explanation for this growth inhibition. The toxicity of the herbicide isoproturon is later tested on the algae S. vacuolatus and P. subcapitata cultured alone and in the co-culture. Despite the supplementary stress on the algae in the co-culture competing for nutrients, the toxicity of the herbicide is lower for the two algae when they are in the co-culture than when they are in separated culture. A model is adapted and used to predict the cell-density inhibition on the alga S. vacuolatus in the co-culture with the alga P. subcapitata exposed to a pulse concentration of isoproturon. Four laboratory experiments are performed to validate the model. The comparison between the laboratory and the modelled effects shows good agreement. The differences can be considered minor most of time. For future studies, it is important to ensure that the cell count is precise, as it is used to determine the parameters of the model. The differences can be also induced by the fact that the cell number of the alga P

  7. Co-culture with infrapatellar fat pad differentially stimulates proteoglycan synthesis and accumulation in cartilage and meniscus tissues.

    Science.gov (United States)

    Nishimuta, James F; Bendernagel, Monica F; Levenston, Marc E

    2017-09-01

    Although osteoarthritis is widely viewed as a disease of the whole joint, relatively few studies have focused on interactions among joint tissues in joint homeostasis and degeneration. In particular, few studies have examined the effects of the infrapatellar fat pad (IFP) on cartilaginous tissues. The aim of this study was to test the hypothesis that co-culture with healthy IFP would induce degradation of cartilage and meniscus tissues. Bovine articular cartilage, meniscus, and IFP were cultured isolated or as cartilage-fat or meniscus-fat co-cultures for up to 14 days. Conditioned media were assayed for sulfated glycosaminoglycan (sGAG) content, nitrite content, and matrix metalloproteinase (MMP) activity, and explants were assayed for sGAG and DNA contents. Co-cultures exhibited increased cumulative sGAG release and sGAG release rates for both cartilage and meniscus, and the cartilage (but not meniscus) exhibited a substantial synergistic effect of co-culture (sGAG release in co-culture was significantly greater than the summed release from isolated cartilage and fat). Fat co-culture did not significantly alter the sGAG content of either cartilage or meniscus explants, indicating that IFP co-culture stimulated net sGAG production by cartilage. Nitrite release was increased relative to isolated tissue controls in co-cultured meniscus, but not the cartilage, with no synergistic effect of co-culture. Interestingly, MMP-2 production was decreased by co-culture for both cartilage and meniscus. This study demonstrates that healthy IFP may modulate joint homeostasis by stimulating sGAG production in cartilage. Counter to our hypothesis, healthy IFP did not promote degradation of either cartilage or meniscus tissues.

  8. Delineating fibronectin bioadhesive micropatterns by photochemical immobilization of polystyrene and poly(vinylpyrrolidone).

    Science.gov (United States)

    Sterner, Olof; Giazzon, Marta; Zürcher, Stefan; Tosatti, Samuele; Liley, Martha; Spencer, Nicholas D

    2014-01-01

    Bioadhesive micropatterns, capable of laterally confining cells to a 2D lattice, have proven effective in simulating the in vivo tissue environment. They reveal fundamental aspects of the role of adhesion in cell mechanics, proliferation, and differentiation. Here we present an approach based on photochemistry for the fabrication of synthetic polymer micropatterns. Perfluorophenyl azide (PFPA), upon deep-UV exposure, forms a reactive nitrene capable of covalently linking to a molecule that is in close proximity. PFPA has been grafted onto a backbone of poly(allyl amine), which readily forms a self-assembled monolayer on silicon wafers or glass. A film of polystyrene was applied by spin-coating, and by laterally confining the UV exposure through a chromium-on-quartz photomask, monolayers of polymers could be immobilized in circular microdomains. Poly(vinylpyrrolidone) (PVP) was attached to the background to form a barrier to nonspecific protein adsorption and cell adhesion. Micropatterns were characterized with high-lateral-resolution time-of-flight secondary ion mass spectrometry (TOF-SIMS), which confirmed the formation of polystyrene domains within a PVP background. Fluorescence-microscopy adsorption assays with rhodamine-labeled bovine serum albumin demonstrated the nonfouling efficiency of PVP and, combined with TOF-SIMS, allowed for a comprehensive characterization of the pattern geometry. The applicability of the micropatterned platform in single-cell assays was tested by culturing two cell types, WM 239 melanoma cells and SaOs-2 osteoblasts, on micropatterned glass, either with or without backfilling of the patterns with fibronectin. It was demonstrated that the platform was efficient in confining cells to the fibronectin-backfilled micropatterns for at least 48 h. PVP is thus proposed as a viable, highly stable alternative to poly(ethylene glycol) for nonfouling applications. Due to the versatility of the nitrene-insertion reaction, the platform could be

  9. Increased methane production in cyanobacteria and methanogenic microbe co-cultures.

    Science.gov (United States)

    Yeung, Tracey; Kwan, Matthew; Adler, Lewis; Mills, Toby J; Neilan, Brett A; Conibeer, Gavin; Patterson, Robert

    2017-11-01

    A novel light-to-bioenergy system produced 3.5 times the baseline methane output using a co-culture of cyanobacteria (Oscillatoria sp.) and a methanogenic microbial community. Analysis of micronutrients in the system during the growth phase indicated that cobalt, iron, nickel and zinc were not appreciably consumed. The stable consumption and return of macronutrients calcium and magnesium were also observed. Essential macronutrients nitrogen, in the form of nitrate, and phosphorus showed no cycling during the growth phase and were depleted at rates of 0.35mg/L/day and 0.40µg/L/day, respectively. Biofilm formation increased the resilience of biomass to bacterial degradation in an anaerobic digester, as shown by viability assays of cyanobacterial biofilms in the co-culture. Copyright © 2017 Elsevier Ltd. All rights reserved.

  10. Triple co-culture cell model as an in vitro model for oral particulate vaccine systems

    DEFF Research Database (Denmark)

    Nielsen, Line Hagner; De Rossi, C.; Lehr, C.-M.

    the immunostimulatory ability of particulate vaccine formulations designed for oral delivery. Levels of cytokine production in response to vaccine administration were measured following particulate vaccine administration, as an indication of dendritic cell and macrophage activation. Precursors of cubosomes containing......; this was not observed with ovalbumin and blank solution. An example of the results is shown in Figure 2 for IL-17A. An established co-culture of Caco-2, THP-1 and MUTZ-3 cells showed promise as an in vitro model for testing of oral vaccine formulations. Mobility of co-culture immune cells as well as cytokine production...... with particle formulations. This was not the case when incubating with ovalbumin solution or blank. The ELISA screening assay showed production of a wide range of cytokines following culture incubation with cubosomes (with and without ovalbumin) and LPS solutions, indicative of a stimulatory effect...

  11. 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.

  12. Co-culturing of Fungal Strains Against Botrytis cinerea as a Model for the Induction of Chemical Diversity and Therapeutic Agents

    Directory of Open Access Journals (Sweden)

    Olga Genilloud

    2017-04-01

    Full Text Available New fungal SMs (SMs have been successfully described to be produced by means of in vitro-simulated microbial community interactions. Co-culturing of fungi has proved to be an efficient way to induce cell–cell interactions that can promote the activation of cryptic pathways, frequently silent when the strains are grown in laboratory conditions. Filamentous fungi represent one of the most diverse microbial groups known to produce bioactive natural products. Triggering the production of novel antifungal compounds in fungi could respond to the current needs to fight health compromising pathogens and provide new therapeutic solutions. In this study, we have selected the fungus Botrytis cinerea as a model to establish microbial interactions with a large set of fungal strains related to ecosystems where they can coexist with this phytopathogen, and to generate a collection of extracts, obtained from their antagonic microbial interactions and potentially containing new bioactive compounds. The antifungal specificity of the extracts containing compounds induced after B. cinerea interaction was determined against two human fungal pathogens (Candida albicans and Aspergillus fumigatus and three phytopathogens (Colletotrichum acutatum, Fusarium proliferatum, and Magnaporthe grisea. In addition, their cytotoxicity was also evaluated against the human hepatocellular carcinoma cell line (HepG2. We have identified by LC-MS the production of a wide variety of known compounds induced from these fungal interactions, as well as novel molecules that support the potential of this approach to generate new chemical diversity and possible new therapeutic agents.

  13. [Experimental study on co-culture of human fibroblasts on decellularized Achilles tendon].

    Science.gov (United States)

    Wang, Zhibing; Zhang, Xia; Guo, Xinyu; Qin, Chuan

    2013-07-01

    To investigate the preparation of decellularized Achilles tendons and the effect of co-culture of human fibroblasts on the scaffold so as to provide a scaffold for the tissue engineered ligament reconstruction. Achilles tendons of both hind limbs were harvested from 10 male New Zealand white rabbits (5-month-old; weighing, 4-5 kg). The Achilles tendons were decellularized using trypsin, Triton X-100, and sodium dodecyl sulfate (SDS), and then gross observation, histological examination, and scanning electron microscope (SEM) observation were performed; the human fibroblasts were seeded on the decellularized Achilles tendon, and then cytocompatibility was tested using the cell counting kit 8 method at 1, 3, 5, 7, and 9 days after co-culture. At 4 weeks after co-culture, SEM, HE staining, and biomechanical test were performed for observing cell-scaffold composite, and a comparison was made with before and after decellularization. After decellularization, the tendons had integrated aponeurosis and enlarged volume with soft texture and good toughness; there was no loose connective tissue and tendon cells between tendon bundles, the collagen fibers arranged loosely with three-dimensional network structure and more pores between tendon bundles; and it had good cytocompatibility. At 4 weeks after co-culture, cells migrated into the pores, and three-dimensional network structure disappeared. By biomechanical test, the tensile strength and Young's elastic modulus of the decellularized Achilles tendon group decreased significantly when compared with normal Achilles tendons group and cell-scaffold composite group (P Achilles tendons group and cell-scaffold composite group (P > 0.05). There was no significant difference in elongation at break among 3 groups (P > 0.05). The decellularized Achilles tendon is biocompatible to fibroblasts. It is suit for the scaffold for tissue engineered ligament reconstruction.

  14. Co-cultures provide a new tool to probe communication between adult sensory neurons and urothelium.

    Science.gov (United States)

    O'Mullane, Lauren M; Keast, Janet R; Osborne, Peregrine B

    2013-08-01

    Recent evidence suggests that the urothelium functions as a sensory transducer of chemical, mechanical or thermal stimuli and signals to nerve terminals and other cells in the bladder wall. The cellular and molecular basis of neuro-urothelial communication is not easily studied in the intact bladder. This led us to establish a method of co-culturing dorsal root ganglion sensory neurons and bladder urothelial cells. Sensory neurons and urothelial cells obtained from dorsal root ganglia and bladders dissected from adult female Sprague-Dawley® rats were isolated by enzyme treatment and mechanical dissociation. They were plated together or separately on collagen coated substrate and cultured in keratinocyte medium for 48 to 72 hours. Retrograde tracer labeling was performed to identify bladder afferents used for functional testing. Neurite growth and complexity in neurons co-cultured with urothelial cells was increased relative to that in neuronal monocultures. The growth promoting effect of urothelial cells was reduced by the tyrosine kinase inhibitor K252a but upstream inhibition of nerve growth factor signaling with TrkA-Fc had no effect. Fura-2 calcium imaging of urothelial cells showed responses to adenosine triphosphate (100 μM) and activation of TRPV4 (4α-PDD, 10 μM) but not TRPV1 (capsaicin, 1 μM), TRPV3 (farnesyl pyrophosphate, 1 μM) or TRPA1 (mustard oil, 100 μM). In contrast, co-cultured neurons were activated by all agonists except farnesyl pyrophosphate. Co-culturing provides a new methodology for investigating neuro-urothelial interactions in animal models of urological conditions. Results suggest that neuronal properties are maintained in the presence of urothelium and neurite growth is potentiated by a nerve growth factor independent mechanism. Copyright © 2013 American Urological Association Education and Research, Inc. Published by Elsevier Inc. All rights reserved.

  15. Human neuron-astrocyte 3D co-culture-based assay for evaluation of neuroprotective compounds.

    Science.gov (United States)

    Terrasso, Ana Paula; Silva, Ana Carina; Filipe, Augusto; Pedroso, Pedro; Ferreira, Ana Lúcia; Alves, Paula Marques; Brito, Catarina

    Central nervous system drug development has registered high attrition rates, mainly due to the lack of efficacy of drug candidates, highlighting the low reliability of the models used in early-stage drug development and the need for new in vitro human cell-based models and assays to accurately identify and validate drug candidates. 3D human cell models can include different tissue cell types and represent the spatiotemporal context of the original tissue (co-cultures), allowing the establishment of biologically-relevant cell-cell and cell-extracellular matrix interactions. Nevertheless, exploitation of these 3D models for neuroprotection assessment has been limited due to the lack of data to validate such 3D co-culture approaches. In this work we combined a 3D human neuron-astrocyte co-culture with a cell viability endpoint for the implementation of a novel in vitro neuroprotection assay, over an oxidative insult. Neuroprotection assay robustness and specificity, and the applicability of Presto Blue, MTT and CytoTox-Glo viability assays to the 3D co-culture were evaluated. Presto Blue was the adequate endpoint as it is non-destructive and is a simpler and reliable assay. Semi-automation of the cell viability endpoint was performed, indicating that the assay setup is amenable to be transferred to automated screening platforms. Finally, the neuroprotection assay setup was applied to a series of 36 test compounds and several candidates with higher neuroprotective effect than the positive control, Idebenone, were identified. The robustness and simplicity of the implemented neuroprotection assay with the cell viability endpoint enables the use of more complex and reliable 3D in vitro cell models to identify and validate drug candidates. Copyright © 2016 Elsevier Inc. All rights reserved.

  16. Calcitriol enhances fat synthesis factors and calpain activity in co-cultured cells.

    Science.gov (United States)

    Choi, Hyuck; Myung, Kyuho

    2014-08-01

    We have conducted an in vitro experiment to determine whether calcitriol can act as a fat synthesizer and/or meat tenderizer when skeletal muscle cells, adipose tissue, and macrophages are co-cultured. When co-cultured, pro-inflammatory cytokine tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) expression increased, whereas decreased anti-inflammatory cytokine (IL-10 and IL-15) expression decreased in both C2C12 and 3T3-L1 cells. Calcitriol increased reactive oxygen species (ROS) production in the media. While adiponectin gene expression decreased, leptin, resistin, CCAAT-enhancer-binding protein-beta (C/EBP-β), and peroxisome proliferator-activated receptor gamma (PPAR-γ) gene expression was significantly (P cultured with two different cell types. Inducible nitric oxide synthase (iNOS) protein levels were also stimulated in the C2C12 and 3T3-L1 cells, but arginase l was attenuated by calcitriol. Cacitriol highly amplified (P = 0.008) µ-calpain gene expression in co-cultured C2C12 cells. The results showed an overall increase in pro-inflammatory cytokines and a decrease in anti-inflammatory cytokines of C2C12 and 3T3-L1 cells with calcitriol in co-culture systems. µ-Calpain protein was also augmented in differentiated C2C12 cells with calcitriol. These findings suggest that calcitriol can be used as not only fat synthesizer, but meat tenderizer, in meat-producing animals. © 2014 International Federation for Cell Biology.

  17. Establishment of mouse neuron and microglial cell co-cultured models and its action mechanism.

    Science.gov (United States)

    Zhang, Bo; Yang, Yunfeng; Tang, Jun; Tao, Yihao; Jiang, Bing; Chen, Zhi; Feng, Hua; Yang, Liming; Zhu, Gang

    2017-06-27

    The objective of this study is to establish a co-culture model of mouse neurons and microglial cells, and to analyze the mechanism of action of oxygen glucose deprivation (OGD) and transient oxygen glucose deprivation (tOGD) preconditioning cell models. Mouse primary neurons and BV2 microglial cells were successfully cultured, and the OGD and tOGD models were also established. In the co-culture of mouse primary neurons and microglial cells, the cell number of tOGD mouse neurons and microglial cells was larger than the OGD cell number, observed by a microscope. CCK-8 assay result showed that at 1h after treatment, the OD value in the control group is lower compared to all the other three groups (P control group compared to other three groups (P neurons cells were cultured. In the meantime mouse BV2 microglia cells were cultured. Two types of cells were co-cultured, and OGD and tOGD cell models were established. There were four groups in the experiment: control group (OGD), treatment group (tOGD+OGD), placebo group (tOGD+OGD+saline) and minocycline intervention group (tOGD+OGD+minocycline). CCK-8 kit was used to detect cell viability and flow cytometry was used to detect apoptosis. In this study, mouse primary neurons and microglial cells were co-cultured. The OGD and tOGD models were established successfully. tOGD was able to effectively protect neurons and microglial cells from damage, and inhibit the apoptosis caused by oxygen glucose deprivation.

  18. Effect of co-culture with enterocinogenic E. faecium on L. monocytogenes key virulence gene expression

    Directory of Open Access Journals (Sweden)

    Eleftherios H. Drosinos

    2016-08-01

    Full Text Available The aim of the present study was to assess the expression of key virulence genes during co-culture of L. monocytogenes with a bacteriocinogenic E. faecium strain in liquid growth medium. For that purpose, BHI broth was inoculated with 7 log CFU·mL–1 L. monocytogenes and 4, 5 or 6 log CFU·mL–1 E. faecium. Sampling took place after 8 and 24 h of incubation, corresponding to the maximum and minimum of enterocin production, respectively. The RNA was extracted, stabilized and expression of prfA, sigB, hly, plcA, plcB, inlA, inlB, inlC and inlJ, was assessed by RT-qPCR. Most of the genes were downregulated during co-culture at 5 °C. Moreover, a statistically significant effect of the inoculum level was evident in most of the cases. On the contrary, no effect on the transcription level of most of the genes was observed during co-culture at 37 °C.

  19. Prevascularization of 3D printed bone scaffolds by bioactive hydrogels and cell co-culture.

    Science.gov (United States)

    Kuss, Mitchell A; Wu, Shaohua; Wang, Ying; Untrauer, Jason B; Li, Wenlong; Lim, Jung Yul; Duan, Bin

    2017-09-13

    Vascularization is a fundamental prerequisite for large bone construct development and remains one of the main challenges of bone tissue engineering. Our current study presents the combination of 3D printing technique with a hydrogel-based prevascularization strategy to generate prevascularized bone constructs. Human adipose derived mesenchymal stem cells (ADMSC) and human umbilical vein endothelial cells (HUVEC) were encapsulated within our bioactive hydrogels, and the effects of culture conditions on in vitro vascularization were determined. We further generated composite constructs by forming 3D printed polycaprolactone/hydroxyapatite scaffolds coated with cell-laden hydrogels and determined how the co-culture affected vascularization and osteogenesis. It was demonstrated that 3D co-cultured ADMSC-HUVEC generated capillary-like networks within the porous 3D printed scaffold. The co-culture systems promoted in vitro vascularization, but had no significant effects on osteogenesis. The prevascularized constructs were subcutaneously implanted into nude mice to evaluate the in vivo vascularization capacity and the functionality of engineered vessels. The hydrogel systems facilitated microvessel and lumen formation and promoted anastomosis of vascular networks of human origin with host murine vasculature. These findings demonstrate the potential of prevascularized 3D printed scaffolds with anatomical shape for the healing of larger bone defects. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2017. © 2017 Wiley Periodicals, Inc.

  20. A porcine astrocyte/endothelial cell co-culture model of the blood-brain barrier.

    Science.gov (United States)

    Jeliazkova-Mecheva, Valentina V; Bobilya, Dennis J

    2003-10-01

    A method for the isolation of porcine atrocytes as a simple extension of a previously described procedure for isolation of brain capillary endothelial cells from adolescent pigs [Methods Cell Sci. 17 (1995) 2] is described. The obtained astroglial culture purified through two passages and by the method of the selective detachment was validated by a phase contrast microscopy and through an immunofluorescent assay for the glial fibrillary acidic protein (GFAP). Porcine astrocytes were co-cultivated with porcine brain capillary endothelial cells (PBCEC) for the development of an in vitro blood-brain barrier (BBB) model. The model was visualized by an electron microscopy and showed elevated transendothellial electrical resistance and reduced inulin permeability. To our knowledge, this is the first report for the establishment of a porcine astrocyte/endothelial cell co-culture BBB model, which avoids interspecies and age differences between the two cell types, usually encountered in the other reported co-culture BBB models. Considering the availability of the porcine brain tissue and the close physiological and anatomical relation between the human and pig brain, the porcine astrocyte/endothelial cell co-culture system can serve as a reliable and easily reproducible model for different in vitro BBB studies.

  1. Semipermeable Capsules Wrapping a Multifunctional and Self-regulated Co-culture Microenvironment for Osteogenic Differentiation

    Science.gov (United States)

    Correia, Clara R.; Pirraco, Rogério P.; Cerqueira, Mariana T.; Marques, Alexandra P.; Reis, Rui L.; Mano, João F.

    2016-02-01

    A new concept of semipermeable reservoirs containing co-cultures of cells and supporting microparticles is presented, inspired by the multi-phenotypic cellular environment of bone. Based on the deconstruction of the “stem cell niche”, the developed capsules are designed to drive a self-regulated osteogenesis. PLLA microparticles functionalized with collagen I, and a co-culture of adipose stem (ASCs) and endothelial (ECs) cells are immobilized in spherical liquified capsules. The capsules are coated with multilayers of poly(L-lysine), alginate, and chitosan nano-assembled through layer-by-layer. Capsules encapsulating ASCs alone or in a co-culture with ECs are cultured in endothelial medium with or without osteogenic differentiation factors. Results show that osteogenesis is enhanced by the co-encapsulation, which occurs even in the absence of differentiation factors. These findings are supported by an increased ALP activity and matrix mineralization, osteopontin detection, and the up regulation of BMP-2, RUNX2 and BSP. The liquified co-capsules also act as a VEGF and BMP-2 cytokines release system. The proposed liquified capsules might be a valuable injectable self-regulated system for bone regeneration employing highly translational cell sources.

  2. Dynamic CT of hepatocellular carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Fujita, Nobuyuki; Shirato, Hiroki; Shinohara, Masahiro; Miyasaka, Kazuo; Morita, Yutaka; Irie, Goro

    1983-03-01

    We performed dynamic CT in 30 cases of hepatocellular carcinoma, and concluded as below. Detecting the stain in the early phase of the dynamic series, it is possible to make a diagnosis of hepatocellular carcinoma. The dynamic CT is effective in a case of small hepatocellular carcinoma in which it is difficult to gain an accurate diagnosis in the routine CT study. The dynamic CT is also effective in the differential diagnosis of hepatic lesions, as other hepatic lesions such as hemangioma and metastatic liver cancer show different patterns compared with hepatocellular carcinoma.

  3. Dynamic CT of hepatocellular carcinoma

    International Nuclear Information System (INIS)

    Fujita, Nobuyuki; Shirato, Hiroki; Shinohara, Masahiro; Miyasaka, Kazuo; Morita, Yutaka; Irie, Goro

    1983-01-01

    We performed dynamic CT in 30 cases of hepatocellular carcinoma, and concluded as below. 1 Decting the stain in the early phase of the dynamic series, it is possible to make a diagnosis of hepatocellular carcinoma. 2 The dinamic CT is effective in a case of small hepatocellular carcinoma in which it is difficult to gain an accurate diagnosis in the routine CT study. 3 The dynamic CT is also effective in the differential diagnosis of hepatic lesions, as other hepatic lesions such as hemangioma and metastatic liver cancer show different patterns compared with hepatocellular carcinoma. (author)

  4. Micro-patterned Nafion membranes for direct methanol fuel cell applications

    NARCIS (Netherlands)

    Yildirim, M.H.; te Braake, J.; Aran, H.C.; Stamatialis, Dimitrios; Wessling, Matthias

    2010-01-01

    In this work, we report the direct methanol fuel cell (DMFC) performance of micro-patterned (μp) Nafion® 117 (N117) membranes prepared by hot embossing and compare them with that of normal N117 and heat and pressure treated (hp) N117 non-patterned (smooth) membranes. Our results suggest that the

  5. MAPLE deposition of 3D micropatterned polymeric substrates for cell culture

    Energy Technology Data Exchange (ETDEWEB)

    Paun, Irina Alexandra, E-mail: irina.paun@physics.pub.ro [National Institute for Laser, Plasma and Radiation Physics, RO-077125, Magurele, Bucharest (Romania); Faculty of Applied Sciences, University Politehnica of Bucharest, RO-060042, Bucharest (Romania); Mihailescu, Mona [Faculty of Applied Sciences, University Politehnica of Bucharest, RO-060042, Bucharest (Romania); Calenic, Bogdan [Department of Biochemistry, Faculty of Dentistry, UMF Carol Davila, Bucharest (Romania); Luculescu, Catalin Romeo [National Institute for Laser, Plasma and Radiation Physics, RO-077125, Magurele, Bucharest (Romania); Greabu, Maria [Department of Biochemistry, Faculty of Dentistry, UMF Carol Davila, Bucharest (Romania); Dinescu, Maria, E-mail: dinescum@nipne.ro [National Institute for Laser, Plasma and Radiation Physics, RO-077125, Magurele, Bucharest (Romania)

    2013-08-01

    3D micropatterned poly(lactide-co-glycolide)/polyurethane (PLGA/PU) substrates were produced by MAPLE deposition through masks and used for regulating the behavior of oral keratinocyte stem cells in response to topography. Flat PLGA/PU substrates were produced for comparison. 3D imaging of the PLGA/PU substrates and of the cultured cells was performed by Digital Holographic Microscopy. The micropatterns were in the shape of squares of 50 × 50 and 80 × 80 μm{sup 2} areas, ∼1.8 μm in height and separated by 20 μm wide channels. It was found that substrate topography guided the adhesion of the cultured cells: on the smooth substrates the cells adhered randomly and showed no preferred orientation; in contrast, on the micropatterned substrates the cells adhered preferentially onto the squares and not in the separating channels. Furthermore, key properties of the cells (size, viability, proliferation rate and stem cell marker expression) did not show any dependence on substrate topography. The size of the cultured cells, their viability, the proportions of actively/slow proliferating cells, as well as the stem cell markers expressions, were similar for both flat and micropatterned substrates. Finally, it was found that the cells cultured on the PLGA/PU substrates deposited by MAPLE exhibited similar properties as the controls (i.e. cells cultured on glass slides), indicating the capability of the former to preserve the properties of the keratinocyte stem cells.

  6. Continuous droplet removal upon dropwise condensation of humid air on a hydrophobic micropatterned surface.

    Science.gov (United States)

    Zamuruyev, Konstantin O; Bardaweel, Hamzeh K; Carron, Christopher J; Kenyon, Nicholas J; Brand, Oliver; Delplanque, Jean-Pierre; Davis, Cristina E

    2014-08-26

    Combination of two physical phenomena, capillary pressure gradient and wettability gradient, allows a simple two-step fabrication process that yields a reliable hydrophobic self-cleaning condenser surface. The surface is fabricated with specific microscopic topography and further treatment with a chemically inert low-surface-energy material. This process does not require growth of nanofeatures (nanotubes) or hydrophilic-hydrophobic patterning of the surface. Trapezoidal geometry of the microfeatures facilitates droplet transfer from the Wenzel to the Cassie state and reduces droplet critical diameter. The geometry of the micropatterns enhances local coalescence and directional movement for droplets with diameter much smaller than the radial length of the micropatterns. The hydrophobic self-cleaning micropatterned condenser surface prevents liquid film formation and promotes continuous dropwise condensation cycle. Upon dropwise condensation, droplets follow a designed wettability gradient created with micropatterns from the most hydrophobic to the least hydrophobic end of the surface. The surface has higher condensation efficiency, due to its directional self-cleaning property, than a plain hydrophobic surface. We explain the self-actuated droplet collection mechanism on the condenser surface and demonstrate experimentally the creation of an effective wettability gradient over a 6 mm radial distance. In spite of its fabrication simplicity, the fabricated surface demonstrates self-cleaning property, enhanced condensation performance, and reliability over time. Our work enables creation of a hydrophobic condenser surface with the directional self-cleaning property that can be used for collection of biological (chemical, environmental) aerosol samples or for condensation enhancement.

  7. Micropatterned 2D Hybrid Perovskite Thin Films with Enhanced Photoluminescence Lifetimes

    NARCIS (Netherlands)

    Kamminga, Machteld E.; Fang, Hong Hua; Loi, Maria Antonietta; Ten Brink, Gert H.; Blake, Graeme R.; Palstra, Thomas T.M.; Ten Elshof, Johan E.

    2018-01-01

    The application of luminescent materials in display screens and devices requires micropatterned structures. In this work, we have successfully printed microstructures of a two-dimensional (2D), orange-colored organic/inorganic hybrid perovskite ((C6H5CH2NH3)2PbI4) using two different soft

  8. Inkjet printing for direct micropatterning of a superhydrophobic surface: Toward biomimetic fog harvesting surfaces

    KAUST Repository

    Zhang, Lianbin

    2015-01-01

    The preparation of biomimetic superhydrophobic surfaces with hydrophilic micro-sized patterns is highly desirable, but a one-step, mask-free method to produce such surfaces has not previously been reported. We have developed a direct method to produce superhydrophilic micropatterns on superhydrophobic surfaces based on inkjet printing technology. This work was inspired by the efficient fog-harvesting behavior of Stenocara beetles in the Namib Desert. A mussel-inspired ink consisting of an optimized solution of dopamine was applied directly by inkjet printing to superhydrophobic surfaces. Stable Wenzel\\'s microdroplets of the dopamine solution with well-defined micropatterns were obtained on these surfaces. Superhydrophilic micropatterns with well-controlled dimensions were then readily achieved on the superhydrophobic surfaces by the formation of polydopamine via in situ polymerization. The micropatterned superhydrophobic surfaces prepared by this inkjet printing method showed enhanced water collection efficiency compared with uniform superhydrophilic and superhydrophobic surfaces. This method can be used for the facile large-scale patterning of superhydrophobic surfaces with high precision and superior pattern stability and is therefore a key step toward patterning superhydrophobic surfaces for practical applications. This journal is

  9. Reprogramming hMSCs morphology with silicon/porous silicon geometric micro-patterns.

    Science.gov (United States)

    Ynsa, M D; Dang, Z Y; Manso-Silvan, M; Song, J; Azimi, S; Wu, J F; Liang, H D; Torres-Costa, V; Punzon-Quijorna, E; Breese, M B H; Garcia-Ruiz, J P

    2014-04-01

    Geometric micro-patterned surfaces of silicon combined with porous silicon (Si/PSi) have been manufactured to study the behaviour of human Mesenchymal Stem Cells (hMSCs). These micro-patterns consist of regular silicon hexagons surrounded by spaced columns of silicon equilateral triangles separated by PSi. The results show that, at an early culture stage, the hMSCs resemble quiescent cells on the central hexagons with centered nuclei and actin/β-catenin and a microtubules network denoting cell adhesion. After 2 days, hMSCs adapted their morphology and cytoskeleton proteins from cell-cell dominant interactions at the center of the hexagonal surface. This was followed by an intermediate zone with some external actin fibres/β-catenin interactions and an outer zone where the dominant interactions are cell-silicon. Cells move into silicon columns to divide, migrate and communicate. Furthermore, results show that Runx2 and vitamin D receptors, both specific transcription factors for skeleton-derived cells, are expressed in cells grown on micropatterned silicon under all observed circumstances. On the other hand, non-phenotypic alterations are under cell growth and migration on Si/PSi substrates. The former consideration strongly supports the use of micro-patterned silicon surfaces to address pending questions about the mechanisms of human bone biogenesis/pathogenesis and the study of bone scaffolds.

  10. Micropatterning of biomolecules on a glass substrate in fused silica microchannels by using photolabile linker-based surface activation

    International Nuclear Information System (INIS)

    Jang, K.; Mawatari, K.; Kitamori, T.; Xu, Y.; Sato, K.; Tanaka, Y.

    2012-01-01

    We report on a straightforward method for creating micropatterns of multiple biomolecules. The anti-fouling agent 2-methacryloyloxyethyl phosphorylcholine (MPC) polymer and a photolabile linker (PL) were covalently linked to an amino-terminated silane surface. Patterns were generated by selective removal of the MPC polymer via UV irradiation. Multiple micropatterns of fluorescein isothiocyanate (FITC)-labeled bovine serum albumin (BSA) and rhodamine-labeled goat fragment antigen-binding fragments (FAB) were deposited on a same glass substrate. We also employed micropatterning of multiple biomolecules in that Texas red-labeled BSA and FITC-labeled rabbit anti-mouse IgG were placed inside a microchannel. (author)

  11. Micropatterning stretched and aligned DNA for sequence-specific nanolithography

    Science.gov (United States)

    Petit, Cecilia Anna Paulette

    Techniques for fabricating nanostructured materials can be categorized as either "top-down" or "bottom-up". Top-down techniques use lithography and contact printing to create patterned surfaces and microfluidic channels that can corral and organize nanoscale structures, such as molecules and nanorods in contrast; bottom-up techniques use self-assembly or molecular recognition to direct the organization of materials. A central goal in nanotechnology is the integration of bottom-up and top-down assembly strategies for materials development, device design; and process integration. With this goal in mind, we have developed strategies that will allow this integration by using DNA as a template for nanofabrication; two top-down approaches allow the placement of these templates, while the bottom-up technique uses the specific sequence of bases to pattern materials along each strand of DNA. Our first top-down approach, termed combing of molecules in microchannels (COMMIC), produces microscopic patterns of stretched and aligned molecules of DNA on surfaces. This process consists of passing an air-water interface over end adsorbed molecules inside microfabricated channels. The geometry of the microchannel directs the placement of the DNA molecules, while the geometry of the airwater interface directs the local orientation and curvature of the molecules. We developed another top-down strategy for creating micropatterns of stretched and aligned DNA using surface chemistry. Because DNA stretching occurs on hydrophobic surfaces, this technique uses photolithography to pattern vinyl-terminated silanes on glass When these surface-, are immersed in DNA solution, molecules adhere preferentially to the silanized areas. This approach has also proven useful in patterning protein for cell adhesion studies. Finally, we describe the use of these stretched and aligned molecules of DNA as templates for the subsequent bottom-up construction of hetero-structures through hybridization

  12. Viral hepatitis and hepatocellular carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Juei-Low, Sung [ed.; National Taiwan University College of Medicine, Taipei (Republic of China Taiwan). Department of Internal Medicine; Ding-Shinn, Chen [ed.; National Taiwan University College of Medicine, Taipei (Republic of China Taiwan). Hepatitis Research Center National Taiwan University College of Medicine, Taipei (Republic of China Taiwan). Graduate Institute of Clinical Medicine

    1990-01-01

    Two papers in this volume are in INIS scope, respectively dealing with MRI in the study of viral hepatitis and hepatocellular carcinoma, and The use of {sup 131}I-labeled Lipidol in the diagnosis of hepato-cellular carcinoma. (H.W.). refs.; figs.; tabs.

  13. Viral hepatitis and hepatocellular carcinoma

    International Nuclear Information System (INIS)

    Sung Juei-Low; Chen Ding-Shinn

    1990-01-01

    Two papers in this volume are in INIS scope, respectively dealing with MRI in the study of viral hepatitis and hepatocellular carcinoma, and The use of 131 I-labeled Lipidol in the diagnosis of hepato-cellular carcinoma. (H.W.). refs.; figs.; tabs

  14. Co-culture of chondrons and mesenchymal stromal cells reduces the loss of collagen VI and improves extracellular matrix production.

    Science.gov (United States)

    Owida, H A; De Las Heras Ruiz, T; Dhillon, A; Yang, Y; Kuiper, N J

    2017-12-01

    Adult articular chondrocytes are surrounded by a pericellular matrix (PCM) to form a chondron. The PCM is rich in hyaluronan, proteoglycans, and collagen II, and it is the exclusive location of collagen VI in articular cartilage. Collagen VI anchors the chondrocyte to the PCM. It has been suggested that co-culture of chondrons with mesenchymal stromal cells (MSCs) might enhance extracellular matrix (ECM) production. This co-culture study investigates whether MSCs help to preserve the PCM and increase ECM production. Primary bovine chondrons or chondrocytes or rat MSCs were cultured alone to establish a baseline level for ECM production. A xenogeneic co-culture monolayer model using rat MSCs (20, 50, and 80%) was established. PCM maintenance and ECM production were assessed by biochemical assays, immunofluorescence, and histological staining. Co-culture of MSCs with chondrons enhanced ECM matrix production, as compared to chondrocyte or chondron only cultures. The ratio 50:50 co-culture of MSCs and chondrons resulted in the highest increase in GAG production (18.5 ± 0.54 pg/cell at day 1 and 11 ± 0.38 pg/cell at day 7 in 50:50 co-culture versus 16.8 ± 0.61 pg/cell at day 1 and 10 ± 0.45 pg/cell at day 7 in chondron monoculture). The co-culture of MSCs with chondrons appeared to decelerate the loss of the PCM as determined by collagen VI expression, whilst the expression of high-temperature requirement serine protease A1 (HtrA1) demonstrated an inverse relationship to that of the collagen VI. Together, this implies that MSCs directly or indirectly inhibited HtrA1 activity and the co-culture of MSCs with chondrons enhanced ECM synthesis and the preservation of the PCM.

  15. 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.

  16. 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.

  17. Microbial reductive dehalogenation of trihalomethanes by a Dehalobacter-containing co-culture.

    Science.gov (United States)

    Zhao, Siyan; Rogers, Matthew J; He, Jianzhong

    2017-07-01

    Trihalomethanes such as chloroform and bromoform, although well-known as a prominent class of disinfection by-products, are ubiquitously distributed in the environment due to widespread industrial usage in the past decades. Chloroform and bromoform are particularly concerning, of high concentrations detected and with long half-lives up to several hundred days in soils and groundwater. In this study, we report a Dehalobacter- and Desulfovibrio-containing co-culture that exhibits dehalogenation of chloroform (~0.61 mM) to dichloromethane and bromoform (~0.67 mM) to dibromomethane within 10-15 days. This co-culture was further found to dechlorinate 1,1,1-trichloroethane (1,1,1-TCA) (~0.65 mM) to 1,1-dichloroethane within 12 days. The Dehalobacter species present in this co-culture, designated Dehalobacter sp. THM1, was found to couple growth with dehalogenation of chloroform, bromoform, and 1,1,1-TCA. Strain THM1 harbors a newly identified reductive dehalogenase (RDase), ThmA, which catalyzes chloroform, bromoform, and 1,1,1-TCA dehalogenation. Additionally, based on the sequences of thmA and other identified chloroform RDase genes, ctrA, cfrA, and tmrA, a pair of chloroform RDase gene-specific primers were designed and successfully applied to investigate the chloroform dechlorinating potential of microbial communities. The comparative analysis of chloroform RDases with tetrachloroethene RDases suggests a possible approach in predicting the substrate specificity of uncharacterized RDases in the future.

  18. Assessment of growth factor treatment on fibrochondrocyte and chondrocyte co-cultures for TMJ fibrocartilage engineering.

    Science.gov (United States)

    Kalpakci, Kerem N; Kim, Eric J; Athanasiou, Kyriacos A

    2011-04-01

    Treatments for patients suffering from severe temporomandibular joint (TMJ) dysfunction are limited, motivating the development of strategies for tissue regeneration. In this study, co-cultures of fibrochondrocytes (FCs) and articular chondrocytes (ACs) were seeded in agarose wells, and supplemented with growth factors, to engineer tissue with biomechanical properties and extracellular matrix composition similar to native TMJ fibrocartilage. In the first phase, growth factors were applied alone and in combination, in the presence or absence of serum, while in the second phase, the best overall treatment was applied at intermittent dosing. Continuous treatment of AC/FC co-cultures with TGF-β1 in serum-free medium resulted in constructs with glycosaminoglycan/wet weight ratios (12.2%), instantaneous compressive moduli (790 kPa), relaxed compressive moduli (120 kPa) and Young's moduli (1.87 MPa) that overlap with native TMJ disc values. Among co-culture groups, TGF-β1 treatment increased collagen deposition ∼20%, compressive stiffness ∼130% and Young's modulus ∼170% relative to controls without growth factor. Serum supplementation, though generally detrimental to functional properties, was identified as a powerful mediator of FC construct morphology. Finally, both intermittent and continuous TGF-β1 treatment showed positive effects, though continuous treatment resulted in greater enhancement of construct functional properties. This work proposes a strategy for regeneration of TMJ fibrocartilage and its future application will be realized through translation of these findings to clinically viable cell sources. Copyright © 2011 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  19. Review of vascularised bone tissue-engineering strategies with a focus on co-culture systems.

    Science.gov (United States)

    Liu, Yuchun; Chan, Jerry K Y; Teoh, Swee-Hin

    2015-02-01

    Poor angiogenesis within tissue-engineered grafts has been identified as a main challenge limiting the clinical introduction of bone tissue-engineering (BTE) approaches for the repair of large bone defects. Thick BTE grafts often exhibit poor cellular viability particularly at the core, leading to graft failure and lack of integration with host tissues. Various BTE approaches have been explored for improving vascularisation in tissue-engineered constructs and are briefly discussed in this review. Recent investigations relating to co-culture systems of endothelial and osteoblast-like cells have shown evidence of BTE efficacy in increasing vascularization in thick constructs. This review provides an overview of key concepts related to bone formation and then focuses on the current state of engineered vascularized co-culture systems using bone repair as a model. It will also address key questions regarding the generation of clinically relevant vascularized bone constructs as well as potential directions and considerations for research with the objective of pursuing engineered co-culture systems in other disciplines of vascularized regenerative medicine. The final objective is to generate serious and functional long-lasting vessels for sustainable angiogenesis that will enable enhanced cellular survival within thick voluminous bone grafts, thereby aiding in bone formation and remodelling in the long term. However, more evidence about the quality of blood vessels formed and its associated functional improvement in bone formation as well as a mechanistic understanding of their interactions are necessary for designing better therapeutic strategies for translation to clinical settings. Copyright © 2012 John Wiley & Sons, Ltd.

  20. Allogeneic human dermal fibroblasts are viable in peripheral blood mononuclear co-culture

    Directory of Open Access Journals (Sweden)

    Restu Syamsul Hadi

    2014-08-01

    Full Text Available Background Transplanted allogeneic dermal fibroblasts retain stem cell subpopulations, and are easily isolated, expanded and stored using standard techniques. Their potential for regenerative therapy of chronic wounds should be evaluated. The aim of this study was to determine allogeneic fibroblast viability in the presence of peripheral blood mononuclear cells (PBMC. Methods In this experimental study, fibroblasts were isolated from foreskin explants, expanded in the presence of serum, and stored using slow-freezing. We used one intervention group of allogeneic fibroblasts co-cultured with PBMC and 2 control groups of separate fibroblast and PBMC cultures.Fibroblasts were characterized by their collagen secretion and octamer-binding transcription factor 4 (OCT4 expression. Viability was evaluated using water soluble tetrazolium-1 (WST-1 proliferation assay. Absorbances were measured at 450 nm. Data analysis was performed by student’s paired t-test. Results Dermal fibroblasts were shown to secrete collagen, express OCT4, be recoverable after cryopreservation, and become attached to the culture dish in a co-culture with PBMC. Co-cultured and control fibroblasts had no significantly different cell viabilities (p>0.05. Calculated viable cell numbers increased 1.8 and 5.1-fold, respectively, at days 2 and 4 in vitro. Both groups showed comparable doubling times at days 2 and 4 in vitro. PBMC did not interfere with allogeneic fibroblast viability and proliferative capacity Conclusions Allogeneic fibroblasts remain viable and proliferate in the presence of host PBMC. Future research should evaluate allogeneic human dermal fibroblast competency in clinical settings. Dermal fibroblasts are a potential source for cell therapy in chronic wound management.

  1. Effects of silver nanoparticles and ions on a co-culture model for the gastrointestinal epithelium.

    Science.gov (United States)

    Georgantzopoulou, Anastasia; Serchi, Tommaso; Cambier, Sébastien; Leclercq, Céline C; Renaut, Jenny; Shao, Jia; Kruszewski, Marcin; Lentzen, Esther; Grysan, Patrick; Eswara, Santhana; Audinot, Jean-Nicolas; Contal, Servane; Ziebel, Johanna; Guignard, Cédric; Hoffmann, Lucien; Murk, AlberTinka J; Gutleb, Arno C

    2016-02-17

    The increased incorporation of silver nanoparticles (Ag NPs) into consumer products makes the characterization of potential risk for humans and other organisms essential. The oral route is an important uptake route for NPs, therefore the study of the gastrointestinal tract in respect to NP uptake and toxicity is very timely. The aim of the present study was to evaluate the effects of Ag NPs and ions on a Caco-2/TC7:HT29-MTX intestinal co-culture model with mucus secretion, which constitutes an important protective barrier to exogenous agents in vivo and may strongly influence particle uptake. The presence of the mucus layer was confirmed with staining techniques (alcian blue and toluidine blue). Mono and co-cultures of Caco-2/TC7 and HT29-MTX cells were exposed to Ag NPs (Ag 20 and 200 nm) and AgNO3 and viability (alamar blue), ROS induction (DCFH-DA assay) and IL-8 release (ELISA) were measured. The particle agglomeration in the media was evaluated with DLS and the ion release with ultrafiltration and ICP-MS. The effects of the Ag NPs and AgNO3 on cells in co-culture were studied at a proteome level with two-dimensional difference in gel electrophoresis (2D-DIGE) followed by Matrix Assisted Laser Desorption Ionization - Time Of Flight/ Time Of Flight (MALDI-TOF/TOF) mass spectrometry (MS). Intracellular localization was assessed with NanoSIMS and TEM. The presence of mucus layer led to protection against ROS and decrease in IL-8 release. Both Ag 20 and 200 nm NPs were taken up by the cells and Ag NPs 20 nm were mainly localized in organelles with high sulfur content. A dose- and size-dependent increase in IL-8 release was observed with a lack of cytotoxicity and oxidative stress. Sixty one differentially abundant proteins were identified involved in cytoskeleton arrangement and cell cycle, oxidative stress, apoptosis, metabolism/detoxification and stress. The presence of mucus layer had an impact on modulating the induced toxicity of NPs. NP-specific effects were

  2. Dopaminergic differentiation of human neural stem cells mediated by co-cultured rat striatal brain slices

    DEFF Research Database (Denmark)

    Anwar, Mohammad Raffaqat; Andreasen, Christian Maaløv; Lippert, Solvej Kølvraa

    2008-01-01

    differentiation, we co-cultured cells from a human neural forebrain-derived stem cell line (hNS1) with rat striatal brain slices. In brief, coronal slices of neonatal rat striatum were cultured on semiporous membrane inserts placed in six-well trays overlying monolayers of hNS1 cells. After 12 days of co......Properly committed neural stem cells constitute a promising source of cells for transplantation in Parkinson's disease, but a protocol for controlled dopaminergic differentiation is not yet available. To establish a setting for identification of secreted neural compounds promoting dopaminergic...

  3. A New In Vitro Co-Culture Model Using Magnetic Force-Based Nanotechnology.

    Science.gov (United States)

    Takanari, Hiroki; Miwa, Keiko; Fu, XianMing; Nakai, Junichi; Ito, Akira; Ino, Kousuke; Honda, Hiroyuki; Tonomura, Wataru; Konishi, Satoshi; Opthof, Tobias; van der Heyden, Marcel Ag; Kodama, Itsuo; Lee, Jong-Kook

    2016-10-01

    Skeletal myoblast (SkMB) transplantation has been conducted as a therapeutic strategy for severe heart failure. However, arrhythmogenicity following transplantation remains unsolved. We developed an in vitro model of myoblast transplantation with "patterned" or "randomly-mixed" co-culture of SkMBs and cardiomyocytes enabling subsequent electrophysiological, and arrhythmogenic evaluation. SkMBs were magnetically labeled with magnetite nanoparticles and co-cultured with neonatal rat ventricular myocytes (NRVMs) on multi-electrode arrays. SkMBs were patterned by a magnet beneath the arrays. Excitation synchronicity was evaluated by Ca(2+) imaging using a gene-encoded Ca(2+) indicator, G-CaMP2. In the monoculture of NRVMs (control), conduction was well-organized. In the randomly-mixed co-culture of NRVMs and SkMBs (random group), there was inhomogeneous conduction from multiple origins. In the "patterned" co-culture where an en bloc SKMB-layer was inserted into the NRVM-layer, excitation homogenously propagated although conduction was distorted by the SkMB-area. The 4-mm distance conduction time (CT) in the random group was significantly longer (197 ± 126 ms) than in control (17 ± 3 ms). In the patterned group, CT through NRVM-area did not change (25 ± 3 ms), although CT through the SkMB-area was significantly longer (132 ± 77 ms). The intervals between spontaneous excitation varied beat-to-beat in the random group, while regular beating was recorded in the control and patterned groups. Synchronized Ca(2+) transients of NRVMs were observed in the patterned group, whereas those in the random group were asynchronous. Patterned alignment of SkMBs is feasible with magnetic nanoparticles. Using the novel in vitro model mimicking cell transplantation, it may become possible to predict arrhythmogenicity due to heterogenous cell transplantation. J. Cell. Physiol. 231: 2249-2256, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  4. Improving the Yield of Glucoamylase and α-amylase in Solid-state Co-culture

    Directory of Open Access Journals (Sweden)

    Takefuji Haruka

    2016-01-01

    Full Text Available Steamed rice inoculated with Aspergillus oryzae, called Koji, is an essential ingredient for making amazake or brewing sake. However, A. oryzae usually offers low enzyme yield, especially in case of glucoamylase. Co-culture of Aspergillus and Rhizopus strains in Koji increased glucoamylase and α-amylase activities. The ratio of initial spore counts of A. oryzae and R. oryzae influenced the activity of amylolytic enzymes. When this ratio was 1:1, α-amylase showed maximum activity (573 U/g-substrate, 95 h, and when this ratio was 200:1, glucoamylase showed maximum activity (180 U/g-substrate, 95 h.

  5. Improving the Yield of Glucoamylase and α-amylase in Solid-state Co-culture

    OpenAIRE

    Takefuji Haruka; Ninomiya Junko; Morita Hiroshi

    2016-01-01

    Steamed rice inoculated with Aspergillus oryzae, called Koji, is an essential ingredient for making amazake or brewing sake. However, A. oryzae usually offers low enzyme yield, especially in case of glucoamylase. Co-culture of Aspergillus and Rhizopus strains in Koji increased glucoamylase and α-amylase activities. The ratio of initial spore counts of A. oryzae and R. oryzae influenced the activity of amylolytic enzymes. When this ratio was 1:1, α-amylase showed maximum activity (573 U/g-subs...

  6. Profuse and Selective Growth in vitro of Rat Spinal Axons on a Micro-Patterned Poly (ethylene imine) Grid

    National Research Council Canada - National Science Library

    Mouveroux, J

    2001-01-01

    To distribute neurites (axons) along a surface and to guide them towards specific point targets we cultured spinal cord explants on coverslips printed with a micro-patterned grid of poly (ethylene imine) (PEI) lanes...

  7. 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

  8. Micro-patterned ZnO semiconductors for high performance thin film transistors via chemical imprinting with a PDMS stamp.

    Science.gov (United States)

    Seong, Kieun; Kim, Kyongjun; Park, Si Yun; Kim, Youn Sang

    2013-04-07

    Chemical imprinting was conducted on ZnO semiconductor films via a chemical reaction at the contact regions between a micro-patterned PDMS stamp and ZnO films. In addition, we applied the chemical imprinting on Li doped ZnO thin films for high performance TFTs fabrication. The representative micro-patterned Li doped ZnO TFTs showed a field effect mobility of 4.2 cm(2) V(-1) s(-1) after sintering at 300 °C.

  9. Ultrasound manifestation of hepatocellular carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Hwang, M S; Yoo, H S; Park, C Y; Choi, H J; Moon, Y M; Lee, S I [Yonsei University College of Medicine, Seoul (Korea, Republic of)

    1982-06-15

    With the advent of gray scale ultrasonographic equipment, the parenchymal disease of liver is more easily evaluated. Ultrasonography is a non-invasive technique, different from angiography, and performed without discomfort to patient. And also ultrasonography can be used in assessing the liver in cases showing equivocal scintigraphy and in differentiation of solid and cystic masses, first detected on scintigrams. Therefore, the complementary use of ultrasonography, Tc-99m-sulfur colloid scan and angiography provides better diagnostic accuracy for the detection of hepatocellular carcinoma, and moreover, sequential ultrasonographic studies in the same patient are valuable of following the course of hepatocellular carcinoma and monitoring the effectiveness of therapy for hepatocellular carcinoma. In thirty patients with histologically proven hepatocellular carcinoma, an analysis of ultrasound manifestation is made and the results are as follows; 1. Ultrasound manifestation of hepatocellular carcinoma by gray scale showed four different sonographic patterns including discrete echo free, discrete echogenic, ill defined echogenic and mixed patterns. 2. The size of hepatocellular carcinoma by ultrasonographic measurement was larger than 5 cm in diameter in 28 cases. 3. In 7 cases performed with angiography, all echogenicities of hepatocellualr carcinoma were correlated with the findings of vascularity of angiography. 4. In cases combined with liver cirrhosis, the sonographic pattern of hepatocellular carcinoma appeared to be discrete or ill defined echogenic patterns.

  10. Ultrasound manifestation of hepatocellular carcinoma

    International Nuclear Information System (INIS)

    Hwang, M. S.; Yoo, H. S.; Park, C. Y.; Choi, H. J.; Moon, Y. M.; Lee, S. I.

    1982-01-01

    With the advent of gray scale ultrasonographic equipment, the parenchymal disease of liver is more easily evaluated. Ultrasonography is a non-invasive technique, different from angiography, and performed without discomfort to patient. And also ultrasonography can be used in assessing the liver in cases showing equivocal scintigraphy and in differentiation of solid and cystic masses, first detected on scintigrams. Therefore, the complementary use of ultrasonography, Tc-99m-sulfur colloid scan and angiography provides better diagnostic accuracy for the detection of hepatocellular carcinoma, and moreover, sequential ultrasonographic studies in the same patient are valuable of following the course of hepatocellular carcinoma and monitoring the effectiveness of therapy for hepatocellular carcinoma. In thirty patients with histologically proven hepatocellular carcinoma, an analysis of ultrasound manifestation is made and the results are as follows; 1. Ultrasound manifestation of hepatocellular carcinoma by gray scale showed four different sonographic patterns including discrete echo free, discrete echogenic, ill defined echogenic and mixed patterns. 2. The size of hepatocellular carcinoma by ultrasonographic measurement was larger than 5 cm in diameter in 28 cases. 3. In 7 cases performed with angiography, all echogenicities of hepatocellualr carcinoma were correlated with the findings of vascularity of angiography. 4. In cases combined with liver cirrhosis, the sonographic pattern of hepatocellular carcinoma appeared to be discrete or ill defined echogenic patterns

  11. Effects of Electrical Stimulation in Sympathetic Neuron-Cardiomyocyte Co-cultures

    Science.gov (United States)

    Takeuchi, Akimasa; Tani, Hiromasa; Mori, Masahide; Moriguchi, Hiroyuki; Kotani, Kiyoshi; Lee, Jong-Kook; Noshiro, Makoto; Jimbo, Yasuhiko

    The sympathetic nervous system is one of the principal sources for regulating cardiovascular functions. Little is known, however, about the network-level interactions between sympathetic neurons and cardiomyocytes. In this study, a semi-separated co-culture system of superior cervical ganglion (SCG) neurons and ventricular myocytes (VMs) was developed by using a polydimethylsyloxane (PDMS) chamber placed on a microelectrode-array (MEA) substrate. Neurites of SCG neurons passed through a conduit of the chamber and reached VMs. Evoked activities of SCG neurons were observed from several electrodes immediately after applying constant-voltage stimulation (1 V, 1 ms, biphasic square pulses) to SCG neurons by using 32 electrodes. Furthermore, this stimulation was applied to SCG neurons at the frequency of 1, 5 and 10 Hz. After applying these three kinds of stimulations, mean minute contraction rate of VMs increased with an increase in the frequency of stimulation. These results suggest that changes in contraction rate of VMs after applying electrical stimulations to SCG neurons depend on frequencies of these stimulations and that the heart-regulating mechanisms as well as that in the body were formed in this co-culture system.

  12. Stimulating effect of gamma radiation on haploid wheat production through microscope over co-culture system

    International Nuclear Information System (INIS)

    Naseri, M.; Rahimi, M.; Faramarz, M.

    2004-01-01

    Haploid production focuses on low plant regeneration in some wheat genotypes. Haploid application gamma ray as an electromagnetic ray has ionizing properties which can produce ions when passing through biological matter. It can produce genetic variation therefore, is applied in crop and ornamental improvement to enhance agronomic traits. The most important changes caused by gamma radiation is in DNA structure existing in the nucleus of cell. The desirable agronomic changes then will be passed on through generations. Another property of gamma ray can be it's stimulating effect which is the aim in this investigation. Microspore-overy co-culture of wheat along with application of low doses of gamma radiation 2,3 and 4 Gy as absorbing doses were implemented with the aim to evaluate wheat haploid production. Modified Morashig and Skoog medium was used as induction medium and 190-2 medium for regeneration. Two winter and two spring wheat cultivars were used as genetic material. Low doses of gamma radiation simulated microspore cell division and produced more calli relative to non-irradiated microspores only in winter type wheats. In microspore overy co-culture, filtered microspores were centrifuged and then plated in Petri dishes containing MMS+500 mg/I glutamine with 25 overies

  13. CT of hepatocellular carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Nakamura, H; Tanaka, T; Sai, H; Kawamoto, S; Morimoto, K [Osaka Univ. (Japan). Faculty of Medicine

    1982-06-01

    CT was investigated in 125 cases of hepatocelluar carcinoma and 47 cases of metastatic hepatic neoplasm. The entire contour of each tumor was traced and the average CT value in the tumor was estimated. As a result, the CT value for hepatocellular carcinoma tended to be higher on plain CT and also after contrast enhancement. The CT findings seen frequently were as follows: capsule in 76 cases (60.8%) and septum in 67 cases (53.6%); tumor thrombus in portal vein in 39 cases (31.2%) and that in inferior vena cava in 3 cases (2.4%); localized enlargement of hepatic bile duct in 24 cases (19.2%). These findings were rarely seen in the cases of metastatic hepatic neoplasm. As a relatively outstanding feature of hepatic metastases, a double contour, like concentric circles or contour lines, with a relatively large inner circle or contour line, was found in 21 cases (44.7%). By paying attention to the change of CT value on contrast enhancement and the characteristic image of each case, hepatocellular carcinoma could be differentiated from metastatic hepatic neoplasm with high probability.

  14. Histopathology of hepatocellular carcinoma.

    Science.gov (United States)

    Schlageter, Manuel; Terracciano, Luigi Maria; D'Angelo, Salvatore; Sorrentino, Paolo

    2014-11-21

    Hepatocellular carcinoma (HCC) is currently the sixth most common type of cancer with a high mortality rate and an increasing incidence worldwide. Its etiology is usually linked to environmental, dietary or life-style factors. HCC most commonly arises in a cirrhotic liver but interestingly an increasing proportion of HCCs develop in the non-fibrotic or minimal fibrotic liver and a shift in the underlying etiology can be observed. Although this process is yet to be completely understood, this changing scenario also has impact on the material seen by pathologists, presenting them with new diagnostic dilemmas. Histopathologic criteria for diagnosing classical, progressed HCC are well established and known, but with an increase in detection of small and early HCCs due to routine screening programs, the diagnosis of these small lesions in core needle biopsies poses a difficult challenge. These lesions can be far more difficult to distinguish from one another than progressed HCC, which is usually a clear cut hematoxylin and eosin diagnosis. Furthermore lesions thought to derive from progenitor cells have recently been reclassified in the WHO. This review summarizes recent developments and tries to put new HCC biomarkers in context with the WHOs reclassification. Furthermore it also addresses the group of tumors known as combined hepatocellular-cholangiocellular carcinomas.

  15. Human pericyte-endothelial cell interactions in co-culture models mimicking the diabetic retinal microvascular environment.

    Science.gov (United States)

    Tarallo, Sonia; Beltramo, Elena; Berrone, Elena; Porta, Massimo

    2012-12-01

    Pericytes regulate vascular tone, perfusion pressure and endothelial cell (EC) proliferation in capillaries. Thiamine and benfotiamine counteract high glucose-induced damage in vascular cells. We standardized two human retinal pericyte (HRP)/EC co-culture models to mimic the diabetic retinal microvascular environment. We aimed at evaluating the interactions between co-cultured HRP and EC in terms of proliferation/apoptosis and the possible protective role of thiamine and benfotiamine against high glucose-induced damage. EC and HRP were co-cultured in physiological glucose and stable or intermittent high glucose, with or without thiamine/benfotiamine. No-contact model: EC were plated on a porous membrane suspended into the medium and HRP on the bottom of the same well. Cell-to-cell contact model: EC and HRP were plated on the opposite sides of the same membrane. Proliferation (cell counts and DNA synthesis), apoptosis and tubule formation in Matrigel were assessed. In the no-contact model, stable high glucose reduced proliferation of co-cultured EC/HRP and EC alone and increased co-cultured EC/HRP apoptosis. In the contact model, both stable and intermittent high glucose reduced co-cultured EC/HRP proliferation and increased apoptosis. Stable high glucose had no effects on HRP in separate cultures. Both EC and HRP proliferated better when co-cultured. Thiamine and benfotiamine reversed high glucose-induced damage in all cases. HRP are sensitive to soluble factors released by EC when cultured in high glucose conditions, as suggested by conditioned media assays. In the Matrigel models, addition of thiamine and benfotiamine re-established the high glucose-damaged interactions between EC/HRP and stabilized microtubules.

  16. A facile micropatterning method for a highly flexible PEDOT:PSS on SU-8

    KAUST Repository

    Cho, Nam Chul; Diekhans, Justin; Steward, Malia; Bakr, Osman; Choi, Seungkeun

    2016-01-01

    We report the micropatterning of conducting polymer on the epoxy-based photoresist to demonstrate fully organic, conducting and flexible electrodes. We show that polystyrene sulfonic acid can be covalently linked to the surface of the photoresist (SU-8) by forming sulfonyl ester at the interfaces. We also present an application of the patterned PEDOT:PSS (poly(3,4-ethylenedioxythiophene) polystyrene sulfonate)/SU-8 to the electroplating of metal electrodes. © 2016 Elsevier B.V.

  17. Scalable fabrication of strongly textured organic semiconductor micropatterns by capillary force lithography.

    Science.gov (United States)

    Jo, Pil Sung; Vailionis, Arturas; Park, Young Min; Salleo, Alberto

    2012-06-26

    Strongly textured organic semiconductor micropatterns made of the small molecule dioctylbenzothienobenzothiophene (C(8)-BTBT) are fabricated by using a method based on capillary force lithography (CFL). This technique provides the C(8)-BTBT solution with nucleation sites for directional growth, and can be used as a scalable way to produce high quality crystalline arrays in desired regions of a substrate for OFET applications. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. A facile micropatterning method for a highly flexible PEDOT:PSS on SU-8

    KAUST Repository

    Cho, Nam Chul

    2016-04-17

    We report the micropatterning of conducting polymer on the epoxy-based photoresist to demonstrate fully organic, conducting and flexible electrodes. We show that polystyrene sulfonic acid can be covalently linked to the surface of the photoresist (SU-8) by forming sulfonyl ester at the interfaces. We also present an application of the patterned PEDOT:PSS (poly(3,4-ethylenedioxythiophene) polystyrene sulfonate)/SU-8 to the electroplating of metal electrodes. © 2016 Elsevier B.V.

  19. Novel co-culture plate enables growth dynamic-based assessment of contact-independent microbial interactions.

    Directory of Open Access Journals (Sweden)

    Thomas J Moutinho

    Full Text Available Interactions between microbes are central to the dynamics of microbial communities. Understanding these interactions is essential for the characterization of communities, yet challenging to accomplish in practice. There are limited available tools for characterizing diffusion-mediated, contact-independent microbial interactions. A practical and widely implemented technique in such characterization involves the simultaneous co-culture of distinct bacterial species and subsequent analysis of relative abundance in the total population. However, distinguishing between species can be logistically challenging. In this paper, we present a low-cost, vertical membrane, co-culture plate to quantify contact-independent interactions between distinct bacterial populations in co-culture via real-time optical density measurements. These measurements can be used to facilitate the analysis of the interaction between microbes that are physically separated by a semipermeable membrane yet able to exchange diffusible molecules. We show that diffusion across the membrane occurs at a sufficient rate to enable effective interaction between physically separate cultures. Two bacterial species commonly found in the cystic fibrotic lung, Pseudomonas aeruginosa and Burkholderia cenocepacia, were co-cultured to demonstrate how this plate may be implemented to study microbial interactions. We have demonstrated that this novel co-culture device is able to reliably generate real-time measurements of optical density data that can be used to characterize interactions between microbial species.

  20. Glia co-culture with neurons in microfluidic platforms promotes the formation and stabilization of synaptic contacts.

    Science.gov (United States)

    Shi, Mingjian; Majumdar, Devi; Gao, Yandong; Brewer, Bryson M; Goodwin, Cody R; McLean, John A; Li, Deyu; Webb, Donna J

    2013-08-07

    Two novel microfluidic cell culture schemes, a vertically-layered set-up and a four chamber set-up, were developed for co-culturing central nervous system (CNS) neurons and glia. The cell chambers in these devices were separated by pressure-enabled valve barriers, which permitted us to control communication between the two cell types. The unique design of these devices facilitated the co-culture of glia with neurons in close proximity (∼50-100 μm), differential transfection of neuronal populations, and dynamic visualization of neuronal interactions, such as the development of synapses. With these co-culture devices, initial synaptic contact between neurons transfected with different fluorescent markers, such as green fluorescent protein (GFP) and mCherry-synaptophysin, was imaged using high-resolution fluorescence microscopy. The presence of glial cells had a profound influence on synapses by increasing the number and stability of synaptic contacts. Interestingly, as determined by liquid chromatography-ion mobility-mass spectrometry, neuron-glia co-cultures produced elevated levels of soluble factors compared to that secreted by individual neuron or glia cultures, suggesting a potential mechanism by which neuron-glia interactions could modulate synaptic function. Collectively, these results show that communication between neurons and glia is critical for the formation and stability of synapses and point to the importance of developing neuron-glia co-culture systems such as the microfluidic platforms described in this study.

  1. 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.

  2. Ureolytic/Non-Ureolytic Bacteria Co-Cultured Self-Healing Agent for Cementitious Materials Crack Repair

    Directory of Open Access Journals (Sweden)

    Hyeong Min Son

    2018-05-01

    Full Text Available The present study investigated the CaCO3 precipitation performance of ureolytic and non-ureolytic bacteria co-cultured as a self-healing agent for cementitious materials crack repair. Three different inoculum ratios of ureolytic Sporosarcina pasteurii and non-ureolytic Bacillus thuringiensis (10:0, 8:2, or 5:5 were used. The effect of coculturing ureolytic and non-ureolytic bacteria on microbial metabolism was investigated by measuring the rate of growth in urea-containing medium and the rate of NH4+ and CaCO3 production in urea–calcium lactate medium. The self-healing efficiency of co-cultured bacteria was examined by exposing cement mortar specimens with predefined cracks to media containing single urease-producing or co-cultured bacteria. The obtained results provide new findings, where CaCO3 precipitation is improved by co-culturing ureolytic and non-ureolytic bacteria, owing to the relatively faster growth rate of non-ureolytic bacteria. The crack filling rate correlated with the width of crack, in particular, specimens with a smaller crack width showed the faster filling effect, indicating that the crack width can be a dominant factor influencing the CaCO3 precipitation capacity of co-cultured bacteria.

  3. Discovery of novel xylosides in co-culture of basidiomycetes Trametes versicolor and Ganoderma applanatum by integrated metabolomics and bioinformatics

    Science.gov (United States)

    Yao, Lu; Zhu, Li-Ping; Xu, Xiao-Yan; Tan, Ling-Ling; Sadilek, Martin; Fan, Huan; Hu, Bo; Shen, Xiao-Ting; Yang, Jie; Qiao, Bin; Yang, Song

    2016-09-01

    Transcriptomic analysis of cultured fungi suggests that many genes for secondary metabolite synthesis are presumably silent under standard laboratory condition. In order to investigate the expression of silent genes in symbiotic systems, 136 fungi-fungi symbiotic systems were built up by co-culturing seventeen basidiomycetes, among which the co-culture of Trametes versicolor and Ganoderma applanatum demonstrated the strongest coloration of confrontation zones. Metabolomics study of this co-culture discovered that sixty-two features were either newly synthesized or highly produced in the co-culture compared with individual cultures. Molecular network analysis highlighted a subnetwork including two novel xylosides (compounds 2 and 3). Compound 2 was further identified as N-(4-methoxyphenyl)formamide 2-O-β-D-xyloside and was revealed to have the potential to enhance the cell viability of human immortalized bronchial epithelial cell line of Beas-2B. Moreover, bioinformatics and transcriptional analysis of T. versicolor revealed a potential candidate gene (GI: 636605689) encoding xylosyltransferases for xylosylation. Additionally, 3-phenyllactic acid and orsellinic acid were detected for the first time in G. applanatum, which may be ascribed to response against T.versicolor stress. In general, the described co-culture platform provides a powerful tool to discover novel metabolites and help gain insights into the mechanism of silent gene activation in fungal defense.

  4. β-Carotene from Yeasts Enhances Laccase Production of Pleurotus eryngii var. ferulae in Co-culture.

    Science.gov (United States)

    Guo, Chaolin; Zhao, Liting; Wang, Feng; Lu, Jian; Ding, Zhongyang; Shi, Guiyang

    2017-01-01

    Laccase is widely used in several industrial applications and co-culture is a common method for enhancing laccase production in submerged fermentation. In this study, the co-culture of four yeasts with Pleurotus eryngii var. ferulae was found to enhance laccase production. An analysis of sterilization temperatures and extraction conditions revealed that the stimulatory compound in yeasts was temperature-sensitive, and that it was fat-soluble. An LC-MS analysis revealed that the possible stimulatory compound for laccase production in the four yeast extracts was β-carotene. Moreover, the addition of 4 mg β-carotene to 150 mL of P. eryngii var. ferulae culture broth improved laccase production by 2.2-fold compared with the control (i.e., a monoculture), and was similar to laccase production in co-culture. In addition, the enhanced laccase production was accompanied by an increase of lac gene transcription, which was 6.2-time higher than the control on the fifth day. Therefore, it was concluded that β-carotene from the co-cultured yeasts enhanced laccase production in P. eryngii var. ferulae , and strains that produce β-carotene could be selected to enhance fungal laccase production in a co-culture. Alternatively, β-carotene or crude extracts of β-carotene could be used to induce high laccase production in large scale.

  5. Morphologic Subtypes of Hepatocellular Carcinoma.

    Science.gov (United States)

    Torbenson, Michael S

    2017-06-01

    Hepatocellular carcinomas can be further divided into distinct subtypes that provide important clinical information and biological insights. These subtypes are distinct from growth patterns and are on based on morphologic and molecular findings. There are 12 reasonably well-defined subtypes as well as 6 provisional subtypes, together making up 35% of all hepatocellular carcinomas. These subtypes are discussed, with an emphasis on their definitions and the key morphologic findings. Copyright © 2017 Elsevier Inc. All rights reserved.

  6. Facile approach to the fabrication of a micropattern possessing nanoscale substructure.

    Science.gov (United States)

    Ji, Qiang; Jiang, Xuesong; Yin, Jie

    2007-12-04

    On the basis of the combined technologies of photolithography and reaction-induced phase separation (RIPS), a facile approach has been successfully developed for the fabrication of a micropattern possessing nanoscale substructure on the thin film surface. This approach involves three steps. In the first step, a thin film was prepared by spin coating from a solution of a commercial random copolymer, polystyrene-r-poly(methyl methacrylate) (PS-r-PMMA) and a commercial crosslinker, trimethylolpropane triacrylate (TMPTA). In the second step, photolithograph was performed with the thin film using a 250 W high-pressure mercury lamp to produce the micropattern. Finally, the resulting micropattern was annealed at 200 degrees C for a certain time, and reaction-induced phase separation occurred. After soaking in chloroform for 4 h, nanoscale substructure was obtained. The whole processes were traced by atomic force microscopy (AFM), X-ray photoelectron spectrometry (XPS), and Fourier transform infrared (FTIR) spectroscopy, and the results supported the proposed structure.

  7. Effect of micropatterning induced surface hydrophobicity on drug release from electrospun cellulose acetate nanofibers

    Science.gov (United States)

    Adepu, Shivakalyani; Gaydhane, Mrunalini K.; Kakunuri, Manohar; Sharma, Chandra S.; Khandelwal, Mudrika; Eichhorn, Stephen J.

    2017-12-01

    Sustained release and prevention of burst release for low half-life drugs like Diclofenac sodium is crucial to prevent drug related toxicity. Electrospun nanofibers have emerged recently as potential carrier materials for controlled and sustained drug release. Here, we present a facile method to prevent burst release by tuning the surface wettability through template assisted micropatterning of drug loaded electrospun cellulose acetate (CA) nanofibers. A known amount of drug (Diclofenac sodium) was first mixed with CA and then electrospun in the form of a nanofabric. This as-spun network was hydrophilic in nature. However, when electrospinning was carried out through non-conducting templates, viz nylon meshes with 50 and 100 μm size openings, two kinds of hydrophobic micro-patterned CA nanofabrics were produced. In vitro transdermal testing of our nanofibrous mats was carried out; these tests were able to show that it would be possible to create a patch for transdermal drug release. Further, our results show that with optimized micro-patterned dimensions, a zero order sustained drug release of up to 12 h may be achieved for the transdermal system when compared to non-patterned samples. This patterning caused a change in the surface wettability, to a hydrophobic surface, resulting in a controlled diffusion of the hydrophilic drug. Patterning assisted in controlling the initial burst release, which is a significant finding especially for low half-life drugs.

  8. Construction of Modular Hydrogel Sheets for Micropatterned Macro-scaled 3D Cellular Architecture.

    Science.gov (United States)

    Son, Jaejung; Bae, Chae Yun; Park, Je-Kyun

    2016-01-11

    Hydrogels can be patterned at the micro-scale using microfluidic or micropatterning technologies to provide an in vivo-like three-dimensional (3D) tissue geometry. The resulting 3D hydrogel-based cellular constructs have been introduced as an alternative to animal experiments for advanced biological studies, pharmacological assays and organ transplant applications. Although hydrogel-based particles and fibers can be easily fabricated, it is difficult to manipulate them for tissue reconstruction. In this video, we describe a fabrication method for micropatterned alginate hydrogel sheets, together with their assembly to form a macro-scale 3D cell culture system with a controlled cellular microenvironment. Using a mist form of the calcium gelling agent, thin hydrogel sheets are easily generated with a thickness in the range of 100 - 200 µm, and with precise micropatterns. Cells can then be cultured with the geometric guidance of the hydrogel sheets in freestanding conditions. Furthermore, the hydrogel sheets can be readily manipulated using a micropipette with an end-cut tip, and can be assembled into multi-layered structures by stacking them using a patterned polydimethylsiloxane (PDMS) frame. These modular hydrogel sheets, which can be fabricated using a facile process, have potential applications of in vitro drug assays and biological studies, including functional studies of micro- and macrostructure and tissue reconstruction.

  9. Co-Culturing of Multipotent Mesenchymal Stromal Cells with Autological and Allogenic Lymphocytes.

    Science.gov (United States)

    Kapranov, N M; Davydova, Yu O; Gal'tseva, I V; Petinati, N A; Bakshinskaitė, M V; Drize, N I; Kuz'mina, L A; Parovichnikova, E N; Savchenko, V G

    2018-03-01

    We studied the effect of autologous and allogeneic lymphocytes on multipotent mesenchymal stromal cells in co-culture. It is shown that changes in multipotent mesenchymal stromal cells and in lymphocytes did not depend on the source of lymphocytes. Contact with lymphocytes triggers expression of HLA-DR molecules on multipotent mesenchymal stromal cells and these cells lose their immune privilege. In multipotent mesenchymal stromal cells, the relative level of expression of factors involved in immunomodulation (IDO1, PTGES, and IL-6) and expression of adhesion molecule ICAM1 increased, while expression of genes involved in the differentiation of multipotent mesenchymal stromal cells remained unchanged. Priming of multipotent mesenchymal stromal cells with IFN did not affect these changes. In turn, lymphocytes underwent activation, expression of HLA-DR increased, subpopulation composition of lymphocytes changed towards the increase in the content of naïve T cells. These findings are important for cell therapy.

  10. Effect of diisopropylfluorophosphate on synaptic transmission and acetylcholine sensitivity in neuroblastoma-myotube co-culture

    International Nuclear Information System (INIS)

    Adler, M.; Chang, F.C.T.; Foster, R.E.; Glenn, J.F.; Mark, G.; Maxwell, D.

    1986-01-01

    The authors investigate the effects of the irreversible organophosphorous cholinesterase inhibitor, DFP, on clonal G8-1 myotubes co-cultured with ACh- secreting NG108-15 neuroblastoma x glioma hybrid cells. The enzyme activity is shown, plotted as a function of time in culture. The enzyme activity remained low over four days. At the end of this time, the cultures were nearly confluent with myoblasts but contained less than 2% multinucleated myotubes. The AChE activity increased gradually after horse serum was added to the growth medium to promote myotube formation, reaching a maximum of 1.1 nmole. C 14 ACh/min/mg protein on the 15th day

  11. A 3D co-culture microtissue model of the human placenta for nanotoxicity assessment.

    Science.gov (United States)

    Muoth, Carina; Wichser, Adrian; Monopoli, Marco; Correia, Manuel; Ehrlich, Nicky; Loeschner, Katrin; Gallud, Audrey; Kucki, Melanie; Diener, Liliane; Manser, Pius; Jochum, Wolfram; Wick, Peter; Buerki-Thurnherr, Tina

    2016-10-06

    There is increasing evidence that certain nanoparticles (NPs) can overcome the placental barrier, raising concerns on potential adverse effects on the growing fetus. But even in the absence of placental transfer, NPs may pose a risk to proper fetal development if they interfere with the viability and functionality of the placental tissue. The effects of NPs on the human placenta are not well studied or understood, and predictive in vitro placenta models to achieve mechanistic insights on NP-placenta interactions are essentially lacking. Using the scaffold-free hanging drop technology, we developed a well-organized and highly reproducible 3D co-culture microtissue (MT) model consisting of a core of placental fibroblasts surrounded by a trophoblast cell layer, which resembles the structure of the in vivo placental tissue. We could show that secretion levels of human chorionic gonadotropin (hCG) were significantly higher in 3D than in 2D cell cultures, which indicates an enhanced differentiation of trophoblasts grown on 3D MTs. NP toxicity assessment revealed that cadmium telluride (CdTe) and copper oxide (CuO) NPs but not titanium dioxide (TiO 2 ) NPs decreased MT viability and reduced the release of hCG. NP acute toxicity was significantly reduced in 3D co-culture MTs compared to 2D monocultures. Taken together, 3D placental MTs provide a new and promising model for the fast generation of tissue-relevant acute NP toxicity data, which are indispensable for the safe development of NPs for industrial, commercial and medical applications.

  12. Mesenchymal stem cells support neuronal fiber growth in an organotypic brain slice co-culture model.

    Science.gov (United States)

    Sygnecka, Katja; Heider, Andreas; Scherf, Nico; Alt, Rüdiger; Franke, Heike; Heine, Claudia

    2015-04-01

    Mesenchymal stem cells (MSCs) have been identified as promising candidates for neuroregenerative cell therapies. However, the impact of different isolation procedures on the functional and regenerative characteristics of MSC populations has not been studied thoroughly. To quantify these differences, we directly compared classically isolated bulk bone marrow-derived MSCs (bulk BM-MSCs) to the subpopulation Sca-1(+)Lin(-)CD45(-)-derived MSCs(-) (SL45-MSCs), isolated by fluorescence-activated cell sorting from bulk BM-cell suspensions. Both populations were analyzed with respect to functional readouts, that are, frequency of fibroblast colony forming units (CFU-f), general morphology, and expression of stem cell markers. The SL45-MSC population is characterized by greater morphological homogeneity, higher CFU-f frequency, and significantly increased nestin expression compared with bulk BM-MSCs. We further quantified the potential of both cell populations to enhance neuronal fiber growth, using an ex vivo model of organotypic brain slice co-cultures of the mesocortical dopaminergic projection system. The MSC populations were cultivated underneath the slice co-cultures without direct contact using a transwell system. After cultivation, the fiber density in the border region between the two brain slices was quantified. While both populations significantly enhanced fiber outgrowth as compared with controls, purified SL45-MSCs stimulated fiber growth to a larger degree. Subsequently, we analyzed the expression of different growth factors in both cell populations. The results show a significantly higher expression of brain-derived neurotrophic factor (BDNF) and basic fibroblast growth factor in the SL45-MSCs population. Altogether, we conclude that MSC preparations enriched for primary MSCs promote neuronal regeneration and axonal regrowth, more effectively than bulk BM-MSCs, an effect that may be mediated by a higher BDNF secretion.

  13. Porphyromonas gingivalis displays a competitive advantage over Aggregatibacter actinomycetemcomitans in co-cultured biofilm.

    Science.gov (United States)

    Takasaki, K; Fujise, O; Miura, M; Hamachi, T; Maeda, K

    2013-06-01

    Biofilm formation occurs through the events of cooperative growth and competitive survival among multiple species. Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans are important periodontal pathogens. The aim of this study was to demonstrate competitive or cooperative interactions between these two species in co-cultured biofilm. P. gingivalis strains and gingipain mutants were cultured with or without A. actinomycetemcomitans. Biofilms formed on glass surfaces were analyzed by crystal violet staining and colony counting. Preformed A. actinomycetemcomitans biofilms were treated with P. gingivalis culture supernatants. Growth and proteolytic activities of gingipains were also determined. Monocultured P. gingivalis strains exhibited a range of biofilm-formation abilities and proteolytic activities. The ATCC33277 strain, noted for its high biofilm-formation ability and proteolytic activity, was found to be dominant in biofilm co-cultured with A. actinomycetemcomitans. In a time-resolved assay, A. actinomycetemcomitans was primarily the dominant colonizer on a glass surface and subsequently detached in the presence of increasing numbers of ATCC33277. Detachment of preformed A. actinomycetemcomitans biofilm was observed by incubation with culture supernatants from highly proteolytic strains. These results suggest that P. gingivalis possesses a competitive advantage over A. actinomycetemcomitans. As the required biofilm-formation abilities and proteolytic activities vary among P. gingivalis strains, the diversity of the competitive advantage is likely to affect disease recurrence during periodontal maintenance. © 2012 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  14. Inference of Interactions in Cyanobacterial-Heterotrophic Co-Cultures via Transcriptome Sequencing

    Energy Technology Data Exchange (ETDEWEB)

    Beliaev, Alex S.; Romine, Margaret F.; Serres, Margaret; Bernstein, Hans C.; Linggi, Bryan E.; Markillie, Lye Meng; Isern, Nancy G.; Chrisler, William B.; Kucek, Leo A.; Hill, Eric A.; Pinchuk, Grigoriy; Bryant, Donald A.; Wiley, H. S.; Fredrickson, Jim K.; Konopka, Allan

    2014-04-29

    We employed deep sequencing technology to identify transcriptional adaptation of the euryhaline unicellular cyanobacterium Synechococcus sp. PCC 7002 and the marine facultative aerobe Shewanella putrefaciens W3-18-1 to growth in a co-culture and infer the effect of carbon flux distributions on photoautotroph-heterotroph interactions. The overall transcriptome response of both organisms to co-cultivation was shaped by their respective physiologies and growth constraints. Carbon limitation resulted in the expansion of metabolic capacities which was manifested through the transcriptional upregulation of transport and catabolic pathways. While growth coupling occurred via lactate oxidation or secretion of photosynthetically fixed carbon, there was evidence of specific metabolic interactions between the two organisms. On one hand, the production and excretion of specific amino acids (methionine and alanine) by the cyanobacterium correlated with the putative downregulation of the corresponding biosynthetic machinery of Shewanella W3-18-1. On the other hand, the broad and consistent decrease of mRNA levels for many Fe-regulated Synechococcus 7002 genes during co-cultivation suggested increased Fe availability as well as more facile and energy-efficient mechanisms for Fe acquisition by the cyanobacterium. Furthermore, evidence pointed at potentially novel interactions between oxygenic photoautotrophs and heterotrophs related to the oxidative stress response as transcriptional patterns suggested that Synechococcus 7002 rather than Shewanella W3-18-1 provided scavenging functions for reactive oxygen species under co-culture conditions. This study provides an initial insight into the complexity of photoautotrophic-heterotrophic interactions and brings new perspectives of their role in the robustness and stability of the association.

  15. 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.

  16. Antitumor Activity of Rat Mesenchymal Stem Cells during Direct or Indirect Co-Culturing with C6 Glioma Cells.

    Science.gov (United States)

    Gabashvili, A N; Baklaushev, V P; Grinenko, N F; Mel'nikov, P A; Cherepanov, S A; Levinsky, A B; Chehonin, V P

    2016-02-01

    The tumor-suppressive effect of rat mesenchymal stem cells against low-differentiated rat C6 glioma cells during their direct and indirect co-culturing and during culturing of C6 glioma cells in the medium conditioned by mesenchymal stem cells was studied in an in vitro experiment. The most pronounced antitumor activity of mesenchymal stem cells was observed during direct co-culturing with C6 glioma cells. The number of live C6 glioma cells during indirect co-culturing and during culturing in conditioned medium was slightly higher than during direct co-culturing, but significantly differed from the control (C6 glioma cells cultured in medium conditioned by C6 glioma cells). The cytotoxic effect of medium conditioned by mesenchymal stem cells was not related to medium depletion by glioma cells during their growth. The medium conditioned by other "non-stem" cells (rat astrocytes and fibroblasts) produced no tumor-suppressive effect. Rat mesenchymal stem cells, similar to rat C6 glioma cells express connexin 43, the main astroglial gap junction protein. During co-culturing, mesenchymal stem cells and glioma C6 cells formed functionally active gap junctions. Gap junction blockade with connexon inhibitor carbenoxolone attenuated the antitumor effect observed during direct co-culturing of C6 glioma cells and mesenchymal stem cells to the level produced by conditioned medium. Cell-cell signaling mediated by gap junctions can be a mechanism of the tumor-suppressive effect of mesenchymal stem cells against C6 glioma cells. This phenomenon can be used for the development of new methods of cell therapy for high-grade malignant gliomas.

  17. Bottom-up fabrication of artery-mimicking tubular co-cultures in collagen-based microchannel scaffolds.

    Science.gov (United States)

    Tan, A; Fujisawa, K; Yukawa, Y; Matsunaga, Y T

    2016-10-20

    We developed a robust bottom-up approach to construct open-ended, tubular co-culture constructs that simulate the human vascular morphology and microenvironment. By design, these three-dimensional artificial vessels mimic the basic architecture of an artery: a collagen-rich extracellular matrix (as the tunica externa), smooth muscle cells (SMCs) (as the tunica media), and an endothelial cell (EC) lining (as the tunica interna). A versatile needle-based fabrication technique was employed to achieve controllable arterial layouts within a PDMS-hosted collagen microchannel scaffold (330 ± 10 μm in diameter): (direct co-culture) a SMC/EC bilayer to follow the structure of an arteriole-like segment; and (encapsulated co-culture) a lateral SMC multilayer covered by an EC monolayer lining to simulate the architecture of a larger artery. Optical and fluorescence microscopy images clearly evidenced the progressive cell elongation and sprouting behavior of SMCs and ECs along the collagen gel contour and within the gel matrix under static co-culture conditions. The progressive cell growth patterns effectively led to the formation of a tubular co-culture with an internal endothelial lining expressing prominent CD31 (cluster of differentiation 31) intercellular junction markers. During a 4-day static maturation period, the artery constructs showed modest alteration in the luminal diameters (i.e. less than 10% changes from the initial measurements). This argues in favor of stable and predictable arterial architecture achieved via the proposed fabrication protocols. Both co-culture models showed a high glucose metabolic rate during the initial proliferation phase, followed by a temporary quiescent (and thus, mature) stage. These proof-of-concept models with a controllable architecture create an important foundation for advanced vessel manipulations such as the integration of relevant physiological functionality or remodeling into a vascular disease-mimicking tissue.

  18. Hepatocellular carcinoma: a review

    Directory of Open Access Journals (Sweden)

    Balogh J

    2016-10-01

    Full Text Available Julius Balogh,1,2 David Victor III,1,3,4 Emad H Asham,1,2 Sherilyn Gordon Burroughs,1,2 Maha Boktour,1,2 Ashish Saharia,1,2 Xian Li,1,2 R Mark Ghobrial,1,2 Howard P Monsour Jr,1,3,4 1Sherrie and Alan Conover Center for Liver Disease and Transplantation, 2Division of Transplantation, Department of Surgery, 3Department of Gastroenterology and Transplant Hepatology, 4Department of Medicine, Houston Methodist Hospital, Houston, TX, USA Abstract: Hepatocellular carcinoma (HCC is the most common primary liver malignancy and is a leading cause of cancer-related death worldwide. In the United States, HCC is the ninth leading cause of cancer deaths. Despite advances in prevention techniques, screening, and new technologies in both diagnosis and treatment, incidence and mortality continue to rise. Cirrhosis remains the most important risk factor for the development of HCC regardless of etiology. Hepatitis B and C are independent risk factors for the development of cirrhosis. Alcohol consumption remains an important additional risk factor in the United States as alcohol abuse is five times higher than hepatitis C. Diagnosis is confirmed without pathologic confirmation. Screening includes both radiologic tests, such as ultrasound, computerized tomography, and magnetic resonance imaging, and serological markers such as α-fetoprotein at 6-month intervals. Multiple treatment modalities exist; however, only orthotopic liver transplantation (OLT or surgical resection is curative. OLT is available for patients who meet or are downstaged into the Milan or University of San Francisco criteria. Additional treatment modalities include transarterial chemoembolization, radiofrequency ablation, microwave ablation, percutaneous ethanol injection, cryoablation, radiation therapy, systemic chemotherapy, and molecularly targeted therapies. Selection of a treatment modality is based on tumor size, location, extrahepatic spread, and underlying liver function. HCC is an

  19. Phototrophic hydrogen production from glucose by pure and co-cultures of Clostridium butyricum and Rhodobacter sphaeroides

    Energy Technology Data Exchange (ETDEWEB)

    Fang, Herbert H.P.; Zhu, Heguang; Zhang, Tong [Centre for Environmental Engineering Research, Department of Civil Engineering, University of Hong Kong, Pokfulam Road, Hong Kong (China)

    2006-12-15

    Phototrophic hydrogen production from glucose by pure and co-cultures of Clostridium butyricum and Rhodobacter sphaeroides was studied in batch experiments. Results showed that in all batches hydrogen was produced after a lag phase of about 10h; pure culture of R. sphaeroides produced hydrogen at rates substantially lower than C. butyricum. In co-culture systems, R. sphaeroides even with cell populations 5.9 times higher still could not compete with C. butyricum for glucose. In co-culture systems, R. sphaeroides syntrophically interacted with C. butyricum, using the acetate and butyrate produced by the latter as substrate for hydrogen production. Hydrogen production was ceased in all batches when the pH was lowered to the level of pH 6.5, resulting from the accumulation of fatty acids. It was also demonstrated in this study that fluorescence in situ hybridization (FISH) was an effective means for the quantification of the relative abundance of individual bacteria in a co-culture system. (author)

  20. Syntrophomonas zehnderi sp. nov., an anaerobe that degrades long chain fatty acids in co-culture with Methanobacterium formicicum

    NARCIS (Netherlands)

    Sousa, D.Z.; Smidt, H.; Alves, M.M.; Stams, A.J.M.

    2007-01-01

    An anaerobic, mesophilic, syntrophic fatty-acid-oxidizing bacterium, designated strain OL-4T, was isolated as a co-culture with Methanobacterium formicicum DSM 1535NT from an anaerobic expanded granular sludge bed reactor used to treat an oleate-based effluent. Strain OL-4T degraded oleate, a

  1. 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

  2. Mass culture strategy for bacterial yeast co-culture for degradation of petroleum hydrocarbons in marine environment.

    Science.gov (United States)

    Priya, Anchal; Mandal, Ajoy K; Ball, Andrew S; Manefield, Mike; Lal, Banwari; Sarma, Priyangshu M

    2015-11-15

    In the present study a metabolically versatile co-culture with two Bacilli and one yeast strain was developed using enrichment culture techniques. The developed co-culture had affinity to degrade both aliphatic and aromatic fractions of petroleum crude oil. Degradation kinetics was established for designing the fermentation protocol of the co-culture. The developed mass culture strategy led to achieve the reduction in surface tension (26dynescm(-1) from 69 dynescm(-1)) and degradation of 67% in bench scale experiments. The total crude oil degradation of 96% was achieved in 4000l of natural seawater after 28days without adding any nutrients. The survival of the augmented co-culture was maintained (10(9)cellsml(-1)) in contaminated marine environment. The mass culture protocol devised for the bioaugmentation was a key breakthrough that was subsequently used for pilot scale studies with 100l and 4000l of natural seawater for potential application in marine oil spills. Copyright © 2015 Elsevier Ltd. All rights reserved.

  3. Retinal pigment epithelial cells upregulate expression of complement factors after co-culture with activated T cells

    DEFF Research Database (Denmark)

    Juel, Helene Bæk; Kaestel, Charlotte; Folkersen, Lasse

    2011-01-01

    In this study we examined the effect of T cell-derived cytokines on retinal pigment epithelial (RPE) cells with respect to expression of complement components. We used an in vitro co-culture system in which CD3/CD28-activated human T cells were separated from the human RPE cell line (ARPE-19...

  4. Trophic Effects of Mesenchymal Stem Cells in Chondrocyte Co-Cultures are Independent of Culture Conditions and Cell Sources

    NARCIS (Netherlands)

    Wu, Ling; Prins, H.J.; Helder, M.; van Blitterswijk, Clemens; Karperien, Hermanus Bernardus Johannes

    2012-01-01

    Earlier, we have shown that the increased cartilage production in pellet co-cultures of chondrocytes and bone marrow-derived mesenchymal stem cells (BM-MSCs) is due to a trophic role of the MSC in stimulating chondrocyte proliferation and matrix production rather than MSCs actively undergoing

  5. Trophic effects of mesenchymal stem cells in chondrocyte co-cultures are independent of culture conditions and cell sources

    NARCIS (Netherlands)

    Wu, L.; Prins, H.J.; Helder, M.N.; van Blitterswijk, C.A.; Karperien, M.

    2012-01-01

    Earlier, we have shown that the increased cartilage production in pellet co-cultures of chondrocytes and bone marrow-derived mesenchymal stem cells (BM-MSCs) is due to a trophic role of the MSC in stimulating chondrocyte proliferation and matrix production rather than MSCs actively undergoing

  6. In vitro co-cultures of Pinus pinaster with Bursaphelenchus xylophilus: a biotechnological approach to study pine wilt disease.

    Science.gov (United States)

    Faria, Jorge M S; Sena, Inês; Vieira da Silva, Inês; Ribeiro, Bruno; Barbosa, Pedro; Ascensão, Lia; Bennett, Richard N; Mota, Manuel; Figueiredo, A Cristina

    2015-06-01

    Co-cultures of Pinus pinaster with Bursaphelenchus xylophilus were established as a biotechnological tool to evaluate the effect of nematotoxics addition in a host/parasite culture system. The pinewood nematode (PWN), Bursaphelenchus xylophilus, the causal agent of pine wilt disease (PWD), was detected for the first time in Europe in 1999 spreading throughout the pine forests in Portugal and recently in Spain. Plant in vitro cultures may be a useful experimental system to investigate the plant/nematode relationships in loco, thus avoiding the difficulties of field assays. In this study, Pinus pinaster in vitro cultures were established and compared to in vivo 1 year-old plantlets by analyzing shoot structure and volatiles production. In vitro co-cultures were established with the PWN and the effect of the phytoparasite on in vitro shoot structure, water content and volatiles production was evaluated. In vitro shoots showed similar structure and volatiles production to in vivo maritime pine plantlets. The first macroscopic symptoms of PWD were observed about 4 weeks after in vitro co-culture establishment. Nematode population in the culture medium increased and PWNs were detected in gaps of the callus tissue and in cavities developed from the degradation of cambial cells. In terms of volatiles main components, plantlets, P. pinaster cultures, and P. pinaster with B. xylophilus co-cultures were all β- and α-pinene rich. Co-cultures may be an easy-to-handle biotechnological approach to study this pathology, envisioning the understanding of and finding ways to restrain this highly devastating nematode.

  7. A dual-color luciferase assay system reveals circadian resetting of cultured fibroblasts by co-cultured adrenal glands.

    Directory of Open Access Journals (Sweden)

    Takako Noguchi

    Full Text Available In mammals, circadian rhythms of various organs and tissues are synchronized by pacemaker neurons in the suprachiasmatic nucleus (SCN of the hypothalamus. Glucocorticoids released from the adrenal glands can synchronize circadian rhythms in other tissues. Many hormones show circadian rhythms in their plasma concentrations; however, whether organs outside the SCN can serve as master synchronizers to entrain circadian rhythms in target tissues is not well understood. To further delineate the function of the adrenal glands and the interactions of circadian rhythms in putative master synchronizing organs and their target tissues, here we report a simple co-culture system using a dual-color luciferase assay to monitor circadian rhythms separately in various explanted tissues and fibroblasts. In this system, circadian rhythms of organs and target cells were simultaneously tracked by the green-emitting beetle luciferase from Pyrearinus termitilluminans (ELuc and the red-emitting beetle luciferase from Phrixothrix hirtus (SLR, respectively. We obtained tissues from the adrenal glands, thyroid glands, and lungs of transgenic mice that expressed ELuc under control of the promoter from a canonical clock gene, mBmal1. The tissues were co-cultured with Rat-1 fibroblasts as representative target cells expressing SLR under control of the mBmal1 promoter. Amplitudes of the circadian rhythms of Rat-1 fibroblasts were potentiated when the fibroblasts were co-cultured with adrenal gland tissue, but not when co-cultured with thyroid gland or lung tissue. The phases of Rat-1 fibroblasts were reset by application of adrenal gland tissue, whereas the phases of adrenal gland tissue were not influenced by Rat-1 fibroblasts. Furthermore, the effect of the adrenal gland tissue on the fibroblasts was blocked by application of a glucocorticoid receptor (GR antagonist. These results demonstrate that glucocorticoids are strong circadian synchronizers for fibroblasts and that

  8. Co-culture of Adult Mesenchymal Stem Cells and Nucleus Pulposus Cells in Bilaminar Pellets for Intervertebral Disc Regeneration.

    Science.gov (United States)

    Allon, Aliza A; Schneider, Richard A; Lotz, Jeffrey C

    2009-01-01

    Our goal is to optimize stem cell-based tissue engineering strategies in the context of the intervertebral disc environment. We explored the benefits of co-culturing nucleus pulposus cells (NPC) and adult mesenchymal stem cells (MSC) using a novel spherical bilaminar pellet culture system where one cell type is enclosed in a sphere of the other cell type. Our 3D system provides a structure that exploits embryonic processes such as tissue induction and condensation. We observed a unique phenomenon: the budding of co-culture pellets and the formation of satellite pellets that separate from the main pellet. MSC and NPC co-culture pellets were formed with three different structural organizations. The first had random organization. The other two had bilaminar organization with either MSC inside and NPC outside or NPC inside and MSC outside. By 14 days, all co-culture pellets exhibited budding and spontaneously generated satellite pellets. The satellite pellets were composed of both cell types and, surprisingly, all had the same bilaminar organization with MSC on the inside and NPC on the outside. This organization was independent of the structure of the main pellet that the satellites stemmed from. The main pellets generated satellite pellets that spontaneously organized into a bilaminar structure. This implies that structural organization occurs naturally in this cell culture system and may be inherently favorable for cell-based tissue engineering strategies. The occurrence of budding and the organization of satellite pellets may have important implications for the use of co-culture pellets in cell-based therapies for disc regeneration. From a therapeutic point of view, the generation of satellite pellets may be a beneficial feature that would serve to spread donor cells throughout the host matrix and restore normal matrix composition in a sustainable way, ultimately renewing tissue function.

  9. Effects of external radiation in a co-culture model of endothelial cells and adipose-derived stem cells

    International Nuclear Information System (INIS)

    Haubner, Frank; Leyh, Michaela; Ohmann, Elisabeth; Pohl, Fabian; Prantl, Lukas; Gassner, Holger G

    2013-01-01

    The inflammatory response clinically observed after radiation has been described to correlate with elevated expression of cytokines and adhesion molecules by endothelial cells. Therapeutic compensation for this microvascular compromise could be an important approach in the treatment of irradiated wounds. Clinical reports describe the potential of adipose-derived stem cells to enhance wound healing, but the underlying cellular mechanisms remain largely unclear. Human dermal microvascular endothelial cells (HDMEC) and human adipose-derived stem cells (ASC) were cultured in a co-culture setting and irradiated with sequential doses of 2 to 12 Gy. Cell count was determined 48 h after radiation using a semi-automated cell counting system. Levels of interleukin-6 (IL-6), basic fibroblast growth factor (FGF), intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) were determined in the supernatants using enzyme-linked immunosorbent assay (ELISA). Irradiated HDMEC and ASC as well as non-irradiated co-cultures, HDMEC or ASC respectively were used as controls. Cell count was significantly reduced in irradiated co-cultures of HDMEC and ASC compared to non-irradiated controls. Levels of IL-6, FGF, ICAM-1 and VCAM-1 in the supernatants of the co-cultures were significantly less affected by external radiation in comparison to HDMEC. The increased expression of cytokines and adhesion molecules by HDMEC after external radiation is mitigated in the co-culture setting with ASC. These in vitro changes seem to support the clinical observation that ASC may have a stabilizing effect when injected into irradiated wounds

  10. Skin equivalent tissue-engineered construct: co-cultured fibroblasts/ keratinocytes on 3D matrices of sericin hope cocoons.

    Directory of Open Access Journals (Sweden)

    Sunita Nayak

    Full Text Available The development of effective and alternative tissue-engineered skin replacements to autografts, allografts and xenografts has became a clinical requirement due to the problems related to source of donor tissue and the perceived risk of disease transmission. In the present study 3D tissue engineered construct of sericin is developed using co-culture of keratinocytes on the upper surface of the fabricated matrices and with fibroblasts on lower surface. Sericin is obtained from "Sericin Hope" silkworm of Bombyx mori mutant and is extracted from cocoons by autoclave. Porous sericin matrices are prepared by freeze dried method using genipin as crosslinker. The matrices are characterized biochemically and biophysically. The cell proliferation and viability of co-cultured fibroblasts and keratinocytes on matrices for at least 28 days are observed by live/dead assay, Alamar blue assay, and by dual fluorescent staining. The growth of the fibroblasts and keratinocytes in co-culture is correlated with the expression level of TGF-β, b-FGF and IL-8 in the cultured supernatants by enzyme-linked immunosorbent assay. The histological analysis further demonstrates a multi-layered stratified epidermal layer of uninhibited keratinocytes in co-cultured constructs. Presence of involucrin, collagen IV and the fibroblast surface protein in immuno-histochemical stained sections of co-cultured matrices indicates the significance of paracrine signaling between keratinocytes and fibroblasts in the expression of extracellular matrix protein for dermal repair. No significant amount of pro inflammatory cytokines (TNF-α, IL-1β and nitric oxide production are evidenced when macrophages grown on the sericin matrices. The results all together depict the potentiality of sericin 3D matrices as skin equivalent tissue engineered construct in wound repair.

  11. Stick-Jump (SJ) Evaporation of Strongly Pinned Nanoliter Volume Sessile Water Droplets on Quick Drying, Micropatterned Surfaces.

    Science.gov (United States)

    Debuisson, Damien; Merlen, Alain; Senez, Vincent; Arscott, Steve

    2016-03-22

    We present an experimental study of stick-jump (SJ) evaporation of strongly pinned nanoliter volume sessile water droplets drying on micropatterned surfaces. The evaporation is studied on surfaces composed of photolithographically micropatterned negative photoresist (SU-8). The micropatterning of the SU-8 enables circular, smooth, trough-like features to be formed which causes a very strong pinning of the three phase (liquid-vapor-solid) contact line of an evaporating droplet. This is ideal for studying SJ evaporation as it contains sequential constant contact radius (CCR) evaporation phases during droplet evaporation. The evaporation was studied in nonconfined conditions, and forced convection was not used. Micropatterned concentric circles were defined having an initial radius of 1000 μm decreasing by a spacing ranging from 500 to 50 μm. The droplet evaporates, successively pinning and depinning from circle to circle. For each pinning radius, the droplet contact angle and volume are observed to decrease quasi-linearly with time. The experimental average evaporation rates were found to decrease with decreasing pining radii. In contrast, the experimental average evaporation flux is found to increase with decreasing droplet radii. The data also demonstrate the influence of the initial contact angle on evaporation rate and flux. The data indicate that the total evaporation time of a droplet depends on the specific micropattern spacing and that the total evaporation time on micropatterned surfaces is always less than on flat, homogeneous surfaces. Although the surface patterning is observed to have little effect on the average droplet flux-indicating that the underlying evaporation physics is not significantly changed by the patterning-the total evaporation time is considerably modified by patterning, up to a factor or almost 2 compared to evaporation on a flat, homogeneous surface. The closely spaced concentric circle pinning maintains a large droplet radius and

  12. Metabolomic analysis of cooperative adaptation between co-cultured Bacillus cereus and Ketogulonicigenium vulgare.

    Directory of Open Access Journals (Sweden)

    Ming-Zhu Ding

    Full Text Available The cooperative adaptation of subcultivated Bacillus cereus and Ketogulonicigenium vulgare significantly increased the productivity of 2-keto-L-gulonic acid, the precursor of vitamin C. The mechanism of cooperative adaptation of the serial subcultivated B. cereus and K. vulgare was investigated in this study by culturing the two strains orthogonally on agar plates. It was found that the swarming distance of B. cereus along the trace of K. vulgare on the plate decreased after 150 days' subcultivation. Metabolomic analysis on these co-cultured B. cereus and K. vulgare strains showed that their cooperative adaptation was accomplished by three key events: (i the ability of nutrients (e.g., amino acids and purines searching and intaking, and proteins biosynthesis is increased in the evolved B. cereus; (ii the capability of protein degradation and amino acids transportation is enhanced in evolved K. vulgare; (iii the evolved B. cereus was found to provide more nutrients (mostly amino acids and purines to K. vulgare, thus strengthening the oxidation and energy generation of K. vulgare. Our results provided novel insights into the systems-level understanding of the cooperative adaptation between strains in synergistic consortium.

  13. Proteomic profiling of halloysite clay nanotube exposure in intestinal cell co-culture.

    Science.gov (United States)

    Lai, Xianyin; Agarwal, Mangilal; Lvov, Yuri M; Pachpande, Chetan; Varahramyan, Kody; Witzmann, Frank A

    2013-11-01

    Halloysite is aluminosilicate clay with a hollow tubular structure with nanoscale internal and external diameters. Assessment of halloysite biocompatibility has gained importance in view of its potential application in oral drug delivery. To investigate the effect of halloysite nanotubes on an in vitro model of the large intestine, Caco-2/HT29-MTX cells in monolayer co-culture were exposed to nanotubes for toxicity tests and proteomic analysis. Results indicate that halloysite exhibits a high degree of biocompatibility characterized by an absence of cytotoxicity, in spite of elevated pro-inflammatory cytokine release. Exposure-specific changes in expression were observed among 4081 proteins analyzed. Bioinformatic analysis of differentially expressed protein profiles suggest that halloysite stimulates processes related to cell growth and proliferation, subtle responses to cell infection, irritation and injury, enhanced antioxidant capability, and an overall adaptive response to exposure. These potentially relevant functional effects warrant further investigation in in vivo models and suggest that chronic or bolus occupational exposure to halloysite nanotubes may have unintended outcomes. Copyright © 2013 John Wiley & Sons, Ltd.

  14. Phenotypic responses to interspecies competition and commensalism in a naturally-derived microbial co-culture

    Energy Technology Data Exchange (ETDEWEB)

    Khan, Nymul; Maezato, Yukari; McClure, Ryan S.; Brislawn, Colin J.; Mobberley, Jennifer M.; Isern, Nancy; Chrisler, William B.; Markillie, Lye Meng; Barney, Brett M.; Song, Hyun-Seob; Nelson, William C.; Bernstein, Hans C.

    2018-01-10

    The fundamental question of whether different microbial species will co-exist or compete in a given environment depends on context, composition and environmental constraints. Model microbial systems can yield some general principles related to this question. In this study we employed a naturally occurring co-culture composed of heterotrophic bacteria, Halomonas sp. HL-48 and Marinobacter sp. HL-58, to ask two fundamental scientific questions: 1) how do the phenotypes of two naturally co-existing species respond to partnership as compared to axenic growth? and 2) how do growth and molecular phenotypes of these species change with respect to competitive and commensal interactions? We hypothesized – and confirmed – that co-cultivation under glucose as the sole carbon source would result in a competitive interactions. Similarly, when glucose was swapped with xylose, the interactions became commensal because Marinobacter HL-58 was supported by metabolites derived from Halomonas HL-48. Each species responded to partnership by changing both its growth and molecular phenotype as assayed via batch growth kinetics and global transcriptomics. These phenotypic responses depended nutrient availability and so the environment ultimately controlled how they responded to each other. This simplified model community revealed that microbial interactions are context-specific and different environmental conditions dictate how interspecies partnerships will unfold.

  15. Follicle stimulating hormone increases spermatogonial stem cell colonization during in vitro co - culture

    Directory of Open Access Journals (Sweden)

    Reza Narenji Sani

    2013-03-01

    Full Text Available The complex process of spermatogenesis is regulated by various factors. Studies onspermatogonial stem cells(SCCshave provided very important tool to improve herd geneticand different field. 0.2 to 0.3 percent of total cells of seminiferous tubules is consist ofspermatogonial stem cells. To investigate and biomanipulation of these cells, proliferationand viability rate of cells should be increasedin vitro, at first. Follicle stimulating hormone(FSH has been suggested to play a determinant role in the survival of germ cells in additionto increasing spermatogonial proliferation. In this study, thein vitroeffects ofFSHonspermatogonial cell colony formation were investigated. Sertoli and spermatogonial cellswere isolated from 3-5 months old calves. The identity of theSertoli cells and spermatogonialstem cells were confirmed through immunocytochemistry and colony morphology,respectively. Co-cultured Sertoli and spermatogonial cells were treatedwithFSHin differentdose of10, 20 and 40 IU mL-1FSH, before colony assay.Results indicated that,FSHincreasedin vitrocolonization of spermatogonial cells in comparison with control group. In conclusion,usingFSHprovided proper bovine spermatogonial stem cell culture medium forin vitrostudy of these cells.

  16. Follicle stimulating hormone increases spermatogonial stem cell colonization during in vitro co-culture.

    Science.gov (United States)

    Narenji Sani, Reza; Tajik, Parviz; Yousefi, Mohammad Hassan; Movahedin, Mansoureh; Qasemi-Panahi, Babak; Shafiei, Shiva; Ahmadi Hamedani, Mahmood

    2013-01-01

    The complex process of spermatogenesis is regulated by various factors. Studies on spermatogonial stem cells (SCCs) have provided very important tool to improve herd genetic and different field. 0.2 to 0.3 percent of total cells of seminiferous tubules is consist of spermatogonial stem cells. To investigate and biomanipulation of these cells, proliferation and viability rate of cells should be increased in vitro, at first. Follicle stimulating hormone (FSH) has been suggested to play a determinant role in the survival of germ cells in addition to increasing spermatogonial proliferation. In this study, the in vitro effects of FSH on spermatogonial cell colony formation were investigated. Sertoli and spermatogonial cells were isolated from 3-5 months old calves. The identity of the Sertoli cells and spermatogonial stem cells were confirmed through immunocytochemistry and colony morphology, respectively. Co-cultured Sertoli and spermatogonial cells were treated with FSH in different dose of 10, 20 and 40 IU mL(-1) FSH, before colony assay. Results indicated that, FSH increased in vitro colonization of spermatogonial cells in comparison with control group. In conclusion, using FSH provided proper bovine spermatogonial stem cell culture medium for in vitro study of these cells.

  17. Cell-friendly inverse opal-like hydrogels for a spatially separated co-culture system.

    Science.gov (United States)

    Kim, Jaeyun; Bencherif, Sidi A; Li, Weiwei Aileen; Mooney, David J

    2014-09-01

    Three-dimensional macroporous scaffolds have extensively been studied for cell-based tissue engineering but their use is mostly limited to mechanical support for cell adhesion and growth on the surface of macropores. Here, a templated fabrication method is described to prepare cell-friendly inverse opal-like hydrogels (IOHs) allowing both cell encapsulation within the hydrogel matrix and cell seeding on the surface of macropores. Ionically crosslinked alginate microbeads and photocrosslinkable biocompatible polymers are used as a sacrificial template and as a matrix, respectively. The alginate microbeads are easily removed by a chelating agent, with minimal toxicity for the encapsulated cells during template removal. The outer surface of macropores in IOHs can also provide a space for cell adherence. The cells encapsulated or attached in IOHs are able to remain viable and to proliferate over time. The elastic modulus and cell-adhesion properties of IOHs can be easily controlled and tuned. Finally, it is demonstrated that IOH can be used to co-culture two distinct cell populations in different spatial positions. This cell-friendly IOH system provides a 3D scaffold for organizing different cell types in a controllable microenvironment to investigate biological processes such as stem cell niches or tumor microenvironments. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Microalgae Harvest through Fungal Pelletization—Co-Culture of Chlorella vulgaris and Aspergillus niger

    Directory of Open Access Journals (Sweden)

    Sarman Oktovianus Gultom

    2014-07-01

    Full Text Available Microalgae harvesting is a labor- and energy-intensive process and new approaches to harvesting microalgae need to be developed in order to decrease the costs. In this study; co-cultivatation of filamentous fungus (Aspergillus niger and microalgae (Chlorella vulgaris to form cell pellets was evaluated under different conditions, including organic carbon source (glucose; glycerol; and sodium acetate concentration; initial concentration of fungal spores and microalgal cells and light. Results showed that 2 g/L of glucose with a 1:300 ratio of fungi to microalgae provided the best culturing conditions for the process to reach >90% of cell harvest efficiency. The results also showed that an organic carbon source was required to sustain the growth of fungi and form the cell pellets. The microalgae/fungi co-cultures at mixotrophic conditions obtained much higher total biomass than pure cultures of each individual strains; indicating the symbiotic relationship between two strains. This can benefit the microbial biofuel production in terms of cell harvest and biomass production.

  19. Emergence of organized structure in co-culture spheroids: Experiments and Theory

    Science.gov (United States)

    Sanford, Roland; Kolbman, Dan; Song, Wei; Wu, Mingming; Ma, Minglin; Das, Moumita

    During tissue morphogenesis, from formation of embryos to tumor progression, cells often live and migrate in a heterogeneous environment consisting of many types of cells. To understand how differences in cell mechanobiological properties impact cellular self-organization and migration, we study a co-culture model composed of two distinct cell types confined in a three-dimensional spherical capsule. The cells are modeled as deformable, interacting, self-propelled particles that proliferate at specified timescales. A disordered potential is introduced to mimic the effect of the extracellular matrix (ECM). By varying the mechano-adhesive properties of each type, we investigate how differences in cell stiffness, cell-cell adhesion, and cell-ECM interaction influence collective properties of the binary cell population, such as self-assembly and migration. The predictions of the model are compared to experimental results on co-cutures of breast cancer cells and non-tumorigenic breast epithelial cells. This work was partially supported by a Cottrell College Science Award from the Research Corporation for Science Advancement.

  20. Conformal ZnO nanocomposite coatings on micro-patterned surfaces for superhydrophobicity

    International Nuclear Information System (INIS)

    Steele, Adam; Bayer, Ilker; Moran, Stephen; Cannon, Andrew; King, William P.; Loth, Eric

    2010-01-01

    A conformal coating process is presented to transform surfaces with inherent micro-morphology into superhydrophobic surfaces with hierarchical surface structure using wet chemical spray casting. Nanocomposite coatings composed of zinc oxide nanoparticles and organosilane quaternary nitrogen compound are dispersed in solution for application. The coating is applied to a micro-patterned polydimethylsiloxane substrate with a regular array of cylindrical microposts as well as a surface with random micro-structure for the purpose of demonstrating improved non-wettability and a superhydrophobic state for water droplets. Coating surface morphology is investigated with an environmental scanning electron microscope and surface wettability performance is characterized by static and dynamic contact angle measurements.

  1. High-throughput creation of micropatterned PDMS surfaces using microscale dual roller casting

    International Nuclear Information System (INIS)

    DiBartolomeo, Franklin J; Ge, Ning; Trinkle, Christine A

    2012-01-01

    This work introduces microscale dual roller casting (MDRC), a novel high-throughput fabrication method for creating continuous micropatterned surfaces using thermosetting polymers. MDRC utilizes a pair of rotating, heated cylindrical molds with microscale surface patterns to cure a continuous microstructured film. Using unmodified polydimethylsiloxane as the thermosetting polymer, we were able to create optically transparent, biocompatible surfaces with submicron patterning fidelity. Compared to other roll-to-roll fabrication processes, this method offers increased flexibility in the types of materials and topography that can be generated, including dual-sided patterning, embedded materials and tunable film thickness. (paper)

  2. Directing functional chemistries on micropatterned conducting polymers for all-polymer cell analysis microsystems

    DEFF Research Database (Denmark)

    Lind, Johan Ulrik; Daugaard, Anders Egede; Andresen, Thomas Lars

    Micrometer scale electrical circuits of PEDOT (poly(3,4-dioxythiophene)) were created by locally oxidizing PEDOT thin films with an agarose stamp containing the oxidizing agent NaOCl. The oxidized PEDOT was removed completely by applying detergents. The process was sufficiently mild that chemical...... groups on the underlying substrate, such as azides or alkynes, were preserved for subsequent specific functionalization. Moreover entire PMOXA (poly(2-methyl-2-oxazoline)) films preventing cell binding could be hidden below the PEDOT and be re-exposed upon stamping, allowing for cell capturing...... microelectrodes on a cell non-adhesive background. Chemically functionalized PEDOT types permitted the introduction of multiple additional types of micropatterned chemistry....

  3. Radioembolization of hepatocellular carcinoma.

    Science.gov (United States)

    Van de Wiele, Christophe

    2010-12-01

    In this review paper, available data on radioembolization of unresectable hepatocellular carcinoma (HCC) using commercially available radiopharmaceuticals, respectively (131)I-Lipiodol, Therasphere (glass-microspheres) and SIRspheres (resin-microspheres) are reviewed. In the palliative setting, (131)I-Lipiodol was shown to yield response rates of 17-92% which in patients with portal vein thrombosis (PVT) translate into a survival benefit as evidenced by a phase III randomized trial. Furthermore, in terms of efficacy, (131)I-Lipiodol is as efficacious as trans-arterial chemoembolization (TACE) but far better tolerated. In the adjuvant setting, improved recurrence-free and overall survival when compared to surgery alone have been reported but these results warrant confirmation by randomized prospective trials. Similar to (131)I-Lipiodol, when administered in a palliative setting, radioembolization using (90)Y microspheres was proven effective for selected cases of non-resectable HCC and well tolerated. Available data suggest that Therasphere treatment outperforms TACE both in terms of response as in terms of event-free survival in unresectable HCC. However, this finding needs confirmation by randomized prospective trials. Therasphere treatment was also shown to limit progression of HCC allowing potential candidates for orthotopic liver transplantation (OLT) more time to wait for donor organs as well as to downstage the HCC disease to such an extent that patients that were initially not, as yet become eligible for OLT with a gain in survival. Finally, Therasphere was shown to be safe and efficacious in HCC patients presenting with PVT, reason for which approval was granted for this indication by the FDA.

  4. Primary chondrocytes enhance cartilage tissue formation upon co-culture with expanded chondrocytes, dermal fibroblasts, 3T3 feeder cells and embryonic stem cells

    NARCIS (Netherlands)

    Hendriks, J.A.A.; Miclea, Razvan L.; Schotel, Roka; de Bruijn, Ewart; Moroni, Lorenzo; Karperien, Hermanus Bernardus Johannes; Riesle, J.U.; van Blitterswijk, Clemens

    2010-01-01

    Co-culture models have been increasingly used in tissue engineering applications to understand cell–cell interactions and consequently improve regenerative medicine strategies. Aiming at further elucidating cartilage tissue formation, we co-cultured bovine primary chondrocytes (BPCs) with human

  5. Hepatocellular carcinoma in Danish patients

    DEFF Research Database (Denmark)

    Stefansdottir, Jenna; Christensen, Erik; Schiødt, Frank Vinholt

    2017-01-01

    OBJECTIVE: Hepatocellular carcinoma (HCC) is a common cause of cancer, and most HCC patients have underlying cirrhosis. Retrospectively, we aimed to characterize patients with newly diagnosed HCC at a Danish hospital and to investigate survival and identify predictive factors for survival. METHODS...

  6. Glutathione treatment of hepatocellular carcinoma

    DEFF Research Database (Denmark)

    Dalhoff, K; Ranek, L; Mantoni, M

    1992-01-01

    This prospective study was undertaken to substantiate observations that glutathione (GSH) inhibits or reverses tumor growth in humans with hepatocellular carcinoma (HCC), a neoplasm with an extremely poor prognosis. Eight patients with biopsy-proven HCC not amenable to surgery were given 5 g of GSH...

  7. Huperzine A protects neural stem cells against Aβ-induced apoptosis in a neural stem cells and microglia co-culture system

    Science.gov (United States)

    Zhu, Ning; Lin, Jizong; Wang, Kewan; Wei, Meidan; Chen, Qingzhuang; Wang, Yong

    2015-01-01

    Objectives: This study aims to explore whether Huperzine A (HupA) could protect neural stem cells against amyloid beta-peptide Aβ induced apoptosis in a neural stem cells (NSCs) and microglia co-culture system. Methods: Rat NSCs and microglial cells were isolated, cultured and identified with immunofluorescence Assays (IFA). Co-culture systems of NSCs and microglial cells were employed using Transwell Permeable Supports. The effects of Aβ1-42 on NSCs were studied in 4 groups using co-culture systems: NSCs, Aβ+NSCs, co-culture and Aβ+co-culture groups. Bromodeoxyuridine (BrdU) incorporation and flow cytometry were utilized to assess the differences of proliferation, differentiation and apoptosis of NSCs between the groups. LQ test was performed to assess the amounts of IL-6, TNF-α and MIP-α secreted, and flow cytometry and Western blotting were used to assess apoptosis of NSCs and the expressions of Bcl-2 and Bax in each group. Results: IFA results showed that isolated rat NSCs were nestin-positive and microglial cells were CD11b/c-positive. Among all the groups, the Aβ+co-culture group has the lowest BrdU expression level, the lowest MAP2-positive, ChAT-positive cell counts and the highest NSC apoptosis rate. Smaller amounts of IL-6, TNF-α and MIP-α were being secreted by microglial cells in the HupA+Aβ+co-culture group compared with those in the Aβ+ co-culture group. Also the Bcl-2: Bax ratio was much higher in the HupA+Aβ+co-culture group than in the Aβ+co-culture group. Conclusions: HupA inhibits cell apoptosis through restraining microglia’s inflammatory response induced by Aβ1-42. PMID:26261518

  8. Astroglia overexpressing heme oxygenase-1 predispose co-cultured PC12 cells to oxidative injury.

    Science.gov (United States)

    Song, Linyang; Song, Wei; Schipper, Hyman M

    2007-08-01

    The mechanisms responsible for the progressive degeneration of dopaminergic neurons and pathologic iron deposition in the substantia nigra pars compacta of patients with Parkinson's disease (PD) remain unclear. Heme oxygenase-1 (HO-1), the rate-limiting enzyme in the oxidative degradation of heme to ferrous iron, carbon monoxide, and biliverdin, is upregulated in affected PD astroglia and may contribute to abnormal mitochondrial iron sequestration in these cells. To determine whether glial HO-1 hyper-expression is toxic to neuronal compartments, we co-cultured dopaminergic PC12 cells atop monolayers of human (h) HO-1 transfected, sham-transfected, or non-transfected primary rat astroglia. We observed that PC12 cells grown atop hHO-1 transfected astrocytes, but not the astroglia themselves, were significantly more susceptible to dopamine (1 microM) + H(2)O(2) (1 microM)-induced death (assessed by nuclear ethidium monoazide bromide staining and anti-tyrosine hydroxylase immunofluorescence microscopy) relative to control preparations. In the experimental group, PC12 cell death was attenuated significantly by the administration of the HO inhibitor, SnMP (1.5 microM), the antioxidant, ascorbate (200 microM), or the iron chelators, deferoxamine (400 microM), and phenanthroline (100 microM). Exposure to conditioned media derived from HO-1 transfected astrocytes also augmented PC12 cell killing in response to dopamine (1 microM) + H(2)O(2) (1 microM) relative to control media. In PD brain, overexpression of HO-1 in nigral astroglia and accompanying iron liberation may facilitate the bioactivation of dopamine to neurotoxic free radical intermediates and predispose nearby neuronal constituents to oxidative damage. (c) 2007 Wiley-Liss, Inc.

  9. Early bovine embryos regulate oviduct epithelial cell gene expression during in vitro co-culture.

    Science.gov (United States)

    Schmaltz-Panneau, Barbara; Cordova, Amanda; Dhorne-Pollet, Sophie; Hennequet-Antier, Christelle; Uzbekova, Sveltlana; Martinot, Emmanuelle; Doret, Sarah; Martin, Patrice; Mermillod, Pascal; Locatelli, Yann

    2014-10-01

    In mammals, the oviduct may participate to the regulation of early embryo development. In vitro co-culture of early bovine embryos with bovine oviduct epithelial cells (BOEC) has been largely used to mimic the maternal environment. However, the mechanisms of BOEC action have not been clearly elucidated yet. The aim of this study was to determine the response of BOEC cultures to the presence of developing bovine embryos. A 21,581-element bovine oligonucleotide array was used compare the gene expression profiles of confluent BOEC cultured for 8 days with or without embryos. This study revealed 34 differentially expressed genes (DEG). Of these 34 genes, IFI6, ISG15, MX1, IFI27, IFI44, RSAD2, IFITM1, EPSTI1, USP18, IFIT5, and STAT1 expression increased to the greatest extent due to the presence of embryos with a major impact on antiviral and immune response. Among the mRNAs at least 25 are already described as induced by interferons. In addition, transcript levels of new candidate genes involved in the regulation of transcription, modulation of the maternal immune system and endometrial remodeling were found to be increased. We selected 7 genes and confirmed their differential expression by quantitative RT-PCR. The immunofluorescence imaging of cellular localization of STAT1 protein in BOEC showed a nuclear translocation in the presence of embryos, suggesting the activation of interferon signaling pathway. This first systematic study of BOEC transcriptome changes in response to the presence of embryos in cattle provides some evidences that these cells are able to adapt their transcriptomic profile in response to embryo signaling. Copyright © 2014 Elsevier B.V. All rights reserved.

  10. Monte Carlo simulations of protein micropatterning in biomembranes: effects of immobile sticky obstacles

    International Nuclear Information System (INIS)

    Arnold, Andreas M; Sevcsik, Eva; Schütz, Gerhard J

    2016-01-01

    Single molecule trajectories of lipids and proteins can yield valuable information about the nanoscopic organization of the plasma membrane itself. The interpretation of such trajectories, however, is complicated, as the mobility of molecules can be affected by the presence of immobile obstacles, and the transient binding of the tracers to these obstacles. We have previously developed a micropatterning approach that allows for immobilizing a plasma membrane protein and probing the diffusional behavior of a putative interaction partner in living cells. Here, we provide guidelines on how this micropatterning approach can be extended to quantify interaction parameters between plasma membrane constituents in their natural environment. We simulated a patterned membrane system and evaluated the effect of different surface densities of patterned immobile obstacles on the relative mobility as well as the surface density of diffusing tracers. In the case of inert obstacles, the size of the obstacle can be assessed from its surface density at the percolation threshold, which in turn can be extracted from the diffusion behavior of the tracer. For sticky obstacles, 2D dissociation constants can be determined from the tracer diffusion or surface density. (paper)

  11. New technologies for large-scale micropatterning of functional nanocomposite polymers

    Science.gov (United States)

    Khosla, A.; Gray, B. L.

    2012-04-01

    We present a review of different micropatterning technologies for flexible elastomeric functional nanocomposites with a particular emphasis on mold material and processes for production of large size substrates. The functional polymers include electrically conducting and magnetic materials developed at the Micro-instrumentation Laboratory at Simon Fraser University, Canada. We present a chart that compares many of these different conductive and magnetic functional nanocomposites and their measured characteristics. Furthermore, we have previously reported hybrid processes for nanocomposite polymers micromolded against SU-8 photoepoxy masters. However, SU-8 is typically limited to substrate sizes that are compatible with microelectronics processing as a microelectronics uv-patterning step is typically involved, and de-molding problems are observed. Recently, we have developed new processes that address the problems faced with SU-8 molds. These new technologies for micropatterning nanocomposites involve new substrate materials. A low cost Poly(methyl methacrylate) (PMMA) microfabrication technology has been developed, which involves fabrication of micromold via either CO2 laser ablation or deep UV. We have previously reported this large-scale patterning technique using laser ablation. Finally, we compare the two processes for PMMA producing micromolds for nanocomposites.

  12. Novel micro-patterning processes for thin film NiTi vascular devices

    International Nuclear Information System (INIS)

    Chun, Y J; Mohanchandra, K P; Carman, G P; Levi, D S; Fishbein, M C

    2010-01-01

    In order to create microscale features in thin film NiTi for use in vascular endografts, a novel 'lift-off process' was developed for use with deep reactive ion etching. A wet etching approach is compared to two variations of this new 'lift-off' process. The first lift-off process (lift-off I) used Si posts to define the features of NiTi film deposited on the Si substrate. This method produced fractures in the NiTi when the film was released. The lift-off II process used Si islands as substrate for the film while the Si wafer defined the specific geometric features. Lift-off II process allowed for the creation of various shape patterns (i.e., ellipse, diamond, circle, square, etc) in the range of 5–180 µm. The lift-off II process produced smooth and well aligned micro-patterns in thin film NiTi without the undercutting found in wet etching techniques. The micro-patterned thin film NiTi formed from the lift-off II process was used to cover a stent. In vivo tests were performed to evaluate the endothelialization though patterned thin films. Angiography, histopathology and SEM showed patency of the artery and uniformly promoted endothelial layer covering without thrombosis in both a medium and small artery

  13. Adhesion and migration of CHO cells on micropatterned single layer graphene

    Science.gov (United States)

    Keshavan, S.; Oropesa-Nuñez, R.; Diaspro, A.; Canale, C.; Dante, S.

    2017-06-01

    Cell patterning technology on single layer graphene (SLG) is a fairly new field that can find applications in tissue engineering and biomaterial/biosensors development. Recently, we have developed a simple and effective approach for the fabrication of patterned SLG substrates by laser micromachining, and we have successfully applied it for the obtainment of geometrically ordered neural networks. Here, we exploit the same approach to investigate the generalization of the cell response to the surface cues of the fabricated substrates and, contextually, to quantify cell adhesion on the different areas of the patterns. To attain this goal, we tested Chinese hamster ovary (CHO) cells on PDL-coated micropatterned SLG substrates and quantified the adhesion by using single cell force spectroscopy (SCFS). Our results indicate higher cell adhesion on PDL-SLG, and, consequently, an initial CHO cell accumulation on the graphene areas, confirming the neuronal behaviour observed previously; interestingly, at later time point in culture, cell migration was observed towards the adjacent SLG ablated regions, which resulted more favourable for cell proliferation. Therefore, our findings indicate that the mechanism of interaction with the surface cues offered by the micropatterned substrates is strictly cell-type dependent.

  14. Removal of phenanthrene from soil by co-cultures of bacteria and fungi pregrown on sugarcane bagasse pith.

    Science.gov (United States)

    Chávez-Gómez, B; Quintero, R; Esparza-García, F; Mesta-Howard, A M; Zavala Díaz de la Serna, F J; Hernández-Rodríguez, C H; Gillén, T; Poggi-Varaldo, H M; Barrera-Cortés, J; Rodríguez-Vázquez, R

    2003-09-01

    Sixteen co-cultures composed of four bacteria and four fungi grown on sugarcane bagasse pith were tested for phenanthrene degradation in soil. The four bacteria were identified as Pseudomonas aeruginose, Ralstonia pickettii, Pseudomonas sp. and Pseudomonas cepacea. The four fungi were identified as: Penicillium sp., Trichoderma viride, Alternaria tenuis and Aspergillus terrus that were previously isolated from different hydrocarbon-contaminated soils. Fungi had a statistically significant positive (0.0001Co-cultures B. cepacea-Penicillium sp., R. pickettii-Penicillium sp., and P. aeruginose-Penicillium sp. exhibited synergism for phenanthrene removal, reaching 72.84+/-3.85%, 73.61+/-6.38% and 69.47+/-4.91%; in 18 days, respectively.

  15. Oxidative damage to DNA by diesel exhaust particle exposure in co-cultures of human lung epithelial cells and macrophages

    DEFF Research Database (Denmark)

    Jantzen, Kim; Roursgaard, Martin; Madsen, Claus Desler

    2012-01-01

    Studies in mono-culture of cells have shown that diesel exhaust particles (DEPs) increase the production of reactive oxygen species (ROS) and oxidative stress-related damage to DNA. However, the level of particle-generated genotoxicity may depend on interplay between different cell types, e.g. lung...... treatment with standard reference DEPs, SRM2975 and SRM1650b. The exposure to DEPs did not affect the colony-forming ability of A549 cells in co-culture with THP-1a cells. The DEPs generated DNA strand breaks and oxidatively damaged DNA, measured using the alkaline comet assay as formamidopyrimidine...... relationship between levels of respiration and ROS production. In conclusion, exposure of mono-cultured cells to DEPs generated oxidative stress to DNA, whereas co-cultures with macrophages had lower levels of oxidatively damaged DNA than A549 epithelial cells....

  16. Spontaneous decoration of Au nanoparticles on micro-patterned reduced graphene oxide shaped by focused laser beam

    International Nuclear Information System (INIS)

    Wan, Y. C.; Tok, E. S.; Teoh, H. F.; Sow, C. H.

    2015-01-01

    We report a facile, two-step method for the micro-landscaping of Au nanoparticles(NPs) on reduced graphene oxide (rGO) film en route to micro-patterned Au(NPs)-rGO hybrid functional materials. This method employs a focused laser beam to first locally convert GO to rGO before immersing the micro-patterned GO-rGO film into HAuCl 4 solution. The rGO micro-pattern, shaped by the focused laser beam, serves as nucleation sites for the reduction of Au ions. The reduction mechanism that governs the decoration of Au NPs on rGO films is akin to electroless deposition process. In this instance, surface charges that are formed during laser reduction of GO to rGO provide active nucleation sites for Au 3+ ions to form Au NPs when HAuCl 4 solution is introduced. The number density, the size, and size distribution of the Au NPs can thus be directly tuned and preferentially anchored onto the rGO micro-pattern by varying the incident laser power, the scanning speed of the laser, or the concentration of HAuCl 4 . The resulting hybrid materials can be used as a substrate for Surface Enhanced Raman Spectroscopy (SERS). Using Rhodamine 6G as the test subject, we found an improvement of SERS enhancement over bare rGO of up to four times, depending on the size of the Au NPs

  17. Spontaneous decoration of Au nanoparticles on micro-patterned reduced graphene oxide shaped by focused laser beam

    Energy Technology Data Exchange (ETDEWEB)

    Wan, Y. C.; Tok, E. S. [Department of Physics, National University of Singapore, 2 Science Drive 3, Singapore 117542 (Singapore); Teoh, H. F. [Graduate School of Integrative Sciences and Engineering, National University of Singapore, 28 Medical Drive, Singapore 117456 (Singapore); Sow, C. H. [Department of Physics, National University of Singapore, 2 Science Drive 3, Singapore 117542 (Singapore); Graduate School of Integrative Sciences and Engineering, National University of Singapore, 28 Medical Drive, Singapore 117456 (Singapore)

    2015-02-07

    We report a facile, two-step method for the micro-landscaping of Au nanoparticles(NPs) on reduced graphene oxide (rGO) film en route to micro-patterned Au(NPs)-rGO hybrid functional materials. This method employs a focused laser beam to first locally convert GO to rGO before immersing the micro-patterned GO-rGO film into HAuCl{sub 4} solution. The rGO micro-pattern, shaped by the focused laser beam, serves as nucleation sites for the reduction of Au ions. The reduction mechanism that governs the decoration of Au NPs on rGO films is akin to electroless deposition process. In this instance, surface charges that are formed during laser reduction of GO to rGO provide active nucleation sites for Au{sup 3+} ions to form Au NPs when HAuCl{sub 4} solution is introduced. The number density, the size, and size distribution of the Au NPs can thus be directly tuned and preferentially anchored onto the rGO micro-pattern by varying the incident laser power, the scanning speed of the laser, or the concentration of HAuCl{sub 4}. The resulting hybrid materials can be used as a substrate for Surface Enhanced Raman Spectroscopy (SERS). Using Rhodamine 6G as the test subject, we found an improvement of SERS enhancement over bare rGO of up to four times, depending on the size of the Au NPs.

  18. Effects of platelet rich plasma and chondrocyte co-culture on MSC chondrogenesis, hypertrophy and pathological responses

    OpenAIRE

    Ramezanifard, Rouhallah; Kabiri, Mahboubeh; Hanaee Ahvaz, Hana

    2017-01-01

    Regarding the inadequate healing capability of cartilage tissue, cell-based therapy is making the future of cartilage repair and regeneration. Mesenchymal stem cells (MSC) have shown great promise in cartilage regeneration. However, a yet-unresolved issue is the emergence of hypertrophic and pathologic markers during in vitro MSC chondrogenesis. Articular chondrocytes (AC) can suppress the undesired hypertrophy when co-cultured with MSC. On the other hand, platelet rich plasma (PRP), is consi...

  19. Co-cultures and cell sheet engineering as relevant tools to improve the outcome of bone tissue engineering strategies

    OpenAIRE

    Pirraco, Rogério

    2011-01-01

    Taking into consideration the complex biology of bone tissue it is quite clear that the understanding of the cellular interactions that regulate the homeostasis and regeneration of this remarkable tissue is essential for a successful Tissue Engineering strategy. The in vitro study of these cellular interactions relies on co-culture systems, a tremendously useful methodology where two or more cell types are cultured at the same time. Such strategy increases the complexity of typ...

  20. Slight changes in the mechanical stimulation affects osteoblast- and osteoclast-like cells in co-culture.

    Science.gov (United States)

    Kadow-Romacker, Anke; Duda, Georg N; Bormann, Nicole; Schmidmaier, Gerhard; Wildemann, Britt

    2013-12-01

    Osteoblast- and osteoclast-like cells are responsible for coordinated bone maintenance, illustrated by a balanced formation and resorption. Both parameters appear to be influenced by mechanical constrains acting on each of these cell types individually. We hypothesized that the interactions between both cell types are also influenced by mechanical stimulation. Co-cultures of osteoblast- and osteoclast-like cells were stimulated with 1,100 µstrain, 0.1 or 0.3 Hz for 1-5 min/day over 5 days. Two different setups depending on the differentiation of the osteoclast-like cells were used: i) differentiation assay for the fusion of pre-osteoclasts to osteoclasts, ii) resorption assay to determine the activity level of osteoclast-like cells. In the differentiation assay (co-culture of osteoblasts with unfused osteoclast precursor cells) the mechanical stimulation resulted in a significant decrease of collagen-1 and osteocalcin produced by osteoblast-like cells. Significantly more TRAP-iso5b was measured after stimulation for 3 min with 0.1 Hz, indicating enhanced osteoclastogenesis. In the resorption assay (co-culture of osteoblasts with fused osteoclasts) the stimulation for 3 min with 0.3 Hz significantly increased the resorption activity of osteoclasts measured by the pit formation and the collagen resorption. The same mechanical stimulation resulted in an increased collagen-1 production by the osteoblast-like cells. The ratio of RANKL/OPG was not different between the groups. These findings demonstrate that already small changes in duration or frequency of mechanical stimulation had significant consequences for the behavior of osteoblast- and osteoclast-like cells in co-culture, which partially depend on the differentiation status of the osteoclast-like cells.

  1. Transcriptome analysis of Acidovorax avenae subsp. avenae cultivated in vivo and co-culture with Burkholderia seminalis

    OpenAIRE

    Bin Li; Muhammad Ibrahim; Mengyu Ge; Zhouqi Cui; Guochang Sun; Fei Xu; Michael Kube

    2014-01-01

    Response of bacterial pathogen to environmental bacteria and its host is critical for understanding of microbial adaption and pathogenesis. Here, we used RNA-Seq to comprehensively and quantitatively assess the transcriptional response of Acidovorax avenae subsp. avenae strain RS-1 cultivated in vitro, in vivo and in co-culture with rice rhizobacterium Burkholderia seminalis R456. Results revealed a slight response to other bacteria, but a strong response to host. In particular, a large numbe...

  2. Permeability of PEGylated immunoarsonoliposomes through in vitro blood brain barrier-medulloblastoma co-culture models for brain tumor therapy.

    Science.gov (United States)

    Al-Shehri, Abdulghani; Favretto, Marco E; Ioannou, Panayiotis V; Romero, Ignacio A; Couraud, Pierre-Olivier; Weksler, Babette Barbash; Parker, Terry L; Kallinteri, Paraskevi

    2015-03-01

    Owing to restricted access of pharmacological agents into the brain due to blood brain barrier (BBB) there is a need: 1. to develop a more representative 3-D-co-culture model of tumor-BBB interaction to investigate drug and nanoparticle transport into the brain for diagnostic and therapeutic evaluation. 2. to address the lack of new alternative methods to animal testing according to replacement-reduction-refinement principles. In this work, in vitro BBB-medulloblastoma 3-D-co-culture models were established using immortalized human primary brain endothelial cells (hCMEC/D3). hCMEC/D3 cells were cultured in presence and in absence of two human medulloblastoma cell lines on Transwell membranes. In vitro models were characterized for BBB formation, zonula occludens-1 expression and permeability to dextran. Transferrin receptors (Tfr) expressed on hCMEC/D3 were exploited to facilitate arsonoliposome (ARL) permeability through the BBB to the tumor by covalently attaching an antibody specific to human Tfr. The effect of anticancer ARLs on hCMEC/D3 was assessed. In vitro BBB and BBB-tumor co-culture models were established successfully. BBB permeability was affected by the presence of tumor aggregates as suggested by increased permeability of ARLs. There was a 6-fold and 8-fold increase in anti-Tfr-ARL uptake into VC312R and BBB-DAOY co-culture models, respectively, compared to plain ARLs. The three-dimensional models might be appropriate models to study the transport of various drugs and nanocarriers (liposomes and immunoarsonoliposomes) through the healthy and diseased BBB. The immunoarsonoliposomes can be potentially used as anticancer agents due to good tolerance of the in vitro BBB model to their toxic effect.

  3. Immunoglobulin production in human mixed lymphocyte cultures: implications for co-cultures of cells from patients and healthy donors

    International Nuclear Information System (INIS)

    Ruemke, H.C.; Terpstra, F.G.; Huis, B.; Out, T.A.; Zeijlemaker, W.P.

    1982-01-01

    When human peripheral blood lymphocytes (PBL) are cultured in the presence of irradiated allogeneic lymphocytes, the resulting mixed lymphocyte reaction (MLR) leads to the secretion into the supernatant of substantial amounts of IgM and IgG, derived from nonirradiated responder B lymphocytes. Our data indicate that stimulation to Ig production by responder B cells may result from different types of of interactions. First, B cells and monocytes among the irradiated stimulator cells activate T responder B cells to produce Ig; second, ''responder'' B cells activate irradiated ''stimulator'' T cells, leading to a ''helper'' signal, back to the responder B cells and leading to Ig production. The latter system is radiosensitive, because allogeneic T cells, irradiated at a dose of 4000 rad or more, failed to induce Ig production by responder B cells. In some combinations of human allogeneic lymphocytes, the co-culture of the cells leads to inhibition of Ig production, both in the presence and in the absence of PWM. Thus, co-culture of allogeneic cells may cause ''positive'' as well as ''negative'' allogeneic effects. The implications of these findings for the interpretation of co-cultures that are aimed at establishing defects in lymphocytes from patients with, for example, immunodeficiencies, who fail to produce Ig in the presence of PWM are discussed

  4. Yeast Autolysis in Sparkling Wine Aging: Use of Killer and Sensitive Saccharomyces cerevisiae Strains in Co-Culture.

    Science.gov (United States)

    Lombardi, Silvia Jane; De Leonardis, Antonella; Lustrato, Giuseppe; Testa, Bruno; Iorizzo, Massimo

    2015-01-01

    Sparkling wines produced by traditional method owe their characteristics to secondary fermentation and maturation that occur during a slow ageing in bottles. Yeast autolysis plays an important role during the sparkling wine aging. Using a combination of killer and sensitive yeasts is possible to accelerate yeast autolysis and reduce maturing time. killer and sensitive Saccharomyces cerevisiae strains, separately and in co-cultures, were inoculated in base wine and bottled on pilot-plant scale. Commercial Saccaromyces bayanus strain was also investigated. Protein free amino acid and polysaccharides contents and sensory analysis were determined on the wine samples at 3, 6 and 9 months of aging. Yeast autolysis that occurs during the production of sparkling wines, obtained with co-cultures of killer and sensitive strains, has influenced free amino acids, total protein and polysaccharides content after 3 months aging time: sparkling wines, produced without the use of these yeasts, have reached the same results only after 9 months aging time. These results demonstrate that killer and sensitive yeasts in co-culture can accelerate the onset of autolysis in enological conditions, and has a positive effect on the quality of the aroma and flavor of sparkling wine. This paper offers an interesting biotechnological method to reduce production time of sparkling wine with economical benefits for the producers. We revised all patents relating to sparkling wine considering only those of interest for our study.

  5. Nicotine and caffeine alter the effects of the LPS- primed mesenchymal stem cells on the co-cultured neutrophils.

    Science.gov (United States)

    Abbasi, Ardeshir; Kukia, Nasim Rahmani; Froushani, Seyyed Meysam Abtahi; Hashemi, Seyed Mahmoud

    2018-04-15

    Mesenchymal stem cells (MSCs) express some of the nicotinic receptor subunits and adenosine receptors. The communication between tissue MSCs with neutrophils has been shown in previous studies. The aim of the present study is to determine the role of nicotine or caffeine on MSCs and its effects on neutrophils. After the isolation, MSCs were pulsed with LPS (10 ng/ml) for 1 h. Then, MSCs were incubated with different concentrations of caffeine (0.1, 0.5 and 1 mM) and or with different concentrations of nicotine (0.1, 0.5, and 1 μM) for 48 h. Afterwards, the medium was aspirated and the cells were used for co-culture experiment with neutrophil. The obtained data showed that LPS primed MSCs could decrease neutrophil vitality, whereas the treatment of MSCs with nicotine and/or especially a treatment with caffeine reverse this effect. Obtained data showed that when the LPS-primed MSCs were treated with nicotine or caffeine, the vitality of co-cultured neutrophils was significantly increased. The rate of the respiratory burst of neutrophils after co-culture by LPS-primed MSCs was decreased compared to the respiratory burst of neutrophil alone. Nicotine and/or caffeine treatment could reverse this reduction. Generally, these findings provide a new insight into understanding the anti-inflammatory and immunomodulatory effects of nicotine and caffeine. Copyright © 2018 Elsevier Inc. All rights reserved.

  6. Puerarin exhibits greater distribution and longer retention time in neurons than astrocytes in a co-cultured system

    Directory of Open Access Journals (Sweden)

    Shu-Yong Wei

    2015-01-01

    Full Text Available The phytoestrogen puerarin has been shown to protect neurons and astrocytes in the brain, and is therefore an attractive drug in the treatment of Alzheimer′s disease, Parkinson′s disease and cerebral ischemia. Whether puerarin exhibits the same biological processes in neurons and astrocytes in vitro has rarely been reported. In this study, cortical neurons and astrocytes of newborn Sprague-Dawley rats were separated, identified and co-cultured in a system based on Transwell membranes. The retention time and distribution of puerarin in each cell type was detected by fluorescence spectrophotometry and fluorescence microscope. The concentration of puerarin in both co-cultured and separately cultured neurons was greater than that of astrocytes. Puerarin concentration reached a maximum 20 minutes after it was added. At 60 minutes after its addition, a scant amount of drug was detected in astrocytes; however in both separately cultured and co-cultured neurons, the concentration of puerarin achieved a stable level of about 12.8 ng/mL. The results indicate that puerarin had a higher concentration and longer retention time in neurons than that observed in astrocytes.

  7. Temporally coordinated spiking activity of human induced pluripotent stem cell-derived neurons co-cultured with astrocytes.

    Science.gov (United States)

    Kayama, Tasuku; Suzuki, Ikuro; Odawara, Aoi; Sasaki, Takuya; Ikegaya, Yuji

    2018-01-01

    In culture conditions, human induced-pluripotent stem cells (hiPSC)-derived neurons form synaptic connections with other cells and establish neuronal networks, which are expected to be an in vitro model system for drug discovery screening and toxicity testing. While early studies demonstrated effects of co-culture of hiPSC-derived neurons with astroglial cells on survival and maturation of hiPSC-derived neurons, the population spiking patterns of such hiPSC-derived neurons have not been fully characterized. In this study, we analyzed temporal spiking patterns of hiPSC-derived neurons recorded by a multi-electrode array system. We discovered that specific sets of hiPSC-derived neurons co-cultured with astrocytes showed more frequent and highly coherent non-random synchronized spike trains and more dynamic changes in overall spike patterns over time. These temporally coordinated spiking patterns are physiological signs of organized circuits of hiPSC-derived neurons and suggest benefits of co-culture of hiPSC-derived neurons with astrocytes. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. Simultaneous and quantitative monitoring of co-cultured Pseudomonas aeruginosa and Staphylococcus aureus with antibiotics on a diffusometric platform

    Science.gov (United States)

    Chung, Chih-Yao; Wang, Jhih-Cheng; Chuang, Han-Sheng

    2017-04-01

    Successful treatments against bacterial infections depend on antimicrobial susceptibility testing (AST). However, conventional AST requires more than 24 h to obtain an outcome, thereby contributing to high patient mortality. An antibiotic therapy based on experiences is therefore necessary for saving lives and escalating the emergence of multidrug-resistant pathogens. Accordingly, a fast and effective drug screen is necessary for the appropriate administration of antibiotics. The mixed pathogenic nature of infectious diseases emphasizes the need to develop an assay system for polymicrobial infections. On this basis, we present a novel technique for simultaneous and quantitative monitoring of co-cultured microorganisms by coupling optical diffusometry with bead-based immunoassays. This simple integration simultaneously achieves a rapid AST analysis for two pathogens. Triple color particles were simultaneously recorded and subsequently analyzed by functionalizing different fluorescent color particles with dissimilar pathogen-specific antibodies. Results suggested that the effect of the antibiotic, gentamicin, on co-cultured Pseudomonas aeruginosa and Staphylococcus aureus was effectively distinguished by the proposed technique. This study revealed a multiplexed and time-saving (within 2 h) platform with a small sample volume (~0.5 μL) and a low initial bacterial count (50 CFU per droplet, ~105 CFU/mL) for continuously monitoring the growth of co-cultured microorganisms. This technique provides insights into timely therapies against polymicrobial diseases in the near future.

  9. Optimization of micropatterned poly(lactic-co-glycolic acid films for enhancing dorsal root ganglion cell orientation and extension

    Directory of Open Access Journals (Sweden)

    Ching-Wen Li

    2018-01-01

    Full Text Available Nerve conduits have been a viable alternative to the ‘gold standard’ autograft for treating small peripheral nerve gap injuries. However, they often produce inadequate functional recovery outcomes and are ineffective in large gap injuries. Ridge/groove surface micropatterning has been shown to promote neural cell orientation and guide growth. However, optimization of the ratio of ridge/groove parameters to promote orientation and extension for dorsal root ganglion (DRG cells on poly(lactic-co-glycolic acid (PLGA films has not been previously conducted. Photolithography and micro-molding were used to define various combinations of ridge/groove dimensions on PLGA films. The DRG cells obtained from chicken embryos were cultured on micropatterned PLGA films for cell orientation and migration evaluation. Biodegradation of the films occurred during the test period, however, this did not cause deformation or distortion of the micropatterns. Results from the DRG cell orientation test suggest that when the ridge/groove ratio equals 1 (ridge/groove width parameters are equal, i.e., 10 μm/10 μm (even, the degree of alignment depends on the size of the ridges and grooves, when the ratio is smaller than 1 (groove controlled the alignment increases as the ridge size decreases, and when the ratio is larger than 1 (ridge controlled, the alignment is reduced as the width of the grooves decreases. The migration rate and neurite extension of DRG neurons were greatest on 10 μm/10 μm and 30 μm/30 μm micropatterned PLGA films. Based on the data, the 10 μm/10 μm and 30 μm/30 μm micropatterned PLGA films are the optimized ridge/groove surface patterns for the construction of nerve repair devices.

  10. Multifunctional surfaces with biomimetic nanofibres and drug-eluting micro-patterns for infection control and bone tissue formation

    Directory of Open Access Journals (Sweden)

    XN Chen

    2012-09-01

    Full Text Available For long-term orthopaedic implants, the creation of a surface that is repulsive to bacteria while adhesive to tissue cells represents a promising strategy to control infection. To obtain such multifunctional surfaces, two possible approaches were explored to incorporate a model antibiotic, rifampicin (Rf, into the osteogenic polycaprolactone (PCL/chitosan (CHS biomimetic nanofibre meshes by (1 blending Rf into the electrospinning solutions and then electrospinning into nanofibres (i.e., Rf-incorporating fibres, or (2 depositing Rf-containing poly(D,L-lactic-co-glycolic acid (PLGA micro-patterns onto the PCL/chitosan nanofibre meshes via ink-jet printing (i.e., Rf-eluting micro-pattern/fibre. Rapid release of Rf from both meshes was measured even though a relatively slower release rate was obtained from the Rf-eluting micro-pattern ones. Antibacterial assay with Staphylococcus epidermidis showed that both mesh surfaces could effectively kill bacteria and prevent biofilm formation. However, only Rf-eluting micro-pattern meshes favoured the attachment, spreading and metabolic activity of preosteoblasts in the cell culture study. Furthermore, the Rf-eluting micro-pattern meshes could better support the osteogenic differentiation of preosteoblasts by up-regulating the gene expression of bone markers (type I collagen and alkaline phosphatase. Clearly, compared to Rf-incorporating nanofibre meshes, Rf-eluting micro-patterns could effectively prevent biofilm formation without sacrificing the osteogenic properties of PCL/chitosan nanofibre surfaces. This finding provides an innovative avenue to design multifunctional surfaces for enhancing bone tissue formation while controlling infection.

  11. Investigation of Legionella Contamination in Bath Water Samples by Culture, Amoebic Co-Culture, and Real-Time Quantitative PCR Methods.

    Science.gov (United States)

    Edagawa, Akiko; Kimura, Akio; Kawabuchi-Kurata, Takako; Adachi, Shinichi; Furuhata, Katsunori; Miyamoto, Hiroshi

    2015-10-19

    We investigated Legionella contamination in bath water samples, collected from 68 bathing facilities in Japan, by culture, culture with amoebic co-culture, real-time quantitative PCR (qPCR), and real-time qPCR with amoebic co-culture. Using the conventional culture method, Legionella pneumophila was detected in 11 samples (11/68, 16.2%). Contrary to our expectation, the culture method with the amoebic co-culture technique did not increase the detection rate of Legionella (4/68, 5.9%). In contrast, a combination of the amoebic co-culture technique followed by qPCR successfully increased the detection rate (57/68, 83.8%) compared with real-time qPCR alone (46/68, 67.6%). Using real-time qPCR after culture with amoebic co-culture, more than 10-fold higher bacterial numbers were observed in 30 samples (30/68, 44.1%) compared with the same samples without co-culture. On the other hand, higher bacterial numbers were not observed after propagation by amoebae in 32 samples (32/68, 47.1%). Legionella was not detected in the remaining six samples (6/68, 8.8%), irrespective of the method. These results suggest that application of the amoebic co-culture technique prior to real-time qPCR may be useful for the sensitive detection of Legionella from bath water samples. Furthermore, a combination of amoebic co-culture and real-time qPCR might be useful to detect viable and virulent Legionella because their ability to invade and multiply within free-living amoebae is considered to correlate with their pathogenicity for humans. This is the first report evaluating the efficacy of the amoebic co-culture technique for detecting Legionella in bath water samples.

  12. Investigation of Legionella Contamination in Bath Water Samples by Culture, Amoebic Co-Culture, and Real-Time Quantitative PCR Methods

    Directory of Open Access Journals (Sweden)

    Akiko Edagawa

    2015-10-01

    Full Text Available We investigated Legionella contamination in bath water samples, collected from 68 bathing facilities in Japan, by culture, culture with amoebic co-culture, real-time quantitative PCR (qPCR, and real-time qPCR with amoebic co-culture. Using the conventional culture method, Legionella pneumophila was detected in 11 samples (11/68, 16.2%. Contrary to our expectation, the culture method with the amoebic co-culture technique did not increase the detection rate of Legionella (4/68, 5.9%. In contrast, a combination of the amoebic co-culture technique followed by qPCR successfully increased the detection rate (57/68, 83.8% compared with real-time qPCR alone (46/68, 67.6%. Using real-time qPCR after culture with amoebic co-culture, more than 10-fold higher bacterial numbers were observed in 30 samples (30/68, 44.1% compared with the same samples without co-culture. On the other hand, higher bacterial numbers were not observed after propagation by amoebae in 32 samples (32/68, 47.1%. Legionella was not detected in the remaining six samples (6/68, 8.8%, irrespective of the method. These results suggest that application of the amoebic co-culture technique prior to real-time qPCR may be useful for the sensitive detection of Legionella from bath water samples. Furthermore, a combination of amoebic co-culture and real-time qPCR might be useful to detect viable and virulent Legionella because their ability to invade and multiply within free-living amoebae is considered to correlate with their pathogenicity for humans. This is the first report evaluating the efficacy of the amoebic co-culture technique for detecting Legionella in bath water samples.

  13. Impact of commercial cigarette smoke condensate on brain tissue co-cultured with astrocytes and blood-brain barrier endothelial cells.

    Science.gov (United States)

    Lee, Seon-Bong; Kim, Ju-Hyeong; Cho, Myung-Haing; Choe, Eun-Sang; Kim, Kwang-Sik; Shim, Soon-Mi

    2017-01-01

    The purpose of the current study was to investigate the effect of two commercial cigarette smoke condensates (CCSC) on oxidative stress and cell cytotoxicity in human brain (T98G) or astrocytes (U-373 MG) in the presence of human brain microvascular endothelial cells (HBMEC). Cell viability of mono-culture of T98G or U-373 MG was markedly decreased in a concentration-dependent manner, and T98G was more susceptible than U-373 MG to CCSC exposure. Cytotoxicity was less prominent when T98G was co-cultured with HBMEC than when T98G was co-cultured with U-373 MG. Significant reduction in trans-epithelial electric resistance (TEER), a biomarker of cellular integrity was noted in HBMEC co-cultured with T98G (HBMEC-T98G co-culture) and U-373 MG co-cultured with T98G (U-373 MG-T98G co-culture) after 24 or 48 hr CCSC exposure, respectively. TEER value of U-373 MG co-cultured with T98G (79-84%) was higher than HBMEC co-cultured with T98G (62-63%) within 120-hr incubation with CCSC. Reactive oxygen species (ROS) generated by CCSC in mono-culture of T98G and U-373 MG reached highest levels at 4 and 16 mg/ml, respectively. ROS production by T98G fell when co-cultured with HBMEC or U-373MG. These findings suggest that adverse consequences of CCSC treatment on brain cells may be protected by blood-brain barrier or astrocytes, but with chronic exposure toxicity may be worsened due to destruction of cellular integrity.

  14. Chondrogenesis of human adipose derived stem cells for future microtia repair using co-culture technique.

    Science.gov (United States)

    Goh, Bee See; Che Omar, Siti Nurhadis; Ubaidah, Muhammad Azhan; Saim, Lokman; Sulaiman, Shamsul; Chua, Kien Hui

    2017-04-01

    In conclusion, these result showed HADSCs could differentiate into chondrocytes-like cells, dependent on signaling induced by TGF-β3 and chondrocytes. This is a promising result and showed that HADSCs is a potential source for future microtia repair. The technique of co-culture is a positive way forward to assist the microtia tissue. Reconstructive surgery for the repair of microtia still remains the greatest challenge among the surgeons. Its repair is associated with donor-site morbidity and the degree of infection is inevitable when using alloplastic prosthesis with uncertain long-term durability. Thus, human adipose derived stem cells (HADSCs) can be an alternative cell source for cartilage regeneration. This study aims to evaluate the chondrogenic potential of HADSCs cultured with transforming growth factor-beta (TGF-β) and interaction of auricular chondrocytes with HADSCs for new cartilage generation. Multi-lineages differentiation features of HADSCs were monitored by Alcian Blue, Alizarin Red, and Oil Red O staining for chondrogenic, adipogenic, and osteogenic differentiation capacity, respectively. Further, HADSCs alone were culture in medium added with TGF-β3; and human auricular chondrocytes were interacted indirectly in the culture with and without TGF-βs for up to 21 days, respectively. Cell morphology and chondrogenesis were monitored by inverted microscope. For cell viability, Alamar Blue assay was used to measure the cell viability and the changes in gene expression of auricular chondrocyte markers were determined by real-time polymerase chain reaction analysis. For the induction of chondrogenic differentiation, HADSCs showed a feature of aggregation and formed a dense matrix of proteoglycans. Staining results from Alizirin Red and Oil Red O indicated the HADSCs also successfully differentiated into adipogenic and osteogenic lineages after 21 days. According to a previous study, HADSCs were strongly positive for the mesenchymal markers CD90, CD73

  15. Breast fibroblasts modulate epithelial cell proliferation in three-dimensional in vitro co-culture

    International Nuclear Information System (INIS)

    Sadlonova, Andrea; Novak, Zdenek; Johnson, Martin R; Bowe, Damon B; Gault, Sandra R; Page, Grier P; Thottassery, Jaideep V; Welch, Danny R; Frost, Andra R

    2005-01-01

    Stromal fibroblasts associated with in situ and invasive breast carcinoma differ phenotypically from fibroblasts associated with normal breast epithelium, and these alterations in carcinoma-associated fibroblasts (CAF) may promote breast carcinogenesis and cancer progression. A better understanding of the changes that occur in fibroblasts during carcinogenesis and their influence on epithelial cell growth and behavior could lead to novel strategies for the prevention and treatment of breast cancer. To this end, the effect of CAF and normal breast-associated fibroblasts (NAF) on the growth of epithelial cells representative of pre-neoplastic breast disease was assessed. NAF and CAF were grown with the nontumorigenic MCF10A epithelial cells and their more transformed, tumorigenic derivative, MCF10AT cells, in direct three-dimensional co-cultures on basement membrane material. The proliferation and apoptosis of MCF10A cells and MCF10AT cells were assessed by 5-bromo-2'-deoxyuridine labeling and TUNEL assay, respectively. Additionally, NAF and CAF were compared for expression of insulin-like growth factor II as a potential mediator of their effects on epithelial cell growth, by ELISA and by quantitative, real-time PCR. In relatively low numbers, both NAF and CAF suppressed proliferation of MCF10A cells. However, only NAF and not CAF significantly inhibited proliferation of the more transformed MCF10AT cells. The degree of growth inhibition varied among NAF or CAF from different individuals. In greater numbers, NAF and CAF have less inhibitory effect on epithelial cell growth. The rate of epithelial cell apoptosis was not affected by NAF or CAF. Mean insulin-like growth factor II levels were not significantly different in NAF versus CAF and did not correlate with the fibroblast effect on epithelial cell proliferation. Both NAF and CAF have the ability to inhibit the growth of pre-cancerous breast epithelial cells. NAF have greater inhibitory capacity than CAF

  16. Effects of Substrate and Co-Culture on Neural Progenitor Cell Differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Jones, Erin Boote [Iowa State Univ., Ames, IA (United States)

    2008-01-01

    In recent years the study of stem and progenitor cells has moved to the forefront of research. Since the isolation of human hematopoietic stem cells in 1988 and the subsequent discovery of a self renewing population of multipotent cells in many tissues, many researchers have envisioned a better understanding of development and potential clinical usage in intractable diseases. Both these goals, however, depend on a solid understanding of the intracellular and extracellular forces that cause stem cells to differentiate to a specific cell fate. Many diseases of large scale cell loss have been suggested as candidates for stem cell based treatments. It is proposed that replacing the function of the damaged or defective cells by specific differentiation of stem or progenitor cells could treat the disease. Before cells can be directed to specific lineages, the mechanisms of differentiation must be better understood. Differentiation in vivo is an intensively complex system that is difficult to study. The goal of this research is to develop further understanding of the effects of soluble and extracellular matrix (ECM) cues on the differentiation of neural progenitor cells with the use of a simplified in vitro culture system. Specific research objectives are to study the differentiation of neural progenitor cells in response to astrocyte conditioned medium and protein substrate composition and concentration. In an effort to reveal the mechanism of the conditioned medium interaction, a test for the presence of a feedback loop between progenitor cells and astrocytes is presented along with an examination of conditioned medium storage temperature, which can reveal enzymatic dependencies. An examination of protein substrate composition and concentration will help to reveal the role of any ECM interactions on differentiation. This thesis is organized into a literature review covering recent advances in use of external modulators of differentiation such as surface coatings, co-culture

  17. Feasibility of biohydrogen production from industrial wastes using defined microbial co-culture

    Directory of Open Access Journals (Sweden)

    Peng Chen

    2015-01-01

    Full Text Available BACKGROUND: The development of clean or novel alternative energy has become a global trend that will shape the future of energy. In the present study, 3 microbial strains with different oxygen requirements, including Clostridium acetobutylicum ATCC 824, Enterobacter cloacae ATCC 13047 and Kluyveromyces marxianus 15D, were used to construct a hydrogen production system that was composed of a mixed aerobic-facultative anaerobic-anaerobic consortium. The effects of metal ions, organic acids and carbohydrate substrates on this system were analyzed and compared using electrochemical and kinetic assays. It was then tested using small-scale experiments to evaluate its ability to convert starch in 5 L of organic wastewater into hydrogen. For the one-step biohydrogen production experiment, H1 medium (nutrient broth and potato dextrose broth was mixed directly with GAM broth to generate H2 medium (H1 medium and GAM broth. Finally, Clostridium acetobutylicum ATCC 824, Enterobacter cloacae ATCC 13047 and Kluyveromyces marxianus 15D of three species microbial co-culture to produce hydrogen under anaerobic conditions. For the two-step biohydrogen production experiment, the H1 medium, after cultured the microbial strains Enterobacter cloacae ATCC 13047 and Kluyveromyces marxianus 15D, was centrifuged to remove the microbial cells and then mixed with GAM broth (H2 medium. Afterward, the bacterial strain Clostridium acetobutylicum ATCC 824 was inoculated into the H2 medium to produce hydrogen by anaerobic fermentation. RESULTS: The experimental results demonstrated that the optimum conditions for the small-scale fermentative hydrogen production system were at pH 7.0, 35°C, a mixed medium, including H1 medium and H2 medium with 0.50 mol/L ferrous chloride, 0.50 mol/L magnesium sulfate, 0.50 mol/L potassium chloride, 1% w/v citric acid, 5% w/v fructose and 5% w/v glucose. The overall hydrogen production efficiency in the shake flask fermentation group was 33.7 m

  18. Effect and Mechanism of EGFL7 Downregulation in Human Osteosarcoma Cells on the Biological Function of Co-cultured HUVEC

    Directory of Open Access Journals (Sweden)

    Xia Li

    2018-03-01

    Full Text Available Background: Even though epidermal growth factor-like domain 7 is known to be overexpressed in osteosarcoma and is associated with poor clinical outcome, few reports are available regarding its mechanism. Aims: The objective of this study was to explore the effect and mechanism of downregulating epidermal growth factor-like domain 7 expression in a human osteosarcoma cell line on the biological function of co-cultured human umbilical vein endothelial cells. Study Design: Cell study. Methods: In the present study, human osteosarcoma cell lines U2OS, Saos-2, HOS, and MG63, and normal human osteoblasts were cultured in Dulbecco’s Modified Eagle Medium containing 10% fetal bovine serum and 1x antibiotics at 37 °C and 5% CO2 in an incubator. Of the four osteosarcoma cell lines, U2OS expresses the highest level of epidermal growth factor-like domain 7 mRNA as determined using quantitative reverse transcription polymerase chain reaction. With the knockdown of epidermal growth factor-like domain 7 in U2OS and human umbilical vein endothelial cells by lentivirus, the proliferation and apoptosis of U2OS and human umbilical vein endothelial cells were investigated using MTT and flow cytometry assays. After the co-culture of human umbilical vein endothelial cells and epidermal growth factor-like domain 7-knockdown U2OS, the in vitro effects on cell proliferation, apoptosis, adhesion, migration, and the angiogenic ability of human umbilical vein endothelial cells were detected using MTT, flow cytometry, Transwell, and tube formation assays, respectively. The expressions of phosphoinositide 3-kinase, phospho-Akt, total Akt, and vascular endothelial growth factor in human umbilical vein endothelial cells were detected using western blot assay. Results: Lentivirus with epidermal growth factor-like domain 7 shRNA could not significantly affect the proliferation and apoptosis of both U2OS and human umbilical vein endothelial cells, whereas the knockdown of

  19. Carbon nanoparticle doped micro-patternable nano-composites for wearable sensing applications (Conference Presentation)

    Science.gov (United States)

    Khosla, Ajit

    2017-04-01

    This talk focuses on preparation, characterization and micropatterning of electrically conducting KETJENBLACK carbon black nanoparticle (80 nm-diameter) doped Polydimethylsiloxane (PDMS) by employing extrusion mixing. Previously, we had reported fabrication of various micropatternable nanocomposites for wearable sensing applications vis solvent assisted ultrasonic mixing technique[1-16] . Extrusion mixing has an advantage as no organic solvents are used and homogenous dispersion of carbon nanoparticles is observed, which is confirmed by SEM analysis. The developed nanocomposite can be micropatterened using standard microfabrication techniques. It is also observed that percolation threshold occurs at 0.51 wt% of carbon nanoparticles in polymer matrix. Examples of developed nano-composites for wearable sensing applications for precision medicine will also be discussed. References: 1.http://summit.sfu.ca/item/12017 A. Khosla. Micropatternable multifunctional nanocomposite polymers for flexible soft MEMS applications. Diss. Applied Science: School of Engineering Science, 2011. 2. A. Khosla ; B. L. Gray; Fabrication of multiwalled carbon nanotube polydimethylsiloxne nanocomposite polymer flexible microelectrodes for microfluidics and MEMS. Proc. SPIE 7642, Electroactive Polymer Actuators and Devices (EAPAD) 2010, 76421V (April 09, 2010); doi:10.1117/12.847292. 3. Ang Li ; Ajit Khosla ; Connie Drewbrook ; Bonnie L. Gray; Fabrication and testing of thermally responsive hydrogel-based actuators using polymer heater elements for flexible microvalves. Proc. SPIE 7929, Microfluidics, BioMEMS, and Medical Microsystems IX, 79290G (February 14, 2011); doi:10.1117/12.873197. 4. Khosla, A. and Gray, B. L. (2010), Preparation, Micro-Patterning and Electrical Characterization of Functionalized Carbon-Nanotube Polydimethylsiloxane Nanocomposite Polymer. Macromol. Symp., 297: 210-218. doi:10.1002/masy.200900165 5. A. Khosla ; D. Hilbich ; C. Drewbrook ; D. Chung ; B. L. Gray; Large

  20. The development of a micropatterned electrode for studies of zinc electrodeposition

    Energy Technology Data Exchange (ETDEWEB)

    Sutija, Dave P. [Univ. of California, Berkeley, CA (United States); Muller, Rolf H. [Univ. of California, Berkeley, CA (United States); Tobias, Charles W. [Univ. of California, Berkeley, CA (United States)

    1986-12-01

    A micropatterned electrode was prepared for the study of electrocrystallization. Using microphotolithography, in conjunction with evaporation and pulse electrodeposition of thin films, a set of artificially roughened electrodes with hemispherical surface features five microns in diameter was developed. Voltammetric studies were conducted to determine the best electrode material. Gold, platinum, and various carbon surfaces were evaluated for zinc nucleation density and hydrogen overpotential. Surface homogeneity was examined by both light and scanning electron microscopy. Gold was determined to possess the best combination of material properties: chemical inertness, low melting point, and a high work function allowing underpotential deposition of zinc which reduces the rate of hydrogen evolution. Stripping coulometry was employed to determine zinc limiting currents, and evaluate effective diffusion coefficients in concentrated zinc chloride solutions. Although the method worked well for dilute zinc chloride and copper sulfate solutions, it failed at higher current densities; the emergence of surface roughness obscured actual limiting current plateaus.

  1. "Imaging" LEIS of micro-patterned solid oxide fuel cell electrodes

    Science.gov (United States)

    Druce, John; Simrick, Neil; Ishihara, Tatsumi; Kilner, John

    2014-08-01

    Understanding the kinetics of oxygen exchange between the gas phase and a ceramic electrode is key to optimising the performance of electrochemical energy conversion devices such as Solid Oxide Fuel Cells. Clearly the surface chemistry of these materials is important, and surface sensitive techniques such as Low Energy Ion Scattering (LEIS) can provide important compositional information key to unravelling electrode kinetics. In this work, we use high lateral resolution LEIS to perform local analyses of a micropatterned electrode structure, of the type often used for studies of the geometrical dependences of electrode performance. We find that the results are comparable to those for bulk materials, but detect evidence of cation interdiffusion from the electrode to the electrolyte. Finally, we note that this preliminary study could open the prospect of in situ measurements of cells near operating conditions.

  2. “Imaging” LEIS of micro-patterned solid oxide fuel cell electrodes

    Energy Technology Data Exchange (ETDEWEB)

    Druce, John, E-mail: john.druce@i2cner.kyushu-u.ac.jp [International Institute for Carbon Neutral Energy Research (wpi-I2CNER), Kyushu University, Fukuoka 819-0395 (Japan); Simrick, Neil [Department of Materials, Imperial College London, London SW7 2BP (United Kingdom); Ishihara, Tatsumi [International Institute for Carbon Neutral Energy Research (wpi-I2CNER), Kyushu University, Fukuoka 819-0395 (Japan); Kilner, John [International Institute for Carbon Neutral Energy Research (wpi-I2CNER), Kyushu University, Fukuoka 819-0395 (Japan); Department of Materials, Imperial College London, London SW7 2BP (United Kingdom)

    2014-08-01

    Understanding the kinetics of oxygen exchange between the gas phase and a ceramic electrode is key to optimising the performance of electrochemical energy conversion devices such as Solid Oxide Fuel Cells. Clearly the surface chemistry of these materials is important, and surface sensitive techniques such as Low Energy Ion Scattering (LEIS) can provide important compositional information key to unravelling electrode kinetics. In this work, we use high lateral resolution LEIS to perform local analyses of a micropatterned electrode structure, of the type often used for studies of the geometrical dependences of electrode performance. We find that the results are comparable to those for bulk materials, but detect evidence of cation interdiffusion from the electrode to the electrolyte. Finally, we note that this preliminary study could open the prospect of in situ measurements of cells near operating conditions.

  3. Design and Characterization of the VMM1 ASIC for Micropattern Gas Detectors

    CERN Document Server

    Metcalfe, J; The ATLAS collaboration; Fried, J; Li, S; Nambiar, N; Polychronakos, V; Vernon, E

    2013-01-01

    We present here the measurements of the first prototype VMM1 ASIC designed at Brookhaven National Laboratory in 130 nm CMOS and fabricated in spring 2012. The 64-channel ASIC features a novel design for use with several types of micropattern gas detectors. The data driven system measures peak amplitude and timing information in tracking mode and first channel hit address in trigger mode. Several programmable gain and integration times allows the flexibility to work with Micromegas, Thin Gap Chambers (TGCs), and Gas Electron Multiplier (GEM) detectors. The IC design architecture and features will be presented along with measurements characterizing the performance of the VMM1 such as noise, linearity of the response, time walk, and calibration range. The concept for use with Micromegas in ATLAS Upgrade will also be covered including characterization under test beam conditions.

  4. A Flexible and Highly Sensitive Piezoresistive Pressure Sensor Based on Micropatterned Films Coated with Carbon Nanotubes

    Directory of Open Access Journals (Sweden)

    Jia-lin Yao

    2016-01-01

    Full Text Available Excellent flexibility, high sensitivity, and low consumption are essential characteristics in flexible microtube pressure sensing occasion, for example, implantable medical devices, industrial pipeline, and microfluidic chip. This paper reports a flexible, highly sensitive, and ultrathin piezoresistive pressure sensor for fluid pressure sensing, whose sensing element is micropatterned films with conductive carbon nanotube layer. The flexible pressure sensor, the thickness of which is 40 ± 10 μm, could be economically fabricated by using biocompatible polydimethylsiloxane (PDMS. Experimental results show that the flexible pressure sensor has high sensitivity (0.047 kPa−1 in gas sensing and 5.6 × 10−3 kPa−1 in liquid sensing and low consumption (<180 μW, and the sensor could be used to measure the pressure in curved microtubes.

  5. 172 nm excimer VUV-triggered photodegradation and micropatterning of aminosilane films

    International Nuclear Information System (INIS)

    Elsner, Christian; Naumov, Sergej; Zajadacz, Joachim; Buchmeiser, Michael R.

    2009-01-01

    Emission from Xe 2 * excimers exhibiting photon energies between 7 and 10 eV can be used to induce strong surface modification effects on polymeric materials in the top 100 nm layer. In order to identify suitable monomers for this VUV-based process, the photodegradation mechanism of different organosilanes of the general structure R-CH 2 -Si(OCH 3 ) 3 was elucidated by quantum chemical calculations. Herein, the photodegradation of 3-aminopropyltrimethoxysilane films by the use of a 172 nm excimer lamp under different irradiation conditions is described and completed by micropatterning experiments. The presence of 1000-5000 ppm oxygen was found to promote the transformation process to an inorganic-like surface. The films obtained were analyzed by X-ray photoelectron spectroscopy, contact angle measurements and fluorescence microscopy after covalent attachment of a fluorescent dye to the remaining amino groups. Complementary, silver staining was used to visualize photopatterning.

  6. Micropatterning of poly(4-hydroxystyrene) by ion beam contact lithography for the control of cell adhesion

    International Nuclear Information System (INIS)

    Hwang, In Tae; Jung, Chan Hee; Choi, Jae Hak; Nho, Young Chang; Lee, Byoung Min; Hong, Sung Kwon

    2009-01-01

    In this study, we report on a simple method of micropatterning of cells by using ion beam contact lithography. Thin poly(4-hydroxystyrene) (Phds) films spin-coated on a silicon wafer were irradiated through a pattern mask in a contact mode with proton ions and then developed to generate the patterns of the Phds. Well-defined 50 μm line (pitch 150 μm) patterns were obtained without using any additives. The remaining thickness after development was increased with an increasing fluence up to 3 x 10 14 ions cm -2 after which it leveled off. The in-vitro cell culture test revealed that the cells were preferentially adhered to and proliferated only on the space regions between the Phds line patterns. Inhibition of cell adhesion on the Phds patterns could be due to antifouling property of the irradiated PHS

  7. Fast formation of hydrophilic and reactive polymer micropatterns by photocatalytic lithography method

    International Nuclear Information System (INIS)

    Chang, Chi-Jung; Wang, Chih-Feng; Chen, Jem-Kun; Hsieh, Chih-Chiao; Chen, Po-An

    2013-01-01

    An approach is developed for the fast formation of a hydrophilic pattern on superhydrophobic substrates with good contrast due to the large wettability contrast between superhydrophobic and superhydrophilic areas. It can be used for forming a polymer pattern with reactive functional groups. TiO 2 nanoparticles were grafted with long alkyl chains and then coated on substrates to produce superhydrophobic films. Photocatalytic degradation of the grafted alkyl chains was effected with UV light irradiation and resulted in transition from superhydrophobicity to superhydrophilicity. After UV light irradiation through a mask for 30 s, dyes or polymers were adsorbed on the photoinduced superhydrophilic areas to make micropatterns. The photoinduced superhydrophilic switching properties can be tuned by changing the alkyl chain length. The ninhydrin assay was adapted to identify free amino groups of polymers on the patterned area. Polymer patterns with free amino groups can be achieved.

  8. Development and characterization of micro-pattern gas detectors for intense beams of hadrons

    Energy Technology Data Exchange (ETDEWEB)

    Vandenbroucke, Maxence

    2012-07-02

    This thesis work is dedicated to the design, development and characterization of Micro-Pattern Gas Detectors. The performances of a Time Projection Chamber (TPC) equipped with a triple Gas Electron Multiplier (GEM) amplification structure are reported. The intrinsic ion backflow suppression of GEM foils drastically reduces the space charge produced by wire readout in traditional TPC. The GEM solution allows the operation of a TPC at much higher event rate. The second part of this thesis describes the development of a 40 x 40 cm{sup 2} Micromegas detector with a highly segmented central area. A reduction of discharges compared to conventional Micromegas detectors is needed for stable operation in intense beams of hadrons. Spark reduction technologies have been successfully studied and results are presented.

  9. Polymeric Shape-Memory Micro-Patterned Surface for Switching Wettability with Temperature

    Directory of Open Access Journals (Sweden)

    Nuria García-Huete

    2015-09-01

    Full Text Available An innovative method to switch the wettability of a micropatterned polymeric surface by thermally induced shape memory effect is presented. For this purpose, first polycyclooctene (PCO is crosslinked with dycumil peroxide (DCP and its melting temperature, which corresponds with the switching transition temperature (Ttrans, is measured by Dynamic Mechanical Thermal Analysis (DMTA in tension mode. Later, the shape memory behavior of the bulk material is analyzed under different experimental conditions employing a cyclic thermomechanical analysis (TMA. Finally, after creating shape memory micropillars by laser ablation of crosslinked thermo-active polycyclooctene (PCO, shape memory response and associated effect on water contact angle is analyzed. Thus, deformed micropillars cause lower contact angle on the surface from reduced roughness, but the original hydrophobicity is restored by thermally induced recovery of the original surface structure.

  10. Nano-structured micropatterns by combination of block copolymer self-assembly and UV photolithography

    International Nuclear Information System (INIS)

    Gorzolnik, B; Mela, P; Moeller, M

    2006-01-01

    A procedure for the fabrication of nano-structured micropatterns by direct UV photo-patterning of a monolayer of a self-assembled block copolymer/transition metal hybrid structure is described. The method exploits the selective photochemical modification of a self-assembled monolayer of hexagonally ordered block copolymer micelles loaded with a metal precursor salt. Solvent development of the monolayer after irradiation results in the desired pattern of micelles on the surface. Subsequent plasma treatment of the pattern leaves ordered metal nanodots. The presented technique is a simple and low-cost combination of 'top-down' and 'bottom-up' approaches that allows decoration of large areas with periodic and aperiodic patterns of nano-objects, with good control over two different length scales: nano- and micrometres

  11. Performance test of a micro-pattern stereo detector with two gas electron multipliers

    International Nuclear Information System (INIS)

    Barvich, T.; Bluem, P.; Erdmann, M.; Fahrer, M.; Kaercher, K.; Kuehn, F.; Moermann, D.; Mueller, Th.; Neuberger, D.; Roederer, F.; Simonis, H.J.; Skiba, A.; Thuemmel, W.H.; Weiler, Th.; Weseler, S.

    2002-01-01

    We report on the performance of a large micro-pattern detector with two gas electron multiplier foils and a two-layer readout structure at ground potential. The two readout layers each have a 406 μm pitch and cross at an effective angle of 6.7 deg. . This structure allows for two orthogonal coordinates to be determined. Using a muon beam at CERN together with a silicon tracking system, the position resolutions of the two coordinates are measured to be 50 μm and 1 mm respectively (1 standard deviation). The muon detection efficiency for the two-dimensional space points reaches 96%. The detector was found to be well operational over a wide range in the settings of the different electrical fields

  12. Fast formation of hydrophilic and reactive polymer micropatterns by photocatalytic lithography method

    Energy Technology Data Exchange (ETDEWEB)

    Chang, Chi-Jung, E-mail: changcj@fcu.edu.tw [Department of Chemical Engineering, Feng Chia University, 100, Wenhwa Road, Seatwen, Taichung 407, Taiwan (China); Wang, Chih-Feng [Department of Materials Science and Engineering, I-Shou University, 1, Syuecheng Road, Dashu District, Kaohsiung 840, Taiwan (China); Chen, Jem-Kun [Department of Materials Science and Engineering, National Taiwan University of Science and Technology, 43, Sec. 4, Keelung Road, Taipei 106, Taiwan (China); Hsieh, Chih-Chiao; Chen, Po-An [Department of Chemical Engineering, Feng Chia University, 100, Wenhwa Road, Seatwen, Taichung 407, Taiwan (China)

    2013-12-01

    An approach is developed for the fast formation of a hydrophilic pattern on superhydrophobic substrates with good contrast due to the large wettability contrast between superhydrophobic and superhydrophilic areas. It can be used for forming a polymer pattern with reactive functional groups. TiO{sub 2} nanoparticles were grafted with long alkyl chains and then coated on substrates to produce superhydrophobic films. Photocatalytic degradation of the grafted alkyl chains was effected with UV light irradiation and resulted in transition from superhydrophobicity to superhydrophilicity. After UV light irradiation through a mask for 30 s, dyes or polymers were adsorbed on the photoinduced superhydrophilic areas to make micropatterns. The photoinduced superhydrophilic switching properties can be tuned by changing the alkyl chain length. The ninhydrin assay was adapted to identify free amino groups of polymers on the patterned area. Polymer patterns with free amino groups can be achieved.

  13. Development and characterization of micro-pattern gas detectors for intense beams of hadrons

    International Nuclear Information System (INIS)

    Vandenbroucke, Maxence

    2012-01-01

    This thesis work is dedicated to the design, development and characterization of Micro-Pattern Gas Detectors. The performances of a Time Projection Chamber (TPC) equipped with a triple Gas Electron Multiplier (GEM) amplification structure are reported. The intrinsic ion backflow suppression of GEM foils drastically reduces the space charge produced by wire readout in traditional TPC. The GEM solution allows the operation of a TPC at much higher event rate. The second part of this thesis describes the development of a 40 x 40 cm 2 Micromegas detector with a highly segmented central area. A reduction of discharges compared to conventional Micromegas detectors is needed for stable operation in intense beams of hadrons. Spark reduction technologies have been successfully studied and results are presented.

  14. Direct writing of conductive silver micropatterns on flexible polyimide film by laser-induced pyrolysis of silver nanoparticle-dispersed film

    International Nuclear Information System (INIS)

    Aminuzzaman, Mohammod; Watanabe, Akira; Miyashita, Tokuji

    2010-01-01

    This article describes fabrication of Ag micropatterns on a flexible polyimide (PI) film by laser direct writing using an Ag nanoparticle-dispersed film as a precursor. Ag micropatterns are characterized by optical microscopy, atomic force microscopy (AFM), field emission scanning electron microscopy (FE-SEM), surface profilometry, and resistivity measurements. The line width of Ag micropatterns can be effectively controlled by altering the experimental parameters of laser direct writing especially laser intensity, objective lens, and laser beam scanning speed etc. Using an objective lens of 100x and laser intensity of 170.50 kW/cm 2 , Ag micropatterns with a line width of about 6 μm have been achieved. The Ag micropatterns show strong adhesion to polyimide surface as evaluated by Scotch-tape test. The resistivity of the Ag micropatterns is determined to be 4.1 x 10 -6 Ω cm using two-point probe method. This value is comparable with the resistivity of bulk Ag (1.6 x 10 -6 Ω cm).

  15. Primary Neuron/Astrocyte Co-Culture on Polyelectrolyte Multilayer Films: A Template for Studying Astrocyte-Mediated Oxidative Stress in Neurons**

    OpenAIRE

    Kidambi, Srivatsan; Lee, Ilsoon; Chan, Christina

    2008-01-01

    We engineered patterned co-cultures of primary neurons and astrocytes on polyelectrolyte multilayer (PEM) films without the aid of adhesive proteins/ligands to study the oxidative stress mediated by astrocytes on neuronal cells. A number of studies have explored engineering co-culture of neurons and astrocytes predominantly using cell lines rather than primary cells owing to the difficulties involved in attaching primary cells onto synthetic surfaces. To our knowledge this is the first demons...

  16. Characterization of A Three-Dimensional Organotypic Co-Culture Skin Model for Epidermal Differentiation of Rat Adipose-Derived Stem Cells.

    Science.gov (United States)

    Ghanavati, Zeinab; Orazizadeh, Mahmoud; Bayati, Vahid; Abbaspour, Mohammad Reza; Khorsandi, Layasadat; Mansouri, Esrafil; Neisi, Niloofar

    2016-01-01

    The organotypic co-culture is a well-known technique to examine cellular interactions and their roles in stem cell proliferation and differentiation. This study aims to evaluate the effects of dermal fibroblasts (DFs) on epidermal differentiation of adipose-derived stem cells (ASCs) using a three-dimensional (3D) organotypic co- culture technique. In this experimental research study, rat DFs and ASCs were isolated and cultured separately on electrospun polycaprolactone (PCL) matrices. The PCL matrices seeded by ASCs were superimposed on to the matrices seeded by DFs in order to create a 3D organotypic co-culture. In the control groups, PCL matrices seeded by ASCs were placed on matrices devoid of DFs. After 10 days, we assessed the expressions of keratinocyte-related genes by real-time reverse transcriptase-polymerase chain reaction (RT-PCR) and expression of pan-cytokeratin protein by immunofluorescence in the differentiated keratinocyte-like cells from co- culture and control groups. Keratinocyte-like cell morphologies were also observed by scanning electron microscopy (SEM). The early, intermediate, and terminal differentiation keratinocyte markers-Cytokeratin14, Filaggrin, and Involucrin significantly expressed in the co-culture groups com- pared to the control ones (P<0.05). We observed pan-cytokeratin in keratinocyte-like cells of both groups by immunofluorescence. SEM observation of the co-culture groups showed that the differentiated keratinocyte-like cells developed a polygonal cobblestone shape, considered characteristic of keratinocytes. The 3D organotypic co-culture bilayered construct that consisted of DFs and ASCs was an effective technique for epidermal differentiation of ASCs. This co-culture might be useful for epidermal differentiation of stem cells for future applications in skin regeneration.

  17. 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

  18. Contacting co-culture of human retinal microvascular endothelial cells alters barrier function of human embryonic stem cell derived retinal pigment epithelial cells.

    Science.gov (United States)

    Skottman, H; Muranen, J; Lähdekorpi, H; Pajula, E; Mäkelä, K; Koivusalo, L; Koistinen, A; Uusitalo, H; Kaarniranta, K; Juuti-Uusitalo, K

    2017-10-01

    Here we evaluated the effects of human retinal microvascular endothelial cells (hREC) on mature human embryonic stem cell (hESC) derived retinal pigment epithelial (RPE) cells. The hESC-RPE cells (Regea08/017, Regea08/023 or Regea11/013) and hREC (ACBRI 181) were co-cultured on opposite sides of transparent membranes for up to six weeks. Thereafter barrier function, small molecule permeability, localization of RPE and endothelial cell marker proteins, cellular fine structure, and growth factor secretion of were evaluated. After co-culture, the RPE specific CRALBP and endothelial cell specific von Willebrand factor were appropriately localized. In addition, the general morphology, pigmentation, and fine structure of hESC-RPE cells were unaffected. Co-culture increased the barrier function of hESC-RPE cells, detected both with TEER measurements and cumulative permeability of FD4 - although the differences varied among the cell lines. Co-culturing significantly altered VEGF and PEDF secretion, but again the differences were cell line specific. The results of this study showed that co-culture with hREC affects hESC-RPE functionality. In addition, co-culture revealed drastic cell line specific differences, most notably in growth factor secretion. This model has the potential to be used as an in vitro outer blood-retinal barrier model for drug permeability testing. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  19. 3D printed scaffolds of calcium silicate-doped β-TCP synergize with co-cultured endothelial and stromal cells to promote vascularization and bone formation.

    Science.gov (United States)

    Deng, Yuan; Jiang, Chuan; Li, Cuidi; Li, Tao; Peng, Mingzheng; Wang, Jinwu; Dai, Kerong

    2017-07-17

    Synthetic bone scaffolds have potential application in repairing large bone defects, however, inefficient vascularization after implantation remains the major issue of graft failure. Herein, porous β-tricalcium phosphate (β-TCP) scaffolds with calcium silicate (CS) were 3D printed, and pre-seeded with co-cultured human umbilical cord vein endothelial cells (HUVECs) and human bone marrow stromal cells (hBMSCs) to construct tissue engineering scaffolds with accelerated vascularization and better bone formation. Results showed that in vitro β-TCP scaffolds doped with 5% CS (5%CS/β-TCP) were biocompatible, and stimulated angiogenesis and osteogenesis. The results also showed that 5%CS/β-TCP scaffolds not only stimulated co-cultured cells angiogenesis on Matrigel, but also stimulated co-cultured cells to form microcapillary-like structures on scaffolds, and promoted migration of BMSCs by stimulating co-cultured cells to secrete PDGF-BB and CXCL12 into the surrounding environment. Moreover, 5%CS/β-TCP scaffolds enhanced vascularization and osteoinduction in comparison with β-TCP, and synergized with co-cultured cells to further increase early vessel formation, which was accompanied by earlier and better ectopic bone formation when implanted subcutaneously in nude mice. Thus, our findings suggest that porous 5%CS/β-TCP scaffolds seeded with co-cultured cells provide new strategy for accelerating tissue engineering scaffolds vascularization and osteogenesis, and show potential as treatment for large bone defects.

  20. 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

  1. Modeling long-term host cell-Giardia lamblia interactions in an in vitro co-culture system.

    Directory of Open Access Journals (Sweden)

    Bridget S Fisher

    Full Text Available Globally, there are greater than 700,000 deaths per year associated with diarrheal disease. The flagellated intestinal parasite, Giardia lamblia, is one of the most common intestinal pathogens in both humans and animals throughout the world. While attached to the gastrointestinal epithelium, Giardia induces epithelial cell apoptosis, disrupts tight junctions, and increases intestinal permeability. The underlying cellular and molecular mechanisms of giardiasis, including the role lamina propria immune cells, such as macrophages, play in parasite control or clearance are poorly understood. Thus far, one of the major obstacles in ascertaining the mechanisms of Giardia pathology is the lack of a functionally relevant model for the long-term study of the parasite in vitro. Here we report on the development of an in vitro co-culture model which maintains the basolateral-apical architecture of the small intestine and allows for long-term survival of the parasite. Using transwell inserts, Caco-2 intestinal epithelial cells and IC-21 macrophages are co-cultured in the presence of Giardia trophozoites. Using the developed model, we show that Giardia trophozoites survive over 21 days and proliferate in a combination media of Caco-2 cell and Giardia medium. Giardia induces apoptosis of epithelial cells through caspase-3 activation and macrophages do not abrogate this response. Additionally, macrophages induce Caco-2 cells to secrete the pro-inflammatory cytokines, GRO and IL-8, a response abolished by Giardia indicating parasite induced suppression of the host immune response. The co-culture model provides additional complexity and information when compared to a single-cell model. This model will be a valuable tool for answering long-standing questions on host-parasite biology that may lead to discovery of new therapeutic interventions.

  2. Dynamic flux balance modeling of microbial co-cultures for efficient batch fermentation of glucose and xylose mixtures.

    Science.gov (United States)

    Hanly, Timothy J; Henson, Michael A

    2011-02-01

    Sequential uptake of pentose and hexose sugars that compose lignocellulosic biomass limits the ability of pure microbial cultures to efficiently produce value-added bioproducts. In this work, we used dynamic flux balance modeling to examine the capability of mixed cultures of substrate-selective microbes to improve the utilization of glucose/xylose mixtures and to convert these mixed substrates into products. Co-culture simulations of Escherichia coli strains ALS1008 and ZSC113, engineered for glucose and xylose only uptake respectively, indicated that improvements in batch substrate consumption observed in previous experimental studies resulted primarily from an increase in ZSC113 xylose uptake relative to wild-type E. coli. The E. coli strain ZSC113 engineered for the elimination of glucose uptake was computationally co-cultured with wild-type Saccharomyces cerevisiae, which can only metabolize glucose, to determine if the co-culture was capable of enhanced ethanol production compared to pure cultures of wild-type E. coli and the S. cerevisiae strain RWB218 engineered for combined glucose and xylose uptake. Under the simplifying assumption that both microbes grow optimally under common environmental conditions, optimization of the strain inoculum and the aerobic to anaerobic switching time produced an almost twofold increase in ethanol productivity over the pure cultures. To examine the effect of reduced strain growth rates at non-optimal pH and temperature values, a break even analysis was performed to determine possible reductions in individual strain substrate uptake rates that resulted in the same predicted ethanol productivity as the best pure culture. © 2010 Wiley Periodicals, Inc.

  3. Effect of Bifidobacterium upon Clostridium difficile growth and toxicity when co-cultured in different prebiotic substrates

    Directory of Open Access Journals (Sweden)

    Lorena Valdés Varela

    2016-05-01

    Full Text Available The intestinal overgrowth of Clostridium difficile, often after disturbance of the gut microbiota by antibiotic treatment, leads to C. difficile infection (CDI which manifestation ranges from mild diarrhoea to life-threatening conditions. The increasing CDI incidence, not only in compromised subjects but also in traditionally considered low-risk populations, together with the frequent relapses of the disease, has attracted the interest for prevention/therapeutic options. Among these, probiotics, prebiotics or synbiotics constitute a promising approach. In this study we determined the potential of selected Bifidobacterium strains for the inhibition of C. difficile growth and toxicity in different carbon sources. We conducted co-cultures of the toxigenic strain C. difficile LMG21717 with four Bifidobacterium strains (Bifidobacterium longum IPLA20022, Bifidobacterium breve IPLA20006, Bifidobacterium bifidum IPLA20015, and Bifidobacterium animalis subsp. lactis Bb12 in the presence of various prebiotic substrates (Inulin, Synergy and Actilight or glucose, and compared the results with those obtained for the corresponding mono-cultures. C. difficile and bifidobacteria levels were quantified by qPCR; the pH and the production of short chain fatty acids was also determined. Moreover, supernatants of the cultures were collected to evaluate their toxicity using a recently developed model. Results showed that co-culture with B. longum IPLA20022 and B. breve IPLA20006 in the presence of short-chain fructooligosaccharides, but not of Inulin, as carbon source significantly reduced the growth of the pathogen. With the sole exception of B. animalis Bb12, whose growth was enhanced, the presence of C. difficile did not show major effects upon the growth of the bifidobacteria. In accordance with the growth data, B. longum and B. breve were the strains showing higher reduction in the toxicity of the co-culture supernatants.

  4. Comparison of the Growth of Lactobacillus delbrueckii, L. paracasei and L. plantarum on Inulin in Co-culture Systems.

    Science.gov (United States)

    Takagi, Risa; Tsujikawa, Yuji; Nomoto, Ryohei; Osawa, Ro

    2014-01-01

    Lactobacillus delbrueckii TU-1, which apparently takes intact inulin into its cells and then degrades it intracellularly, was co-cultured in vitro with L. paracasei KTN-5, an extracellular inulin degrader; or L. plantarum 22A-3, a strain that is able to utilize fructose but not inulin; or both in order to prequalify inulin as a prebiotic agent in vivo. When L. delbrueckii TU-1 was co-cultured with L. paracasei KTN-5 on fructose or inulin, the growth of L. delbrueckii TU-1 on inulin was markedly higher than that of L. paracasei KTN-5, whereas the growth of L. delbrueckii TU-1 on fructose was much lower than that of L. paracasei KTN-5. These results suggest that L. delbrueckii TU-1 and L. paracasei KTN-5 were efficient at utilizing inulin and fructose, respectively. When L. plantarum 22A-3 was co-cultured with L. delbrueckii TU-1 on inulin, the growth of L. plantarum 22A-3 was enhanced by L. paracasei KTN-5 but not by L. delbrueckii TU-1, suggesting that the fructose moiety that L. paracasei KTN-5 released temporarily into the medium was "scavenged" by L. plantarum 22A-3. Thus, L. delbrueckii TU-1, L. paracasei KTN-5, and L. plantarum 22A-3 were then cultured altogether on inulin. The growth of L. delbrueckii TU-1 was unaffected but that of L. paracasei KTN-5 was markedly suppressed. This evidence suggests that prebiotic use of inulin supported the selective growth of intracellular inulin degraders such as L. delbrueckii rather than extracellular inulin degraders such as L. paracasei in the host microbiota.

  5. Long-term culture and differentiation of porcine red bone marrow hematopoietic cells co-cultured with immortalized mesenchymal cells.

    Science.gov (United States)

    Garba, Abubakar; Acar, Delphine D; Roukaerts, Inge D M; Desmarets, Lowiese M B; Devriendt, Bert; Nauwynck, Hans J

    2017-09-01

    Mesenchymal cells are multipotent stromal cells with self-renewal, differentiation and immunomodulatory capabilities. We aimed to develop a co-culture model for differentiating hematopoietic cells on top of immortalized mesenchymal cells for studying interactions between hematopoietic and mesenchymal cells, useful for adequately exploring the therapeutic potential of mesenchymal cells. In this study, we investigated the survival, proliferation and differentiation of porcine red bone marrow hematopoietic cells co-cultured with immortalized porcine bone marrow mesenchymal cells for a period of five weeks. Directly after collection, primary porcine bone marrow mesenchymal cells adhered firmly to the bottom of the culture plates and showed a fibroblast-like appearance, one week after isolation. Upon immortalization, porcine bone marrow mesenchymal cells were continuously proliferating. They were positive for simian virus 40 (SV40) large T antigen and the mesenchymal cell markers CD44 and CD55. Isolated red bone marrow cells were added to these immortalized mesenchymal cells. Five weeks post-seeding, 92±6% of the red bone marrow hematopoietic cells were still alive and their number increased 3-fold during five weekly subpassages on top of the immortalized mesenchymal cells. The red bone marrow hematopoietic cells were originally small and round; later, the cells increased in size. Some of them became elongated, while others remained round. Tiny dendrites appeared attaching hematopoietic cells to the underlying immortalized mesenchymal cells. Furthermore, weekly differential-quick staining of the cells indicated the presence of monoblasts, monocytes, macrophages and lymphocytes in the co-cultures. At three weeks of co-culture, flow cytometry analysis showed an increased surface expression of CD172a, CD14, CD163, CD169, CD4 and CD8 up to 37±0.8%, 40±8%, 41±4%, 23±3% and 19±5% of the hematopoietic cells, respectively. In conclusion, continuous mesenchymal cell

  6. Three-Dimensional Organotypic Co-Culture Model of Intestinal Epithelial Cells and Macrophages to Study "Salmonella Enterica" Colonization Patterns

    Science.gov (United States)

    Ott, Mark; Yang, J; Barilla, J.; Crabbe, A.; Sarker, S. F.; Liu, Y.

    2017-01-01

    Three-dimensional/3-D organotypic models of human intestinal epithelium mimic the differentiated form and function of parental tissues often not exhibited by 2-D monolayers and respond to Salmonella in ways that reflect in vivo infections. To further enhance the physiological relevance of 3-D models to more closely approximate in vivo intestinal microenvironments during infection, we developed and validated a novel 3-D intestinal co-culture model containing multiple epithelial cell types and phagocytic macrophages, and applied to study enteric infection by different Salmonella pathovars.

  7. 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

  8. An improved in vitro blood-brain barrier model: rat brain endothelial cells co-cultured with astrocytes.

    Science.gov (United States)

    Abbott, N Joan; Dolman, Diana E M; Drndarski, Svetlana; Fredriksson, Sarah M

    2012-01-01

    In vitro blood-brain barrier (BBB) models using primary cultured brain endothelial cells are important for establishing cellular and molecular mechanisms of BBB function. Co-culturing with BBB-associated cells especially astrocytes to mimic more closely the in vivo condition leads to upregulation of the BBB phenotype in the brain endothelial cells. Rat brain endothelial cells (RBECs) are a valuable tool allowing ready comparison with in vivo studies in rodents; however, it has been difficult to obtain pure brain endothelial cells, and few models achieve a transendothelial electrical resistance (TEER, measure of tight junction efficacy) of >200 Ω cm(2), i.e. the models are still relatively leaky. Here, we describe methods for preparing high purity RBECs and neonatal rat astrocytes, and a co-culture method that generates a robust, stable BBB model that can achieve TEER >600 Ω cm(2). The method is based on >20 years experience with RBEC culture, together with recent improvements to kill contaminating cells and encourage BBB differentiation.Astrocytes are isolated by mechanical dissection and cell straining and are frozen for later co-culture. RBECs are isolated from 3-month-old rat cortices. The brains are cleaned of meninges and white matter and enzymatically and mechanically dissociated. Thereafter, the tissue homogenate is centrifuged in bovine serum albumin to separate vessel fragments from other cells that stick to the myelin plug. The vessel fragments undergo a second enzyme digestion to separate pericytes from vessels and break down vessels into shorter segments, after which a Percoll gradient is used to separate capillaries from venules, arterioles, and single cells. To kill remaining contaminating cells such as pericytes, the capillary fragments are plated in puromycin-containing medium and RBECs grown to 50-60% confluence. They are then passaged onto filters for co-culture with astrocytes grown in the bottom of the wells. The whole procedure takes ∼2

  9. Liquid chromatography-mass spectrometry for metabolic footprinting of co-cultures of lactic and propionic acid bacteria

    DEFF Research Database (Denmark)

    Honore, Anders H.; Thorsen, Michael; Skov, Thomas

    2013-01-01

    (UPLC) coupled to high-resolution mass spectrometry (MS) via electrospray ionisation (ESI) operated in both positive and negative modes is regarded as the optimum instrumental technique. The applicability of a range of liquid chromatographic techniques ranging from ion-pair (IPC) and hydrophilic...... could be a potent approach to elucidation of the mechanism. The purpose of this review is to discuss the two pre-requisites for such a study-the compound classes expected in the co-cultures, and on the basis of these, the most suitable analytical technique(s). Ultrahigh-performance liquid chromatography...

  10. Development of striatal patch/matrix organization in organotypic co-cultures of perinatal striatum, cortex and substantia nigra.

    Science.gov (United States)

    Snyder-Keller, A; Costantini, L C; Graber, D J

    2001-01-01

    Organotypic cultures of fetal or early postnatal striatum were used to assess striatal patch formation and maintenance in the presence or absence of dopaminergic and glutamatergic influences. Vibratome-cut slices of the striatum prepared from embryonic day 19 to postnatal day 4 rat pups were maintained in static culture on clear membrane inserts in Dulbecco's modified Eagle's medium/F12 (1:1) with 20% horse serum. Some were co-cultured with embryonic day 12-16 ventral mesencephalon and/or embryonic day 19 to postnatal day 4 cortex, which produced a dense dopaminergic innervation and a modest cortical innervation. Donors of striatal and cortical tissue were previously injected with bromo-deoxyuridine (BrdU) on embryonic days 13 and 14 in order to label striatal neurons destined to populate the patch compartment of the striatum. Patches of BrdU-immunoreactive cells were maintained in organotypic cultures of late prenatal (embryonic days 20-22) or early postnatal striatum in the absence of nigral dopaminergic or cortical glutamatergic influences. In slices taken from embryonic day 19 fetuses prior to the time of in vivo patch formation, patches were observed to form after 10 days in vitro, in 39% of nigral-striatal co-cultures compared to 6% of striatal slices cultured alone or in the presence of cortex only. Patches of dopaminergic fibers, revealed by tyrosine hydroxylase immunoreactivity, were observed in the majority of nigral-striatal co-cultures. Immunostaining for the AMPA-type glutamate receptor GluR1 revealed a dense patch distribution in nearly all cultures, which developed in embryonic day 19 cultures after at least six days in vitro. These findings indicate that striatal patch/matrix organization is maintained in organotypic culture, and can be induced to form in vitro in striatal slices removed from fetuses prior to the time of in vivo patch formation. Furthermore, dopaminergic innervation from co-cultured pieces of ventral mesencephalon enhances patch

  11. In vitro differentiation of bone marrow stromal cells into neurons and glial cells and differential protein expression in a two-compartment bone marrow stromal cell/neuron co-culture system.

    Science.gov (United States)

    Qi, Xu; Shao, Ming; Peng, Haisheng; Bi, Zhenggang; Su, Zhiqiang; Li, Hulun

    2010-07-01

    This study was performed to establish a bone marrow stromal cell (BMSC)/neuron two-compartment co-culture model in which differentiation of BMSCs into neurons could occur without direct contact between the two cell types, and to investigate protein expression changes during differentiation of this entirely BMSC-derived population. Cultured BMSCs isolated from Wistar rats were divided into three groups: BMSC culture, BMSC/neuron co-culture and BMSC/neuron two-compartment co-culture. Cells were examined for neuron-specific enolase (NSE) and glial fibrillary acidic protein (GFAP) expression. The electrophysiological behavior of the BMSCs was examined using patch clamping. Proteins that had significantly different expression levels in BMSCs cultured alone and co-cultured with neurons were studied using a protein chip-mass spectroscopy technique. Expression of NSE and GFAP were significantly higher in co-culture cells than in two-compartment co-culture cells, and significantly higher in both co-culture groups than in BMSCs cultured alone. Five proteins showed significant changes in expression during differentiation: TIP39_RAT and CALC_RAT underwent increases, and INSL6_RAT, PNOC_RAT and PCSK1_RAT underwent decreases in expression. We conclude that BMSCs can differentiate into neurons during both contact co-culture with neurons and two-compartment co-culture with neurons. The rate at which BMSCs differentiated into neurons was higher in contact co-culture than in non-contact co-culture.

  12. Current management of hepatocellular carcinoma

    Science.gov (United States)

    Tabrizian, Parissa; Roayaie, Sasan; Schwartz, Myron E

    2014-01-01

    Hepatocellular carcinoma (HCC) is the sixth most common cancer worldwide and leading cause of death among patients with cirrhosis. Treatment guidelines are based according to the Barcelona Clinic Liver Cancer staging system. The choice among therapeutic options that include liver resection, liver transplantation, locoregional, and systemic treatments must be individualized for each patient. The aim of this paper is to review the outcomes that can be achieved in the treatment of HCC with the heterogeneous therapeutic options currently available in clinical practice. PMID:25132740

  13. Transhemangioma Ablation of Hepatocellular Carcinoma

    International Nuclear Information System (INIS)

    Pua, Uei

    2012-01-01

    Radiofrequency ablation (RFA) is a well-established treatment modality in the treatment of early hepatocellular carcinoma (HCC) [1]. Safe trajectory of the RFA probe is crucial in decreasing collateral tissue damage and unwarranted probe transgression. As a percutaneous technique, however, the trajectory of the needle is sometimes constrained by the available imaging plane. The presence of a hemangioma beside an HCC is uncommon but poses the question of safety related to probe transgression. We hereby describe a case of transhemangioma ablation of a dome HCC.

  14. Transhemangioma Ablation of Hepatocellular Carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Pua, Uei, E-mail: druei@yahoo.com [Tan Tock Seng Hospital, Department of Diagnostic Radiology (Singapore)

    2012-12-15

    Radiofrequency ablation (RFA) is a well-established treatment modality in the treatment of early hepatocellular carcinoma (HCC) [1]. Safe trajectory of the RFA probe is crucial in decreasing collateral tissue damage and unwarranted probe transgression. As a percutaneous technique, however, the trajectory of the needle is sometimes constrained by the available imaging plane. The presence of a hemangioma beside an HCC is uncommon but poses the question of safety related to probe transgression. We hereby describe a case of transhemangioma ablation of a dome HCC.

  15. Assessment of cellular materials generated by co-cultured ‘inflamed’ and healthy periodontal ligament stem cells from patient-matched groups

    Energy Technology Data Exchange (ETDEWEB)

    Tang, Hao-Ning [State Key Laboratory of Military Stomatology & National Clinical Research Center for Oral Diseases & Shaanxi Key Laboratory of Stomatology, Department of Periodontology, School of Stomatology, Fourth Military Medical University, Xi' an (China); Department of Stomatology, The First Affiliated Hospital of the Chinese PLA General Hospital, Beijing 100048 (China); Xia, Yu [State Key Laboratory of Military Stomatology & National Clinical Research Center for Oral Diseases & Shaanxi Key Laboratory of Stomatology, Department of Periodontology, School of Stomatology, Fourth Military Medical University, Xi' an (China); Department of Stomatology, The 309th Hospital of Chinese People' s Liberation Army, Beijing 100091 (China); Xu, Jie; Tian, Bei-Min; Zhang, Xi-Yu [State Key Laboratory of Military Stomatology & National Clinical Research Center for Oral Diseases & Shaanxi Key Laboratory of Stomatology, Department of Periodontology, School of Stomatology, Fourth Military Medical University, Xi' an (China); Chen, Fa-Ming, E-mail: cfmsunhh@fmmu.edu.cn [State Key Laboratory of Military Stomatology & National Clinical Research Center for Oral Diseases & Shaanxi Key Laboratory of Stomatology, Department of Periodontology, School of Stomatology, Fourth Military Medical University, Xi' an (China)

    2016-08-01

    Recently, stem cells derived from the'inflamed’ periodontal ligament (PDL) tissue of periodontally diseased teeth (I-PDLSCs) have been increasingly suggested as a more readily accessible source of cells for regenerative therapies than those derived from healthy PDL tissue (H-PDLSCs). However, substantial evidence indicates that I-PDLSCs exhibit impaired functionalities compared with H-PDLSCs. In this study, patient-matched I-PDLSCs and H-PDLSCs were co-cultured at various ratios. Cellular materials derived from these cultures were investigated regarding their osteogenic potential in vitro and capacity to form new bone following in vivo transplantation. While patient-matched I-PDLSCs and H-PDLSCs could co-exist in co-culture systems, the proportion of I-PDLSCs tended to increase during in vitro incubation. Compared with H-PDLSC monoculture, the presence of I-PDLSCs in the co-cultures appeared to enhance the overall cell proliferation. Although not completely rescued, the osteogenic and regenerative potentials of the cellular materials generated by co-cultured I-PDLSCs and H-PDLSCs were significantly improved compared with those derived from I-PDLSC monocultures. Notably, cells in co-cultures containing either 50% I-PDLSCs plus 50% H-PDLSCs or 25% I-PDLSCs plus 75% H-PDLSCs expressed osteogenesis-related proteins and genes at levels similar to those expressed in H-PDLSC monocultures (P>0.05). Irrespective of the percentage of I-PDLSCs, robust cellular materials were obtained from co-cultures with 50% or more H-PDLSCs, which exhibited equivalent potential to form new bone in vivo compared with sheets generated by H-PDLSC monocultures. These data suggest that the co-culture of I-PDLSCs with patient-matched H-PDLSCs is a practical and effective method for increasing the overall osteogenic and regenerative potentials of resultant cellular materials. - Highlights: • Co-culturing H-PDLSCs with I-PDLSCs led to rapid cell expansion. • H-PDLSCs and I-PDLSCs co-cultured

  16. Assessment of cellular materials generated by co-cultured ‘inflamed’ and healthy periodontal ligament stem cells from patient-matched groups

    International Nuclear Information System (INIS)

    Tang, Hao-Ning; Xia, Yu; Xu, Jie; Tian, Bei-Min; Zhang, Xi-Yu; Chen, Fa-Ming

    2016-01-01

    Recently, stem cells derived from the'inflamed’ periodontal ligament (PDL) tissue of periodontally diseased teeth (I-PDLSCs) have been increasingly suggested as a more readily accessible source of cells for regenerative therapies than those derived from healthy PDL tissue (H-PDLSCs). However, substantial evidence indicates that I-PDLSCs exhibit impaired functionalities compared with H-PDLSCs. In this study, patient-matched I-PDLSCs and H-PDLSCs were co-cultured at various ratios. Cellular materials derived from these cultures were investigated regarding their osteogenic potential in vitro and capacity to form new bone following in vivo transplantation. While patient-matched I-PDLSCs and H-PDLSCs could co-exist in co-culture systems, the proportion of I-PDLSCs tended to increase during in vitro incubation. Compared with H-PDLSC monoculture, the presence of I-PDLSCs in the co-cultures appeared to enhance the overall cell proliferation. Although not completely rescued, the osteogenic and regenerative potentials of the cellular materials generated by co-cultured I-PDLSCs and H-PDLSCs were significantly improved compared with those derived from I-PDLSC monocultures. Notably, cells in co-cultures containing either 50% I-PDLSCs plus 50% H-PDLSCs or 25% I-PDLSCs plus 75% H-PDLSCs expressed osteogenesis-related proteins and genes at levels similar to those expressed in H-PDLSC monocultures (P>0.05). Irrespective of the percentage of I-PDLSCs, robust cellular materials were obtained from co-cultures with 50% or more H-PDLSCs, which exhibited equivalent potential to form new bone in vivo compared with sheets generated by H-PDLSC monocultures. These data suggest that the co-culture of I-PDLSCs with patient-matched H-PDLSCs is a practical and effective method for increasing the overall osteogenic and regenerative potentials of resultant cellular materials. - Highlights: • Co-culturing H-PDLSCs with I-PDLSCs led to rapid cell expansion. • H-PDLSCs and I-PDLSCs co-cultured

  17. A pilot-scale study of biohydrogen production from distillery effluent using defined bacterial co-culture

    Energy Technology Data Exchange (ETDEWEB)

    Vatsala, T.M.; Raj, S. Mohan; Manimaran, A. (Shri AMM Murugappa Chettiar Research Centre, Photosynthesis and Energy Division, Tharamani, Chennai, India, 600)

    2008-10-15

    We evaluated the feasibility of improving the scale of hydrogen (H{sub 2}) production from sugar cane distillery effluent using co-cultures of Citrobacter freundii 01, Enterobacter aerogenes E10 and Rhodopseudomonas palustris P2 at 100 m{sup 3} scale. The culture conditions at 100 ml and 2 L scales were optimized in minimal medium and we observed that the co-culture of the above three strains enhanced H{sub 2} productivity significantly. Results at the 100 m{sup 3} scale revealed a maximum of 21.38 kg of H{sub 2}, corresponding to 10692.6 mol, which was obtained through batch method at 40 h from reducing sugar (3862.3 mol) as glucose. The average yield of H{sub 2} was 2.76 mol mol{sup -1} glucose, and the rate of H{sub 2} production was estimated as 0.53 kg/100 m{sup 3}/h. Our results demonstrate the utility of distillery effluent as a source of clean alternative energy and provide insights into treatment for industrial exploitation. (author)

  18. Syntrophic co-culture of aerobic Bacillus and anaerobic Clostridium for bio-fuels and bio-hydrogen production

    Energy Technology Data Exchange (ETDEWEB)

    Chang, Jui-Jen; Ho, Cheng-Yu.; Chen, Wei-En; Huang, Chieh-Chen [Department of Life Sciences, National Chung Hsing University, Taichung (China); Chou, Chia-Hung; Lay, Jiunn-Jyi [Department of Science and Technology, National Kaohsiung First University, Kaohsiung (China)

    2008-10-15

    By using brewery yeast waste and microflora from rice straw compost, an anaerobic semi-solid bio-hydrogen-producing system has been established. For the purpose of industrialization, the major players of both aerobic and anaerobic bacterial strains in the system were isolated and their combination for an effective production of bio-hydrogen and other bio-fuels was examined in this study. The phylogenetic analysis found that four anaerobic isolates (Clostridium beijerinckii L9, Clostridium diolis Z2, Clostridium roseum Z5-1, and C. roseum W8) were highly related with each other and belongs to the cluster I clostridia family, the family that many of solvent-producing strains included. On the other hand, one of the aerobic isolates, the Bacillus thermoamylovorans strain I, shown multiple extracellular enzyme activities including lipase, protease, {alpha}-amylase, pectinase and cellulase, was suggested as a good partner for creating an anaerobic environment and pre-saccharification of substrate for those co-cultured solventogenic clostridial strain. Among these clostridial strains, though C. beijerinckii L9 do not show as many extracellular enzyme activities as Bacillus, but it performs the highest hydrogen-producing ability. The original microflora can be updated to a syntrophic bacterial co-culture system contended only with B. thermoamylovorans I and C. beijerinckii L9. The combination of aerobic Bacillus and anaerobic Clostridium may play the key role for developing the industrialized bio-fuels and bio-hydrogen-producing system from biomass. (author)

  19. Abilities of Co-cultures of Brown-Rot Fungus Fomitopsis pinicola and Bacillus subtilis on Biodegradation of DDT.

    Science.gov (United States)

    Sariwati, Atmira; Purnomo, Adi Setyo; Kamei, Ichiro

    2017-09-01

    DDT (1,1,1-trichloro-2,2-bis(4-chlorophenyl)ethane) is one of the pesticides that are hazardous for the environment and human health. Effective environmental-friendly treatment using co-cultures of fungi and bacteria is needed. In this study, the bacteria Bacillus subtilis at various volumes of 1, 3, 5, 7, and 10 mL (1 mL ≈ 6.7 × 10 8  CFU) were mixed into 10 mL of the brown-rot fungus Fomitopsis pinicola culture for degrading DDT during a 7-days incubation period. DDT was degraded by approximately 42% by F. pinicola during the 7-days incubation period. The addition of 10 mL of B. subtilis into F. pinicola culture showed the highest DDT degradation of approximately 86% during the 7-days incubation period. DDD (1,1-dichloro-2,2-bis(4-chlorophenyl)ethane), DDE (1,1-dichloro-2,2-bis(4-chlorophenyl)ethylene), and DDMU (1-chloro-2,2-bis(4-chlorophenyl)ethylene) were detected as metabolic products from DDT degradation by co-cultures of F. pinicola and B. subtilis. Transformation pathway was proposed in which DDT was transformed into three pathways as follows: (1) dechlorination to DDD, (2) dehydrochlorination to DDE, and (3) formation of DDMU.

  20. Interplay of differential cell mechanical properties, motility, and proliferation in emergent collective behavior of cell co-cultures

    Science.gov (United States)

    Sutter, Leo; Kolbman, Dan; Wu, Mingming; Ma, Minglin; Das, Moumita

    The biophysics of cell co-cultures, i.e. binary systems of cell populations, is of great interest in many biological processes including formation of embryos, and tumor progression. During these processes, different types of cells with different physical properties are mixed with each other, with important consequences for cell-cell interaction, aggregation, and migration. The role of the differences in their physical properties in their collective behavior remains poorly understood. Furthermore, until recently most theoretical studies of collective cell migration have focused on two dimensional systems. Under physiological conditions, however, cells often have to navigate three dimensional and confined micro-environments. We study a confined, three-dimensional binary system of interacting, active, and deformable particles with different physical properties such as deformability, motility, adhesion, and division rates using Langevin Dynamics simulations. Our findings may provide insights into how the differences in and interplay between cell mechanical properties, division, and motility influence emergent collective behavior such as cell aggregation and segregation experimentally observed in co-cultures of breast cancer cells and healthy breast epithelial cells. This work was partially supported by a Cottrell College Science Award.

  1. Modelling the tumour microenvironment in long-term microencapsulated 3D co-cultures recapitulates phenotypic features of disease progression.

    Science.gov (United States)

    Estrada, Marta F; Rebelo, Sofia P; Davies, Emma J; Pinto, Marta T; Pereira, Hugo; Santo, Vítor E; Smalley, Matthew J; Barry, Simon T; Gualda, Emilio J; Alves, Paula M; Anderson, Elizabeth; Brito, Catarina

    2016-02-01

    3D cell tumour models are generated mainly in non-scalable culture systems, using bioactive scaffolds. Many of these models fail to reflect the complex tumour microenvironment and do not allow long-term monitoring of tumour progression. To overcome these limitations, we have combined alginate microencapsulation with agitation-based culture systems, to recapitulate and monitor key aspects of the tumour microenvironment and disease progression. Aggregates of MCF-7 breast cancer cells were microencapsulated in alginate, either alone or in combination with human fibroblasts, then cultured for 15 days. In co-cultures, the fibroblasts arranged themselves around the tumour aggregates creating distinct epithelial and stromal compartments. The presence of fibroblasts resulted in secretion of pro-inflammatory cytokines and deposition of collagen in the stromal compartment. Tumour cells established cell-cell contacts and polarised around small lumina in the interior of the aggregates. Over the culture period, there was a reduction in oestrogen receptor and membranous E-cadherin alongside loss of cell polarity, increased collective cell migration and enhanced angiogenic potential in co-cultures. These phenotypic alterations, typical of advanced stages of cancer, were not observed in the mono-cultures of MCF-7 cells. The proposed model system constitutes a new tool to study tumour-stroma crosstalk, disease progression and drug resistance mechanisms. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

  2. Role of cell division and self-propulsion in self-organization of 2D cell co-cultures

    Science.gov (United States)

    Das, Moumita; Dey, Supravat; Wu, Mingming; Ma, Minglin

    Self-organization of cells is a key process in developmental and cancer biology. The differential adhesion hypothesis (DAH), which assumes cells as equilibrium liquid droplets and relates the self-assembly of cells to differences in inter-cellular adhesiveness, has been very successful in explaining cellular organization during morphogenesis where neighboring cells have the same non-equilibrium properties (motility, proliferation rate). However, recently it has been experimentally shown that for a co-culture of two different cell types proliferating at different rates, the resulting spatial morphologies cannot be explained using the DAH alone. Motivated by this, we develop and study a two-dimensional model of a cell co-culture that includes cell division and self-propulsion in addition to cell-cell adhesion, and systemically study how cells with significantly different adhesion, motility, and proliferation rate dynamically organize themselves in a spatiotemporal and context-dependent manner. Our results may help to understand how differential equilibrium and non-equilibrium properties cooperate and compete leading to different morphologies during tumor development, with important consequences for invasion and metastasis

  3. Astrocyte-neuron co-culture on microchips based on the model of SOD mutation to mimic ALS.

    Science.gov (United States)

    Kunze, Anja; Lengacher, Sylvain; Dirren, Elisabeth; Aebischer, Patrick; Magistretti, Pierre J; Renaud, Philippe

    2013-07-24

    Amyotrophic lateral sclerosis (ALS) is the most common motor neuron disease. ALS is believed to be a non-cell autonomous condition, as other cell types, including astrocytes, have been implicated in disease pathogenesis. Hence, to facilitate the development of therapeutics against ALS, it is crucial to better understand the interactions between astrocytes and neural cells. Furthermore, cell culture assays are needed that mimic the complexity of cell to cell communication at the same time as they provide control over the different microenvironmental parameters. Here, we aim to validate a previously developed microfluidic system for an astrocyte-neuron cell culture platform, in which astrocytes have been genetically modified to overexpress either a human wild-type (WT) or a mutated form of the super oxide dismutase enzyme 1 (SOD1). Cortical neural cells were co-cultured with infected astrocytes and studied for up to two weeks. Using our microfluidic device that prevents direct cell to cell contact, we could evaluate neural cell response in the vicinity of astrocytes. We showed that neuronal cell density was reduced by about 45% when neurons were co-cultured with SOD-mutant astrocytes. Moreover, we demonstrated that SOD-WT overexpressing astrocytes reduced oxidative stress on cortical neurons that were in close metabolic contact. In contrast, cortical neurons in metabolic contact with SOD-mutant astrocytes lost their synapsin protein expression after severe glutamate treatment, an indication of the toxicity potentiating effect of the SOD-mutant enzyme.

  4. Decreased hypertrophic differentiation accompanies enhanced matrix formation in co-cultures of outer meniscus cells with bone marrow mesenchymal stromal cells

    Science.gov (United States)

    2012-01-01

    Introduction The main objective of this study was to determine whether meniscus cells from the outer (MCO) and inner (MCI) regions of the meniscus interact similarly to or differently with mesenchymal stromal stem cells (MSCs). Previous study had shown that co-culture of meniscus cells with bone marrow-derived MSCs result in enhanced matrix formation relative to mono-cultures of meniscus cells and MSCs. However, the study did not examine if cells from the different regions of the meniscus interacted similarly to or differently with MSCs. Methods Human menisci were harvested from four patients undergoing total knee replacements. Tissue from the outer and inner regions represented pieces taken from one third and two thirds of the radial distance of the meniscus, respectively. Meniscus cells were released from the menisci after collagenase treatment. Bone marrow MSCs were obtained from the iliac crest of two patients after plastic adherence and in vitro culture until passage 2. Primary meniscus cells from the outer (MCO) or inner (MCI) regions of the meniscus were co-cultured with MSCs in three-dimensional (3D) pellet cultures at 1:3 ratio, respectively, for 3 weeks in the presence of serum-free chondrogenic medium containing TGF-β1. Mono-cultures of MCO, MCI and MSCs served as experimental control groups. The tissue formed after 3 weeks was assessed biochemically, histochemically and by quantitative RT-PCR. Results Co-culture of inner (MCI) or outer (MCO) meniscus cells with MSCs resulted in neo-tissue with increased (up to 2.2-fold) proteoglycan (GAG) matrix content relative to tissues formed from mono-cultures of MSCs, MCI and MCO. Co-cultures of MCI or MCO with MSCs produced the same amount of matrix in the tissue formed. However, the expression level of aggrecan was highest in mono-cultures of MSCs but similar in the other four groups. The DNA content of the tissues from co-cultured cells was not statistically different from tissues formed from mono-cultures of

  5. Transdifferentiation of mouse adipose-derived stromal cells into acinar cells of the submandibular gland using a co-culture system

    International Nuclear Information System (INIS)

    Lee, Jingu; Park, Sangkyu; Roh, Sangho

    2015-01-01

    A loss of salivary gland function often occurs after radiation therapy in head and neck tumors, though secretion of saliva by the salivary glands is essential for the health and maintenance of the oral environment. Transplantation of salivary acinar cells (ACs), in part, may overcome the side effects of therapy. Here we directly differentiated mouse adipose-derived stromal cells (ADSCs) into ACs using a co-culture system. Multipotent ADSCs can be easily collected from stromal vascular fractions of adipose tissues. The isolated ADSCs showed positive expression of markers such as integrin beta-1 (CD29), cell surface glycoprotein (CD44), endoglin (CD105), and Nanog. The cells were able to differentiate into adipocytes, osteoblasts, and neural-like cells after 14 days in culture. ADSCs at passage 2 were co-cultured with mouse ACs in AC culture medium using the double-chamber (co-culture system) to avoid mixing the cell types. The ADSCs in this co-culture system expressed markers of ACs, such as α-amylases and aquaporin5, in both mRNA and protein. ADSCs cultured in AC-conditioned medium also expressed AC markers. Cellular proliferation and senescence analyses demonstrated that cells in the co-culture group showed lower senescence and a higher proliferation rate than the AC-conditioned medium group at Days 14 and 21. The results above imply direct conversion of ADSCs into ACs under the co-culture system; therefore, ADSCs may be a stem cell source for the therapy for salivary gland damage. - Highlights: • ADSCs could transdifferentiate into acinar cells (ACs) using ACs co-culture (CCA). • Transdifferentiated ADSCs expressed ACs markers such as α-amylase and aquaporin5. • High proliferation and low senescence were presented in CCA at Day 14. • Transdifferentiation of ADSCs into ACs using CCA may be an appropriate method for cell-based therapy

  6. Transdifferentiation of mouse adipose-derived stromal cells into acinar cells of the submandibular gland using a co-culture system

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Jingu; Park, Sangkyu; Roh, Sangho, E-mail: sangho@snu.ac.kr

    2015-05-15

    A loss of salivary gland function often occurs after radiation therapy in head and neck tumors, though secretion of saliva by the salivary glands is essential for the health and maintenance of the oral environment. Transplantation of salivary acinar cells (ACs), in part, may overcome the side effects of therapy. Here we directly differentiated mouse adipose-derived stromal cells (ADSCs) into ACs using a co-culture system. Multipotent ADSCs can be easily collected from stromal vascular fractions of adipose tissues. The isolated ADSCs showed positive expression of markers such as integrin beta-1 (CD29), cell surface glycoprotein (CD44), endoglin (CD105), and Nanog. The cells were able to differentiate into adipocytes, osteoblasts, and neural-like cells after 14 days in culture. ADSCs at passage 2 were co-cultured with mouse ACs in AC culture medium using the double-chamber (co-culture system) to avoid mixing the cell types. The ADSCs in this co-culture system expressed markers of ACs, such as α-amylases and aquaporin5, in both mRNA and protein. ADSCs cultured in AC-conditioned medium also expressed AC markers. Cellular proliferation and senescence analyses demonstrated that cells in the co-culture group showed lower senescence and a higher proliferation rate than the AC-conditioned medium group at Days 14 and 21. The results above imply direct conversion of ADSCs into ACs under the co-culture system; therefore, ADSCs may be a stem cell source for the therapy for salivary gland damage. - Highlights: • ADSCs could transdifferentiate into acinar cells (ACs) using ACs co-culture (CCA). • Transdifferentiated ADSCs expressed ACs markers such as α-amylase and aquaporin5. • High proliferation and low senescence were presented in CCA at Day 14. • Transdifferentiation of ADSCs into ACs using CCA may be an appropriate method for cell-based therapy.

  7. Dynamics of bone marrow-derived endothelial progenitor cell/mesenchymal stem cell interaction in co-culture and its implications in angiogenesis

    International Nuclear Information System (INIS)

    Aguirre, A.; Planell, J.A.; Engel, E.

    2010-01-01

    Research highlights: → BM-EPCs and MSCs establish complex, self-organizing structures in co-culture. → Co-culture decreases proliferation by cellular self-regulatory mechanisms. → Co-cultured cells present an activated proangiogenic phenotype. → qRT-PCR and cluster analysis identify new target genes playing important roles. -- Abstract: Tissue engineering aims to regenerate tissues and organs by using cell and biomaterial-based approaches. One of the current challenges in the field is to promote proper vascularization in the implant to prevent cell death and promote host integration. Bone marrow endothelial progenitor cells (BM-EPCs) and mesenchymal stem cells (MSCs) are bone marrow resident stem cells widely employed for proangiogenic applications. In vivo, they are likely to interact frequently both in the bone marrow and at sites of injury. In this study, the physical and biochemical interactions between BM-EPCs and MSCs in an in vitro co-culture system were investigated to further clarify their roles in vascularization. BM-EPC/MSC co-cultures established close cell-cell contacts soon after seeding and self-assembled to form elongated structures at 3 days. Besides direct contact, cells also exhibited vesicle transport phenomena. When co-cultured in Matrigel, tube formation was greatly enhanced even in serum-starved, growth factor free medium. Both MSCs and BM-EPCs contributed to these tubes. However, cell proliferation was greatly reduced in co-culture and morphological differences were observed. Gene expression and cluster analysis for wide panel of angiogenesis-related transcripts demonstrated up-regulation of angiogenic markers but down-regulation of many other cytokines. These data suggest that cross-talk occurs in between BM-EPCs and MSCs through paracrine and direct cell contact mechanisms leading to modulation of the angiogenic response.

  8. Ultrasonographic finding of hepatocellular carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Ryu, Han Soo; Woo, Seong Ku; Lim, Jae Hoon; Ko, Young Tae; Kim, Ho Kyun; Kim, Soon Yong [Kyung Hee University Hospital, Seoul (Korea, Republic of)

    1983-12-15

    With the development of gray scale ultrasonography, detection and evaluation of hepatic parenchymal disease including space occupying lesion are easily performed and frequently used in the world. Thrity five cases of histopathologically proven and ultrasonographically suggested hepatocellular carcinoma are retrospectively studied. The results were as follows; 1. Ultrasonographic findings of hepatocellular carcinoma show hyperechoic pattern in 22 cases (63%), hypoechoic pattern in 2 cases (6%), and mixed pattern in 11 cases (31%). 2. The margin of tumor is ill-defined in 19 cases (54%) and well defined in16 cases (46%). 3. The size of tumor by sonographic measurement was large than 5 cm in diameter in 33 cases (94%). 4. The number of tumor is solitary in 19 cases and multiple in 16 cases. The sites of involved lobe were right lobe in 22 cases (63%), left lobe in 2 cases (6%), and both lobes in 11 cases (31%). 5. Associated sonographic findings were hepatomegaly with focal contour change in 25 cases (71%), splenomegaly in 16 cases (46%), cirrhosis of liver in 15 cases (43%), ascites in 11 cases (31%) and tumoral thrombosis in portal vein in 8 cases (23%). 6. The sex ratio is 6 : 1 male predominence and the age ranges from 32 to 76 years with highest incidence in 5th and 6th decades.

  9. Synchronous gastric neuroendocrine carcinoma and hepatocellular carcinoma

    DEFF Research Database (Denmark)

    Ewertsen, Caroline; Henriksen, Birthe Merete; Hansen, Carsten Palnæs

    2009-01-01

    of synchronous gastric NEC and hepatocellular carcinoma in a patient with several other precancerous lesions is presented. The patient had anaemia, and a gastric tumour and two duodenal polyps were identified on upper endoscopy. A CT scan of the abdomen revealed several lesions in the liver. The lesions were...... invisible on B-mode sonography and real-time sonography fused with CT was used to identify and biopsy one of the lesions. Histology showed hepatocellular carcinoma. A literature search showed that only one case of a hepatocellular carcinoma synchronous with a gastric NEC has been reported previously. TRIAL...

  10. Tribological improvement of moving microparts by application of thin films and micropatterning

    International Nuclear Information System (INIS)

    Bandorf, R; Paulkowski, D M; Schiffmann, K I; Kuester, R L A

    2008-01-01

    The relevance of active microsystems continually increases and with the expansion of applications also aspects of reliability and durability of the moving microparts. In particular, the microtribological improvement of active MEMS therefore was investigated and improved. On the one hand, the influence of different thin films was investigated; on the other hand, improvement by micropatterning of the tribological surfaces in combination with coatings was studied. Most promising results were gained using thin carbon-based films. The typical film thickness of the coatings was in a range of several tens up to a few hundred nanometres. The films were deposited by plasma processes varying different process parameters, mainly the applied bias voltage. Different doping elements (Si, Au and W) were added to the diamond-like carbon (DLC) films (a-C, a-C:H, a-C:H:Si, a-C:H:Au, and a-C:H:W). Different methods were used for characterizing the thin films. Under single asperity contact indentation and scratch tests were performed to determine microhardness, microwear and microfriction. An influence of the applied bias voltage on the micromechanical and microtribological properties was found. At low constant loads of 100 μN the more brittle a-C films showed an initially higher microwear volume than the more elastic a-C:H films. With increasing load the microwear of the a-C films showed less increase of microwear compared to a-C:H films. At low load in the range from 50 μN to a few 100 μN the friction coefficient in single-asperity contact decreased with increasing load. Reaching a critical load the behaviour changed and due to inelastic effects, the friction coefficient increased with further increasing load. For investigation of microabrasive wear under multi-asperity contact a specifically developed tester was used. Besides a ranking of different materials regarding their abrasive microwear resistance the influence of the substrate material on the resulting wear behaviour was

  11. A Highly Efficient Human Pluripotent Stem Cell Microglia Model Displays a Neuronal-Co-culture-Specific Expression Profile and Inflammatory Response

    Directory of Open Access Journals (Sweden)

    Walther Haenseler

    2017-06-01

    Full Text Available Microglia are increasingly implicated in brain pathology, particularly neurodegenerative disease, with many genes implicated in Alzheimer's, Parkinson's, and motor neuron disease expressed in microglia. There is, therefore, a need for authentic, efficient in vitro models to study human microglial pathological mechanisms. Microglia originate from the yolk sac as MYB-independent macrophages, migrating into the developing brain to complete differentiation. Here, we recapitulate microglial ontogeny by highly efficient differentiation of embryonic MYB-independent iPSC-derived macrophages then co-culture them with iPSC-derived cortical neurons. Co-cultures retain neuronal maturity and functionality for many weeks. Co-culture microglia express key microglia-specific markers and neurodegenerative disease-relevant genes, develop highly dynamic ramifications, and are phagocytic. Upon activation they become more ameboid, releasing multiple microglia-relevant cytokines. Importantly, co-culture microglia downregulate pathogen-response pathways, upregulate homeostatic function pathways, and promote a more anti-inflammatory and pro-remodeling cytokine response than corresponding monocultures, demonstrating that co-cultures are preferable for modeling authentic microglial physiology.

  12. Human nasal turbinates as a viable source of respiratory epithelial cells using co-culture system versus dispase-dissociation technique.

    Science.gov (United States)

    Noruddin, Nur Adelina Ahmad; Saim, Aminuddin B; Chua, Kien Hui; Idrus, Ruszymah

    2007-12-01

    To compare a co-culture system with a conventional dispase-dissociation method for obtaining functional human respiratory epithelial cells from the nasal turbinates for tissue engineering application. Human respiratory epithelial cells were serially passaged using a co-culture system and a conventional dispase-dissociation technique. The growth kinetics and gene expression levels of the cultured respiratory epithelial cells were compared. Four genes were investigated, namely cytokeratin-18, a marker for ciliated and secretory epithelial cells; cytokeratin-14, a marker for basal epithelial cells; MKI67, a proliferation marker; and MUC5B, a marker for mucin secretion. Immunocytochemical analysis was performed using monoclonal antibodies against the high molecular-weight cytokeratin 34 beta E12, cytokeratin 18, and MUC5A to investigate the protein expression from cultured respiratory epithelial cells. Respiratory epithelial cells cultured using both methods maintained polygonal morphology throughout the passages. At passage 1, co-cultured respiratory epithelial showed a 2.6-times higher growth rate compared to conventional dispase dissociation technique, and 7.8 times higher at passage 2. Better basal gene expression was observed by co-cultured respiratory epithelial cells compared to dispase dissociated cells. Immunocytochemical analyses were positive for the respiratory epithelial cells cultured using both techniques. Co-culture system produced superior quality of cultured human respiratory epithelial cells from the nasal turbinates as compared to dispase dissociation technique.

  13. Expression of human immunodeficiency virus (HIV) in naturally infected peripheral blood mononuclear cells: comparison of a standard co-culture technique with a newly developed microculture method.

    Science.gov (United States)

    Eberlein, B; Baur, A; Neundorfer, M; Jahn, G

    1991-05-01

    Peripheral blood mononuclear cells (PBMCs) from 29 patients infected with human immunodeficiency virus (HIV) were cultured by two different methods. One was the standard co-culture technique, the other a newly developed microculture method. In this assay 10(6) PBMCs were cultivated in 250 microliters medium, no activating agents or allogeneic cells were present. P24 antigen production measured by this method was found in 7 out of 11 PBMC cultures of patients in the Walter Reed (WR) stage 1 or 2, whereas only 4 samples were positive by the co-culture procedure. Cultures from patients in the later stages of the disease (WR 5/6) showed a higher p24 production by the co-culture method than by the microculture assay. It is assumed that rapidly growing HIV strains can be better assessed by the co-culture method which may select for these strains. P24 expression can be more easily obtained by the microculture technique even in cases where slowly replicating strains may be present. In conclusion, results from the microculture procedure described may be a useful supplementation to findings observed by the co-culture method.

  14. The effects of biomimetically conjugated VEGF on osteogenesis and angiogenesis of MSCs (human and rat) and HUVECs co-culture models.

    Science.gov (United States)

    Lü, Lanxin; Deegan, Anthony; Musa, Faiza; Xu, Tie; Yang, Ying

    2018-07-01

    The purpose of this work was to investigate if the biomimetically conjugated VEGF and HUVECs co-culture could modulate the osteogenic and angiogenic differentiation of MSCs derived from rat and human bone marrow (rMSCs and hMSCs). After treated by ammonia plasma, Poly(lactic-co-glycolic acid) (PLGA) electrospun nanofibers were immobilized with VEGF through heparin to fulfil the sustained release. The proliferation capacity of rMSCs and hMSCs on neat PLGA nanofibers (NF) and VEGF immobilized NF (NF-VEGF) surfaces were assessed by CCK-8 and compared when MSCs were mono-cultured and co-cultured with HUVECs. The effect of VEGF and HUVECs co-culturing on osteogenic and angiogenic differentiation of rMSCs and hMSCs were investigated by calcium deposits and CD31 expression on NF and NF-VEGF surfaces. The results indicated that VEGF has been biomimetically immobilized onto PLGA nanofibers surface and kept sustained release successfully. The CD31 staining results showed that both VEGF and HUVECs co-culture could enhance the angiogenesis of rMSCs and hMSCs. However, the proliferation and osteogenic differentiation of MSCs when cultured with VEGF and HUVECs showed a species dependent response. Taken together, VEGF immobilization and co-culture with HUVECs promoted angiogenesis of MSCs, indicating a good strategy for vascularization in bone tissue engineering. Copyright © 2018. Published by Elsevier B.V.

  15. Development of a co-culture of keratinocytes and immune cells for in vitro investigation of cutaneous sulfur mustard toxicity.

    Science.gov (United States)

    Balszuweit, Frank; Menacher, Georg; Bloemeke, Brunhilde; Schmidt, Annette; Worek, Franz; Thiermann, Horst; Steinritz, Dirk

    2014-11-05

    Sulfur mustard (SM) is a chemical warfare agent causing skin blistering, ulceration and delayed wound healing. Inflammation and extrinsic apoptosis are known to have an important role in SM-induced cytotoxicity. As immune cells are involved in those processes, they may significantly modulate SM toxicity, but the extent of those effects is unknown. We adapted a co-culture model of immortalized keratinocytes (HaCaT) and immune cells (THP-1) and exposed this model to SM. Changes in necrosis, apoptosis and inflammation, depending on SM challenge, absence or presence and number of THP-1 cells were investigated. THP-1 were co-cultured for 24h prior to SM exposure in order to model SM effects on immune cells continuously present in the skin. Our results indicate that the presence of THP-1 strongly increased necrosis, apoptosis and inflammation. This effect was already significant when the ratio of THP-1 and HaCaT cells was similar to the ratio of Langerhans immune cells and keratinocytes in vivo. Any further increases in the number of THP-1 had only slight additional effects on SM-induced cytotoxicity. In order to assess the effects of immune cells migrating into skin areas damaged by SM, we added non-exposed THP-1 to SM-exposed HaCaT. Those THP-1 had only slight effects on SM-induced cytotoxicity. Notably, in HaCaT exposed to 300μM SM, necrosis and inflammation were slightly reduced by adding intact THP-1. This effect was dependent on the number of immune cells, steadily increasing with the number of unexposed THP-1 added. In summary, we have demonstrated that (a) the presented co-culture is a robust model to assess SM toxicity and can be used to test the efficacy of potential antidotes in vitro; (b) immune cells, damaged by SM strongly amplified cytotoxicity, (c) in contrast, unexposed THP-1 (simulating migration of immune cells into affected areas after exposure in vivo) had no pronounced adverse, but exhibited some protective effects. Thus, protecting immune cells

  16. Biofilm formation by designed co-cultures of Caldicellulosiruptor species as a means to improve hydrogen productivity.

    Science.gov (United States)

    Pawar, Sudhanshu S; Vongkumpeang, Thitiwut; Grey, Carl; van Niel, Ed Wj

    2015-01-01

    Caldicellulosiruptor species have gained a reputation as being among the best microorganisms to produce hydrogen (H2) due to possession of a combination of appropriate features. However, due to their low volumetric H2 productivities (Q H2), Caldicellulosiruptor species cannot be considered for any viable biohydrogen production process yet. In this study, we evaluate biofilm forming potential of pure and co-cultures of Caldicellulosiruptor saccharolyticus and Caldicellulosiruptor owensensis in continuously stirred tank reactors (CSTR) and up-flow anaerobic (UA) reactors. We also evaluate biofilms as a means to retain biomass in the reactor and its influence on Q H2. Moreover, we explore the factors influencing the formation of biofilm. Co-cultures of C. saccharolyticus and C. owensensis form substantially more biofilm than formed by C. owensensis alone. Biofilms improved substrate conversion in both of the reactor systems, but improved the Q H2 only in the UA reactor. When grown in the presence of each other's culture supernatant, both C. saccharolyticus and C. owensensis were positively influenced on their individual growth and H2 production. Unlike the CSTR, UA reactors allowed retention of C. saccharolyticus and C. owensensis when subjected to very high substrate loading rates. In the UA reactor, maximum Q H2 (approximately 20 mmol · L(-1)  · h(-1)) was obtained only with granular sludge as the carrier material. In the CSTR, stirring negatively affected biofilm formation. Whereas, a clear correlation was observed between elevated (>40 μM) intracellular levels of the secondary messenger bis-(3'-5')-cyclic dimeric guanosine monophosphate (c-di-GMP) and biofilm formation. In co-cultures C. saccharolyticus fortified the trade of biofilm formation by C. owensensis, which was mediated by elevated levels of c-di-GMP in C. owensensis. These biofilms were effective in retaining biomass of both species in the reactor and improving Q H2 in a UA reactor using

  17. In vivo osteogenic differentiation of stem cells inside compartmentalized capsules loaded with co-cultured endothelial cells.

    Science.gov (United States)

    Correia, Clara R; Santos, Tírcia C; Pirraco, Rogério P; Cerqueira, Mariana T; Marques, Alexandra P; Reis, Rui L; Mano, João F

    2017-04-15

    Capsules coated with polyelectrolytes and co-encapsulating adipose stem (ASCs) and endothelial (ECs) cells with surface modified microparticles are developed. Microparticles and cells are freely dispersed in a liquified core, responsible to maximize the diffusion of essential molecules and allowing the geometrical freedom for the autonomous three-dimensional (3D) organization of cells. While the membrane wraps all the instructive cargo elements within a single structure, the microparticles provide a solid 3D substrate for the encapsulated cells. Our hypothesis is that inside this isolated biomimetic 3D environment, ECs would lead ASCs to differentiate into the osteogenic lineage to ultimately generate a mineralized tissue in vivo. For that, capsules encapsulating only ASCs (MONO capsules) or co-cultured with ECs (CO capsules) are subcutaneously implanted in nude mice up to 6weeks. Capsules implanted immediately after production or after 21days of in vitro osteogenic stimulation are tested. The most valuable outcome of the present study is the mineralized tissue in CO capsules without in vitro pre-differentiation, with similar levels compared to the pre-stimulated capsules in vitro. We believe that the proposed bioencapsulation strategy is a potent self-regulated system, which might find great applicability in bone tissue engineering. The diffusion efficiency of essential molecules for cell survival is a main issue in cell encapsulation. Former studies reported the superior biological outcome of encapsulated cells within liquified systems. However, most cells used in TE are anchorage-dependent, requiring a solid substrate to perform main cellular processes. We hypothesized that liquified capsules encapsulating microparticles are a promising attempt. Inspired by the multiphenotypic cellular environment of bone, we combine the concept of liquified capsules with co-cultures of stem and endothelial cells. After implantation, results show that co-cultured capsules

  18. Risks of Liver (Hepatocellular) Cancer Screening

    Science.gov (United States)

    ... cancer. Having hepatitis or cirrhosis can increase the risk of developing liver cancer. Anything that increases the ... clinical trials is available from the NCI website . Risks of Liver (Hepatocellular) Cancer Screening Key Points Screening ...

  19. Capability of Sputtered Micro-patterned NiTi Thick Films

    Science.gov (United States)

    Bechtold, Christoph; Lima de Miranda, Rodrigo; Quandt, Eckhard

    2015-09-01

    Today, most NiTi devices are manufactured by a combination of conventional metal fabrication steps, e.g., melting, extrusion, cold working, etc., and are subsequently structured by high accuracy laser cutting. This combination has been proven to be very successful; however, there are several limitations to this fabrication route, e.g., in respect to the fabrication of more complex device designs, device miniaturization or the combination of different materials for the integration of further functionality. These issues have to be addressed in order to develop new devices and applications. The fabrication of micro-patterned films using magnetron sputtering, UV lithography, and wet etching has great potential to overcome limitations of conventional device manufacturing. Due to its fabrication characteristics, this method allows the production of devices with complex designs, high structural accuracy, smooth edge profile, at layer thicknesses up to 75 µm. The aim of this study is to present recent developments in the field of NiTi thin film technology, its advantages and limitations, as well as new possible applications in the medical and in non-medical fields. These developments include among others NiTi scaffold structures covered with NiTi membranes for their potential use as filters, heart valve components or aneurysm treatments, as well as micro-actuators for consumable electronics or automotive applications.

  20. Effects of UV laser micropatterning on frictional performance of diamond-like nanocomposite films

    Science.gov (United States)

    Zavedeev, Evgeny V.; Zilova, Olga S.; Shupegin, Mikhail L.; Barinov, Alexej D.; Arutyunyan, Natalia R.; Roch, Teja; Pimenov, Sergei M.

    2016-11-01

    We report on UV laser modification and micropatterning of diamond-like nanocomposite (DLN) films (a-C:H,Si:O) with nanosecond pulses and effects of laser surface microstructuring on the frictional performance of DLN films on the nano- and macroscale. A technique of direct laser interference patterning was applied to produce arrays of periodic linear microstructures on the DLN films. The UV laser irradiation was performed at low fluences corresponding to the regime of surface graphitization and incipient ablation. At the initial stage of the thin film modification, the laser-induced spallation and graphitization in the surface layers were found to strongly influence the nanoscale topography and mechanical properties of the DLN surface. Frictional properties of the laser-patterned DLN films were studied using (1) atomic force microscopy in lateral force mode and (2) a ball-on-flat tribometer under linear reciprocating sliding against a 100Cr6 steel ball. The lateral force microscopy measurements revealed that the laser-irradiated regions were characterized by increased friction forces due to microspallation effects and enhanced surface roughness, correlating with tribotests at the initial stage of sliding. During prolonged sliding in ambient air, both the original and laser-patterned DLN surfaces exhibited low-friction performance at the friction coefficient of 0.07-0.08.

  1. Enhancement of silicon using micro-patterned surfaces of thin films

    Directory of Open Access Journals (Sweden)

    E Kaivosoja

    2010-04-01

    Full Text Available Micro-textured biomaterials might enhance cytocompatibility of silicon-based micro-electro-mechanical system (bio-MEMS dummies. Photolithography-physical vapour deposition was used to produce diamond-like carbon (DLC or Ti squares and circles on silicon, and also their inverse replicas; then DLC and Ti were compared for their guiding potential, using a SaOS-2 cell model. Scanning electron microscopy at 48 hours indicated cells were well-spread on large-sized patterns (several cells on one pattern and assumed the geometrical architecture of underlying features. Medium-sized patterns (slightly smaller than solitary indicator cells were inhabited by singular cells, which stretched from one island to another, assuming longitudinal or branching morphologies. On small-sized patterns (much smaller than individual cells cells covered large micro-textured areas, but cellular filopodia bypassed the bare silicon. Immunofluorescence and confocal laser scanning microscopy indicated that the actin cytoskeleton and vinculin-containing adhesion junctions were present on the patterned areas, but not on the bare silicon. Cell density/coverage disclosed a 3.4-3.7-fold preference for the biomaterial patterns over silicon substrate (p < 0.001. Differences in the cellular response between materials were lost at 120 hours when cells were confluent. The working hypothesis was proven; enhancement by micro-patterning depends on the pattern size, shape and material and can be used to improve biocompatibility during the initial integration phase of the device.

  2. Micropatterned 2D Hybrid Perovskite Thin Films with Enhanced Photoluminescence Lifetimes.

    Science.gov (United States)

    Kamminga, Machteld E; Fang, Hong-Hua; Loi, Maria Antonietta; Ten Brink, Gert H; Blake, Graeme R; Palstra, Thomas T M; Ten Elshof, Johan E

    2018-04-18

    The application of luminescent materials in display screens and devices requires micropatterned structures. In this work, we have successfully printed microstructures of a two-dimensional (2D), orange-colored organic/inorganic hybrid perovskite ((C 6 H 5 CH 2 NH 3 ) 2 PbI 4 ) using two different soft lithography techniques. Notably, both techniques yield microstructures with very high aspect ratios in the range of 1.5-1.8. X-ray diffraction reveals a strong preferential orientation of the crystallites along the c-axis in both patterned structures, when compared to nonpatterned, drop-casted thin films. Furthermore, (time-resolved) photoluminescence (PL) measurements reveal that the optical properties of (C 6 H 5 CH 2 NH 3 ) 2 PbI 4 are conserved upon patterning. We find that the larger grain sizes of the patterned films with respect to the nonpatterned film give rise to an enhanced PL lifetime. Thus, our results demonstrate easy and cost-effective ways to manufacture patterns of 2D organic/inorganic hybrid perovskites, while even improving their optical properties. This demonstrates the potential use of color-tunable 2D hybrids in optoelectronic devices.

  3. High rate behavior and discharge limits in micro-pattern detectors

    CERN Document Server

    Bressan, A; Pagano, P; Ropelewski, Leszek; Sauli, Fabio; Biagi, S F; Buzulutskov, A F; Gruwé, M; De Lentdecker, G; Mörmann, D; Sharma, A

    1999-01-01

    We present and discuss a set of systematic measurements, carried out with gaseous proportional micro-pattern detectors, in order to assess their maximum gain when irradiated with high-rate soft X-rays and heavily ionizing alpha particles. The inventory of detectors tested includes: micro-strip micromegas, micro-dot, gas electron multiplier, CAT (compteur à trous), trench (or groove), micro-CAT (or WELL) detectors, as well as systems with two elements of gaseous amplification in cascade. We confirm the general trend of all single-stage detectors to follow Raether's criterion, i.e. a spontaneous transition from avalanche to streamer, followed by a discharge, when the avalanche size reaches a value of a few 10 7 ; a noticeable exception is the micro-dot counter holding more than 10 8. In multiple structures, where the gain under irradiation is increased by at least one order of magnitude; we speculate this to be a consequence of a voltage dependence of Raether's limit, larger for low operating potentials. Our c...

  4. Cubic Gallium Nitride on Micropatterned Si (001) for Longer Wavelength LEDs

    Energy Technology Data Exchange (ETDEWEB)

    Durniak, Mark T. [Rensselaer Polytechnic Inst., Troy, NY (United States). Dept. of Materials Science and Engineering; Chaudhuri, Anabil [Univ. of New Mexico, Albuquerque, NM (United States). Center for High Technology Materials; Smith, Michael L. [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States). Advanced Material Sciences; Allerman, Andrew A. [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States). Advanced Material Sciences; Lee, S. C. [Univ. of New Mexico, Albuquerque, NM (United States). Center for High Technology Materials; Brueck, S. R. J. [Univ. of New Mexico, Albuquerque, NM (United States). Center for High Technology Materials; Wetzel, Christian [Rensselaer Polytechnic Inst., Troy, NY (United States). Dept. of Physics, Applied Physics, and Astronomy and Dept. of Materials Science and Engineering

    2016-03-01

    GaInN/GaN heterostructures of cubic phase have the potential to overcome the limitations of wurtzite structures commonly used for light emitting and laser diodes. Wurtzite GaInN suffers from large internal polarization fields, which force design compromises ( 0001 ) towards ultra-narrow quantum wells and reduce recombination volume and efficiency. Cubic GaInN microstripes grown at Rensselaer Polytechnic Institute by metal organic vapor phase epitaxy on micropatterned Si , with {111} v-grooves oriented along Si ( 001 ) , offer a system free of internal polarization fields, wider quantum wells, and smaller <00$\\bar1$> bandgap energy. We prepared 6 and 9 nm Ga x In 1-x N/GaN single quantum well structures with peak wavelength ranges from 520 to 570 nm with photons predominately polarized perpendicular to the grooves. We estimate a cubic InN composition range of 0 < x < 0.5 and an upper limit of the internal quantum efficiency of 50%. Stripe geometry and polarization may be suitable for mode confinement and reduced threshold stimulated emission.

  5. Improving LED CCT uniformity using micropatterned films optimized by combining ray tracing and FDTD methods.

    Science.gov (United States)

    Ding, Xinrui; Li, Jiasheng; Chen, Qiu; Tang, Yong; Li, Zongtao; Yu, Binhai

    2015-02-09

    Although the light-emitting diode (LED) has revolutionized lighting, the non-uniformity of its correlated color temperature (CCT) still remains a major concern. In this context, to improve the light distribution performance of remote phosphor LED lamps, we employ a micropatterned array (MPA) optical film fabricated using a low-cost molding process. The parameters of the MPA, including different installation configurations, positioning, and diameters, are optimized by combining the finite-difference time-domain and ray-tracing methods. Results show that the sample with the upward-facing convex-cone MPA film that has a diameter of half of that of the remote phosphor glass, and is tightly affixed to the inward surface of the remote phosphor glass renders a superior light distribution performance. When compared with the case in which no MPA film is used, the deviation of the CCT distribution decreases from 1033 K to 223 K, and the corresponding output power of the sample is an acceptable level of 85.6%. We perform experiments to verify our simulation results, and the two sets of results exhibit a close agreement. We believe that our approach can be used to optimize MPA films for various lighting applications.

  6. Effect of Flow on Cultured Cell at Micro-Pattern of Ridge Lines

    Directory of Open Access Journals (Sweden)

    Haruka Hino

    2017-10-01

    Full Text Available A flow channel with a micro-pattern of ridge lines of a scaffold has been designed to study quantitatively the effect of flow on an oriented cell in vitro. The lines of parallel micro ridges (0.001 mm height, 0.003 mm width, and 0.003 mm interval are made by the lithography technique on the lower surface of the channel as the scaffold to make orientation of each cell. Variation is made about the angle between the longitudinal direction of the ridge line and the direction of the flow: zero, 0.79 and 1.6 rad. The suspension of C2C12 (mouse myoblast cell line was injected to the channel, and incubated for two hours on the micro ridges before the flow test for four hours. The flow rate of 3/hour is controlled by a syringe pump to make variation of the wall shear stress of < 3 Pa. The action of each cell adhered on the micro pattern was analyzed at the time lapse images. The experimental results show that both the migration and the deformation of each myoblast along the micro ridge are restricted by the wall shear stress higher than 3 Pa.

  7. Microfluidic impact printer with interchangeable cartridges for versatile non-contact multiplexed micropatterning.

    Science.gov (United States)

    Ding, Yuzhe; Huang, Eric; Lam, Kit S; Pan, Tingrui

    2013-05-21

    Biopatterning has been increasingly used for well-defined cellular microenvironment, patterned surface topology, and guided biological cues; however, it meets challenges on biocompatibility, thermal and chemical sensitivity, as well as limited availability of reagents. In this paper, we aim at combining the desired features from non-contact inkjet printing and dot-matrix impact printing to establish a versatile multiplexed micropatterning platform, referred to as Microfluidic Impact Printer (MI-Printer), for emerging biomedical applications. Using this platform, we can achieve the distinct features of no cross-contamination, sub-microliter ink loading with a minimal dead volume, high-throughput printing, biocompatible non-contact processing, sequential patterning with self-alignment, wide adaptability for complex media (e.g., cell suspension or colloidal solutions), interchangeable/disposable cartridge design, and simple assembly and configuration, all highly desirable towards laboratory-based research and development. Specifically, the printing resolution of the MI-printer platform has been experimentally characterized and theoretically analysed. Optimal printing resolution of 80 μm has been repeatedly obtained. Furthermore, two useful functions of the MI-printer, multiplexed printing and combinatorial printing, have been experimentally demonstrated with less than 10 μm misalignment. Moreover, molecular and biological patterning, utilizing the multiplexed and combinatorial printing, has been implemented to illustrate the utility of this versatile printing technique for emerging biomedical applications.

  8. Tests of Micro-Pattern Gaseous Detectors for active target time projection chambers in nuclear physics

    Energy Technology Data Exchange (ETDEWEB)

    Pancin, J., E-mail: pancin@ganil.fr [GANIL, CEA/DSM-CNRS/IN2P3, Bvd H. Becquerel, Caen (France); Damoy, S.; Perez Loureiro, D. [GANIL, CEA/DSM-CNRS/IN2P3, Bvd H. Becquerel, Caen (France); Chambert, V.; Dorangeville, F. [IPNO, CNRS/IN2P3, Orsay (France); Druillole, F. [CEA, DSM/Irfu/SEDI, Gif-Sur-Yvette (France); Grinyer, G.F. [GANIL, CEA/DSM-CNRS/IN2P3, Bvd H. Becquerel, Caen (France); Lermitage, A.; Maroni, A.; Noël, G. [IPNO, CNRS/IN2P3, Orsay (France); Porte, C.; Roger, T. [GANIL, CEA/DSM-CNRS/IN2P3, Bvd H. Becquerel, Caen (France); Rosier, P. [IPNO, CNRS/IN2P3, Orsay (France); Suen, L. [GANIL, CEA/DSM-CNRS/IN2P3, Bvd H. Becquerel, Caen (France)

    2014-01-21

    Active target detection systems, where the gas used as the detection medium is also a target for nuclear reactions, have been used for a wide variety of nuclear physics applications since the eighties. Improvements in Micro-Pattern Gaseous Detectors (MPGDs) and in micro-electronics achieved in the last decade permit the development of a new generation of active targets with higher granularity pad planes that allow spatial and time information to be determined with unprecedented accuracy. A novel active target and time projection chamber (ACTAR TPC), that will be used to study reactions and decays of exotic nuclei at facilities such as SPIRAL2, is presently under development and will be based on MPGD technology. Several MPGDs (Micromegas and Thick GEM) coupled to a 2×2 mm{sup 2} pixelated pad plane have been tested and their performances have been determined with different gases over a wide range of pressures. Of particular interest for nuclear physics experiments are the angular and energy resolutions. The angular resolution has been determined to be better than 1° FWHM for short traces of about 4 cm in length and the energy resolution deduced from the particle range was found to be better than 5% for 5.5 MeV α particles. These performances have been compared to Geant4 simulations. These experimental results validate the use of these detectors for several applications in nuclear physics.

  9. High rate behavior and discharge limits in micro-pattern detectors

    International Nuclear Information System (INIS)

    Bressan, A.; Hoch, M.; Pagano, P.; Ropelewski, L.; Sauli, F.; Biagi, S.; Buzulutskov, A.; Gruwe, M.; De Lentdecker, G.; Moermann, D.; Sharma, A.

    1999-01-01

    We present and discuss a set of systematic measurements, carried out with gaseous proportional micro-pattern detectors, in order to assess their maximum gain when irradiated with high-rate soft X-rays and heavily ionizing alpha particles. The inventory of detectors tested includes: micro-strips, micromegas, micro-dot, gas electron multiplier, CAT (compteur a trous), trench (or groove), micro-CAT (or WELL) detectors, as well as systems with two elements of gaseous amplification in cascade. We confirm the general trend of all single-stage detectors to follow Raether's criterion, i.e. a spontaneous transition from avalanche to streamer, followed by a discharge, when the avalanche size reaches a value of a few 10 7 ; a noticeable exception is the micro-dot counter holding more than 10 8 . In multiple structures, where the gain is shared between two devices in cascade, the maximum overall gain under irradiation is increased by at least one order of magnitude; we speculate this to be a consequence of a voltage dependence of Raether's limit, larger for low operating potentials. Our conclusion is that only multiple devices can guarantee a sufficient margin of reliability for operation in harsh LHC running conditions

  10. Tests of Micro-Pattern Gaseous Detectors for active target time projection chambers in nuclear physics

    International Nuclear Information System (INIS)

    Pancin, J.; Damoy, S.; Perez Loureiro, D.; Chambert, V.; Dorangeville, F.; Druillole, F.; Grinyer, G.F.; Lermitage, A.; Maroni, A.; Noël, G.; Porte, C.; Roger, T.; Rosier, P.; Suen, L.

    2014-01-01

    Active target detection systems, where the gas used as the detection medium is also a target for nuclear reactions, have been used for a wide variety of nuclear physics applications since the eighties. Improvements in Micro-Pattern Gaseous Detectors (MPGDs) and in micro-electronics achieved in the last decade permit the development of a new generation of active targets with higher granularity pad planes that allow spatial and time information to be determined with unprecedented accuracy. A novel active target and time projection chamber (ACTAR TPC), that will be used to study reactions and decays of exotic nuclei at facilities such as SPIRAL2, is presently under development and will be based on MPGD technology. Several MPGDs (Micromegas and Thick GEM) coupled to a 2×2 mm 2 pixelated pad plane have been tested and their performances have been determined with different gases over a wide range of pressures. Of particular interest for nuclear physics experiments are the angular and energy resolutions. The angular resolution has been determined to be better than 1° FWHM for short traces of about 4 cm in length and the energy resolution deduced from the particle range was found to be better than 5% for 5.5 MeV α particles. These performances have been compared to Geant4 simulations. These experimental results validate the use of these detectors for several applications in nuclear physics

  11. Cell adhesion and spreading on polymer surfaces micropatterned by ion beams

    International Nuclear Information System (INIS)

    Satriano, C.; Carnazza, S.; Licciardello, A.; Guglielmino, S.; Marletta, G.

    2003-01-01

    The cell adhesion and spreading behavior on surfaces of poly(ethyleneterephtalate) and poly(hydroxymethylsiloxane) micropatterned by focused 15 keV Ga + beams has been studied. It has been found that while no modification in the cell adhesion process could be observed for unirradiated and irradiated areas on the patterned surfaces, in the case of polyhydroxymethylsiloxane the cell adhesion process is basically confined within the irradiated areas and a clear dependence of the cell ordering on the lateral size of the irradiated areas is observed. The results are discussed in terms of the specific spatially resolved chemical modification induced by Ga + irradiation onto the two different polymers. Thus, the irradiation-induced modification of composition, functional groups concentration, surface free energy, and nanoscale morphology have been studied by means of x-ray photoelectron spectroscopy, time-of-flight secondary ion mass spectrometry, contact angle, and atomic force microscopy. The cell adhesion and spreading behavior was found to nicely correlate with the increase of the acid-base component γ AB of the surface free energy and more particularly with the dramatic increase of the Lewis basic electron-donor term

  12. Oral fibroblasts produce more HGF and KGF than skin fibroblasts in response to co-culture with keratinocytes

    DEFF Research Database (Denmark)

    Grøn, Birgitte; Stoltze, Kaj; Andersson, Anders

    2002-01-01

    The production of hepatocyte growth factor (HGF) and keratinocyte growth factor (KGF) in subepithelial fibroblasts from buccal mucosa, periodontal ligament, and skin was determined after co-culture with keratinocytes. The purpose was to detect differences between the fibroblast subpopulations...... days by ELISA. When cultured on polystyrene, the constitutive level of KGF and HGF in periodontal fibroblasts was higher than the level in buccal and skin fibroblasts. In the presence of keratinocytes, all three types of fibroblasts in general increased their HGF and KGF production 2-3 times. When...... cells were maintained in collagen, the level of HGF and KGF was decreased mainly in skin cultures. However, in oral fibroblasts, induction after stimulation was at a similar level in collagen compared to on polystyrene. Skin fibroblasts maintained in collagen produced almost no HGF whether...

  13. Interaction of cruciferin-based nanoparticles with Caco-2 cells and Caco-2/HT29-MTX co-cultures.

    Science.gov (United States)

    Akbari, Ali; Lavasanifar, Afsaneh; Wu, Jianping

    2017-12-01

    The objective of this work was to assess the potential of Cruciferin/Calcium (Cru/Ca) and Cruciferin/Chitosan (Cru/Cs) nanoparticles for oral drug delivery. For this purpose, Cru/Ca and Cru/Cs nanoparticles were developed through cold gelation of Cruciferin, a major canola protein, and in interaction with calcium and chitosan, respectively. The extent and rate of particle uptake in Caco-2 cells and Caco-2/HT29 co-culture was then evaluated by fluorescence spectroscopy as well as flow cytometry. Through pre-incubation of Caco-2 cell monolayer with specific endocytosis inhibitors, the mechanism of cell uptake was investigated. Our results showed that the uptake of negatively-charged Cru/Ca particles to be ∼3 times higher than positively-charged Cru/Cs ones by Caco-2 cells. Presence of mucus secreted by HT29 cells in their co-culture with Caco-2 had negligible influence on the uptake and transport of both particles. In contrast to Cru/Ca particles which were dissociated in the simulated gastrointestinal conditions, digestion of Cru/Cs particles resulted in 6- and 2-fold increase in the cellular uptake and transport of encapsulated coumarin in the latter particles, respectively. While the presence of mucus in Caco-2/HT29 co-culture caused 40-50% decrease of cellular uptake and transport for coumarin encapsulated in digested Cru/Cs particles, it had no significant effect on the cell uptake and transport of coumarin associated with Cru/Ca particles after digestion. Energy-dependent mechanisms were the dominant mechanism for uptake of both undigested and digested particles. Therefore, in Caco-2/HT29 co-culture which closely simulated intestinal epithelial cells, undigested Cru/Ca and Cru/Cs particles had the ability to penetrate mucus layers, while digested Cru/Cs particles showed mucoadhesive property, and digested Cru/Ca particles were dissociated. Our results points to a potential for cruciferin based nanoparticles for oral drug delivery. The long-term objective of

  14. A multifunctional 3D co-culture system for studies of mammary tissue morphogenesis and stem cell biology.

    Directory of Open Access Journals (Sweden)

    Jonathan J Campbell

    Full Text Available Studies on the stem cell niche and the efficacy of cancer therapeutics require complex multicellular structures and interactions between different cell types and extracellular matrix (ECM in three dimensional (3D space. We have engineered a 3D in vitro model of mammary gland that encompasses a defined, porous collagen/hyaluronic acid (HA scaffold forming a physiologically relevant foundation for epithelial and adipocyte co-culture. Polarized ductal and acinar structures form within this scaffold recapitulating normal tissue morphology in the absence of reconstituted basement membrane (rBM hydrogel. Furthermore, organoid developmental outcome can be controlled by the ratio of collagen to HA, with a higher HA concentration favouring acinar morphological development. Importantly, this culture system recapitulates the stem cell niche as primary mammary stem cells form complex organoids, emphasising the utility of this approach for developmental and tumorigenic studies using genetically altered animals or human biopsy material, and for screening cancer therapeutics for personalised medicine.

  15. Computed tomographic findings of hepatocellular carcinoma

    International Nuclear Information System (INIS)

    Jo, In Su; Jong, Woo Yung; Lee, Jong Yul; Choi, Han Yong; Kim, Bong Ki

    1987-01-01

    With Development of Computed Tomography, detection of the Hepatocellular Carcinoma are easily performed and frequently used in the world. During 15 months, from December 1985 to February 1987, 59 patients with hepatocellular carcinoma were evaluated with computed tomography in department of radiology at Wallace Memorial Baptist Hospital. The results were as follow: 1. The most prevalent age group was 5th to 7th decades, male to female ratio was 4.9:1. 2. Classification with incidence of computed tomographic appearance of the hepatocellular carcinoma were solitary type 28 cases (48%), multinodular type 24 cases (40%), and diffuse type 7 cases (12%), Association with liver cirrhosis was noted in 22 cases (38%). 3. Inhomogenous internal consistency of hepatocellular carcinoma due to central necrosis were 35 cases (60%). Portal vein invasion by hepatocellular carcinoma was noted in 15 cases (25%), and particularly most common in diffuse type 4 cases (55%). 4. On precontrast scan, all hepatocellular carcinoma were seen as area of low density except for 3 cases(0.5%) of near isodensity which turned out to be remarkable low density on postcontrast scan. 5. In solitary type, posterior segment of right lobe was most common site of involvement 12 cases (43%). In diffuse type, bilobar involvement was most common, 6 cases (85%)

  16. Percutaneous cryoablation for hepatocellular carcinoma

    Directory of Open Access Journals (Sweden)

    Kyoung Doo Song

    2016-12-01

    Full Text Available Local ablation therapy is considered as a conventional treatment option for patients with early stage hepatocellular carcinoma (HCC. Although radiofrequency (RF ablation is widely used for HCC, the use of cryoablation has been increasing as newer and safer cryoablation systems have developed. The thermodynamic mechanism of freezing and thawing used in cryoablation is the Joule-Thomson effect. Cryoablation destroys tissue via direct tissue destruction and vascular-related injury. A few recent comparative studies have shown that percutaneous cryoablation for HCCs is comparable to percutaneous RF ablation in terms of long term therapeutic outcomes and complications. Cryoablation has several advantages over RF ablation such as well visualization of iceball, no causation of severe pain, and lack of severe damage to great vessels and gallbladder. It is important to know the advantages and disadvantages of cryoablation compared with RF ablation for improvement of therapeutic efficacy and safety.

  17. Medical treatment of hepatocellular carcinoma.

    Science.gov (United States)

    Granito, Alessandro; Bolondi, Luigi

    2009-12-16

    Hepatocellular carcinoma (HCC) is the fifth most common neoplasm and the third leading cause of cancer-related deaths worldwide. Cirrhosis, most often due to viral hepatitis, is the predominant risk factors for HCC and geographical differences in both risk factors and incidence are largely due to epidemiological variations in hepatitis B and C infection. Hepatic function is a relevant parameter in selecting therapy in HCC. The current clinical classification of HCC split patients into 5 stages, with a specific treatment schedule for any stage. As patients with early stages can receive curative treatments, such as surgical resection, liver transplantation or local ablation, surveillance program in high-risk populations has become mandatory. Sorafenib, a multikinase inhibitor, has recently shown survival benefits in patients at advanced stage of disease. Hopefully, new molecular targeted therapies and their combination with sorafenib or interventional and surgical procedures, should expand the therapeutic armamentarium against HCC.

  18. [Radiofrequency ablation of hepatocellular carcinoma].

    Science.gov (United States)

    Widmann, Gerlig; Schullian, Peter; Bale, Reto

    2013-03-01

    Percutaneous radiofrequency ablation (RFA) is well established in the treatment of hepatocellular carcinoma (HCC). Due to its curative potential, it is the method of choice for non resectable BCLC (Barcelona Liver Clinic) 0 and A. RFA challenges surgical resection for small HCC and is the method of choice in bridging for transplantation and recurrence after resection or transplantation. The technical feasibility of RFA depends on the size and location of the HCC and the availability of ablation techniques (one needle techniques, multi-needle techniques). More recently, stereotactic multi-needle techniques with 3D trajectory planning and guided needle placement substantially improve the spectrum of treatable lesions including large volume tumors. Treatment success depends on the realization of ablations with large intentional margins of tumor free tissue (A0 ablation in analogy to R0 resection), which has to be documented by fusion of post- with pre-ablation images, and confirmed during follow-up imaging.

  19. Gene mutations in hepatocellular adenomas

    DEFF Research Database (Denmark)

    Raft, Marie B; Jørgensen, Ernö N; Vainer, Ben

    2015-01-01

    is associated with bi-allelic mutations in the TCF1 gene and morphologically has marked steatosis. β-catenin activating HCA has increased activity of the Wnt/β-catenin pathway and is associated with possible malignant transformation. Inflammatory HCA is characterized by an oncogene-induced inflammation due...... to alterations in the Janus kinase/signal transducer and activator of transcription (JAK/STAT) pathway. In the diagnostic setting, sub classification of HCA is based primarily on immunohistochemical analyzes, and has had an increasing impact on choice of treatment and individual prognostic assessment....... This review offers an overview of the reported gene mutations associated with hepatocellular adenomas together with a discussion of the diagnostic and prognostic value....

  20. Bi-directional exchange of membrane components occurs during co-culture of mesenchymal stem cells and nucleus pulposus cells.

    Science.gov (United States)

    Strassburg, Sandra; Hodson, Nigel W; Hill, Patrick I; Richardson, Stephen M; Hoyland, Judith A

    2012-01-01

    Mesenchymal stem cell (MSC)-based therapies have been proposed as novel treatments for intervertebral disc (IVD) degeneration. We have previously demonstrated that when MSCs are co-cultured with nucleus pulposus (NP) cells with direct cell-cell contact, they differentiate along the NP lineage and simultaneously stimulate the degenerate NP cell population to regain a normal (non-degenerate) phenotype, an effect which requires cell-cell communication. However, the mechanisms by which NP cells and MSCs interact in this system are currently unclear. Thus, in this study we investigated a range of potential mechanisms for exchange of cellular components or information that may direct these changes, including cell fusion, gap-junctional communication and exchange of membrane components by direct transfer or via microvesicle formation. Flow cytometry of fluorescently labeled MSCs and NP cells revealed evidence of some cell fusion and formation of gapjunctions, although at the three timepoints studied these phenomena were detectable only in a small proportion of cells. While these mechanisms may play a role in cell-cell communication, the data suggests they are not the predominant mechanism of interaction. However, flow cytometry of fluorescently dual-labeled cells showed that extensive bi-directional transfer of membrane components is operational during direct co-culture of MSCs and NP cells. Furthermore, there was also evidence for secretion and internalization of membrane-bound microvesicles by both cell types. Thus, this study highlights bi-directional intercellular transfer of membrane components as a possible mechanism of cellular communication between MSC and NP cells.

  1. Engineered monoculture and co-culture of methylotrophic yeast for de novo production of monacolin J and lovastatin from methanol.

    Science.gov (United States)

    Liu, Yiqi; Tu, Xiaohu; Xu, Qin; Bai, Chenxiao; Kong, Chuixing; Liu, Qi; Yu, Jiahui; Peng, Qiangqiang; Zhou, Xiangshan; Zhang, Yuanxing; Cai, Menghao

    2018-01-01

    As a promising one-carbon renewable substrate for industrial biotechnology, methanol has attracted much attention. However, engineering of microorganisms for industrial production of pharmaceuticals using a methanol substrate is still in infancy. In this study, the methylotrophic yeast Pichia pastoris was used to produce anti-hypercholesterolemia pharmaceuticals, lovastatin and its precursor monacolin J, from methanol. The biosynthetic pathways for monacolin J and lovastatin were first assembled and optimized in single strains using single copies of the relevant biosynthetic genes, and yields of 60.0mg/L monacolin J and 14.4mg/L lovastatin were obtained using methanol following pH controlled monoculture. To overcome limitations imposed by accumulation of intermediates and metabolic stress in monoculture, approaches using pathway splitting and co-culture were developed. Two pathway splitting strategies for monacolin J, and four for lovastatin were tested at different metabolic nodes. Biosynthesis of monacolin J and lovastatin was improved by 55% and 71%, respectively, when the upstream and downstream modules were separately accommodated in two different fluorescent strains, split at the metabolic node of dihydromonacolin L. However, pathway distribution at monacolin J blocked lovastatin biosynthesis in all designs, mainly due to its limited ability of crossing cellular membranes. Bioreactor fermentations were tested for the optimal co-culture strategies, and yields of 593.9mg/L monacolin J and 250.8mg/L lovastatin were achieved. This study provides an alternative method for production of monacolin J and lovastatin and reveals the potential of a methylotrophic yeast to produce complicated pharmaceuticals from methanol. Copyright © 2017 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.

  2. Neuronal differentiation of hair-follicle-bulge-derived stem cells co-cultured with mouse cochlear modiolus explants.

    Directory of Open Access Journals (Sweden)

    Timo Schomann

    Full Text Available Stem-cell-based repair of auditory neurons may represent an attractive therapeutic option to restore sensorineural hearing loss. Hair-follicle-bulge-derived stem cells (HFBSCs are promising candidates for this type of therapy, because they (1 have migratory properties, enabling migration after transplantation, (2 can differentiate into sensory neurons and glial cells, and (3 can easily be harvested in relatively high numbers. However, HFBSCs have never been used for this purpose. We hypothesized that HFBSCs can be used for cell-based repair of the auditory nerve and we have examined their migration and incorporation into cochlear modiolus explants and their subsequent differentiation. Modiolus explants obtained from adult wild-type mice were cultured in the presence of EF1α-copGFP-transduced HFBSCs, constitutively expressing copepod green fluorescent protein (copGFP. Also, modiolus explants without hair cells were co-cultured with DCX-copGFP-transduced HFBSCs, which demonstrate copGFP upon doublecortin expression during neuronal differentiation. Velocity of HFBSC migration towards modiolus explants was calculated, and after two weeks, co-cultures were fixed and processed for immunohistochemical staining. EF1α-copGFP HFBSC migration velocity was fast: 80.5 ± 6.1 μm/h. After arrival in the explant, the cells formed a fascicular pattern and changed their phenotype into an ATOH1-positive neuronal cell type. DCX-copGFP HFBSCs became green-fluorescent after integration into the explants, confirming neuronal differentiation of the cells. These results show that HFBSC-derived neuronal progenitors are migratory and can integrate into cochlear modiolus explants, while adapting their phenotype depending on this micro-environment. Thus, HFBSCs show potential to be employed in cell-based therapies for auditory nerve repair.

  3. In vitro formation of the Merkel cell-neurite complex in embryonic mouse whiskers using organotypic co-cultures.

    Science.gov (United States)

    Ishida, Kentaro; Saito, Tetsuichiro; Mitsui, Toshiyuki

    2018-06-01

    A Merkel cell-neurite complex is a touch receptor composed of specialized epithelial cells named Merkel cells and peripheral sensory nerves in the skin. Merkel cells are found in touch-sensitive skin components including whisker follicles. The nerve fibers that innervate Merkel cells of a whisker follicle extend from the maxillary branch of the trigeminal ganglion. Whiskers as a sensory organ attribute to the complicated architecture of the Merkel cell-neurite complex, and therefore it is intriguing how the structure is formed. However, observing the dynamic process of the formation of a Merkel cell-neurite complex in whiskers during embryonic development is still difficult. In this study, we tried to develop an organotypic co-culture method of a whisker pad and a trigeminal ganglion explant to form the Merkel cell-neurite complex in vitro. We initially developed two distinct culture methods of a single whisker row and a trigeminal ganglion explant, and then combined them. By dissecting and cultivating a single row from a whisker pad, the morphogenesis of whisker follicles could be observed under a microscope. After the co-cultivation of the whisker row with a trigeminal ganglion explant, a Merkel cell-neurite complex composed of Merkel cells, which were positive for both cytokeratin 8 and SOX2, Neurofilament-H-positive trigeminal nerve fibers and Schwann cells expressing Nestin, SOX2 and SOX10 was observed via immunohistochemical analyses. These results suggest that the process for the formation of a Merkel cell-neurite complex can be observed under a microscope using our organotypic co-culture method. © 2018 Japanese Society of Developmental Biologists.

  4. Angiographic findings of hepatocellular carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Han, Man Chung; Cho, Byung Jae; Huh, Seung Jae; Bae, Sang Hoon; Kim, Ung Jin; Kim, Chung Yong; Kim, Noe Kyeong [Yonsei University College of Medicine, Seoul (Korea, Republic of)

    1985-12-15

    From March 1977 to July 1979, 69 cases of angiograms of hepatocellular carcinoma were observed in Seoul National University Hospital. The findings of selective celiac and/or hepatic arteriography in total 69 cases of confirmed hepatocellular carcinoma, with clinical and laboratory findings, were analyzed. The summarized results are as follows; 1. Among 69 cases od hepatoma, 62 were male and 7 were female with sex ratio of 8.9 : 1. Peak incidence is 5th to 7th decades (72.5%). Epigastric pain, indigestion, and palpable mass in right upper quadrant were common symptoms and sign. Laboratory findings showed elevated serum alkaline phosphatase more than 5 Bodansky unit in 75.4%. Alpha-feto protein was positive in 65.2% of all the patients. 2 All 69 cases were classified into 31 cases of massive type, 22 cases of diffuse type, and 16 cases of nodular type, in accordance with angiographic gross anatomy. The frequency of angiographic findings were hypervascularities and tumor vessels (100%), tumor stainings (98.5%), arteriovenous shunt (71.0%), displacement of intrahepatic arteries (66.7%), vascular lakes and channel (59.4%). Encasement of hepatic artery and portal vein regurgitation was respectively 4 cases. Tumor mass in portal vein were 6 cases and tumor mass in hepatic vein was 1 case. 3. Intraarterial infusion of 5-FU was performed in 15 hepatoma patients, and the results were that angiographic improvement was demonstrated in 3 cases, no improvement in 8 cases, and incomplete infusion in 4 cases. 4. The selective celiac and/or hepatic angiograms are excellent diagnostic tools as well as therapeutic management for intraarterial infusion of anticancerous drugs.

  5. Angiographic findings of hepatocellular carcinoma

    International Nuclear Information System (INIS)

    Han, Man Chung; Cho, Byung Jae; Huh, Seung Jae; Bae, Sang Hoon; Kim, Ung Jin; Kim, Chung Yong; Kim, Noe Kyeong

    1985-01-01

    From March 1977 to July 1979, 69 cases of angiograms of hepatocellular carcinoma were observed in Seoul National University Hospital. The findings of selective celiac and/or hepatic arteriography in total 69 cases of confirmed hepatocellular carcinoma, with clinical and laboratory findings, were analyzed. The summarized results are as follows; 1. Among 69 cases od hepatoma, 62 were male and 7 were female with sex ratio of 8.9 : 1. Peak incidence is 5th to 7th decades (72.5%). Epigastric pain, indigestion, and palpable mass in right upper quadrant were common symptoms and sign. Laboratory findings showed elevated serum alkaline phosphatase more than 5 Bodansky unit in 75.4%. Alpha-feto protein was positive in 65.2% of all the patients. 2 All 69 cases were classified into 31 cases of massive type, 22 cases of diffuse type, and 16 cases of nodular type, in accordance with angiographic gross anatomy. The frequency of angiographic findings were hypervascularities and tumor vessels (100%), tumor stainings (98.5%), arteriovenous shunt (71.0%), displacement of intrahepatic arteries (66.7%), vascular lakes and channel (59.4%). Encasement of hepatic artery and portal vein regurgitation was respectively 4 cases. Tumor mass in portal vein were 6 cases and tumor mass in hepatic vein was 1 case. 3. Intraarterial infusion of 5-FU was performed in 15 hepatoma patients, and the results were that angiographic improvement was demonstrated in 3 cases, no improvement in 8 cases, and incomplete infusion in 4 cases. 4. The selective celiac and/or hepatic angiograms are excellent diagnostic tools as well as therapeutic management for intraarterial infusion of anticancerous drugs.

  6. Predictivity of dog co-culture model, primary human hepatocytes and HepG2 cells for the detection of hepatotoxic drugs in humans

    International Nuclear Information System (INIS)

    Atienzar, Franck A.; Novik, Eric I.; Gerets, Helga H.; Parekh, Amit; Delatour, Claude; Cardenas, Alvaro; MacDonald, James; Yarmush, Martin L.; Dhalluin, Stéphane

    2014-01-01

    Drug Induced Liver Injury (DILI) is a major cause of attrition during early and late stage drug development. Consequently, there is a need to develop better in vitro primary hepatocyte models from different species for predicting hepatotoxicity in both animals and humans early in drug development. Dog is often chosen as the non-rodent species for toxicology studies. Unfortunately, dog in vitro models allowing long term cultures are not available. The objective of the present manuscript is to describe the development of a co-culture dog model for predicting hepatotoxic drugs in humans and to compare the predictivity of the canine model along with primary human hepatocytes and HepG2 cells. After rigorous optimization, the dog co-culture model displayed metabolic capacities that were maintained up to 2 weeks which indicates that such model could be also used for long term metabolism studies. Most of the human hepatotoxic drugs were detected with a sensitivity of approximately 80% (n = 40) for the three cellular models. Nevertheless, the specificity was low approximately 40% for the HepG2 cells and hepatocytes compared to 72.7% for the canine model (n = 11). Furthermore, the dog co-culture model showed a higher superiority for the classification of 5 pairs of close structural analogs with different DILI concerns in comparison to both human cellular models. Finally, the reproducibility of the canine system was also satisfactory with a coefficient of correlation of 75.2% (n = 14). Overall, the present manuscript indicates that the dog co-culture model may represent a relevant tool to perform chronic hepatotoxicity and metabolism studies. - Highlights: • Importance of species differences in drug development. • Relevance of dog co-culture model for metabolism and toxicology studies. • Hepatotoxicity: higher predictivity of dog co-culture vs HepG2 and human hepatocytes

  7. Predictivity of dog co-culture model, primary human hepatocytes and HepG2 cells for the detection of hepatotoxic drugs in humans

    Energy Technology Data Exchange (ETDEWEB)

    Atienzar, Franck A., E-mail: franck.atienzar@ucb.com [UCB Pharma SA, Non-Clinical Development, Chemin du Foriest, 1420 Braine-l' Alleud (Belgium); Novik, Eric I. [H mu rel Corporation, 675 U.S. Highway 1, North Brunswick, NJ 08902 (United States); Gerets, Helga H. [UCB Pharma SA, Non-Clinical Development, Chemin du Foriest, 1420 Braine-l' Alleud (Belgium); Parekh, Amit [H mu rel Corporation, 675 U.S. Highway 1, North Brunswick, NJ 08902 (United States); Delatour, Claude; Cardenas, Alvaro [UCB Pharma SA, Non-Clinical Development, Chemin du Foriest, 1420 Braine-l' Alleud (Belgium); MacDonald, James [Chrysalis Pharma Consulting, LLC, 385 Route 24, Suite 1G, Chester, NJ 07930 (United States); Yarmush, Martin L. [Department of Biomedical Engineering, Rutgers University, Piscataway, NJ 08854 (United States); Dhalluin, Stéphane [UCB Pharma SA, Non-Clinical Development, Chemin du Foriest, 1420 Braine-l' Alleud (Belgium)

    2014-02-15

    Drug Induced Liver Injury (DILI) is a major cause of attrition during early and late stage drug development. Consequently, there is a need to develop better in vitro primary hepatocyte models from different species for predicting hepatotoxicity in both animals and humans early in drug development. Dog is often chosen as the non-rodent species for toxicology studies. Unfortunately, dog in vitro models allowing long term cultures are not available. The objective of the present manuscript is to describe the development of a co-culture dog model for predicting hepatotoxic drugs in humans and to compare the predictivity of the canine model along with primary human hepatocytes and HepG2 cells. After rigorous optimization, the dog co-culture model displayed metabolic capacities that were maintained up to 2 weeks which indicates that such model could be also used for long term metabolism studies. Most of the human hepatotoxic drugs were detected with a sensitivity of approximately 80% (n = 40) for the three cellular models. Nevertheless, the specificity was low approximately 40% for the HepG2 cells and hepatocytes compared to 72.7% for the canine model (n = 11). Furthermore, the dog co-culture model showed a higher superiority for the classification of 5 pairs of close structural analogs with different DILI concerns in comparison to both human cellular models. Finally, the reproducibility of the canine system was also satisfactory with a coefficient of correlation of 75.2% (n = 14). Overall, the present manuscript indicates that the dog co-culture model may represent a relevant tool to perform chronic hepatotoxicity and metabolism studies. - Highlights: • Importance of species differences in drug development. • Relevance of dog co-culture model for metabolism and toxicology studies. • Hepatotoxicity: higher predictivity of dog co-culture vs HepG2 and human hepatocytes.

  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. Yeast-yeast interactions revealed by aromatic profile analysis of Sauvignon Blanc wine fermented by single or co-culture of non-Saccharomyces and Saccharomyces yeasts.

    Science.gov (United States)

    Sadoudi, Mohand; Tourdot-Maréchal, Raphaëlle; Rousseaux, Sandrine; Steyer, Damien; Gallardo-Chacón, Joan-Josep; Ballester, Jordi; Vichi, Stefania; Guérin-Schneider, Rémi; Caixach, Josep; Alexandre, Hervé

    2012-12-01

    There has been increasing interest in the use of selected non-Saccharomyces yeasts in co-culture with Saccharomyces cerevisiae. The main reason is that the multistarter fermentation process is thought to simulate indigenous fermentation, thus increasing wine aroma complexity while avoiding the risks linked to natural fermentation. However, multistarter fermentation is characterised by complex and largely unknown interactions between yeasts. Consequently the resulting wine quality is rather unpredictable. In order to better understand the interactions that take place between non-Saccharomyces and Saccharomyces yeasts during alcoholic fermentation, we analysed the volatile profiles of several mono-culture and co-cultures. Candida zemplinina, Torulaspora delbrueckii and Metschnikowia pulcherrima were used to conduct fermentations either in mono-culture or in co-culture with S. cerevisiae. Up to 48 volatile compounds belonging to different chemical families were quantified. For the first time, we show that C. zemplinina is a strong producer of terpenes and lactones. We demonstrate by means of multivariate analysis that different interactions exist between the co-cultures studied. We observed a synergistic effect on aromatic compound production when M. pulcherrima was in co-culture with S. cerevisiae. However a negative interaction was observed between C. zemplinina and S. cerevisiae, which resulted in a decrease in terpene and lactone content. These interactions are independent of biomass production. The aromatic profiles of T. delbrueckii and S. cerevisiae in mono-culture and in co-culture are very close, and are biomass-dependent, reflecting a neutral interaction. This study reveals that a whole family of compounds could be altered by such interactions. These results suggest that the entire metabolic pathway is affected by these interactions. Copyright © 2012 Elsevier Ltd. All rights reserved.

  10. Co-culture with human synovium-derived mesenchymal stem cells inhibits inflammatory activity and increases cell proliferation of sodium nitroprusside-stimulated chondrocytes

    International Nuclear Information System (INIS)

    Ryu, Jae-Sung; Jung, Yeon-Hwa; Cho, Mi-Young; Yeo, Jee Eun; Choi, Yun-Jin; Kim, Yong Il; Koh, Yong-Gon

    2014-01-01

    Highlights: • Co-culture of hSDMSCs with SNP-stimulated chondrocytes improves anti-inflammation. • Co-culture system produces IGF-1. • Co-culture system suppresses inflammatory genes expression. • Co-culture system improves cell proliferation. • Exogenous IGF-1 inhibits inflammatory activity in SNP-stimulated chondrocytes. - Abstract: Rheumatoid arthritis (RA) and osteoarthritis (OA) are primarily chronic inflammatory diseases. Mesenchymal stem cells (MSCs) have the ability to differentiate into cells of the mesodermal lineage, and to regulate immunomodulatory activity. Specifically, MSCs have been shown to secrete insulin-like growth factor 1 (IGF-1). The purpose of the present study was to examine the inhibitory effects on inflammatory activity from a co-culture of human synovium-derived mesenchymal stem cells (hSDMSCs) and sodium nitroprusside (SNP)-stimulated chondrocytes. First, chondrocytes were treated with SNP to generate an in vitro model of RA or OA. Next, the co-culture of hSDMSCs with SNP-stimulated chondrocytes reduced inflammatory cytokine secretion, inhibited expression of inflammation activity-related genes, generated IGF-1 secretion, and increased the chondrocyte proliferation rate. To evaluate the effect of IGF-1 on inhibition of inflammation, chondrocytes pre-treated with IGF-1 were treated with SNP, and then the production of inflammatory cytokines was analyzed. Treatment with IGF-1 was shown to significantly reduce inflammatory cytokine secretion in SNP-stimulated chondrocytes. Our results suggest that hSDMSCs offer a new strategy to promote cell-based cartilage regeneration in RA or OA

  11. Co-culture with human synovium-derived mesenchymal stem cells inhibits inflammatory activity and increases cell proliferation of sodium nitroprusside-stimulated chondrocytes

    Energy Technology Data Exchange (ETDEWEB)

    Ryu, Jae-Sung; Jung, Yeon-Hwa; Cho, Mi-Young; Yeo, Jee Eun; Choi, Yun-Jin; Kim, Yong Il; Koh, Yong-Gon, E-mail: yonseranglab@daum.net

    2014-05-16

    Highlights: • Co-culture of hSDMSCs with SNP-stimulated chondrocytes improves anti-inflammation. • Co-culture system produces IGF-1. • Co-culture system suppresses inflammatory genes expression. • Co-culture system improves cell proliferation. • Exogenous IGF-1 inhibits inflammatory activity in SNP-stimulated chondrocytes. - Abstract: Rheumatoid arthritis (RA) and osteoarthritis (OA) are primarily chronic inflammatory diseases. Mesenchymal stem cells (MSCs) have the ability to differentiate into cells of the mesodermal lineage, and to regulate immunomodulatory activity. Specifically, MSCs have been shown to secrete insulin-like growth factor 1 (IGF-1). The purpose of the present study was to examine the inhibitory effects on inflammatory activity from a co-culture of human synovium-derived mesenchymal stem cells (hSDMSCs) and sodium nitroprusside (SNP)-stimulated chondrocytes. First, chondrocytes were treated with SNP to generate an in vitro model of RA or OA. Next, the co-culture of hSDMSCs with SNP-stimulated chondrocytes reduced inflammatory cytokine secretion, inhibited expression of inflammation activity-related genes, generated IGF-1 secretion, and increased the chondrocyte proliferation rate. To evaluate the effect of IGF-1 on inhibition of inflammation, chondrocytes pre-treated with IGF-1 were treated with SNP, and then the production of inflammatory cytokines was analyzed. Treatment with IGF-1 was shown to significantly reduce inflammatory cytokine secretion in SNP-stimulated chondrocytes. Our results suggest that hSDMSCs offer a new strategy to promote cell-based cartilage regeneration in RA or OA.

  12. Inhibition of glutamine synthesis induces glutamate dehydrogenase-dependent ammonia fixation into alanine in co-cultures of astrocytes and neurons

    DEFF Research Database (Denmark)

    Dadsetan, Sherry; Bak, Lasse Kristoffer; Sørensen, Michael

    2011-01-01

    study it was investigated if the glutamine synthetase (GS) inhibitor methionine sulfoximine (MSO) would enhance alanine synthesis by blocking the GS-dependent ammonia scavenging process. Hence, co-cultures of neurons and astrocytes were incubated for 2.5h with [U-(13)C]glucose to monitor de novo......It has been previously demonstrated that ammonia exposure of neurons and astrocytes in co-culture leads to net synthesis not only of glutamine but also of alanine. The latter process involves the concerted action of glutamate dehydrogenase (GDH) and alanine aminotransferase (ALAT). In the present...

  13. An optical fiber-based flexible readout system for micro-pattern gas detectors

    Science.gov (United States)

    Li, C.; Feng, C. Q.; Zhu, D. Y.; Liu, S. B.; An, Q.

    2018-04-01

    This paper presents an optical fiber-based readout system that is intended to provide a general purpose multi-channel readout solution for various Micro-Pattern Gas Detectors (MPGDs). The proposed readout system is composed of several front-end cards (FECs) and a data collection module (DCM). The FEC exploits the capability of an existing 64-channel generic TPC readout ASIC chip, named AGET, to implement 256 channels readout. AGET offers FEC a large flexibility in gain range (4 options from 120 fC to 10 pC), peaking time (16 options from 50 ns to 1 us) and sampling freqency (100 MHz max.). The DCM contains multiple 1 Gbps optical fiber serial link interfaces that allow the system scaling up to 1536 channels with 6 FECs and 1 DCM. Further scaling up is possible through cascading of multiple DCMs, by configuring one DCM as a master while other DCMs in slave mode. This design offers a rapid readout solution for different application senario. Tests indicate that the nonlinearity of each channel is less than 1%, and the equivalent input noise charge is typically around 0.7 fC in RMS (root mean square), with a noise slope of about 0.01 fC/pF. The system level trigger rate limit is about 700 Hz in all channel readout mode. When in hit channel readout mode, supposing that typically 10 percent of channels are fired, trigger rate can go up to about 7 kHz. This system has been tested with Micromegas detector and GEM detector, confirming its capability in MPGD readout. Details of hardware and FPGA firmware design, as well as system performances, are described in the paper.

  14. Excimer laser micropatterning of freestanding thermo-responsive hydrogel layers for cells-on-chip applications

    International Nuclear Information System (INIS)

    Santaniello, Tommaso; Milani, Paolo; Lenardi, Cristina; Martello, Federico; Tocchio, Alessandro; Gassa, Federico; Webb, Patrick

    2012-01-01

    We report a novel reliable and repeatable technologic manufacturing protocol for the realization of micro-patterned freestanding hydrogel layers based on thermo-responsive poly-(N-isopropyl)acrylamide (PNIPAAm), which have potential to be employed as temperature-triggered smart surfaces for cells-on-chip applications. PNIPAAm-based films with controlled mechanical properties and different thicknesses (100–300 µm thickness) were prepared by injection compression moulding at room temperature. A 9 × 9 array of 20 µm diameter through-holes is machined by means of the KrF excimer laser on dry PNIPAAm films which are physically attached to flat polyvinyl chloride (PVC) substrates. Machining parameters, such as fluence and number of shots, are optimized in order to achieve highly resolved features. Micro-structured freestanding films are then easily obtained after hydrogels are detached from PVC by gradually promoting the film swelling in ethanol. In the PNIPAAm water-swollen state, the machined holes’ diameter approaches a slight larger value (30 µm) according to the measured hydrogel swelling ratio. Thermo-responsive behaviour and through-hole tapering characterization are carried out by metrology measurements using an optical inverted and confocal microscope setup, respectively. After the temperature of freestanding films is raised above 32 °C, we observe that the shrinkage of the whole through-hole array occurs, thus reducing the holes’ diameter to less than a half its original size (about 15 µm) as a consequence of the film dehydration. Different holes’ diameters (10 and 30 µm) are also obtained on dry hydrogel employing suitable projection masks, showing similar shrinking behaviour when hydrated and undergone thermo-response tests. Thermo-responsive PNIPAAm-based freestanding layers could then be integrated with other suitable micro-fabricated thermoplastic components in order to preliminary test their feasibility in operating as temperature

  15. Laser printing of nanoparticle toner enables digital control of micropatterned carbon nanotube growth.

    Science.gov (United States)

    Polsen, Erik S; Stevens, Adam G; Hart, A John

    2013-05-01

    Commercialization of materials utilizing patterned carbon nanotube (CNT) forests, such as hierarchical composite structures, dry adhesives, and contact probe arrays, will require catalyst patterning techniques that do not rely on cleanroom photolithography. We demonstrate the large scale patterning of CNT growth catalyst via adaptation of a laser-based electrostatic printing process that uses magnetic ink character recognition (MICR) toner. The MICR toner contains iron oxide nanoparticles that serve as the catalyst for CNT growth, which are printed onto a flexible polymer (polyimide) and then transferred to a rigid substrate (silicon or alumina) under heat and mechanical pressure. Then, the substrate is processed for CNT growth under an atmospheric pressure chemical vapor deposition (CVD) recipe. This process enables digital control of patterned CNT growth via the laser intensity, which controls the CNT density; and via the grayscale level, which controls the pixelation of the image into arrays of micropillars. Moreover, virtually any pattern can be designed using standard software (e.g., MS Word, AutoCAD, etc.) and printed on demand. Using a standard office printer, we realize isolated CNT microstructures as small as 140 μm and isolated catalyst ″pixels″ as small as 70 μm (one grayscale dot) and determine that individual toner microparticles result in features of approximately 5-10 μm . We demonstrate that grayscale CNT patterns can function as dry adhesives and that large-area catalyst patterns can be printed directly onto metal foils or transferred to ceramic plates. Laser printing therefore shows promise to enable high-speed micropatterning of nanoparticle-containing thin films under ambient conditions, possibly for a wide variety of nanostructures by engineering of toners containing nanoparticles of desired composition, size, and shape.

  16. Fabrication of Biomimetic Fog-Collecting Superhydrophilic-Superhydrophobic Surface Micropatterns Using Femtosecond Lasers.

    Science.gov (United States)

    Kostal, Elisabeth; Stroj, Sandra; Kasemann, Stephan; Matylitsky, Victor; Domke, Matthias

    2018-03-06

    The exciting functionalities of natural superhydrophilic and superhydrophobic surfaces served as inspiration for a variety of biomimetic designs. In particular, the combination of both extreme wetting states to micropatterns opens up interesting applications, as the example of the fog-collecting Namib Desert beetle shows. In this paper, the beetle's elytra were mimicked by a novel three-step fabrication method to increase the fog-collection efficiency of glasses. In the first step, a double-hierarchical surface structure was generated on Pyrex wafers using femtosecond laser structuring, which amplified the intrinsic wetting property of the surface and made it superhydrophilic (water contact angle 150°). In the last step, the Teflon-like coating was selectively removed by fs-laser ablation to uncover superhydrophilic spots below the superhydrophobic surface, following the example of the Namib Desert beetle's fog-collecting elytra. To investigate the influence on the fog-collection behavior, (super)hydrophilic, (super)hydrophobic, and low and high contrast wetting patterns were fabricated on glass wafers using selected combinations of these three processing steps and were exposed to fog in an artificial nebulizer setup. This experiment revealed that high-contrast wetting patterns collected the highest amount of fog and enhanced the fog-collection efficiency by nearly 60% compared to pristine Pyrex glass. The comparison of the fog-collection behavior of the six samples showed that the superior fog-collection efficiency of surface patterns with extreme wetting contrast is due to the combination of water attraction and water repellency: the superhydrophilic spots act as drop accumulation areas, whereas the surrounding superhydrophobic areas allow a fast water transportation caused by gravity. The presented method enables a fast and flexible surface functionalization of a broad range of materials including transparent substrates, which offers exciting possibilities for

  17. Surgical management of spontaneous ruptured hepatocellular adenoma

    Directory of Open Access Journals (Sweden)

    Marcelo Augusto Fontenelle Ribeiro Junior

    2009-01-01

    Full Text Available AIMS: Spontaneous ruptured hepatocellular adenoma (SRHA is a rare life-threatening condition that may require surgical treatment to control hemorrhaging and also stabilize the patient. We report a series of emergency surgeries performed at our institution for this condition. METHODS: We reviewed medical records and radiology files of 28 patients (from 1989 to 2006 with a proven diagnosis of hepatocellular adenoma (HA. Three (10.7% of 28 patients had spontaneous ruptured hepatocellular adenoma, two of which were associated with intrahepatic hemorrhage while one had intraperitoneal bleeding. Two patients were female and one was male. Both female patients had a background history of oral contraceptive use. Sudden abdominal pain associated with hemodynamic instability occurred in all patients who suffered from spontaneous ruptured hepatocellular adenoma. The mean age was 41.6 years old. The preoperative assessment included liver function tests, ultrasonography and computed tomography. RESULTS: The surgical approaches were as follows: right hemihepatectomy for controlling intraperitoneal bleeding, and right extended hepatectomy and non-anatomic resection of the liver for intrahepatic hemorrhage. There were no deaths, and the postoperative complications were bile leakage and wound infection (re-operation, as well as intraperitoneal abscess (re-operation and pleural effusion. CONCLUSION: Spontaneous ruptured hepatocellular adenoma may be treated by surgery for controlling hemorrhages and stabilizing the patient, and the decision to operate depends upon both the patient's condition and the expertise of the surgical team.

  18. Production of plant cell wall degrading enzymes by monoculture and co-culture of Aspergillus niger and Aspergillus terreus under SSF of banana peels.

    Science.gov (United States)

    Rehman, Shazia; Aslam, Hina; Ahmad, Aqeel; Khan, Shakeel Ahmed; Sohail, Muhammad

    2014-01-01

    Filamentous fungi are considered to be the most important group of microorganisms for the production of plant cell wall degrading enzymes (CWDE), in solid state fermentations. In this study, two fungal strains Aspergillus niger MS23 and Aspergillus terreus MS105 were screened for plant CWDE such as amylase, pectinase, xylanase and cellulases (β-glucosidase, endoglucanase and filterpaperase) using a novel substrate, Banana Peels (BP) for SSF process. This is the first study, to the best of our knowledge, to use BP as SSF substrate for plant CWDE production by co-culture of fungal strains. The titers of pectinase were significantly improved in co-culture compared to mono-culture. Furthermore, the enzyme preparations obtained from monoculture and co-culture were used to study the hydrolysis of BP along with some crude and purified substrates. It was observed that the enzymatic hydrolysis of different crude and purified substrates accomplished after 26 h of incubation, where pectin was maximally hydrolyzed by the enzyme preparations of mono and co-culture. Along with purified substrates, crude materials were also proved to be efficiently degraded by the cocktail of the CWDE. These results demonstrated that banana peels may be a potential substrate in solid-state fermentation for the production of plant cell wall degrading enzymes to be used for improving various biotechnological and industrial processes.

  19. Purinergic modulation of adult guinea pig cardiomyocytes in long term cultures and co-cultures with extracardiac or intrinsic cardiac neurones.

    Science.gov (United States)

    Horackova, M; Huang, M H; Armour, J A

    1994-05-01

    To determine the capacity of ATP to modify cardiomyocytes directly or indirectly via peripheral autonomic neurones, the effects of various purinergic agents were studied on long term cultures of adult guinea pig ventricular myocytes and their co-cultures with extracardiac (stellate ganglion) or intrinsic cardiac neurones. Ventricular myocytes and cardiac neurones were enzymatically dissociated and plated together or alone (myocytes only). Myocyte cultures were used for experiments after three to six weeks. The electrical and contractile properties of cultured myocytes and myocyte-neuronal networks were investigated. The spontaneous beating frequency of ventricular myocytes co-cultured with stellate ganglion neurones increased by approximately 140% (p under control conditions, but when beta adrenergic receptors of tetrodotoxin sensitive neural responses were blocked, ATP induced greater augmentation (> 100%). In contrast, ATP induced much smaller effects in non-innervated myocyte cultures (approximately 26%, p UTP > MSATP > beta gamma ATP > alpha beta ATP. Adenosine (10(-4) M) attenuated the beating frequency of myocytes in both types of co-culture, while not significantly affecting non-innervated myocyte cultures. The experimental model used in this study showed that extrinsic and intrinsic cardiac neurones which possess P2 receptors can greatly enhance cardiac myocyte contractile rate when activated by ATP. Since adenosine reduced contractile rate in both types of co-cultures while not affecting non-innervated myocytes, it is concluded that some of these neurones possess P1 receptors.

  20. The allogeneic umbilical cord mesenchymal stem cells regulate the function of T helper 17 cells from patients with rheumatoid arthritis in an in vitro co-culture system

    Directory of Open Access Journals (Sweden)

    Wang Qin

    2012-12-01

    Full Text Available Abstract Background Previous in vivo studies have shown that mesenchymal stem cell (MSC transplantation significantly improves the condition of a number of autoimmune diseases including autoimmune cerebrospinal meningitis, multiple sclerosis, glomerulonephritis and systemic lupus erythematosus. Methods To investigate the immunoregulatory effect of stem cell transplantation, human umbilical cord MSCs were co-cultured with peripheral blood mononuclear cells (PBMCs from patients with rheumatoid arthritis (RA. Orphan nuclear receptor gamma (ROR-γ mRNA and protein expression was detected with real-time PCR and Western blotting. Interleukin (IL-17, IL-6 and tumor necrosis factor (TNF-α in the cell culture supernatant were measured using a flow cytometric bead capture method. Results After 72 hours of co-culture, the mRNA and protein expression levels of ROR-γ in co-cultured PBMCs were decreased compared with that in PBMC of RA patients cultured alone (p  Conclusions In vitro co-culture with MSCs down-regulated the inflammatory response of PBMCs from RA patients with severe disease activity, but had no significant effect on PBMCs from healthy controls or patients with mild disease activity, suggesting that the immunoregulatory role of MSCs may associate with the occurrence of inflammatory mediators.

  1. Production of plant cell wall degrading enzymes by monoculture and co-culture of Aspergillus niger and Aspergillus terreus under SSF of banana peels

    Directory of Open Access Journals (Sweden)

    Shazia Rehman

    2014-12-01

    Full Text Available Filamentous fungi are considered to be the most important group of microorganisms for the production of plant cell wall degrading enzymes (CWDE, in solid state fermentations. In this study, two fungal strains Aspergillus niger MS23 and Aspergillus terreus MS105 were screened for plant CWDE such as amylase, pectinase, xylanase and cellulases (β-glucosidase, endoglucanase and filterpaperase using a novel substrate, Banana Peels (BP for SSF process. This is the first study, to the best of our knowledge, to use BP as SSF substrate for plant CWDE production by co-culture of fungal strains. The titers of pectinase were significantly improved in co-culture compared to mono-culture. Furthermore, the enzyme preparations obtained from monoculture and co-culture were used to study the hydrolysis of BP along with some crude and purified substrates. It was observed that the enzymatic hydrolysis of different crude and purified substrates accomplished after 26 h of incubation, where pectin was maximally hydrolyzed by the enzyme preparations of mono and co-culture. Along with purified substrates, crude materials were also proved to be efficiently degraded by the cocktail of the CWDE. These results demonstrated that banana peels may be a potential substrate in solid-state fermentation for the production of plant cell wall degrading enzymes to be used for improving various biotechnological and industrial processes.

  2. 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.

  3. Enhancement of ligninolytic enzyme activities in a Trametes maxima–Paecilomyces carneus co-culture: Key factors revealed after screening using a Plackett–Burman experimental design

    Directory of Open Access Journals (Sweden)

    Wilberth Chan Cupul

    2014-05-01

    Conclusions: Interaction between indigenous fungi: T. maxima–P. carneus improves laccase and MnP activities. The inoculation time of P. carneus on T. maxima plays an important role in the laccase and MnP enhancement. The nutritional requirements for enzyme improvement in a co-culture system are different from those required for a monoculture system.

  4. Human Wharton’s jelly-derived mesenchymal stem cells express oocyte developmental genes during co-culture with placental cells

    Directory of Open Access Journals (Sweden)

    Hamid Reza Asgari

    2015-01-01

    Conclusion: Placental cell supplementsTransforming growth factor (TGF α, β and basic fibroblast growth factor (bFGF in a co-culture model can provide proper environment for induction of HUMSCs into PGCs and expression of oocyte-like markers.

  5. Chondrocytes co-cultured with Stromal Vascular Fraction of adipose tissue present more intense chondrogenic characteristics than with Adipose Stem Cells

    NARCIS (Netherlands)

    Wu, Ling; Prins, H.J.; Leijten, Jeroen Christianus Hermanus; Helder, M.; Evseenko, D.; Moroni, L; van Blitterswijk, Clemens; Lin, Y.; Karperien, Hermanus Bernardus Johannes

    2016-01-01

    Partly replacement of chondrocytes by stem cells has been proposed to improve the performance of autologous chondrocytes implantation (ACI). Our previous studies showed that the increased cartilage production in pellet co-cultures of chondrocytes and mesenchymal stem cells (MSCs) is due to a trophic

  6. One-pot bioethanol production from cellulose by co-culture of Acremonium cellulolyticus and Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Park Enoch Y

    2012-08-01

    Full Text Available Abstract Background While the ethanol production from biomass by consolidated bioprocess (CBP is considered to be the most ideal process, simultaneous saccharification and fermentation (SSF is the most appropriate strategy in practice. In this study, one-pot bioethanol production, including cellulase production, saccharification of cellulose, and ethanol production, was investigated for the conversion of biomass to biofuel by co-culture of two different microorganisms such as a hyper cellulase producer, Acremonium cellulolyticus C-1 and an ethanol producer Saccharomyces cerevisiae. Furthermore, the operational conditions of the one-pot process were evaluated for maximizing ethanol concentration from cellulose in a single reactor. Results Ethanol production from cellulose was carried out in one-pot bioethanol production process. A. cellulolyticus C-1 and S. cerevisiae were co-cultured in a single reactor. Cellulase producing-medium supplemented with 2.5 g/l of yeast extract was used for productions of both cellulase and ethanol. Cellulase production was achieved by A. cellulolyticus C-1 using Solka-Floc (SF as a cellulase-inducing substrate. Subsequently, ethanol was produced with addition of both 10%(v/v of S. cerevisiae inoculum and SF at the culture time of 60 h. Dissolved oxygen levels were adjusted at higher than 20% during cellulase producing phase and at lower than 10% during ethanol producing phase. Cellulase activity remained 8–12 FPU/ml throughout the one-pot process. When 50–300 g SF/l was used in 500 ml Erlenmeyer flask scale, the ethanol concentration and yield based on initial SF were as 8.7–46.3 g/l and 0.15–0.18 (g ethanol/g SF, respectively. In 3-l fermentor with 50–300 g SF/l, the ethanol concentration and yield were 9.5–35.1 g/l with their yields of 0.12–0.19 (g/g respectively, demonstrating that the one-pot bioethanol production is a reproducible process in a scale-up bioconversion of cellulose to ethanol

  7. Pseudomonads Isolated from Pristine Background Groundwater Proliferate More Effectively in Co-culture than in Monoculture Under Denitrifying Conditions

    Science.gov (United States)

    Aaring, A. B.; Lancaster, A.; Novichkov, P.; Adams, M. W. W.; Deutschbauer, A. M.; Chakraborty, R.

    2016-12-01

    As part of the Ecosystems and Networks Integrated with Genes and Molecular Assemblies (ENIGMA) consortium, we study the microbial community at the U.S. Department of Energy's Field Research Center (FRC) in Oak Ridge. The groundwater at this site contains plumes of nitrate with concentrations up to 14,000mg/L among other contaminants, though molybdenum concentrations are low. Because molybdenum is essential to nitrate reduction, this can be inhibitory to growth. Several strains of Pseudomonas were isolated from the same background groundwater sample. These isolates utilized diverse carbon sources ranging from acetate to glucose while growing under denitrifying conditions. The strains were also screened for nitrate tolerance and a couple of them were shown to be tolerant to 300-400 mM nitrate under anaerobic conditions. In the field site the bacteria live in consortia rather than in isolation, therefore we hypothesized that growth of these strains will be more robust in co-culture, as the denitrification pathway was segmented between the species. Three of the isolates (Pseudomonas fluorescens strains N1B4, N2E2, N2E3) were selected for in-depth analysis based on growth in pairwise co-cultures relative to monocultures, and the availability of the relevant genetic tools, such as transposon mutant libraries. Full genome sequencing showed that strain N2E3 has a truncated dentrification pathway: it lacks nitrous oxide reductase. Our results show strain N2E2 grow to maximum cell density an average of 45 hours more quickly when grown with strain N2E3 than in monoculture. Utilizing RB-TnSeq libraries of our strains, it was also found that some genes involved in nitrate reduction, sulfate permeability, molybdenum utilization, and anaerobic reduction are important for growth under these conditions. In addition, a few unexpected genes were also shown to be positively correlated to growth, such as genes homologous to genes for DNA proofreading or antibiotic production. These

  8. Human induced pluripotent stem cell (hiPSC)-derived neurons respond to convulsant drugs when co-cultured with hiPSC-derived astrocytes.

    Science.gov (United States)

    Ishii, Misawa Niki; Yamamoto, Koji; Shoji, Masanobu; Asami, Asano; Kawamata, Yuji

    2017-08-15

    Accurate risk assessment for drug-induced seizure is expected to be performed before entering clinical studies because of its severity and fatal damage to drug development. Induced pluripotent stem cell (iPSC) technology has allowed the use of human neurons and glial cells in toxicology studies. Recently, several studies showed the advantage of co-culture system of human iPSC (hiPSC)-derived neurons with rodent/human primary astrocytes regarding neuronal functions. However, the application of hiPSC-derived neurons for seizure risk assessment has not yet been fully addressed, and not at all when co-cultured with hiPSC-derived astrocytes. Here, we characterized hiPSC-derived neurons co-cultured with hiPSC-derived astrocytes to discuss how hiPSC-derived neurons are useful to assess seizure risk of drugs. First, we detected the frequency of spikes and synchronized bursts hiPSC-derived neurons when co-cultured with hiPSC-derived astrocytes for 8 weeks. This synchronized burst was suppressed by the treatment with 6-cyano-7-nitroquinoxaline-2,3-dione, α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor antagonist, and D-(-)-2-amino-5-phosphonopentanoic acid, an N-Methyl-d-aspartate (NMDA) receptor antagonist. These data suggested that co-cultured hiPSC-derived neurons formed synaptic connections mediated by AMPA and NMDA receptors. We also demonstrated that co-cultured hiPSC-derived neurons showed epileptiform activity upon treatment with gabazine or kaliotoxin. Finally, we performed single-cell transcriptome analysis in hiPSC-derived neurons and found that hiPSC-derived astrocytes activated the pathways involved in the activities of AMPA and NMDA receptor functions, neuronal polarity, and axon guidance in hiPSC-derived neurons. These data suggested that hiPSC-derived astrocytes promoted the development of action potential, synaptic functions, and neuronal networks in hiPSC-derived neurons, and then these functional alterations result in the epileptiform

  9. Giant hepatocellular adenoma; case report

    Energy Technology Data Exchange (ETDEWEB)

    Pitella, F.A.; Coutinho, A.M.N.; Coura Filho, G.B.; Costa, P.L.A.; Ono, C.R.; Watanabe, T.; Sapienza, M.T.; Hironaka, F.; Cerri, G.G.; Buchpiguel, C.A. [Universidade de Sao Paulo (FM/USP), SP (Brazil). Inst. de Radiologia. Servico de Medicina Nuclear

    2008-07-01

    Full text: Introduction: Hepatocellular adenoma is a benign hepatic tumor identified mainly in women during fertility age, with estimated incidence of 4/1000 inhabitants. It is usually unique, well circumscribed, with or without a capsule, size varying from 1 to 30 cm, with possible central areas of necrosis and hemorrhage. Case Report: A 37-year-old female patient presenting with no comorbities, use of hormonal birth control pills for 18 years, a condition of reduction in the consistency of feces, increase in number of daily defecations, abdominal cramps, and a stuffed sensation after meals for two years. A palpable abdominal mass extending from the right hypochondriac to the right iliac fossa was noticed four months ago. A computerized tomography (CT) showed an extensive hepatic mass on the right which was considered, within the diagnostic hypotheses, hepatic adenomatosis, without ruling out secondary lesions. A hepatic scintillography with {sup 99m}Tc-DISIDA showed an extensive exophytic area from segment V to the right iliac fossa with arterialized blood flow and hepatocytic activity, as well as a hepatic nodule in segment VII with hepatocytic activity consistent with the hepatic adenomas hypothesis. The biopsy confirmed the hepatic adenoma diagnosis and the patient was submitted to a partial hepatectomy and cholecystectomy with good clinical evolution. Conclusion: Nuclear Medicine may supplement the assessment of hepatic nodules, including giant masses, thus suggesting new hypotheses and direction to therapeutic conduct. (author)

  10. Treatment options for hepatocellular carcinoma.

    Science.gov (United States)

    Sandhu, Dalbir S; Tharayil, Vivek S; Lai, Jin-Ping; Roberts, Lewis R

    2008-02-01

    Hepatocellular carcinoma (HCC) is frequently diagnosed at advanced stages and has a high mortality rate. With improved survival of patients with cirrhotic liver disease and increased prevalence of chronic hepatitis C viral infections, a rise in the number of HCC cases is being reported worldwide. Early diagnosis and treatment can significantly improve the prognosis of patients with HCC. Although surgical resection is an important potentially curative therapy for liver tumors, in appropriately selected patients, liver transplantation has been shown to achieve excellent survival rates for a solid tumor. Locally ablative and locoregional therapies in the form of percutaneous ethanol injection, radiofrequency ablation, transcatheter arterial chemoembolization and transcatheter arterial radioembolization (TheraSphere) are viable options in patients with unresectable HCC. Unfortunately, the role of systemic therapy has been very limited in the treatment of these patients. Novel treatment options based on an improved understanding of the molecular pathogenesis of HCC are being explored. These targeted molecular therapies are aimed at growth factors and their receptors, intracellular signal transduction and cell cycle control. A substantial improvement in outcomes of intermediate and advanced stage HCC is expected with the advent of these targeted therapies, used in combination with surgical or locoregional therapies. Recent positive results from a large Phase III study of the receptor tyrosine kinase inhibitor, sorafenib, hold great promise in the treatment of HCC.

  11. Cryopreservation studies of an artificial co-culture between the cobalamin-requiring green alga Lobomonas rostrata and the bacterium Mesorhizobium loti.

    Science.gov (United States)

    Ridley, Christian J A; Day, John G; Smith, Alison G

    2018-01-01

    Algal-bacterial co-cultures, rather than cultures of algae alone, are regarded as having the potential to enhance productivity and stability in industrial algal cultivation. As with other inocula in biotechnology, to avoid loss of production strains, it is important to develop preservation methods for the long-term storage of these cultures, and one of the most commonly used approaches is cryopreservation. However, whilst there are many reports of cryopreserved xenic algal cultures, little work has been reported on the intentional preservation of both algae and beneficial bacteria in xenic cultures. Instead, studies have focused on the development of methods to conserve the algal strain(s) present, or to avoid overgrowth of bacteria in xenic isolates during the post-thaw recovery phase. Here, we have established a co-cryopreservation method for the long-term storage of both partners in a unialgal-bacterial co-culture. This is an artificial model mutualism between the alga Lobomonas rostrata and the bacterium Mesorhizobium loti , which provides vitamin B 12 (cobalamin) to the alga in return for photosynthate. Using a Planer Kryo 360 controlled-rate cooler, post-thaw viability (PTV) values of 72% were obtained for the co-culture, compared to 91% for the axenic alga. The cultures were successfully revived after 6 months storage in liquid nitrogen, and continued to exhibit mutualism. Furthermore, the alga could be cryopreserved with non-symbiotic bacteria, without bacterial overgrowth occurring. It was also possible to use less controllable passive freezer chambers to cryopreserve the co-cultures, although the PTV was lower. Finally, we demonstrated that an optimised cryopreservation method may be used to prevent the overgrowth potential of non-symbiotic, adventitious bacteria in both axenic and co-cultures of L. rostrata after thawing.

  12. 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.

  13. Damage of Neuroblastoma Cell SH-SY5Y Mediated by MPP+ Inhibits Proliferation of T-Cell Leukemia Jurkat by Co-Culture System

    Directory of Open Access Journals (Sweden)

    Fuli Wang

    2014-06-01

    Full Text Available The adaptive immune system has implications in pathology of Parkinson’s disease (PD. Research data demonstrated that the peripheral CD4+ T-cell population decreased in pathogenesis of PD. The effect of damaged dopaminergic neurons on peripheral T cells of PD is still unknown. In this study, we constructed a neuronal and glial cells co-culture model by using human neuroblastoma cells SH-SY5Y and gliomas cells U87. After the co-culture cells were treated with neurotoxin 1-methyl-4-phenylpyridinium (MPP+ for 24 h, the conditioned media was harvested and used to cultivate T-cell leukemia Jurkat cells for another 24 h. We then analyzed the cell proliferation, cell cycle and necrosis effect of Jurkat cells. The results showed that co-culture medium of SH-SY5Y and U87 cells with MPP+ treatment inhibited the proliferation of Jurkat cells compared to control medium without MPP+, even though the same concentration of MPP+ had very little toxicity to the Jurkat cell. Furthermore, co-culture medium with low concentration of MPP+ (100 µM arrested Jurkat cells cycle in G2/M phase through increasing cell cycle division 2 (CDC2 and CyclinB1 expression level, whereas co-culture medium with high concentration of MPP+ (500 µM induced Jurkat cell necrosis through cellular swelling and membrane breakage. Our data implies that damaged dopamine neurons with glial cells can lead to the reduced number or inhibited proliferation activity of peripheral T cells.

  14. IGF-1 promotes angiogenesis in endothelial cells/adipose-derived stem cells co-culture system with activation of PI3K/Akt signal pathway.

    Science.gov (United States)

    Lin, Shiyu; Zhang, Qi; Shao, Xiaoru; Zhang, Tao; Xue, Changyue; Shi, Sirong; Zhao, Dan; Lin, Yunfeng

    2017-12-01

    The aim of this study was to investigate the role of insulin-like growth factor-1 (IGF-1) and crosstalk between endothelial cells (ECs) and adipose-derived stem cells (ASCs) in the process of angiogenesis. A three-dimensional collagen gel used to culture mouse ASCs and mouse ECs in vitro was established. The effects of angiogenesis after exposure to IGF-1 were observed by confocal laser scanning microscopy. Western blotting and qPCR were performed to elucidate the underlying mechanisms. IGF-1 treatment promoted the formation of vessel-like structures and the recruitment of ASCs in the three-dimensional collagen gel. The angiogenic genes and proteins in ECs were up-regulated by IGF-1 and in co-culture. Similar changes in the genes and in the proteins were detected in ASCs after exposure to IGF-1 and co-culture. p-Akt expression levels were high in ECs and ASCs after exposure to IGF-1 and co-culture. IGF-1 and co-culture between cells facilitate the process of angiogenesis via the PI3-kinase/Akt signalling pathway. In ECs, IGF-1 stimulates the expression of angiogenesis-related growth factors with the activation of the PI3-kinase/Akt signalling pathway. Co-cultured ECs exposed to excess VEGF-A and other angiogenesis-related growth factors para-secreted from ASCs exhibit high expression of angiogenesis-related genes and proteins. In ASCs, IGF-1 induces the recruitment and function of ASCs by up-regulating the expression of PDGFB, MMPs and α-SMA. Crosstalk with ECs further facilitates changes in ASCs. © 2017 John Wiley & Sons Ltd.

  15. Influence of epidermal growth factor (EGF) and hydrocortisone on the co-culture of mature adipocytes and endothelial cells for vascularized adipose tissue engineering.

    Science.gov (United States)

    Huber, Birgit; Czaja, Alina Maria; Kluger, Petra Juliane

    2016-05-01

    The composition of vascularized adipose tissue is still an ongoing challenge as no culture medium is available to supply adipocytes and endothelial cells appropriately. Endothelial cell medium is typically supplemented with epidermal growth factor (EGF) as well as hydrocortisone (HC). The effect of EGF on adipocytes is discussed controversially. Some studies say it inhibits adipocyte differentiation while others reported of improved adipocyte lipogenesis. HC is known to have lipolytic activities, which might result in mature adipocyte dedifferentiation. In this study, we evaluated the influence of EGF and HC on the co-culture of endothelial cells and mature adipocytes regarding their cell morphology and functionality. We showed in mono-culture that high levels of HC promoted dedifferentiation and proliferation of mature adipocytes, whereas EGF seemed to have no negative influence. Endothelial cells kept their typical cobblestone morphology and showed a proliferation rate comparable to the control independent of EGF and HC concentration. In co-culture, HC promoted dedifferentiation of mature adipocytes, which was shown by a higher glycerol release. EGF had no negative impact on adipocyte morphology. No negative impact on endothelial cell morphology and functionality could be seen with reduced EGF and HC supplementation in co-culture with mature adipocytes. Taken together, our results demonstrate that reduced levels of HC are needed for co-culturing mature adipocytes and endothelial cells. In co-culture, EGF had no influence on mature adipocytes. Therefore, for the composition of vascularized adipose tissue constructs, the media with low levels of HC and high or low levels of EGF can be used. © 2016 International Federation for Cell Biology.

  16. Co-culture of clonal beta cells with GLP-1 and glucagon-secreting cell line impacts on beta cell insulin secretion, proliferation and susceptibility to cytotoxins.

    Science.gov (United States)

    Green, Alastair D; Vasu, Srividya; Moffett, R Charlotte; Flatt, Peter R

    2016-06-01

    We investigated the direct effects on insulin releasing MIN6 cells of chronic exposure to GLP-1, glucagon or a combination of both peptides secreted from GLUTag L-cell and αTC1.9 alpha-cell lines in co-culture. MIN6, GLUTag and αTC1.9 cell lines exhibited high cellular hormone content and release of insulin, GLP-1 and glucagon, respectively. Co-culture of MIN6 cells with GLUTag cells significantly increased cellular insulin content, beta-cell proliferation, insulin secretory responses to a range of established secretogogues and afforded protection against exposure cytotoxic concentrations of glucose, lipid, streptozotocin or cytokines. Benefits of co-culture of MIN6 cells with αTC1.9 alphacells were limited to enhanced beta-cell proliferation with marginal positive actions on both insulin secretion and cellular protection. In contrast, co-culture of MIN6 with GLUTag cells plus αTC1.9 cells, markedly enhanced both insulin secretory responses and protection against beta-cell toxins compared with co-culture with GLUTag cells alone. These data indicate important long-term effects of conjoint GLP-1 and glucagon exposure on beta-cell function. This illustrates the possible functional significance of alpha-cell GLP-1 production as well as direct beneficial effects of dual agonism at beta-cell GLP-1 and glucagon receptors. Copyright © 2016 Elsevier B.V. and Société française de biochimie et biologie Moléculaire (SFBBM). All rights reserved.

  17. Protective role of klotho protein on epithelial cells upon co-culture with activated or senescent monocytes

    Energy Technology Data Exchange (ETDEWEB)

    Mytych, Jennifer, E-mail: jennifermytych@gmail.com [Institute of Applied Biotechnology and Basic Sciences, University of Rzeszow, Werynia 502, 36-100 Kolbuszowa (Poland); Centre of Applied Biotechnology and Basic Sciences, University of Rzeszow, Werynia 502, 36-100 Kolbuszowa (Poland); Wos, Izabela; Solek, Przemyslaw; Koziorowski, Marek [Institute of Applied Biotechnology and Basic Sciences, University of Rzeszow, Werynia 502, 36-100 Kolbuszowa (Poland); Centre of Applied Biotechnology and Basic Sciences, University of Rzeszow, Werynia 502, 36-100 Kolbuszowa (Poland)

    2017-01-15

    Monocytes ensure proper functioning and maintenance of epithelial cells, while good condition of monocytes is a key factor of these interactions. Although, it was shown that in some circumstances, a population of altered monocytes may appear, there is no data regarding their effect on epithelial cells. In this study, using direct co-culture model with LPS-activated and Dox-induced senescent THP-1 monocytes, we reported for the first time ROS-induced DNA damage, reduced metabolic activity, proliferation inhibition and cell cycle arrest followed by p16-, p21- and p27-mediated DNA damage response pathways activation, premature senescence and apoptosis induction in HeLa cells. Also, we show that klotho protein possessing anti-aging and anti-inflammatory characteristics reduced cytotoxic and genotoxic events by inhibition of insulin/IGF-IR and downregulation of TRF1 and TRF2 proteins. Therefore, klotho protein could be considered as a protective factor against changes caused by altered monocytes in epithelial cells. - Highlights: • Activated and senescent THP-1 monocytes induced cyto- and genotoxicity in HeLa cells. • Altered monocytes provoked oxidative and nitrosative stress-induced DNA damage. • DNA damage activated DDR pathways and lead to premature senescence and apoptosis. • Klotho reduced ROS/RNS-mediated toxicity through insulin/IGF-IR pathway inhibition. • Klotho protects HeLa cells from cyto- and genotoxicity induced by altered monocytes.

  18. Transcriptome analysis of Acidovorax avenae subsp. avenae cultivated in vivo and co-culture with Burkholderia seminalis.

    Science.gov (United States)

    Li, Bin; Ibrahim, Muhammad; Ge, Mengyu; Cui, Zhouqi; Sun, Guochang; Xu, Fei; Kube, Michael

    2014-07-16

    Response of bacterial pathogen to environmental bacteria and its host is critical for understanding of microbial adaption and pathogenesis. Here, we used RNA-Seq to comprehensively and quantitatively assess the transcriptional response of Acidovorax avenae subsp. avenae strain RS-1 cultivated in vitro, in vivo and in co-culture with rice rhizobacterium Burkholderia seminalis R456. Results revealed a slight response to other bacteria, but a strong response to host. In particular, a large number of virulence associated genes encoding Type I to VI secretion systems, 118 putative non-coding RNAs, and 7 genomic islands (GIs) were differentially expressed in vivo based on comparative genomic and transcriptomic analyses. Furthermore, the loss of virulence for knockout mutants of 11 differentially expressed T6SS genes emphasized the importance of these genes in bacterial pathogenicity. In addition, the reliability of expression data obtained by RNA-Seq was supported by quantitative real-time PCR of the 25 selected T6SS genes. Overall, this study highlighted the role of differentially expressed genes in elucidating bacterial pathogenesis based on combined analysis of RNA-Seq data and knockout of T6SS genes.

  19. Co-culture microorganisms with different initial proportions reveal the mechanism of chalcopyrite bioleaching coupling with microbial community succession.

    Science.gov (United States)

    Ma, Liyuan; Wang, Xingjie; Feng, Xue; Liang, Yili; Xiao, Yunhua; Hao, Xiaodong; Yin, Huaqun; Liu, Hongwei; Liu, Xueduan

    2017-01-01

    The effect of co-culture microorganisms with different initial proportions on chalcopyrite bioleaching was investigated. Communities were rebuilt by six typical strains isolated from the same habitat. The results indicated, by community with more sulfur oxidizers at both 30 and 40°C, the final copper extraction rate was 19.8% and 6.5% higher, respectively, than that with more ferrous oxidizers. The variations of pH, redox potential, ferrous and copper ions in leachate also provided evidences that community with more sulfur oxidizers was more efficient. Community succession of free and attached cells revealed that initial proportions played decisive roles on community dynamics at 30°C, while communities shared similar structures, not relevant to initial proportions at 40°C. X-ray diffraction analysis confirmed different microbial functions on mineral surface. A mechanism model for chalcopyrite bioleaching was established coupling with community succession. This will provide theoretical basis for reconstructing an efficient community in industrial application. Copyright © 2016 Elsevier Ltd. All rights reserved.

  20. Heterotrophic cultivation of Chlorella pyrenoidosa using sucrose as the sole carbon source by co-culture with Rhodotorula glutinis.

    Science.gov (United States)

    Wang, Shikai; Wu, Yong; Wang, Xu

    2016-11-01

    Heterotrophic cultivation of microalgae is a feasible alternative strategy to avoid the light limitation of photoautotrophic culture, but the heterotrophic utilization of disaccharides is difficult for microalgae. Aimed at this problem, a co-culture system was developed by mix culture of C. pyrenoidosa and R. glutinis using sucrose as the sole carbon source. In this system, C. pyrenoidosa could utilize glucose and fructose which were hydrolyzed from sucrose by R. glutinis. The highest specific growth rate and final cell number proportion of algae was 1.02day(-1) and 45%, respectively, when cultured at the initial algal cell number proportion of 95.24% and the final algal cell density was 111.48×10(6)cells/mL. In addition, the lipid content was also promoted due to the synergistic effects in mix culture. This study provides a novel approach using sucrose-riched wastes for the heterotrophic culture of microalgae and may effectively decrease the cost of carbon source. Copyright © 2016 Elsevier Ltd. All rights reserved.

  1. Microglia in Glia-Neuron Co-cultures Exhibit Robust Phagocytic Activity Without Concomitant Inflammation or Cytotoxicity.

    Science.gov (United States)

    Adams, Alexandra C; Kyle, Michele; Beaman-Hall, Carol M; Monaco, Edward A; Cullen, Matthew; Vallano, Mary Lou

    2015-10-01

    A simple method to co-culture granule neurons and glia from a single brain region is described, and microglia activation profiles are assessed in response to naturally occurring neuronal apoptosis, excitotoxin-induced neuronal death, and lipopolysaccharide (LPS) addition. Using neonatal rat cerebellar cortex as a tissue source, glial proliferation is regulated by omission or addition of the mitotic inhibitor cytosine arabinoside (AraC). After 7-8 days in vitro, microglia in AraC(-) cultures are abundant and activated based on their amoeboid morphology, expressions of ED1 and Iba1, and ability to phagocytose polystyrene beads and the majority of neurons undergoing spontaneous apoptosis. Microglia and phagocytic activities are sparse in AraC(+) cultures. Following exposure to excitotoxic kainate concentrations, microglia in AraC(-) cultures phagocytose most dead neurons within 24 h without exacerbating neuronal loss or mounting a strong or sustained inflammatory response. LPS addition induces a robust inflammatory response, based on microglial expressions of TNF-α, COX-2 and iNOS proteins, and mRNAs, whereas these markers are essentially undetectable in control cultures. Thus, the functional effector state of microglia is primed for phagocytosis but not inflammation or cytotoxicity even after kainate exposure that triggers death in the majority of neurons. This model should prove useful in studying the progressive activation states of microglia and factors that promote their conversion to inflammatory and cytotoxic phenotypes.

  2. Protective role of klotho protein on epithelial cells upon co-culture with activated or senescent monocytes

    International Nuclear Information System (INIS)

    Mytych, Jennifer; Wos, Izabela; Solek, Przemyslaw; Koziorowski, Marek

    2017-01-01

    Monocytes ensure proper functioning and maintenance of epithelial cells, while good condition of monocytes is a key factor of these interactions. Although, it was shown that in some circumstances, a population of altered monocytes may appear, there is no data regarding their effect on epithelial cells. In this study, using direct co-culture model with LPS-activated and Dox-induced senescent THP-1 monocytes, we reported for the first time ROS-induced DNA damage, reduced metabolic activity, proliferation inhibition and cell cycle arrest followed by p16-, p21- and p27-mediated DNA damage response pathways activation, premature senescence and apoptosis induction in HeLa cells. Also, we show that klotho protein possessing anti-aging and anti-inflammatory characteristics reduced cytotoxic and genotoxic events by inhibition of insulin/IGF-IR and downregulation of TRF1 and TRF2 proteins. Therefore, klotho protein could be considered as a protective factor against changes caused by altered monocytes in epithelial cells. - Highlights: • Activated and senescent THP-1 monocytes induced cyto- and genotoxicity in HeLa cells. • Altered monocytes provoked oxidative and nitrosative stress-induced DNA damage. • DNA damage activated DDR pathways and lead to premature senescence and apoptosis. • Klotho reduced ROS/RNS-mediated toxicity through insulin/IGF-IR pathway inhibition. • Klotho protects HeLa cells from cyto- and genotoxicity induced by altered monocytes.

  3. Co-Cultures of Pseudomonas aeruginosa and Roseobacter denitrificans Reveal Shifts in Gene Expression Levels Compared to Solo Cultures

    Directory of Open Access Journals (Sweden)

    Crystal A. Conway

    2012-01-01

    Full Text Available Consistent biosynthesis of desired secondary metabolites (SMs from pure microbial cultures is often unreliable. In a proof-of-principle study to induce SM gene expression and production, we describe mixed “co-culturing” conditions and monitoring of messages via quantitative real-time PCR (qPCR. Gene expression of model bacterial strains (Pseudomonas aeruginosa PAO1 and Roseobacter denitrificans Och114 was analyzed in pure solo and mixed cocultures to infer the effects of interspecies interactions on gene expression in vitro, Two P. aeruginosa genes (PhzH coding for portions of the phenazine antibiotic pathway leading to pyocyanin (PCN and the RhdA gene for thiosulfate: cyanide sulfurtransferase (Rhodanese and two R. denitrificans genes (BetaLact for metallo-beta-lactamase and the DMSP gene for dimethylpropiothetin dethiomethylase were assessed for differential expression. Results showed that R. denitrificans DMSP and BetaLact gene expression became elevated in a mixed culture. In contrast, P. aeruginosa co-cultures with R. denitrificans or a third species did not increase target gene expression above control levels. This paper provides insight for better control of target SM gene expression in vitro and bypass complex genetic engineering manipulations.

  4. Patterned cell arrays and patterned co-cultures on polydopamine-modified poly(vinyl alcohol) hydrogels

    International Nuclear Information System (INIS)

    Beckwith, Kai M; Sikorski, Pawel

    2013-01-01

    Live cell arrays are an emerging tool that expand traditional 2D in vitro cell culture, increasing experimental precision and throughput. A patterned cell system was developed by combining the cell-repellent properties of polyvinyl alcohol hydrogels with the cell adhesive properties of self-assembled films of dopamine (polydopamine). It was shown that polydopamine could be patterned onto spin-cast polyvinyl alcohol hydrogels by microcontact printing, which in turn effectively patterned the growth of several cell types (HeLa, human embryonic kidney, human umbilical vein endothelial cells (HUVEC) and prostate cancer). The cells could be patterned in geometries down to single-cell confinement, and it was demonstrated that cell patterns could be maintained for at least 3 weeks. Furthermore, polydopamine could be used to modify poly(vinyl alcohol) in situ using a cell-compatible deposition buffer (1 mg mL −1 dopamine in 25 mM tris with a physiological salt balance). The treatment switched the PVA hydrogel from cell repellent to cell adhesive. Finally, by combining microcontact printing and in situ deposition of polydopamine, patterned co-cultures of the same cell type (HeLa/HeLa) and dissimilar cell types (HeLa/HUVEC) were realized through simple chemistry and could be studied over time. The combination of polyvinyl alcohol and polydopamine was shown to be an attractive route to versatile, patterned cell culture experiments with minimal infrastructure requirements and low complexity. (paper)

  5. Effect of micro-patterned fluorine-doped tin oxide films on electrochromic properties of Prussian blue films

    International Nuclear Information System (INIS)

    Lee, Kyuha; Kim, A-Young; Park, Ji Hun; Jung, Hun-Gi; Choi, Wonchang; Lee, Hwa Young; Lee, Joong Kee

    2014-01-01

    Graphical abstract: - Highlights: • PB-based ECD employed micro-patterned FTO electrode was fabricated. • Effect of interface morphology on electrochromic characteristics was examined. • Electrochromic properties were enhanced by employing a patterned interface. - Abstract: The effect of interface morphology on electrochromic characteristics was examined for an electrochromic device (ECD). Micro-patterned fluorine-doped tin oxide (FTO) films were fabricated using a photolithography process. Prussian blue (PB) films were then deposited on the patterned FTO films. The surface areas of both PB films and FTO films were increased by patterning. ECDs were assembled using patterned PB/FTO films as the electrochromic electrode, bare FTO films as the counter electrode, and an electrolyte containing LiClO 4 salt. The increased effective surface area of the patterned PB/FTO electrode boosted the mobility of ions at the interphase between the electrolyte and PB electrode, and the electron transfer between PB films and FTO films. As a result, electrochromic properties such as transmittance and response time were significantly improved by employing the patterned FTO films as the transparent conductive oxide layer of the electrochromic electrode

  6. Effect of micro-patterned fluorine-doped tin oxide films on electrochromic properties of Prussian blue films

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Kyuha [Center for Energy Convergence Research, Green City Technology Institute, Korea Institute of Science and Technology, Hwarangno 14-gil 5, Seongbuk-gu, Seoul 136-791 (Korea, Republic of); Kim, A-Young [Center for Energy Convergence Research, Green City Technology Institute, Korea Institute of Science and Technology, Hwarangno 14-gil 5, Seongbuk-gu, Seoul 136-791 (Korea, Republic of); Department of Material Science and Engineering, Korea University, Seoul 136-701 (Korea, Republic of); Park, Ji Hun; Jung, Hun-Gi; Choi, Wonchang; Lee, Hwa Young [Center for Energy Convergence Research, Green City Technology Institute, Korea Institute of Science and Technology, Hwarangno 14-gil 5, Seongbuk-gu, Seoul 136-791 (Korea, Republic of); Lee, Joong Kee, E-mail: leejk@kist.re.kr [Center for Energy Convergence Research, Green City Technology Institute, Korea Institute of Science and Technology, Hwarangno 14-gil 5, Seongbuk-gu, Seoul 136-791 (Korea, Republic of)

    2014-09-15

    Graphical abstract: - Highlights: • PB-based ECD employed micro-patterned FTO electrode was fabricated. • Effect of interface morphology on electrochromic characteristics was examined. • Electrochromic properties were enhanced by employing a patterned interface. - Abstract: The effect of interface morphology on electrochromic characteristics was examined for an electrochromic device (ECD). Micro-patterned fluorine-doped tin oxide (FTO) films were fabricated using a photolithography process. Prussian blue (PB) films were then deposited on the patterned FTO films. The surface areas of both PB films and FTO films were increased by patterning. ECDs were assembled using patterned PB/FTO films as the electrochromic electrode, bare FTO films as the counter electrode, and an electrolyte containing LiClO{sub 4} salt. The increased effective surface area of the patterned PB/FTO electrode boosted the mobility of ions at the interphase between the electrolyte and PB electrode, and the electron transfer between PB films and FTO films. As a result, electrochromic properties such as transmittance and response time were significantly improved by employing the patterned FTO films as the transparent conductive oxide layer of the electrochromic electrode.

  7. Micro-patterned agarose gel devices for single-cell high-throughput microscopy of E. coli cells.

    Science.gov (United States)

    Priest, David G; Tanaka, Nobuyuki; Tanaka, Yo; Taniguchi, Yuichi

    2017-12-21

    High-throughput microscopy of bacterial cells elucidated fundamental cellular processes including cellular heterogeneity and cell division homeostasis. Polydimethylsiloxane (PDMS)-based microfluidic devices provide advantages including precise positioning of cells and throughput, however device fabrication is time-consuming and requires specialised skills. Agarose pads are a popular alternative, however cells often clump together, which hinders single cell quantitation. Here, we imprint agarose pads with micro-patterned 'capsules', to trap individual cells and 'lines', to direct cellular growth outwards in a straight line. We implement this micro-patterning into multi-pad devices called CapsuleHotel and LineHotel for high-throughput imaging. CapsuleHotel provides ~65,000 capsule structures per mm 2 that isolate individual Escherichia coli cells. In contrast, LineHotel provides ~300 line structures per mm that direct growth of micro-colonies. With CapsuleHotel, a quantitative single cell dataset of ~10,000 cells across 24 samples can be acquired and analysed in under 1 hour. LineHotel allows tracking growth of > 10 micro-colonies across 24 samples simultaneously for up to 4 generations. These easy-to-use devices can be provided in kit format, and will accelerate discoveries in diverse fields ranging from microbiology to systems and synthetic biology.

  8. Integration of 3D Printed and Micropatterned Polycaprolactone Scaffolds for Guidance of Oriented Collagenous Tissue Formation In Vivo.

    Science.gov (United States)

    Pilipchuk, Sophia P; Monje, Alberto; Jiao, Yizu; Hao, Jie; Kruger, Laura; Flanagan, Colleen L; Hollister, Scott J; Giannobile, William V

    2016-03-01

    Scaffold design incorporating multiscale cues for clinically relevant, aligned tissue regeneration has potential to improve structural and functional integrity of multitissue interfaces. The objective of this preclinical study is to develop poly(ε-caprolactone) (PCL) scaffolds with mesoscale and microscale architectural cues specific to human ligament progenitor cells and assess their ability to form aligned bone-ligament-cementum complexes in vivo. PCL scaffolds are designed to integrate a 3D printed bone region with a micropatterned PCL thin film consisting of grooved pillars. The patterned film region is seeded with human ligament cells, fibroblasts transduced with bone morphogenetic protein-7 genes seeded within the bone region, and a tooth dentin segment positioned on the ligament region prior to subcutaneous implantation into a murine model. Results indicate increased tissue alignment in vivo using micropatterned PCL films, compared to random-porous PCL. At week 6, 30 μm groove depth significantly enhances oriented collagen fiber thickness, overall cell alignment, and nuclear elongation relative to 10 μm groove depth. This study demonstrates for the first time that scaffolds with combined hierarchical mesoscale and microscale features can align cells in vivo for oral tissue repair with potential for improving the regenerative response of other bone-ligament complexes. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Current radiologic interventions in hepatocellular carcinoma

    International Nuclear Information System (INIS)

    Masoud, I.; Naeem, M.Q.T.; Saeed, F.; Mirza, S.A.M.; Khan, A.; Bhatti, M.A.

    2006-01-01

    With the rising incidence of chronic liver disease caused by viral hepatitis, hepatocellular carcinoma is showing a corresponding rise worldwide. Surgery remains the mainstay of treatment, but patients unfit for surgery or liver transplantation form the bulk of those presenting with this disease. Palliative treatments are being used to treat those and radiological modalities form the mainstay of the treatment. Radiology plays a major role in the diagnosis, treatment and follow-up of hepatocellular carcinoma. Current radiological treatment modalities include percutaneous ethanol ablation, radiofrequency ablation and trans-arterial chemoembolization. This update highlights the recent advancements in the field and compares their relative merits and demerits. (author)

  10. Surgical Treatment of Hepatocellular Carcinoma

    Science.gov (United States)

    Zamora-Valdes, Daniel; Taner, Timucin; Nagorney, David M.

    2017-01-01

    Hepatocellular carcinoma (HCC) is a major cause of cancer-related death worldwide. In select patients, surgical treatment in the form of either resection or transplantation offers a curative option. The aims of this review are to (1) review the current American Association for the Study of Liver Diseases/European Association for the Study of the Liver guidelines on the surgical management of HCC and (2) review the proposed changes to these guidelines and analyze the strength of evidence underlying these proposals. Three authors identified the most relevant publications in the literature on liver resection and transplantation for HCC and analyzed the strength of evidence according to the Grading of Recommendations Assessment, Development and Evaluation (GRADE) classification. In the United States, the liver allocation system provides priority for liver transplantation to patients with HCC within the Milan criteria. Current evidence suggests that liver transplantation may also be indicated in certain patient groups beyond Milan criteria, such as pediatric patients with large tumor burden or adult patients who are successfully downstaged. Patients with no underlying liver disease may also benefit from liver transplantation if the HCC is unresectable. In patients with no or minimal (compensated) liver disease and solitary HCC ≥2 cm, liver resection is warranted. If liver transplantation is not available or contraindicated, liver resection can be offered to patients with multinodular HCC, provided that the underlying liver disease is not decompensated. Many patients may benefit from surgical strategies adapted to local resources and policies (hepatitis B prevalence, organ availability, etc). Although current low-quality evidence shows better overall survival with aggressive surgical strategies, this approach is limited to select patients. Larger and well-designed prospective studies are needed to better define the benefits and limits of such approach. PMID:28975836

  11. New advances in hepatocellular carcinoma

    Science.gov (United States)

    Pascual, Sonia; Herrera, Iván; Irurzun, Javier

    2016-01-01

    Hepatocellular carcinoma (HCC) is the leading cause of deaths in cirrhotic patients and the third cause of cancer related deaths. Most HCC are associated with well known underlying risk factors, in fact, HCC arise in cirrhotic patients in up to 90% of cases, mainly due to chronic viral hepatitis and alcohol abuse. The worldwide prevention strategies are conducted to avoid the infection of new subjects and to minimize the risk of liver disease progression in infected patients. HCC is a condition which lends itself to surveillance as at-risk individuals can readily be identified. The American and European guidelines recommended implementation of surveillance programs with ultrasound every six months in patient at-risk for developing HCC. The diagnosis of HCC can be based on non-invasive criteria (only in cirrhotic patient) or pathology. Accurately staging patients is essential to oncology practice. The ideal tumour staging system in HCC needs to account for both tumour characteristics and liver function. Treatment allocation is based on several factors: Liver function, size and number of tumours, macrovascular invasion or extrahepatic spread. The recommendations in terms of selection for different treatment strategies must be based on evidence-based data. Resection, liver transplant and interventional radiology treatment are mainstays of HCC therapy and achieve the best outcomes in well-selected candidates. Chemoembolization is the most widely used treatment for unresectable HCC or progression after curative treatment. Finally, in patients with advanced HCC with preserved liver function, sorafenib is the only approved systemic drug that has demonstrated a survival benefit and is the standard of care in this group of patients. PMID:27028578

  12. High-resolution spatiotemporal strain mapping reveals non-uniform deformation in micropatterned elastomers

    Science.gov (United States)

    Aksoy, B.; Rehman, A.; Bayraktar, H.; Alaca, B. E.

    2017-04-01

    Micropatterns are generated on a vast selection of polymeric substrates for various applications ranging from stretchable electronics to cellular mechanobiological systems. When these patterned substrates are exposed to external loading, strain field is primarily affected by the presence of microfabricated structures and similarly by fabrication-related defects. The capturing of such nonhomogeneous strain fields is of utmost importance in cases where study of the mechanical behavior with a high spatial resolution is necessary. Image-based non-contact strain measurement techniques are favorable and have recently been extended to scanning tunneling microscope and scanning electron microscope images for the characterization of mechanical properties of metallic materials, e.g. steel and aluminum, at the microscale. A similar real-time analysis of strain heterogeneity in elastomers is yet to be achieved during the entire loading sequence. The available measurement methods for polymeric materials mostly depend on cross-head displacement or precalibrated strain values. Thus, they suffer either from the lack of any real-time analysis, spatiotemporal distribution or high resolution in addition to a combination of these factors. In this work, these challenges are addressed by integrating a tensile stretcher with an inverted optical microscope and developing a subpixel particle tracking algorithm. As a proof of concept, the patterns with a critical dimension of 200 µm are generated on polydimethylsiloxane substrates and strain distribution in the vicinity of the patterns is captured with a high spatiotemporal resolution. In the field of strain measurement, there is always a tradeoff between minimum measurable strain value and spatial resolution. Current noncontact techniques on elastomers can deliver a strain resolution of 0.001% over a minimum length of 5 cm. More importantly, inhomogeneities within this quite large region cannot be captured. The proposed technique can

  13. High-resolution spatiotemporal strain mapping reveals non-uniform deformation in micropatterned elastomers

    International Nuclear Information System (INIS)

    Aksoy, B; Alaca, B E; Rehman, A; Bayraktar, H

    2017-01-01

    Micropatterns are generated on a vast selection of polymeric substrates for various applications ranging from stretchable electronics to cellular mechanobiological systems. When these patterned substrates are exposed to external loading, strain field is primarily affected by the presence of microfabricated structures and similarly by fabrication-related defects. The capturing of such nonhomogeneous strain fields is of utmost importance in cases where study of the mechanical behavior with a high spatial resolution is necessary. Image-based non-contact strain measurement techniques are favorable and have recently been extended to scanning tunneling microscope and scanning electron microscope images for the characterization of mechanical properties of metallic materials, e.g. steel and aluminum, at the microscale. A similar real-time analysis of strain heterogeneity in elastomers is yet to be achieved during the entire loading sequence. The available measurement methods for polymeric materials mostly depend on cross-head displacement or precalibrated strain values. Thus, they suffer either from the lack of any real-time analysis, spatiotemporal distribution or high resolution in addition to a combination of these factors. In this work, these challenges are addressed by integrating a tensile stretcher with an inverted optical microscope and developing a subpixel particle tracking algorithm. As a proof of concept, the patterns with a critical dimension of 200 µ m are generated on polydimethylsiloxane substrates and strain distribution in the vicinity of the patterns is captured with a high spatiotemporal resolution. In the field of strain measurement, there is always a tradeoff between minimum measurable strain value and spatial resolution. Current noncontact techniques on elastomers can deliver a strain resolution of 0.001% over a minimum length of 5 cm. More importantly, inhomogeneities within this quite large region cannot be captured. The proposed technique can

  14. Metabolomics Investigation of an Association of Induced Features and Corresponding Fungus during the Co-culture of Trametes versicolor and Ganoderma applanatum

    Science.gov (United States)

    Xu, Xiao-Yan; Shen, Xiao-Ting; Yuan, Xiao-Jie; Zhou, Yuan-Ming; Fan, Huan; Zhu, Li-Ping; Du, Feng-Yu; Sadilek, Martin; Yang, Jie; Qiao, Bin; Yang, Song

    2018-01-01

    The co-culture of Trametes versicolor and Ganoderma applanatum is a model of intense basidiomycete interaction, which induces many newly synthesized or highly produced features. Currently, one of the major challenges is an identification of the origin of induced features during the co-culture. Herein, we report a 13C-dynamic labeling analysis used to determine an association of induced features and corresponding fungus even if the identities of metabolites were not available or almost nothing was known of biochemical aspects. After the co-culture of T. versicolor and G. applanatum for 10 days, the mycelium pellets of T. versicolor and G. applanatum were sterilely harvested and then mono-cultured in the liquid medium containing half fresh medium with 13C-labeled glucose as carbon source and half co-cultured supernatants collected on day 10. 13C-labeled metabolome analyzed by LC-MS revealed that 31 induced features including 3-phenyllactic acid and orsellinic acid were isotopically labeled in the mono-culture after the co-culture stimulation. Twenty features were derived from T. versicolor, 6 from G. applanatum, and 5 features were synthesized by both T. versicolor and G. applanatum. 13C-labeling further suggested that 12 features such as previously identified novel xyloside [N-(4-methoxyphenyl)formamide 2-O-beta-D-xyloside] were likely induced through the direct physical interaction of mycelia. Use of molecular network analysis combined with 13C-labeling provided an insight into the link between the generation of structural analogs and producing fungus. Compound 1 with m/z 309.0757, increased 15.4-fold in the co-culture and observed 13C incorporation in the mono-culture of both T. versicolor and G. applanatum, was purified and identified as a phenyl polyketide, 2,5,6-trihydroxy-4, 6-diphenylcyclohex-4-ene-1,3-dione. The biological activity study indicated that this compound has a potential to inhibit cell viability of leukemic cell line U937. The current work sets

  15. Bulky PAH-DNA induced by exposure of a co-culture model of human alveolar macrophages and embryonic epithelial cells to atmospheric particulate pollution

    International Nuclear Information System (INIS)

    Abbas, Imane; Garcon, Guillaume; Billet, Sylvain; Shirali, Pirouz; Andre, Veronique; Le Goff, Jeremie; Sichel, Francois; Roy Saint-Georges, Francoise; Mulliez, Philippe

    2012-01-01

    Because of their deep penetration in human lungs, fine airborne particulate matter were described as mainly responsible for the deleterious effects of exposure to air pollution on health. Organic constituents are adsorbed on particles surface and, after inhalation, some (polycyclic aromatic hydrocarbons, PAHs) can be activated into reactive metabolites and can bind to DNA. The formation of bulky DNA adducts has been researched after exposure of mono-and co-cultures of alveolar macrophages (AM) and human embryonic human lung epithelial (L132), to fine air pollution particulate matter Air samples have been collected with cascade impactor and characterized: size distribution (92.15% 2 /g), inorganic (Fe, AI, Ca, Na, K, Mg, Pb, etc.) and organic compounds (PAHs, etc.). 32 P post-labeling method was applied to detect bulky DNA adducts in AM and L132, in mono-and co-cultures, 72 h after their exposure to atmospheric particles at their Lethals and Effects concentrations or (LC or CE) to 50% (i.e. MA: EC 50 = 74.63 μg/mL and L132: LC-5-0 = 75.36 μg/mL). Exposure to desorbed particles (MA: C1= 61.11 μg/mL and L132 : C2 = 61.71 μg/mL) and B[a]P (1 μM) were included. Bulky PAH-DNA adducts were detected in AM in mono-culture after exposure to total particles (Pt), to B[a]P and desorbed particles (Pd). Whatever the exposure, no DNA adduct was detected in L132 in mono-culture. These results are coherent with the enzymatic activities of cytochrome P450 l Al in AM and L132. Exposure of co-culture to Pt, or Pd induced bulky adducts to DNA in AM but not in L132. Exposure to B[a]P alone has altered the DNA of AM and L132, in co-culture. Exposure to Pt is closer to the environmental conditions, but conferred an exposure to amounts of genotoxic agents compared to studies using organic extracts. The formation of bulky DNA adducts was nevertheless observed in AM exposed to Pt, in mono- or co-culture, indicating that they were competent in terms of metabolic activation of PAHs. The

  16. Hyperkalaemia after radiofrequency ablation of hepatocellular carcinoma

    NARCIS (Netherlands)

    Verhoevena, BH; Haagsma, EB; Appeltans, BMG; Slooff, MJH; de Jong, KP

    Radiofrequency ablation of liver tumours is a useful therapy for otherwise unresectable tumours. The complication rate is said to be low. In this case report we describe hyperkalaemia after radiofrequency ablation of a hepatocellular carcinoma in a patient with end-stage renal insufficiency. (C)

  17. Radioembolisation for treatment of pediatric hepatocellular carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Hawkins, Clifford Matthew; Kukreja, Kamlesh [Cincinnati Children' s Hospital Medical Center, Department of Radiology, Cincinnati, OH (United States); Geller, James I. [Cincinnati Children' s Hospital Medical Center, Department of Hematology/Oncology, Cincinnati, OH (United States); Schatzman, Carmen; Ristagno, Ross [University of Cincinnati, UC Health, Department of Radiology, Division of Interventional Radiology, Cincinnati, OH (United States)

    2013-07-15

    Transarterial radioembolisation with yttrium-90 (TARE-Y90), a catheter-directed therapy, has been used extensively in adults to treat primary and secondary hepatic malignancies. To our knowledge, the use of this palliative technique has not been described in children. We present two children with unresectable hepatocellular carcinoma (HCC) treated with TARE-Y90. (orig.)

  18. Small hepatocellular carcinoma versus small cavernous hemangioma

    International Nuclear Information System (INIS)

    Choi, B.I.; Park, H.W.; Kim, S.H.; Han, M.C.; Kim, C.W.

    1989-01-01

    To determine the optimal pulse sequence for detection and differential diagnosis of small hepatocellular carcinomas and cavernous hemangiomas less than 5 cm in diameter, the authors have analyzed spin-echo (SE) images of 15 small hepatocellular carcinomas and 31 small cavernous hemangiomas obtained at 2.0 T. Pulse sequences used included repetition times (TRs) of 500 and 2,000 msec and echo times (TEs) of 30,60,90,120,150, and 180 msec. Mean tumor-liver contrast-to-noise ratios on the SE 2,000/60 (TR msec/TE msec) sequence were 23.90 ± 16.33 and 62.10 ± 25.94 for small hepatocellular carcinomas and hemangiomas, respectively, and were significantly greater than for all other pulse sequences. Mean tumor-liver signal intensity ratios on the SE 2,000/150 sequence were 2.34 ± 1.72 and 6.04 ± 2.72 for small hepatocellular carcinomas and hemangiomas, respectively, and were significantly greater than for all other pulse sequences in hemangiomas

  19. Liver transplantation in patients with hepatocellular carcinoma

    NARCIS (Netherlands)

    Polak, Wojciech G.; Soyama, Akihiko; Slooff, Maarten J. H.

    2008-01-01

    Liver transplantation has a definitive place in the treatment of patients with hepatocellular carcinoma (HCC) in a cirrhotic liver. Patients with a tumor load within the Milan criteria have excellent survival comparable to survival in patients with benign indications. When tumor load exceeds the

  20. Delayed presentation and diagnosis of metastatic hepatocellular ...

    African Journals Online (AJOL)

    Hepatocellular carcinoma (HCC) is rarely diagnosed in women of reproductive age. ... an adverse effect on HCC, and that high levels of oestrogen associated with pregnancy may ... A 30-year-old pregnant woman presented at 23 weeks' gestation and was diagnosed as HIV-infected, with anaemia. She was initiated on.

  1. Hepatocellular carcinoma: risk groups, surveillance and outcome

    NARCIS (Netherlands)

    van Meer, S

    2016-01-01

    The burden of hepatocellular carcinoma (HCC) has changed in the past few decades. Although the majority of HCC cases develops in East Asia and Sub-Saharan Africa, HCC has become an increasing problem in Western countries such as the Netherlands. Surveillance for HCC is controversial because of

  2. Ecological effects of co-culturing sea cucumber Apostichopus japonicus (Selenka) with scallop Chlamys farreri in earthen ponds

    Science.gov (United States)

    Ren, Yichao; Dong, Shuanglin; Qin, Chuanxin; Wang, Fang; Tian, Xiangli; Gao, Qinfeng

    2012-01-01

    Monthly changes in sedimentation and sediment properties were studied for three different culture treatments: sea cucumber monoculture (Mc), sea cucumber and scallop polyculture (Ps-c) and scallop monoculture (Ms). Results indicated that the survival rate of sea cucumber was significantly higher in Ps-c cultures than in Mc cultures. Sea cucumber yield was 69.6% higher in Ps-c culture than in Mc culture. No significant differences in body weight and scallop shell length were found between Ps-c and Ms cultures. The mean sedimentation rate of total particulate matter (TPM) was 72.2 g/(m2·d) in Ps-c cultures, with a maximum of 119.7 g/(m2·d), which was markedly higher than that of Mc (mean value). Sedimentation rates of organic matter (OM), total organic carbon (TOC), total nitrogen (TN) and total phosphorus (TP) in Ps-c cultures were also significantly higher than those in Mc cultures. TOC and TN contents of sediment increased rapidly in the first 5 months in Ms cultures and remained at a high level. TOC and TN contents in Mc and Ps-c cultures decreased during sea cucumber feeding seasons and increased during sea cucumber dormancy periods (summer and winter). The study demonstrates that co-culture of sea cucumber and scallop in earthen ponds is an alternative way to alleviate nutrient loads and improve water quality in coastal aquaculture systems. Moreover, it provides the additional benefit of an increased sea cucumber yield.

  3. Mouse preantral follicle growth in 3D co-culture system using human menstrual blood mesenchymal stem cell.

    Science.gov (United States)

    Rajabi, Zahra; Yazdekhasti, Hossein; Noori Mugahi, Seyed Mohammad Hossein; Abbasi, Mehdi; Kazemnejad, Somaieh; Shirazi, Abolfazl; Majidi, Masoumeh; Zarnani, Amir-Hassan

    2018-03-01

    Follicle culture provides a condition which can help investigators to evaluate various aspects of ovarian follicle growth and development and impact of different components and supplementations as well as presumably application of follicle culture approach in fertility preservation procedures. Mesenchymal Stem Cells (MSCs), particularly those isolated from menstrual blood has the potential to be used as a tool for improvement of fertility. In the current study, a 3D co-culture system with mice preantral follicles and human Menstrual Blood Mesenchymal Stem Cells (MenSCs) using either collagen or alginate beads was designed to investigate whether this system allows better preantral follicles growth and development. Results showed that MenSCs increase the indices of follicular growth including survival rate, diameter, and antrum formation as well as the rate of in vitro maturation (IVM) in both collagen and alginates beads. Although statistically not significant, alginate was found to be superior in terms of supporting survival rate and antrum formation. Hormone assay demonstrated that the amount of secreted 17 β-estradiol and progesterone in both 3D systems increased dramatically after 12 days, with the highest levels in system employing MenSCs. Data also demonstrated that relative expression of studied genes increased for Bmp15 and Gdf9 and decreased for Mater when follicles were cultured in the presence of MenSCs. Collectively, results of the present study showed that MenSCs could improve indices of follicular growth and maturation in vitro. Further studies are needed before a clinical application of MenSCs-induced IVM is considered. Copyright © 2018 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. All rights reserved.

  4. Paired single cell co-culture microenvironments isolated by two-phase flow with continuous nutrient renewal.

    Science.gov (United States)

    Chen, Yu-Chih; Cheng, Yu-Heng; Kim, Hong Sun; Ingram, Patrick N; Nor, Jacques E; Yoon, Euisik

    2014-08-21

    Cancer-stromal cell interactions are a critical process in tumorigenesis. Conventional dish-based assays, which simply mix two cell types, have limitations in three aspects: 1) limited control of the cell microenvironment; 2) inability to study cell behavior in a single-cell manner; and 3) have difficulties in characterizing single cell behavior within a highly heterogeneous cell population (e.g. tumor). An innovative use of microfluidic technology is for improving the spatial resolution for single cell assays. However, it is challenging to isolate the paired interacting cells while maintaining nutrient renewal. In this work, two-phase flow was used as a simple isolation method, separating the microenvironment of each individual chamber. As nutrients in an isolated chamber are consumed by cells, media exchange is required. To connect the cell culture chamber to the media exchange layer, we demonstrated a 3D microsystem integration technique using vertical connections fabricated by deep reactive-ion etching (DRIE). Compared to previous approaches, the presented process allows area reduction of vertical connections by an order of magnitude, enabling compact 3D integration. A semi-permeable membrane was sandwiched between the cell culture layer and the media exchange layer. The selectivity of the semi-permeable membrane results in the retention of the signaling proteins within the chamber while allowing free diffusion of nutrients (e.g., glucose and amino acids). Thus, paracrine signals are accumulated inside the chamber without cross-talk between cells in other chambers. Utilizing these innovations, we co-cultured UM-SCC-1 (head and neck squamous cell carcinoma) cells and endothelial cells to simulate tumor proliferation enhancement in the vascular endothelial niche.

  5. Role of IFN-gamma and LPS on Neuron/Glial Co-Cultures Infected by Neospora caninum

    Directory of Open Access Journals (Sweden)

    Erica Etelvina Viana De Jesus

    2014-10-01

    Full Text Available Neospora caninum causes cattle abortion and neurological symptoms in dogs. Although infection is usually asymptomatic, classical neurological symptoms of neosporosis may be associated with encephalitis. This parasite can grow in brain endothelial cells without markedly damages, but it can modulate the cellular environment to promote its survival in the brain. In previous studies, we described that IFN-γ decreased the parasite proliferation and down regulated nitric oxide production in astrocyte/microglia cultures. However, it remains unclear how glial cells respond to N. caninum in the presence of neurons. Therefore, we evaluated the effect of 300 IU/mL IFN-γ or 1.0 μg/mL of LPS on infected rat neuron/glial co-cultures. After 72 hours of infection, LPS did not affect the mitochondrial dehydrogenase activity. However, IFN-γ decreased this parameter by 15.5 and 12.0% in uninfected and infected cells, respectively. The number of tachyzoites decreased 54.1 and 44.3% in cells stimulated with IFN-γ and LPS, respectively. Infection or LPS treatment did not change NO production. On the other hand, IFN-γ induced increased nitrite release in 55.7%, but the infection reverted this induction. IL-10 levels increased only in infected cultures (treated or not, meanwhile PGE2 release was improved in IFN-γ/infected or LPS/infected cells. Although IFN-γ significantly reduced the neurite length in uninfected cultures (42.64%; p < 0.001, this inflammatory cytokine reverted the impairment of neurite outgrowth induced by the infection (81.39%. The results suggest a neuroprotective potential response of glia to N. caninum infection under IFN-γ stimulus. This observation contributes to understand the immune mediated mechanisms of neosporosis in CNS

  6. Differential Effect of Medium on the Ratio of ICM/TE of Bovine Embryos in a Co-culture System

    Directory of Open Access Journals (Sweden)

    Mohsen Forouzanfar

    2010-01-01

    Full Text Available Background: This study was undertaken to investigate the efficiency of two differentembryo somatic cell co-culture conditions, tissue culture medium 199 (TCM199–vero cellsand Menezo B2 (B2-vero cells, for the in vitro developmental quantity and quality of bovineembryos.Materials and Methods: Bovine oocytes were allowed to mature and subsequently undergofertilization in vitro. Their presumptive zygotes were cultured in either TCM199 or B2 culturemedia in conjunction with vero cells for up to nine days. The culture media were refreshedevery two days and the proportion of embryos that cleaved and further developed to themorula and blastocyst (early, expand and hatched stages were recorded. Hatched blastocystsunderwent differential staining in order to determine the numbers of inner cell mass (ICMand tropho ectoderm (TE and total cell number (TCN.Results: Of the two groups, no significant difference was seen between the proportions ofthe presumptive zygotes cleaved, those which developed to 8-16 cells, morula and reacheddays 7or 8 blastocyst stage or hatched. However, the values for TCN and TE of the TCM199-vero embryos were significantly greater than those of B2-vero embryos. The values for ICM/TCN and ICM/TE were significantly greater in the B2-vero group versus the TCM199-verogroup.Conclusion: Both TCM199 and B2 culture media in conjunction with vero cells were ofthe same efficiency when used for in vitro development of bovine presumptive zygotes.However, TCM199 was superior in providing embryos with more embryo cell numbers,whereas B2 medium was superior in providing embryos with greater ICM/TE and ICM/TCN ratios.

  7. In vitro gastrointestinal digestion increases the translocation of polystyrene nanoparticles in an in vitro intestinal co-culture model.

    Science.gov (United States)

    Walczak, Agata P; Kramer, Evelien; Hendriksen, Peter J M; Helsdingen, Richard; van der Zande, Meike; Rietjens, Ivonne M C M; Bouwmeester, Hans

    2015-01-01

    The conditions of the gastrointestinal tract may change the physicochemical properties of nanoparticles (NPs) and therewith the bioavailability of orally taken NPs. Therefore, we assessed the impact of in vitro gastrointestinal digestion on the protein corona of polystyrene NPs (PS-NPs) and their subsequent translocation across an in vitro intestinal barrier. A co-culture of intestinal Caco-2 and HT29-MTX cells was exposed to 50 nm PS-NPs of different charges (positive and negative) in two forms: pristine and digested in an in vitro gastrointestinal digestion model. In vitro digestion significantly increased the translocation of all, except the "neutral", PS-NPs. Upon in vitro digestion, translocation was 4-fold higher for positively charged NPs and 80- and 1.7-fold higher for two types of negatively charged NPs. Digestion significantly reduced the amount of protein in the corona of three out of four types of NPs. This reduction of proteins was 4.8-fold for "neutral", 3.5-fold for positively charged and 1.8-fold for one type of negatively charged PS-NPs. In vitro digestion also affected the composition of the protein corona of PS-NPs by decreasing the presence of higher molecular weight proteins and shifting the protein content of the corona to low molecular weight proteins. These findings are the first to report that in vitro gastrointestinal digestion significantly affects the protein corona and significantly increases the in vitro translocation of differently charged PS-NPs. These findings stress the importance of including the in vitro digestion in future in vitro intestinal translocation screening studies for risk assessment of orally taken NPs.

  8. Particles from wood smoke and traffic induce differential pro-inflammatory response patterns in co-cultures

    International Nuclear Information System (INIS)

    Kocbach, Anette; Herseth, Jan Inge; Lag, Marit; Refsnes, Magne; Schwarze, Per E.

    2008-01-01

    The inflammatory potential of particles from wood smoke and traffic has not been well elucidated. In this study, a contact co-culture of monocytes and pneumocytes was exposed to 10-40 μg/cm 2 of particles from wood smoke and traffic for 12, 40 and 64 h to determine their influence on pro-inflammatory cytokine release (TNF-α, IL-1, IL-6, IL-8) and viability. To investigate the role of organic constituents in cytokine release the response to particles, their organic extracts and the washed particles were compared. Antagonists were used to investigate source-dependent differences in intercellular signalling (TNF-α, IL-1). The cytotoxicity was low after exposure to particles from both sources. However, wood smoke, and to a lesser degree traffic-derived particles, induced a reduction in cell number, which was associated with the organic fraction. The release of pro-inflammatory cytokines was similar for both sources after 12 h, but traffic induced a greater release than wood smoke particles with increasing exposure time. The organic fraction accounted for the majority of the cytokine release induced by wood smoke, whereas the washed traffic particles induced a stronger response than the corresponding organic extract. TNF-α and IL-1 antagonists reduced the release of IL-8 induced by particles from both sources. In contrast, the IL-6 release was only reduced by the IL-1 antagonist during exposure to traffic-derived particles. In summary, particles from wood smoke and traffic induced differential pro-inflammatory response patterns with respect to cytokine release and cell number. Moreover, the influence of the organic particle fraction and intercellular signalling on the pro-inflammatory response seemed to be source-dependent

  9. Syntrophomonas zehnderi sp. nov., an anaerobe that degrades long-chain fatty acids in co-culture with Methanobacterium formicicum.

    Science.gov (United States)

    Sousa, Diana Z; Smidt, Hauke; Alves, M Madalena; Stams, Alfons J M

    2007-03-01

    An anaerobic, mesophilic, syntrophic fatty-acid-oxidizing bacterium, designated strain OL-4(T), was isolated as a co-culture with Methanobacterium formicicum DSM 1535(NT) from an anaerobic expanded granular sludge bed reactor used to treat an oleate-based effluent. Strain OL-4(T) degraded oleate, a mono-unsaturated fatty acid, and straight-chain fatty acids C(4 : 0)-C(18 : 0) in syntrophic association with Methanobacterium formicicum DSM 1535(NT). Even-numbered fatty acids were degraded to acetate and methane whereas odd-numbered fatty acids were degraded to acetate, propionate and methane. Branched-chain fatty acids were not degraded. The bacterium could not grow axenically with any other substrate tested and therefore is considered to be obligately syntrophic. Fumarate, sulfate, thiosulfate, sulfur and nitrate could not serve as electron acceptors for strain OL-4(T) to degrade oleate or butyrate. Cells of strain OL-4(T) were curved rods, formed spores and showed a variable response to Gram staining. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain OL-4(T) was most closely related to the fatty-acid-oxidizing, syntrophic bacterium Syntrophomonas sp. TB-6 (95 % similarity), Syntrophomonas wolfei subsp. wolfei DSM 2245(T) (94 % similarity) and Syntrophomonas erecta DSM 16215(T) (93 % similarity). In addition to this moderate similarity, phenotypic and physiological characteristics, such as obligate syntrophy, spore formation and utilization of a broader substrate range, differentiated strain OL-4(T) from these Syntrophomonas species. Therefore strain OL-4(T) represents a novel species, for which the name Syntrophomonas zehnderi sp. nov. is proposed. The type strain is OL-4(T) (=DSM 17840(T)=JCM 13948(T)).

  10. Determination of volatile components in fermented soybean prepared by a co-culture of Bacillus subtilis and Rhizopus oligosporus

    Directory of Open Access Journals (Sweden)

    Chukeatirote, E.,

    2017-07-01

    Full Text Available Fermented soybeans are traditional foods widely consumed in many countries (i.e., Natto in Japan, Jang in Korea, Kinema in India, Douichi in China, and Thua Nao in Thailand. In this study, an attempt was made with an expectation to improve the fermentation process using a co-culture of Bacillus subtilis and Rhizopus oligosporus. Initially, the raw soybeans were washed, sterilized by autoclaving, and inoculated with two inocula; for this, three different ratios between B. subtilis and R. oligosporus used were 100:0, 50:50, and 0:100. The fermentation was then carried out at 30°C for 3 days. The volatile compounds in the non-fermented and the fermented soybean products were determined by gas chromatography/ mass spectrometry (GC/MS and extracted using a solid phase microextraction (SPME technique. In total, 165 compounds were identified in the non-fermented and the fermented soybean products. For the non-fermented products, the predominant volatile compounds were alcohols (25.81%, aldehydes (13.64%, acids and esters (7.57%, furans (6.13% and ketones (0.88%. In contrast, the major volatiles compounds presented in the fermented soybeans were as follows: i The treatment of 100:0 consisted of acids and esters (35.89%, alcohols (14.55%, aldehydes (8.72%, ketones (4.97%, pyrazines (4.87%, and furans (4.22%; ii 50:50 comprised of acids and esters (55.62%, alcohols (16.22%, aldehydes (7.80%, pyrazine (3.65%, ketones (2.55%, furans (1.67%, and aromatic compounds (1.46%; and iii 0:100 included acids and esters (66.50%, alcohols (15.44%, aldehydes (2.59%, ketones (2.72%, furans (1.89%, aromatic compounds (1.80%, pyrazines (1.35%, and sulphur containing compounds (0.24% respectively.

  11. Hepatocellular carcinoma: a clinico pathological study

    International Nuclear Information System (INIS)

    Abbasi, A.; Butt, N.; Bhutto, A.R.; Gulzar, K.; Munir, S.M.

    2010-01-01

    To describe the clinico-pathological and radiological profile of hepatocellular carcinoma. All consecutive patients suspected of having hepatocellular carcinoma (HCC), were admitted and included in this study. Diagnosis of HCC was established by clinical, biochemical, ultrasonographic and histopathologic findings. Patients with primary carcinoma elsewhere in the body, metastatic in the liver, fibrolamellar carcinoma and benign tumours were excluded from the study. At ultrasonography, the details of tumour size and number, portal vein thrombosis and presence of ascites were recorded. Patients were staged according to Okuda staging system. Results were described in mean and percentage values. There were 82 patients with hepatocellular carcinoma including 58 males and 24 females, with male to female ratio of 2.8:1. The mean age of patients was 56.24 +- 13.65 years. Right hypochondrial pain was the main symptom in 52 (63.4%) patients. The duration of symptoms varied from 1 month to 2 years. Tumour size was larger than 50% of liver size in 42 (51.2%) with portal vein thrombosis in 10 (12.19%). Anti HCV was positive in 44 (53.7%), HBsAg in 26 (31.7%) and both were found positive in 2 (2.44%) patients. Ten patients (12.2/%) found negative both for anti-HCV and HBsAg. According to Okuda staging system 18 patients had stage 1, 50 had stage 2 and 14 had stage 3 hepatocellular carcinoma. The mean age of presentation of hepatocellular carcinoma was younger as compared to western countries with potentially large non-resectable lesions. Chronic hepatitis C and B was found to be the major known factors. Patients with chronic hepatitis C and B should undergo vigorous HCC surveillance to detect early, potentially respectable HCC. (author)

  12. In vitro co-cultures of human gut bacterial species as predicted from co-occurrence network analysis

    DEFF Research Database (Denmark)

    Das, Promi; Ji, Boyang; Kovatcheva-Datchary, Petia

    2018-01-01

    Network analysis of large metagenomic datasets generated by current sequencing technologies can reveal significant co-occurrence patterns between microbial species of a biological community. These patterns can be analyzed in terms of pairwise combinations between all species comprising a community...... thetaiotaomicron, as well as Faecalibacterium prausnitzii and Roseburia inulinivorans as model organisms for our study. We then delineate the outcome of the co-cultures when equal distributions of resources were provided. The growth behavior of the co-culture was found to be dependent on the types of microbial...... species present, their specific metabolic activities, and resulting changes in the culture environment. Through this reductionist approach and using novel in vitro combinations of microbial species under anaerobic conditions, the results of this work will aid in the understanding and design of synthetic...

  13. Protective Effects of Calcium on Cadmium Accumulation in Co-Cultured Silver Carp (Hypophthalmichthys molitrix) and Triangle Sail Mussel (Hyriopsis cumingii).

    Science.gov (United States)

    Li, Deliang; Pi, Jie; Wang, Jianping; Zhu, Pengfei; Lei, Liuping; Zhang, Ting; Liu, Deming

    2016-12-01

    Discovering cost effective strategies to reduce cadmium (Cd) uptake is of great concern for consumer food safety in the aquaculture industry. This study investigated the protective effects of calcium (Ca) on Cd uptake in co-cultured silver carp (Hypophthalmichthys molitrix) and triangle sail mussel (Hyriopsis cumingii). The results show that Ca-depending on its applied concentration-caused a significant decrease in the Cd uptake into muscle (by 48 %-72 %), gills (by 51 %-57 %), liver (by 52 %-81 %) and kidney (by 54 %-81 %) of silver carp (p sail mussels (p sail mussel, the quality of co-cultured edible fish might improve as a consequence of the potentially reduced Cd uptake.

  14. Production of rhamnolipids by Pseudomonas aeruginosa is inhibited by H2S but resumes in a co-culture with P. stutzeri: applications for microbial enhanced oil recovery.

    Science.gov (United States)

    Zhao, Feng; Ma, Fang; Shi, Rongjiu; Zhang, Jie; Han, Siqin; Zhang, Ying

    2015-09-01

    Sulfate-reducing bacteria and H2S exist widely in oil production systems, and in situ production of rhamnolipids is promising for microbial enhanced oil recovery (MEOR). However, information of the effect of S(2-) on rhamnolipids production is scarce. Two facultative anaerobic rhamnolipids-producing bacterial strains, Pseudomonas aeruginosa SG and WJ-1, were used. Above 10 mg S(2-)/l, both cell growth and rhamnolipids production were inhibited. A large inoculum (9%, v/v) failed to completely relieve the inhibitory effect of 10 mg S(2-)/l. Below 30 mg S(2-)/l, both strains resumed rhamnolipid production through co-culturing with the denitrifying and sulphide-removing strain Pseudomonas stutzeri DQ1. H2S has a direct but reversible inhibitory effect on rhamnolipids production. Control of H2S in oilfields is indispensable to MEOR, and the co-culture method is effective in restoring rhamnolipid production in presence of S(2-).

  15. File list: Unc.Liv.05.AllAg.Carcinoma,_Hepatocellular [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available Unc.Liv.05.AllAg.Carcinoma,_Hepatocellular mm9 Unclassified Liver Carcinoma, Hepato...cellular http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/Unc.Liv.05.AllAg.Carcinoma,_Hepatocellular.bed ...

  16. File list: His.Liv.10.AllAg.Carcinoma,_Hepatocellular [Chip-atlas[Archive

    Lifescience Database Archive (English)

    Full Text Available His.Liv.10.AllAg.Carcinoma,_Hepatocellular mm9 Histone Liver Carcinoma, Hepatocellu...lar http://dbarchive.biosciencedbc.jp/kyushu-u/mm9/assembled/His.Liv.10.AllAg.Carcinoma,_Hepatocellular.bed ...

  17. Co-culture of chondrocytes and bone marrow mesenchymal stem cells in vitro enhances the expression of cartilaginous extracellular matrix components

    Directory of Open Access Journals (Sweden)

    Chang Qing

    2011-04-01

    Full Text Available Chondrocytes and bone marrow mesenchymal stem cells (BMSCs are frequently used as seed cells in cartilage tissue engineering. In the present study, we determined if the co-culture of rabbit articular chondrocytes and BMSCs in vitro promotes the expression of cartilaginous extracellular matrix and, if so, what is the optimal ratio of the two cell types. Cultures of rabbit articular chondrocytes and BMSCs were expanded in vitro and then cultured individually or at a chondrocyte:BMSC ratio of 4:1, 2:1, 1:1, 1:2, 1:4 for 21 days and cultured in DMEM/F12. BMSCs were cultured in chondrogenic induction medium. Quantitative real-time RT-PCR and Western blot were used to evaluate gene expression. In the co-cultures, type II collagen and aggrecan expression increased on days 14 and 21. At the mRNA level, the expression of type II collagen and aggrecan on day 21 was much higher in the 4:1, 2:1, and 1:1 groups than in either the articular chondrocyte group or the induced BMSC group, and the best ratio of co-culture groups seems to be 2:1. Also on day 21, the expression of type II collagen and aggrecan proteins in the 2:1 group was much higher than in all other groups. The results demonstrate that the co-culture of rabbit chondrocytes and rabbit BMSCs at defined ratios can promote the expression of cartilaginous extracellular matrix. The optimal cell ratio appears to be 2:1 (chondrocytes:BMSCs. This approach has potential applications in cartilage tissue engineering since it provides a protocol for maintaining and promoting seed-cell differentiation and function.

  18. Co-culture of chondrocytes and bone marrow mesenchymal stem cells in vitro enhances the expression of cartilaginous extracellular matrix components.

    Science.gov (United States)

    Qing, Chang; Wei-ding, Cui; Wei-min, Fan

    2011-04-01

    Chondrocytes and bone marrow mesenchymal stem cells (BMSCs) are frequently used as seed cells in cartilage tissue engineering. In the present study, we determined if the co-culture of rabbit articular chondrocytes and BMSCs in vitro promotes the expression of cartilaginous extracellular matrix and, if so, what is the optimal ratio of the two cell types. Cultures of rabbit articular chondrocytes and BMSCs were expanded in vitro and then cultured individually or at a chondrocyte:BMSC ratio of 4:1, 2:1, 1:1, 1:2, 1:4 for 21 days and cultured in DMEM/F12. BMSCs were cultured in chondrogenic induction medium. Quantitative real-time RT-PCR and Western blot were used to evaluate gene expression. In the co-cultures, type II collagen and aggrecan expression increased on days 14 and 21. At the mRNA level, the expression of type II collagen and aggrecan on day 21 was much higher in the 4:1, 2:1, and 1:1 groups than in either the articular chondrocyte group or the induced BMSC group, and the best ratio of co-culture groups seems to be 2:1. Also on day 21, the expression of type II collagen and aggrecan proteins in the 2:1 group was much higher than in all other groups. The results demonstrate that the co-culture of rabbit chondrocytes and rabbit BMSCs at defined ratios can promote the expression of cartilaginous extracellular matrix. The optimal cell ratio appears to be 2:1 (chondrocytes:BMSCs). This approach has potential applications in cartilage tissue engineering since it provides a protocol for maintaining and promoting seed-cell differentiation and function.

  19. A Triple Co-Culture Model of the Human Respiratory Tract to Study Immune-Modulatory Effects of Liposomes and Virosomes.

    Directory of Open Access Journals (Sweden)

    Rebecca A M Blom

    Full Text Available The respiratory tract with its ease of access, vast surface area and dense network of antigen-presenting cells (APCs represents an ideal target for immune-modulation. Bio-mimetic nanocarriers such as virosomes may provide immunomodulatory properties to treat diseases such as allergic asthma. In our study we employed a triple co-culture model of epithelial cells, macrophages and dendritic cells to simulate the human airway barrier. The epithelial cell line 16HBE was grown on inserts and supplemented with human blood monocyte-derived macrophages (MDMs and dendritic cells (MDDCs for exposure to influenza virosomes and liposomes. Additionally, primary human nasal epithelial cells (PHNEC and EpCAM+ epithelial progenitor cell mono-cultures were utilized to simulate epithelium from large and smaller airways, respectively. To assess particle uptake and phenotype change, cell cultures were analyzed by flow cytometry and pro-inflammatory cytokine concentrations were measured by ELISA. All cell types internalized virosomes more efficiently than liposomes in both mono- and co-cultures. APCs like MDMs and MDDCs showed the highest uptake capacity. Virosome and liposome treatment caused a moderate degree of activation in MDDCs from mono-cultures and induced an increased cytokine production in co-cultures. In epithelial cells, virosome uptake was increased compared to liposomes in both mono- and co-cultures with EpCAM+ epithelial progenitor cells showing highest uptake capacity. In conclusion, all cell types successfully internalized both nanocarriers with virosomes being taken up by a higher proportion of cells and at a higher rate inducing limited activation of MDDCs. Thus virosomes may represent ideal carrier antigen systems to modulate mucosal immune responses in the respiratory tract without causing excessive inflammatory changes.

  20. Cultures and co-cultures of human blood mononuclear cells and endothelial cells for the biocompatibility assessment of surface modified AISI 316L austenitic stainless steel

    International Nuclear Information System (INIS)

    Stio, Maria; Martinesi, Maria; Treves, Cristina; Borgioli, Francesca

    2016-01-01

    Samples of AISI 316L austenitic stainless steel were subjected either to grinding and polishing procedure, or to grinding and then low temperature glow-discharge nitriding treatment, or to grinding, nitriding and subsequently coating with collagen-I. Nitrided samples, even if only ground, show a higher corrosion resistance in PBS solution, in comparison with ground and polished AISI 316L. Biocompatibility was evaluated in vitro by incubating the samples with either peripheral blood mononuclear cells (PBMC) or human umbilical vein endothelial cells (HUVEC), tested separately or in co-culture. HUVEC-PBMC co-culture and co-incubation of HUVEC with PBMC culture medium, after the previous incubation of PBMC with metallic samples, allowed to determine whether the incubation of PBMC with the different samples might affect HUVEC behaviour. Many biological parameters were considered: cell proliferation, release of cytokines, matrix metalloproteinases (MMPs) and sICAM-1, gelatinolytic activity of MMPs, and ICAM-1 protein expression. Nitriding treatment, with or without collagen coating of the samples, is able to ameliorate some of the biological parameters taken into account. The obtained results point out that biocompatibility may be successfully tested in vitro, using cultures of normal human cells, as blood and endothelial cells, but more than one cell line should be used, separately or in co-culture, and different parameters should be determined, in particular those correlated with inflammatory phenomena. - Highlights: • Nitriding improves corrosion resistance and biocompatibility of ground AISI 316L. • The metallic samples differently affect different human cell cultures. • PBMC and HUVEC are a suitable model to test in vitro biocompatibility. • Co-cultures show that HUVEC are affected by pre-incubation of PBMC with the samples. • Inflammation parameters must be taken into account for assessing biocompatibility.

  1. Promotion of Vascular Morphogenesis of Endothelial Cells Co-Cultured with Human Adipose-Derived Mesenchymal Stem Cells Using Polycaprolactone/Gelatin Nanofibrous Scaffolds

    Directory of Open Access Journals (Sweden)

    Yun-Min Kook

    2018-02-01

    Full Text Available New blood vessel formation is essential for tissue regeneration to deliver oxygen and nutrients and to maintain tissue metabolism. In the field of tissue engineering, in vitro fabrication of new artificial vessels has been a longstanding challenge. Here we developed a technique to reconstruct a microvascular system using a polycaprolactone (PCL/gelatin nanofibrous structure and a co-culture system. Using a simple electrospinning process, we fabricated three-dimensional mesh scaffolds to support the sprouting of human umbilical vein endothelial cells (HUVECs along the electrospun nanofiber. The co-culture with adipose-derived mesenchymal stem cells (ADSCs supported greater sprouting of endothelial cells (ECs. In a two-dimensional culture system, angiogenic cell assembly produced more effective direct intercellular interactions and paracrine signaling from ADSCs to assist in the vascular formation of ECs, compared to the influence of growth factor. Although vascular endothelial growth factor and sphingosine-1-phosphate were present during the culture period, the presence of ADSCs was the most important factor for the construction of a cell-assembled structure in the two-dimensional culture system. On the contrary, HUVECs co-cultured on PCL/gelatin nanofiber scaffolds produced mature and functional microvessel and luminal structures with a greater expression of vascular markers, including platelet endothelial cell adhesion molecule-1 and podocalyxin. Furthermore, both angiogenic factors and cellular interactions with ADSCs through direct contact and paracrine molecules contributed to the formation of enhanced engineered blood vessel structures. It is expected that the co-culture system of HUVECs and ADSCs on bioengineered PCL/gelatin nanofibrous scaffolds will promote robust and functional microvessel structures and will be valuable for the regeneration of tissue with restored blood vessels.

  2. Cultures and co-cultures of human blood mononuclear cells and endothelial cells for the biocompatibility assessment of surface modified AISI 316L austenitic stainless steel

    Energy Technology Data Exchange (ETDEWEB)

    Stio, Maria; Martinesi, Maria; Treves, Cristina [Dipartimento di Scienze Biomediche, Sperimentali e Cliniche ‘Mario Serio’, Sezione di Scienze Biochimiche, Università di Firenze, viale Morgagni 50, 50134 Firenze (Italy); Borgioli, Francesca, E-mail: francesca.borgioli@unifi.it [Dipartimento di Ingegneria Industriale (DIEF), Università di Firenze, via S. Marta 3, 50139 Firenze (Italy)

    2016-12-01

    Samples of AISI 316L austenitic stainless steel were subjected either to grinding and polishing procedure, or to grinding and then low temperature glow-discharge nitriding treatment, or to grinding, nitriding and subsequently coating with collagen-I. Nitrided samples, even if only ground, show a higher corrosion resistance in PBS solution, in comparison with ground and polished AISI 316L. Biocompatibility was evaluated in vitro by incubating the samples with either peripheral blood mononuclear cells (PBMC) or human umbilical vein endothelial cells (HUVEC), tested separately or in co-culture. HUVEC-PBMC co-culture and co-incubation of HUVEC with PBMC culture medium, after the previous incubation of PBMC with metallic samples, allowed to determine whether the incubation of PBMC with the different samples might affect HUVEC behaviour. Many biological parameters were considered: cell proliferation, release of cytokines, matrix metalloproteinases (MMPs) and sICAM-1, gelatinolytic activity of MMPs, and ICAM-1 protein expression. Nitriding treatment, with or without collagen coating of the samples, is able to ameliorate some of the biological parameters taken into account. The obtained results point out that biocompatibility may be successfully tested in vitro, using cultures of normal human cells, as blood and endothelial cells, but more than one cell line should be used, separately or in co-culture, and different parameters should be determined, in particular those correlated with inflammatory phenomena. - Highlights: • Nitriding improves corrosion resistance and biocompatibility of ground AISI 316L. • The metallic samples differently affect different human cell cultures. • PBMC and HUVEC are a suitable model to test in vitro biocompatibility. • Co-cultures show that HUVEC are affected by pre-incubation of PBMC with the samples. • Inflammation parameters must be taken into account for assessing biocompatibility.

  3. Chip-based human liver-intestine and liver-skin co-cultures--A first step toward systemic repeated dose substance testing in vitro.

    Science.gov (United States)

    Maschmeyer, Ilka; Hasenberg, Tobias; Jaenicke, Annika; Lindner, Marcus; Lorenz, Alexandra Katharina; Zech, Julie; Garbe, Leif-Alexander; Sonntag, Frank; Hayden, Patrick; Ayehunie, Seyoum; Lauster, Roland; Marx, Uwe; Materne, Eva-Maria

    2015-09-01

    Systemic repeated dose safety assessment and systemic efficacy evaluation of substances are currently carried out on laboratory animals and in humans due to the lack of predictive alternatives. Relevant international regulations, such as OECD and ICH guidelines, demand long-term testing and oral, dermal, inhalation, and systemic exposure routes for such evaluations. So-called "human-on-a-chip" concepts are aiming to replace respective animals and humans in substance evaluation with miniaturized functional human organisms. The major technical hurdle toward success in this field is the life-like combination of human barrier organ models, such as intestine, lung or skin, with parenchymal organ equivalents, such as liver, at the smallest biologically acceptable scale. Here, we report on a reproducible homeostatic long-term co-culture of human liver equivalents with either a reconstructed human intestinal barrier model or a human skin biopsy applying a microphysiological system. We used a multi-organ chip (MOC) platform, which provides pulsatile fluid flow within physiological ranges at low media-to-tissue ratios. The MOC supports submerse cultivation of an intact intestinal barrier model and an air-liquid interface for the skin model during their co-culture with the liver equivalents respectively at (1)/100.000 the scale of their human counterparts in vivo. To increase the degree of organismal emulation, microfluidic channels of the liver-skin co-culture could be successfully covered with human endothelial cells, thus mimicking human vasculature, for the first time. Finally, exposure routes emulating oral and systemic administration in humans have been qualified by applying a repeated dose administration of a model substance - troglitazone - to the chip-based co-cultures. Copyright © 2015. Published by Elsevier B.V.

  4. The effect of tobacco smoke exposure on the generation of reactive oxygen species and cellular membrane damage using co-culture model of blood brain barrier with astrocytes.

    Science.gov (United States)

    Seo, Seung-Beom; Choe, Eun Sang; Kim, Kwang-Sik; Shim, Soon-Mi

    2017-06-01

    Brain tissue is known to be vulnerable to the exposure by tobacco smoke. Tobacco smoke can induce generation of reactive oxygen species (ROS), causing inflammatory activity and blood-brain barrier (BBB) impairment. The aim of the present study was to investigate the effect of tobacco smoke on cell cytotoxicity, generation of ROS, and cellular membrane damage in astrocytes and BBB using a co-culture system. Cell viability of U373MG cells was reduced in a dose-dependent manner, ranging from 96.7% to 40.3% by tobacco smoke condensate (TSC). Cell viability of U373MG co-cultured with human brain microvascular endothelial cells (HBMECs) was 104.9% at the IC 50 value of TSC. Trans-epithelial electric resistance values drastically decreased 80% following 12-h incubation. The value was maintained until 48 h and then increased at 72-h incubation (85%). It then decreased to 75% at 120 h. Generation of ROS increased in a dose-dependent manner, ranging from 102.7% to 107.9%, when various concentrations of TSC (4-16 mg/mL) were administered to the U373MG monoculture. When TSC was added into U373MG co-cultured with HBMECs, production of ROS ranged from 101.7% to 102.6%, slightly increasing over 12 h. Maximum exposure-generated ROS of 104.8% was reached at 24 h. Cell cytotoxicity and oxidative stress levels in the U373MG co-culture model system with HBMECs were lower than U373MG monoculture. HBMECs effectively acted as a barrier to protect the astrocytes (U373MG) from toxicity of TSC.

  5. A study of a three-dimensional PLGA sponge containing natural polymers co-cultured with endothelial and mesenchymal stem cells as a tissue engineering scaffold

    International Nuclear Information System (INIS)

    Shim, Jung Bo; Kim, Hyeongseok; Khang, Gilson; Ankeny, Randall F; Nerem, Robert M

    2014-01-01

    The interaction between vascular endothelial cells (ECs) and vascular smooth muscle cells (VSMCs) in a complex hemodynamic and mechanical environment plays an important role in the control of blood vessel growth and function. Despite the importance of VSMCs, substitutes are needed for vascular therapies. A potential VSMC substitute is human adult bone marrow derived mesenchymal stem cells (hMSCs). In this study, the effect of poly(lactic-co-glycolic acid) (PLGA) scaffolds containing three natural polymers (demineralized bone particles, silk, and small intestine submucosa) on the phenotype of MSCs and SMCs cultured with or without ECs was investigated. The study objective was to create a media equivalent for a tissue engineered blood vessel using PLGA, natural polymers, and MSCs co-cultured with ECs. The PLGA containing the natural polymers silk and SIS showed increased proliferation and cell adhesion. The presence of silk and DBP promoted a MSC phenotype change into a SMC-like phenotype at the mRNA level; however these differences at the protein level were not seen. Additionally, PLGA containing SIS did not induce SMC gene or protein upregulation. Finally, the effect of ECs in combination with the natural polymers was tested. When co-cultured with ECs, the mRNA of SMC specific markers in MSCs and SMCs were increased when compared to SMCs or MSCs alone. However, MSCs, when co-cultured with ECs on PLGA containing silk, exhibited significantly increased α-SMA and calponin expression when compared to PLGA only scaffolds. These results indicate that the natural polymer silk in combination with the co-culture of endothelial cells was most effective at increasing cell viability and inducing a SMC-like phenotype at the mRNA and protein level in MSCs. (paper)

  6. , , , , , and Gene Expression in Single- and Co-cultured Bovine Satellite Cells and Intramuscular Preadipocytes Treated with Palmitic, Stearic, Oleic, and Linoleic Acid

    Directory of Open Access Journals (Sweden)

    S. H. Choi

    2015-03-01

    Full Text Available We previously demonstrated that bovine subcutaneous preadipocytes promote adipogenic gene expression in muscle satellite cells in a co-culture system. Herein we hypothesize that saturated fatty acids would promote adipogenic/lipogenic gene expression, whereas mono- and polyunsaturated fatty acids would have the opposite effect. Bovine semimembranosus satellite cells (BSC and intramuscular preadipocytes (IPA were isolated from crossbred steers and cultured with 10% fetal bovine serum (FBS/Dulbecco’s Modified Eagle Medium (DMEM and 1% antibiotics during the 3-d proliferation period. After proliferation, cells were treated for 3 d with 3% horse serum/DMEM (BSC or 5% FBS/DMEM (IPA with antibiotics. Media also contained 10 μg/mL insulin and 10 μg/mL pioglitazone. Subsequently, differentiating BSC and IPA were cultured in their respective media with 40 μM palmitic, stearic, oleic, or linoleic acid for 4 d. Finally, BSC and IPA were single- or co-cultured for an additional 2 h. All fatty acid treatments increased (p = 0.001 carnitine palmitoyltransferase-1 beta (CPT1β gene expression, but the increase in CPT1β gene expression was especially pronounced in IPA incubated with palmitic and stearic acid (6- to 17- fold increases. Oleic and linoleic acid decreased (p = 0.001 stearoyl-CoA desaturase (SCD gene expression over 80% in both BSC and IPA. Conversely, palmitic and stearic acid increased SCD gene expression three fold in co-cultured in IPA, and stearic acid increased AMPKα gene expression in single- and co-cultured BSC and IPA. Consistent with our hypothesis, saturated fatty acids, especially stearic acid, promoted adipogenic and lipogenic gene expression, whereas unsaturated fatty acids decreased expression of those genes associated with fatty acid metabolism.

  7. Differences between co-cultures and monocultures in testing the toxicity of particulate matter derived from log wood and pellet combustion

    Science.gov (United States)

    Happo, Mikko S.; Rönkkö, Teemu J.; Orasche, Jürgen; Jokiniemi, Jorma; Kortelainen, Miika; Tissari, Jarkko; Zimmermann, Ralf; Hirvonen, Maija-Riitta; Jalava, Pasi I.

    2018-01-01

    Background In vitro studies with monocultures of human alveolar cells shed deeper knowledge on the cellular mechanisms by which particulate matter (PM) causes toxicity, but cannot account for mitigating or aggravating effects of cell-cell interactions on PM toxicity. Methods We assessed inflammation, oxidative stress as well as cytotoxic and genotoxic effects induced by PM from the combustion of different types of wood logs and softwood pellets in three cell culture setups: two monocultures of either human macrophage-like cells or human alveolar epithelial cells, and a co-culture of these two cell lines. The adverse effects of the PM samples were compared between these setups. Results We detected clear differences in the endpoints between the mono- and co-cultures. Inflammatory responses were more diverse in the macrophage monoculture and the co-culture compared to the epithelial cells where only an increase of IL-8 was detected. The production of reactive oxygen species was the highest in epithelial cells and macrophages seemed to have protective effects against oxidative stress from the PM samples. With no metabolically active cells at the highest doses, the cytotoxic effects of the PM samples from the wood log combustion were far more pronounced in the macrophages and the co-culture than in the epithelial cells. All samples caused DNA damage in macrophages, whereas only beech and spruce log combustion samples caused DNA damage in epithelial cells. The organic content of the samples was mainly associated with cytotoxicity and DNA damage, while the metal content of the samples correlated with the induction of inflammatory responses. Conclusions All of the tested PM samples induce adverse effects and the chemical composition of the samples determines which pathway of toxicity is induced. In vitro testing of the toxicity of combustion-derived PM in monocultures of one cell line, however, is inadequate to account for all the possible pathways of toxicity. PMID

  8. Quantifying rates of cell migration and cell proliferation in co-culture barrier assays reveals how skin and melanoma cells interact during melanoma spreading and invasion.

    Science.gov (United States)

    Haridas, Parvathi; Penington, Catherine J; McGovern, Jacqui A; McElwain, D L Sean; Simpson, Matthew J

    2017-06-21

    Malignant spreading involves the migration of cancer cells amongst other native cell types. For example, in vivo melanoma invasion involves individual melanoma cells migrating through native skin, which is composed of several distinct subpopulations of cells. Here, we aim to quantify how interactions between melanoma and fibroblast cells affect the collective spreading of a heterogeneous population of these cells in vitro. We perform a suite of circular barrier assays that includes: (i) monoculture assays with fibroblast cells; (ii) monoculture assays with SK-MEL-28 melanoma cells; and (iii) a series of co-culture assays initiated with three different ratios of SK-MEL-28 melanoma cells and fibroblast cells. Using immunostaining, detailed cell density histograms are constructed to illustrate how the two subpopulations of cells are spatially arranged within the spreading heterogeneous population. Calibrating the solution of a continuum partial differential equation to the experimental results from the monoculture assays allows us to estimate the cell diffusivity and the cell proliferation rate for the melanoma and the fibroblast cells, separately. Using the parameter estimates from the monoculture assays, we then make a prediction of the spatial spreading in the co-culture assays. Results show that the parameter estimates obtained from the monoculture assays lead to a reasonably accurate prediction of the spatial arrangement of the two subpopulations in the co-culture assays. Overall, the spatial pattern of spreading of the melanoma cells and the fibroblast cells is very similar in monoculture and co-culture conditions. Therefore, we find no clear evidence of any interactions other than cell-to-cell contact and crowding effects. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. Fabrication of micropatterned alginate-gelatin and k-carrageenan hydrogels of defined shapes using simple wax mould method as a platform for stem cell/induced Pluripotent Stem Cells (iPSC) culture.

    Science.gov (United States)

    Vignesh, S; Gopalakrishnan, Aswathi; M R, Poorna; Nair, Shantikumar V; Jayakumar, R; Mony, Ullas

    2018-06-01

    Micropatterning techniques involve soft lithography, which is laborious, expensive and restricted to a narrow spectrum of biomaterials. In this work we report, first time employment of patterned wax moulds for generation of micropatterned alginate-gelatin and κ-carrageenan (κ-CRG) hydrogel systems by a novel, simple and cost effective method. We generated and characterized uniform and reproducible micropatterned hydrogels of varying sizes and shapes such as square projections, square grooves, and circular grids and crisscrossed hillocks. The rheological analysis showed that κ-carrageenan hydrogels had higher gel strength when compared to alginate-gelatin hydrogels. Human Mesenchymal stem cells (hMSCs) and Human Induced Pluripotent Stem Cells (hiPSCs) were found to be cytocompatible with these hydrogels. This micropatterned hydrogel system may have potential application in tissue engineering and also in understanding the basic biology behind the stem cell/iPSC fate. Copyright © 2018 Elsevier B.V. All rights reserved.

  10. Fluoxetine and its active metabolite norfluoxetine disrupt estrogen synthesis in a co-culture model of the feto-placental unit.

    Science.gov (United States)

    Hudon Thibeault, Andrée-Anne; Laurent, Laetitia; Vo Duy, Sung; Sauvé, Sébastien; Caron, Patrick; Guillemette, Chantal; Sanderson, J Thomas; Vaillancourt, Cathy

    2017-02-15

    The effects of fluoxetine, one of the most prescribed selective serotonin-reuptake inhibitors (SSRIs) during pregnancy, and its active metabolite norfluoxetine were studied on placental aromatase (CYP19) and feto-placental steroidogenesis. Fluoxetine did not alter estrogen secretion in co-culture of fetal-like adrenocortical (H295R) and trophoblast-like (BeWo) cells used as a model of the feto-placental unit, although it induced CYP19 activity, apparently mediated by the serotonin (5-HT) 2A receptor/PKC signaling pathway. Norfluoxetine decreased estrogen secretion in the feto-placental co-culture and competitively inhibited catalytic CYP19 activity in BeWo cells. Decreased serotonin transporter (SERT) activity in the co-culture was comparable to 17β-estradiol treatment of BeWo cells. This work shows that the complex interaction of fluoxetine and norfluoxetine with placental estrogen production, involves 5-HT-dependent and -independent mechanisms. Considering the crucial role of estrogens during pregnancy, our results raise concern about the impact of SSRI treatment on placental function and fetal health. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  11. Enhanced Agarose and Xylan Degradation for Production of Polyhydroxyalkanoates by Co-Culture of Marine Bacterium, Saccharophagus degradans and Its Contaminant, Bacillus cereus

    Directory of Open Access Journals (Sweden)

    Shailesh S. Sawant

    2017-02-01

    Full Text Available Over reliance on energy or petroleum products has raised concerns both in regards to the depletion of their associated natural resources as well as their increasing costs. Bioplastics derived from microbes are emerging as promising alternatives to fossil fuel derived petroleum plastics. The development of a simple and eco-friendly strategy for bioplastic production with high productivity and yield, which is produced in a cost effective manner utilising abundantly available renewable carbon sources, would have the potential to result in an inexhaustible global energy source. Here we report the biosynthesis of bioplastic polyhydroxyalkanoates (PHAs in pure cultures of marine bacterium, Saccharophagus degradans 2-40 (Sde 2-40, its contaminant, Bacillus cereus, and a co-culture of these bacteria (Sde 2-40 and B. cereus degrading plant and algae derived complex polysaccharides. Sde 2-40 degraded the complex polysaccharides agarose and xylan as sole carbon sources for biosynthesis of PHAs. The ability of Sde 2-40 to degrade agarose increased after co-culturing with B. cereus. The association of Sde 2-40 with B. cereus resulted in increased cell growth and higher PHA production (34.5% of dry cell weight from xylan as a carbon source in comparison to Sde 2-40 alone (22.7% of dry cell weight. The present study offers an innovative prototype for production of PHA through consolidated bioprocessing of complex carbon sources by pure and co-culture of microorganisms.

  12. Biological responses of T cells encapsulated with polyelectrolyte-coated gold nanorods and their cellular activities in a co-culture system

    Science.gov (United States)

    Wattanakull, Porntida; Killingsworth, Murray C.; Pissuwan, Dakrong

    2017-11-01

    Currently, human T cell therapy is of considerable scientific interest. In addition, cell encapsulation has become an attractive approach in biomedical applications. Here, we propose an innovative technique of single-cell encapsulation of human T cells using polyelectrolytes combined with gold nanorods. We have demonstrated encapsulation of human Jurkat T cells with poly(sodium 4-styrenesulfonate) (PSS)-coated gold nanorods (PSS-GNRs). Other forms of encapsulation, using polyelectrolytes without GNRs, were also performed. After Jurkat T cells were encapsulated with poly(allylamine hydrochloride) (PAH) and/or PSS-GNRs or PSS, most cells survived and could proliferate. Jurkat T cells encapsulated with a double layer of PSS-GNR/PAH (PSS-GNR/PAH@Jurkat) showed the highest rate of cell proliferation when compared to 24-h encapsulated cells. With the exception of IL-6, no significant induction of inflammatory cytokines (IL-2, IL-1β, and TNF-α) was observed. Interestingly, when encapsulated cells were co-cultured with THP-1 macrophages, co-cultures exhibited TNF-α production enhancement. However, the co-culture of THP-1 macrophage and PSS-GNR/PAH@Jurkat or PSS/PAH@Jurkat did not enhance TNF-α production. No significant inductions of IL-2, IL-1β, and IL-6 were detected. These data provide promising results, demonstrating the potential use of encapsulated PSS-GNR/PAH@Jurkat to provide a more inert T cell population for immunotherapy application and other biomedical applications.

  13. Effect of Co-Culturing of Mice Liver Cells and Embryonic Carcinomatous Stem Cells on the Rate of Differentiation to Hematopoietic Cells

    Directory of Open Access Journals (Sweden)

    AA Pourfatollah

    2005-10-01

    Full Text Available Introduction: Considering the importance of co-culture in differentiation of embryonic stem cells, the aim of this study was evaluation of the effect of co-culturing fetal liver stroma cells with P19 cells on the line of differentiation. Materials and Methods: For this purpose, P19 cells were cultured directly in semisolid medium. These cells proliferated and primarily differentiated to colonies know as embryoid bodies (EBs after 8-12 days. The Ebs cells were trypsinized and dissociated to single or double cells. Then these cells were co-cultured on the mouse fetal liver feeder layer in the absence of exogenous factors. After 14-18 days, the colonies were studied morphologically by benzidine and giemsa staining and also counted under invert microscope. Results: The percentages of benzidine positive (or erythroid and negative colonies were 94% and 6% respectively and also the cells of colonies were studied by Giemsa staining. Results showed that they were myeloid or lymphoid type cells. Thus, the results show that in the presence of mouse fetal liver feeder layer, the number of erythroid colonies was increased. Conclusions: Therefore, this technique may be effective for differentiation of stem cells from different sources into hematopoietic cells and can be used in future for human cell therapy.

  14. A novel co-culture model of murine K12 osteosarcoma cells and S. aureus on common orthopedic implant materials: 'the race to the surface' studied in vitro.

    Science.gov (United States)

    McConda, David B; Karnes, Jonathan M; Hamza, Therwa; Lindsey, Brock A

    2016-07-01

    Infection is a major cause of orthopedic implant failure. There are few studies assessing both tissue cell and bacterial adherence on common orthopedic implant materials in a co-culture environment. An in vitro co-culture model was created using K12 osteosarcoma cells and Staphylococcus aureus in a medium incubated over metal disks for 48 h. The results showed that, in the presence of S. aureus, there were fewer osteosarcoma cells attached to the disks for all substrata tested. There were significantly more osteosarcoma cells adhering to the cobalt chrome than the stainless steel and titanium disks. Overall, in the presence of osteosarcoma cells, there were more bacteria adhering to the disks for all the substrata tested, with significantly more bacteria adhering to the stainless steel disks compared to cobalt chrome and titanium disks. Scanning electron microscopy verified that osteosarcoma cells and bacteria were adherent to the metal disks after incubation for 48 h. Furthermore, the observation that more bacteria were in the co-culture than in the control sample suggests that the osteosarcoma cells serve as a nutrient source for the bacteria. Future models assessing the interaction of osteogenic cells with bacteria on a substratum would be improved if the model accounted for the role of the immune system in secondary bone healing.

  15. Bioethanol production from sweet potato (Ipomoea batatas L. flour using co-culture of Trichoderma sp. and Saccharomyces cerevisiae in solid-state fermentation

    Directory of Open Access Journals (Sweden)

    Manas Ranjan Swain

    2013-04-01

    Full Text Available The aim of this work was to study the optimiation of co-culturing of Trichoderma sp. and Saccharomyces cerevisiae (1:4 ratio on sweet potato (Ipomoea batatas L. flour (SPF for the production of bio-ethanol in solid-state fermentation (SSF. Maximum ethanol (172 g/kg substrate was produced in a medium containing 80% moisture, ammonium sulphate 0.2%, pH 5.0, inoculuted with 10% inoculum size and fermented at 30ºC for 72h. .Concomitant with highest ethanol concentration, maximum ethanol productivity (2.8 g/kg substrate/h, microbial biomass (23×10(8 CFU/ g substrate, ethanol yield (47 g/100g sugar consumed and fermentation efficiency (72% were also obtained under these conditions. Cell interaction was observed familiar between the viable cells of Trichoderma sp. and S. cerevisiae when co-cultured. Ethanol production ability by the co-culture was 65 % higher than the single culture of S. cerevisiae from un-saccharified SPF.

  16. Live Faecalibacterium prausnitzii induces greater TLR2 and TLR2/6 activation than the dead bacterium in an apical anaerobic co-culture system.

    Science.gov (United States)

    Maier, Eva; Anderson, Rachel C; Altermann, Eric; Roy, Nicole C

    2018-02-01

    Inappropriate activation of intestinal innate immune receptors, such as toll-like receptors (TLRs), by pathogenic bacteria is linked to chronic inflammation. In contrast, a "tonic" level of TLR activation by commensal bacteria is required for intestinal homeostasis. A technical challenge when studying this activation in vitro is the co-culturing of oxygen-requiring mammalian cells with obligate anaerobic commensal bacteria. To overcome this, we used a novel apical anaerobic co-culture system to successfully adapt a TLR activation assay to be conducted in conditions optimised for both cell types. Live Faecalibacterium prausnitzii, an abundant obligate anaerobe of the colonic microbiota, induced higher TLR2 and TLR2/6 activation than the dead bacterium. This enhanced TLR induction by live F. prausnitzii, which until now has not previously been described, may contribute to maintenance of gastrointestinal homeostasis. This highlights the importance of using physiologically relevant co-culture systems to decipher the mechanisms of action of live obligate anaerobes. © 2017 John Wiley & Sons Ltd.

  17. Monocarboxylate transporter-dependent mechanism confers resistance to oxygen- and glucose-deprivation injury in astrocyte-neuron co-cultures.

    Science.gov (United States)

    Gao, Chen; Zhou, Liya; Zhu, Wenxia; Wang, Hongyun; Wang, Ruijuan; He, Yunfei; Li, Zhiyun

    2015-05-06

    Hypoxic and low-glucose stressors contribute to neuronal death in many brain diseases. Astrocytes are anatomically well-positioned to shield neurons from hypoxic injury. During hypoxia/ischemia, lactate released from astrocytes is taken up by neurons and stored for energy. This process is mediated by monocarboxylate transporters (MCTs) in the central nervous system. In the present study, we investigated the ability of astrocytes to protect neurons from oxygen- and glucose-deprivation (OGD) injury via an MCT-dependent mechanism in vitro. Primary cultures of neurons, astrocytes, and astrocytes-neurons derived from rat hippocampus were subjected to OGD, MCT inhibition with small interfering (si)RNA. Cell survival and expression of MCT4, MCT2, glial fibrillary acidic protein, and neuronal nuclear antigen were evaluated. OGD significantly increased cell death in neuronal cultures and up-regulated MCT4 expression in astrocyte cultures, but no increased cell death was observed in neuron-astrocyte co-cultures or astrocyte cultures. However, neuronal cell death in co-cultures was increased by exposure to MCT4- or MCT2-specific siRNA, and this effect was attenuated by the addition of lactate into the extracellular medium of neuronal cultures prior to OGD. These findings demonstrate that resistance to OGD injury in astrocyte-neuron co-cultures occurs via an MCT-dependent mechanism. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  18. Micro-patterning of self-supporting layers with conducting polymer wires for 3D-chip interconnection applications

    International Nuclear Information System (INIS)

    Ackermann, J.; Videlot, C.; Nguyen, T.N.; Wang, L.; Sarro, P.M.; Crawley, D.; Nikolic, K.; Forshaw, M.

    2003-01-01

    Highly conducting polymers have attracted much interest because of their potential applications in sensors and electronic devices. By the use of templates like porous membranes during polymerization conducting molecular wires can be formed with highly anisotropic properties which can be used as interconnecting layers in a three-dimensional (3D)-chip stacking. We focussed on two electrochemical polymerization (ECP) techniques to produce molecular wires based on polypyrrole (PPy) embedded in isolating porous polycarbonate membranes as self-supporting layers. The growth of the polymer through the membrane pores was investigated in order to achieve a good conductivity through the pores, but with a small cross-talk between them. A new polymerization technique based on a structured cathode has been developed in order to control the polymerization locally. By that technique micro-patterned membranes with separated conducting polymer wires could be produced

  19. Micro-patterning of resin-bonded NdFeB magnet for a fully integrated electromagnetic actuator

    Science.gov (United States)

    Tao, Kai; Wu, Jin; Kottapalli, Ajay Giri Prakash; Chen, Di; Yang, Zhuoqing; Ding, Guifu; Lye, Sun Woh; Miao, Jianmin

    2017-12-01

    This paper reports a fully-integrated, batch-fabricated electromagnetic actuator which features micro-patterned NdFeB magnets. The entire actuator is fabricated through MEMS-compatible laminated surface micromachining technology, eliminating the requirement for further component assembly processes. The fabrication strategy allowed the entire volume of the actuator to be reduced to a small size of 2.5 × 2.5 × 2 mm3, which is one of the smallest NdFeB-based electromagnetic actuators demonstrated to date. The magnetic properties of NdFeB thin films are further investigated and optimized using different types of lithographically-defined micromolds. By altering the direction of the input current, actuating displacements of approximately ±10 μm are achieved during both the attraction and the repulsion operations. This work demonstrates the viability and compatibility of using polymer-bonded magnets for magnetic MEMS applications.

  20. Peritoneal carcinomatosis: an unusual presentation of fibrolamellar hepatocellular carcinoma

    International Nuclear Information System (INIS)

    Vicente, R.; Garcia-Gutierrez, J. A.; Fernandez, A.; Santalla, F.

    2001-01-01

    Fibrolamellar hepatocellular carcinoma is an uncommon malignant tumor with characteristic clinical, radiological and histopathological features that is usually associated with a more favorable natural course and greater survival than more common variants of hepatocellular carcinoma. We describe an atypical case of a fibrolamellar hepatocellular carcinomas sowing aggressive behaviour in a 20-year-old woman. The lesion presented with massive ascites, and imaging studies revealed extensive peritoneal metastatic spread. (Author) 8 refs

  1. CP-25 attenuates the inflammatory response of fibroblast-like synoviocytes co-cultured with BAFF-activated CD4(+) T cells.

    Science.gov (United States)

    Jia, Xiaoyi; Wei, Fang; Sun, Xiaojing; Chang, Yan; Xu, Shu; Yang, Xuezhi; Wang, Chun; Wei, Wei

    2016-08-02

    Total glucosides of paeony (TGP) is the first anti-inflammatory immune regulatory drug approved for the treatment of rheumatoid arthritis in China. A novel compound, paeoniflorin-6'-O-benzene sulfonate (code CP-25), comes from the structural modification of paeoniflorin (Pae), which is the effective active ingredient of TGP. The aim of the present study is to investigate the effect of CP-25 on adjuvant arthritis (AA) fibroblast-like synoviocytes (FLS) co-cultured with BAFF-activated CD4(+) T cells and the expression of BAFF-R in CD4(+) T cells. The mRNA expression of BAFF and its receptors was assessed by qPCR. The expression of BAFF receptors in CD4(+) T cells was analyzed by flow cytometry. The effect of CP-25 on AA rats was evaluated by their joint histopathology. The cell culture growth of thymocytes and FLS was detected by cell counting kit (CCK-8). The concentrations of IL-1β, TNF-α, and IL-6 were measured by Enzyme-linked immunosorbent assay (ELISA). The mRNA expression levels of BAFF and BAFF-R were enhanced in the mesenteric lymph nodes of AA rats, TACI expression was reduced, and BCMA had no change. The expression of BAFF-R in CD4(+) T cells was also enhanced. CP-25 alleviated the joint histopathology and decreased the expression of BAFF-R in CD4(+) T cells from AA rats in vivo. In vitro, CP-25 inhibited the abnormal cell culture growth of BAFF-stimulated thymocytes and FLS. In the co-culture system, IL-1β, IL-6 and TNF-α production was enhanced by FLS co-cultured with BAFF-activated CD4(+) T cells. Moreover, BAFF-stimulated CD4(+) T cells promoted the cell culture growth of FLS. The addition of CP-25 decreased the expression of BAFF-R in CD4(+) T cells and inhibited the cell culture growth and cytokine secretion ability of FLS co-cultured with BAFF-activated CD4(+) T cells. The present study indicates that CP-25 may repress the cell culture growth and cytokine secretion ability of FLS, and its inhibitory effects might be associated with its ability

  2. Effect of Cyclic Dynamic Compressive Loading on Chondrocytes and Adipose-Derived Stem Cells Co-Cultured in Highly Elastic Cryogel Scaffolds

    Directory of Open Access Journals (Sweden)

    Chih-Hao Chen

    2018-01-01

    Full Text Available In this study, we first used gelatin/chondroitin-6-sulfate/hyaluronan/chitosan highly elastic cryogels, which showed total recovery from large strains during repeated compression cycles, as 3D scaffolds to study the effects of cyclic dynamic compressive loading on chondrocyte gene expression and extracellular matrix (ECM production. Dynamic culture of porcine chondrocytes was studied at 1 Hz, 10% to 40% strain and 1 to 9 h/day stimulation duration, in a mechanical-driven multi-chamber bioreactor for 14 days. From the experimental results, we could identify the optimum dynamic culture condition (20% and 3 h/day to enhance the chondrocytic phenotype of chondrocytes from the expression of marker (Col I, Col II, Col X, TNF-α, TGF-β1 and IGF-1 genes by quantitative real-time polymerase chain reactions (qRT-PCR and production of ECM (GAGs and Col II by biochemical analysis and immunofluorescence staining. With up-regulated growth factor (TGF-β1 and IGF-1 genes, co-culture of chondrocytes with porcine adipose-derived stem cells (ASCs was employed to facilitate chondrogenic differentiation of ASCs during dynamic culture in cryogel scaffolds. By replacing half of the chondrocytes with ASCs during co-culture, we could obtain similar production of ECM (GAGs and Col II and expression of Col II, but reduced expression of Col I, Col X and TNF-α. Subcutaneous implantation of cells/scaffold constructs in nude mice after mono-culture (chondrocytes or ASCs or co-culture (chondrocytes + ASCs and subject to static or dynamic culture condition in vitro for 14 days was tested for tissue-engineering applications. The constructs were retrieved 8 weeks post-implantation for histological analysis by Alcian blue, Safranin O and Col II immunohistochemical staining. The most abundant ectopic cartilage tissue was found for the chondrocytes and chondrocytes + ASCs groups using dynamic culture, which showed similar neo-cartilage formation capability with half of the

  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. Diagnostic Approaches to Metastatic Hepatocellular Carcinoma of the Orbit.

    Science.gov (United States)

    Geske, Michael J; Bloomer, Michele M; Kersten, Robert C; Vagefi, M Reza

    Orbital metastasis of hepatocellular carcinoma is exceedingly rare and caries a grave prognosis. Three cases of metastatic orbital hepatocellular carcinoma in which the primary tumor was initially unknown and the diagnostic challenges encountered are presented. With hepatocellular carcinoma, open biopsy and palliative tumor debulking has an increased bleeding risk due to the highly vascular nature of the tumor and coagulopathy associated with chronic liver disease. As an alternative, fine needle aspiration biopsy should be considered for hepatocellular carcinoma with a readily accessible mass and the availability of an experienced cytopathologist.

  5. Research progress of vascular change after TACE in hepatocellular carcinoma

    International Nuclear Information System (INIS)

    Kang Zhen; Xiao Enhua

    2013-01-01

    Mortality rate of hepatocellular carcinoma is high. The majority of the patients are diagnosed in advanced stage and lose surgical opportunities. Many studies have reported transcatheter arterial chemoembolization (TACE) is an effective treatment for unresectable hepatocellular carcinoma, and recommended TACE as a standard treatment for hepatocellular carcinoma of Barcelona Clinical Liver Cancer staging (BCLC staging) B. However, TACE can hardly fully embolize tumor blood supply, TACE postoperative hemodynamics and angiogenesis can induce tumor recurrence and metastasis. This paper reviewed characteristics of vascular changes, mechanisms, diagnosis and treatment methods, new progress in the field of hepatocellular carcinoma after TACE. (authors)

  6. Computed tomography diagnosis of hepatocellular carcinoma rupture haemorrhage

    International Nuclear Information System (INIS)

    Zhi Weike; Jiang Bin; Liu Jinquan; Li Sixia; Zhu Zhichang

    2004-01-01

    Objective: To evaluate the diagnostic value of hepatocellular carcinoma rupture hemorrhage using Computed Tomography. Methods: Six cases diagnosed hepatocellular carcinoma rupture hemorrhage were analyzed by morphic and histologic method and investigated the key point of scan in diagnosis. Result: The correct rate of hepatocellular carcinoma rupture hemorrhage by Computed Tomography is above 83 percent, it characteristic representation is strip and would high-density shadow after enhancement. Conclusion: The characteristic representation of hepatocellular carcinoma rupture hemorrhage is attain by Computed Tomography, which provides effective operation evidences for clinical operation. (authors)

  7. Radio-embolization for hepatocellular carcinoma

    International Nuclear Information System (INIS)

    Raoul, J.L.; Edeline, J.; Pracht, M.; Boucher, E.; Rolland, Y.; Garin, E.

    2011-01-01

    Hepatocellular carcinoma is now a major public health concern. In intermediate stages (one third of hepatocellular carcinoma patients), chemo-embolization is the standard of care despite a poor tolerance and a moderate efficacy. Moreover, despite recent improvements, this technique seems in a dead end. Radio-embolization could be an excellent tool for such patients. Currently 131 I-Lipiodol, 188 Re-Lipiodol, 90 Y-glass or resin microspheres are available. More recent and promising data come from microspheres, but phase II and III studies are needed before drawing any conclusion. In the future, the combination of radio-embolization with systemic chemotherapy or targeted agents (particularly anti-angiogenic drugs) seems very promising. (authors)

  8. Serum immunoreactive calcitonin concentration in hepatocellular carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Dugard, J; Kew, M C; Da Fonseca, M; Levin, J [University of the Witwatersrand, Johannesburg (South Africa)

    1982-08-21

    Having found raised serum calcitonin concentrations in 94% of patients with hepatocellular carcinoma when using a dextran-coated-charcoal radio-immunoassay, we have now repeated the study, using a double-antibody radio-immunoassay, in 102 further patients with hepatocellular carcinoma and 35 matched controls. Serum immunoreactive calcitonin concentrations (iCT) in the controls ranged from 10 to 310 pg/ml (mean 154,6 pg/ml). Values in the tumour patients ranged from 10 to 1,650 pg/ml (mean 302,6 pg/ml). The mean figures were significantly higher in the tumour patients (P smaller than 0,001), 35.5% of them having values above 310 pg/ml. In 65 of the patients serum iCT concentrations were also determined by dextran-coated-charcoal radio-immunoassay. Values ranged from 10 to 10780 pg/ml (mean 2,179 pg/ml). If 1,000 pg/ml is taken as the upper limit of normal, 69% of the patients had raised iCT concentrations. There was a good correlation (r=0,67; P smaller than 0,001) between serum iCT values measured with both methods in 50 patients. If measured by the double-antibody radio-immunoassay method, the serum calcitonin value is not useful as a marker for hepatocellular carcinoma.

  9. Serum immunoreactive calcitonin concentration in hepatocellular carcinoma

    International Nuclear Information System (INIS)

    Dugard, J.; Kew, M.C.; Da Fonseca, M.; Levin, J.

    1982-01-01

    Having found raised serum calcitonin concentrations is 94% of patients with hepatocellular carcinoma when using a dextran-coated-charcoal radio-immunoassay, we have now repeated the study, using a double-antibody radio-immunoassay, in 102 further patients with hepatocellular carcinoma and 35 matched controls. Serum immunoreactive calcitonin concentrations (iCT) in the controls ranged from 10 to 310 pg/ml (mean 154,6 pg/ml). Values in the tumour patients ranged from 10 to 1 650 pg/ml (mean 302,6 pg/ml). The mean figures were significantly higher in the tumour patients (P smaller than 0,001), 35,5% of them having values above 310 pg/ml. In 65 of the patients serum iCT concentrations were also determined by dextran-coated-charcoal radio-immunoassay. Values ranged from 10 to 10780 pg/ml (mean 2 179 pg/ml). If 1 000 pg/ml is taken as the upper limit of normal, 69% of the patients had raised iCT concentrations. There was a good correlation (r=0,67; P smaller than 0,001) between serum iCT values measured with both methods in 50 patients. If measured by the double-antibody radio-immunoassay method, the serum calcitonin value is not useful as a marker for hepatocellular carcinoma

  10. Hepatocellular carcinoma arising from hepatocellular adenoma in a hepatitis B virus-associated cirrhotic liver

    International Nuclear Information System (INIS)

    Seo, J.M.; Lee, S.J.; Kim, S.H.; Park, C.K.; Ha, S.Y.

    2012-01-01

    Hepatocellular adenoma (HCA) is a rare, benign proliferation of hepatocytes that occurs mostly in a normal liver and in extreme rare cases, occurs in a cirrhotic liver. Hepatocellular carcinomas (HCC) arising within HCA through malignant transformation is rare. The specific incidence and mechanism of malignant transformation has not been established, but the long term use of oral contraceptives is considered a causative agent. We report a case of HCC arising from HCA detected in a hepatitis B-related cirrhotic liver with serial radiologic images.

  11. Hepatocellular carcinoma arising from hepatocellular adenoma in a hepatitis B virus-associated cirrhotic liver

    Energy Technology Data Exchange (ETDEWEB)

    Seo, J.M. [Department of Radiology and Center for Imaging Science, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul (Korea, Republic of); Lee, S.J., E-mail: lucia@skku.edu [Department of Radiology and Center for Imaging Science, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul (Korea, Republic of); Kim, S.H. [Department of Radiology and Center for Imaging Science, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul (Korea, Republic of); Park, C.K.; Ha, S.Y. [Department of Pathology, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul (Korea, Republic of)

    2012-04-15

    Hepatocellular adenoma (HCA) is a rare, benign proliferation of hepatocytes that occurs mostly in a normal liver and in extreme rare cases, occurs in a cirrhotic liver. Hepatocellular carcinomas (HCC) arising within HCA through malignant transformation is rare. The specific incidence and mechanism of malignant transformation has not been established, but the long term use of oral contraceptives is considered a causative agent. We report a case of HCC arising from HCA detected in a hepatitis B-related cirrhotic liver with serial radiologic images.

  12. Modulation of human dermal microvascular endothelial cell and human gingival fibroblast behavior by micropatterned silica coating surfaces for zirconia dental implant applications

    International Nuclear Information System (INIS)

    Laranjeira, Marta S; Carvalho, Ângela; Ferraz, Maria Pia; Monteiro, Fernando Jorge; Pelaez-Vargas, Alejandro; Hansford, Derek; Coimbra, Susana; Costa, Elísio; Santos-Silva, Alice; Fernandes, Maria Helena

    2014-01-01

    Dental ceramic implants have shown superior esthetic behavior and the absence of induced allergic disorders when compared to titanium implants. Zirconia may become a potential candidate to be used as an alternative to titanium dental implants if surface modifications are introduced. In this work, bioactive micropatterned silica coatings were produced on zirconia substrates, using a combined methodology of sol–gel processing and soft lithography. The aim of the work was to compare the in vitro behavior of human gingival fibroblasts (HGFs) and human dermal microvascular endothelial cells (HDMECs) on three types of silica-coated zirconia surfaces: flat and micropatterned (with pillars and with parallel grooves). Our results showed that cells had a higher metabolic activity (HGF, HDMEC) and increased gene expression levels of fibroblast-specific protein-1 (FSP-1) and collagen type I (COL I) on surfaces with pillars. Nevertheless, parallel grooved surfaces were able to guide cell growth. Even capillary tube-like networks of HDMEC were oriented according to the surface geometry. Zirconia and silica with different topographies have shown to be blood compatible and silica coating reduced bacteria adhesion. All together, the results indicated that microstructured bioactive coating seems to be an efficient strategy to improve soft tissue integration on zirconia implants, protecting implants from peri-implant inflammation and improving long-term implant stabilization. This new approach of micropatterned silica coating on zirconia substrates can generate promising novel dental implants, with surfaces that provide physical cues to guide cells and enhance their behavior. (paper)

  13. Live-Cell, Label-Free Identification of GABAergic and Non-GABAergic Neurons in Primary Cortical Cultures Using Micropatterned Surface

    Science.gov (United States)

    Kono, Sho; Kushida, Takatoshi; Hirano-Iwata, Ayumi; Niwano, Michio; Tanii, Takashi

    2016-01-01

    Excitatory and inhibitory neurons have distinct roles in cortical dynamics. Here we present a novel method for identifying inhibitory GABAergic neurons from non-GABAergic neurons, which are mostly excitatory glutamatergic neurons, in primary cortical cultures. This was achieved using an asymmetrically designed micropattern that directs an axonal process to the longest pathway. In the current work, we first modified the micropattern geometry to improve cell viability and then studied the axon length from 2 to 7 days in vitro (DIV). The cell types of neurons were evaluated retrospectively based on immunoreactivity against GAD67, a marker for inhibitory GABAergic neurons. We found that axons of non-GABAergic neurons grow significantly longer than those of GABAergic neurons in the early stages of development. The optimal threshold for identifying GABAergic and non-GABAergic neurons was evaluated to be 110 μm at 6 DIV. The method does not require any fluorescence labelling and can be carried out on live cells. The accuracy of identification was 98.2%. We confirmed that the high accuracy was due to the use of a micropattern, which standardized the development of cultured neurons. The method promises to be beneficial both for engineering neuronal networks in vitro and for basic cellular neuroscience research. PMID:27513933

  14. Bioavailability of mineral-bound iron to a snow algae-bacteria co-culture and implications for albedo-altering snow algae blooms.

    Science.gov (United States)

    Harrold, Z R; Hausrath, E M; Garcia, A H; Murray, A E; Tschauner, O; Raymond, J; Huang, S

    2018-01-26

    Snow algae can form large-scale blooms across the snowpack surface and near-surface environments. These pigmented blooms can decrease snow albedo, increase local melt rates, and may impact the global heat budget and water cycle. Yet, underlying causes for the geospatial occurrence of these blooms remain unconstrained. One possible factor contributing to snow algae blooms is the presence of mineral dust as a micronutrient source. We investigated the bioavailability of iron (Fe) -bearing minerals, including forsterite (Fo 90 , Mg 1.8 Fe 0.2 SiO 4 ), goethite, smectite and pyrite as Fe sources for a Chloromonas brevispina - bacteria co-culture through laboratory-based experimentation. Fo 90 was capable of stimulating snow algal growth and increased the algal growth rate in otherwise Fe-depleted co-cultures. Fo 90 -bearing systems also exhibited a decrease in bacteria:algae ratios compared to Fe-depleted conditions, suggesting a shift in microbial community structure. The C. brevispina co-culture also increased the rate of Fo 90 dissolution relative to an abiotic control. Analysis of 16S rRNA genes in the co-culture identified Gammaproteobacteria , Betaprotoeobacteria and Sphingobacteria , all of which are commonly found in snow and ice environments. Archaea were not detected. Collimonas and Pseudomonas , which are known to enhance mineral weathering rates, comprised two of the top eight (> 1 %) OTUs. These data provide unequivocal evidence that mineral dust can support elevated snow algae growth under otherwise Fe-depleted growth conditions, and that snow algae can enhance mineral dissolution under these conditions. IMPORTANCE Fe, a key micronutrient for photosynthetic growth, is necessary to support the formation of high-density snow algae blooms. The laboratory experiments described herein allow for a systematic investigation of snow algae-bacteria-mineral interactions and their ability to mobilize and uptake mineral-bound Fe. Results provide unequivocal and

  15. Transarterial (chemo)embolisation for unresectable hepatocellular carcinoma

    DEFF Research Database (Denmark)

    Oliveri, Roberto S; Wetterslev, Jørn; Gluud, Christian

    2011-01-01

    Hepatocellular carcinoma (HCC) results in more than 600,000 deaths per year. Transarterial embolisation (TAE) and transarterial chemoembolisation (TACE) have become standard loco-regional treatments for unresectable HCC.......Hepatocellular carcinoma (HCC) results in more than 600,000 deaths per year. Transarterial embolisation (TAE) and transarterial chemoembolisation (TACE) have become standard loco-regional treatments for unresectable HCC....

  16. Selective angiography in fifty patients with primary hepatocellular carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Shou-Zhong, Wang; Xing-Rong, Chen; Gong-Xian, Wang

    1983-06-01

    Selective angiography is of great importance in the diagnosis of primary hepatocellular carcinoma. It offers information on the findings, multicentricity, localisation, extension, and type of growth. This paper discusses angiography from the methodical point of view, the findings to be obtained, the types of hepatocellular carcinoma, and the diagnostic efficiency of selective angiography in the evaluation of this type of tumour.

  17. Orbital Metastasis of Hepatocellular Carcinoma: A Case Report ...

    African Journals Online (AJOL)

    Background: Hepatocellular carcinoma is one of the commonest malignancies in Nigeria, however metastasis to the orbit is a rare presentation. Objective: To present a rare case of orbital metastasis of hepatocellular carcinoma. Case Report: A 25-year-old man presented with a 3-month history of pain, progressive swelling ...

  18. Improved viability and activity of neutrophils differentiated from HL-60 cells by co-culture with adipose tissue-derived mesenchymal stem cells

    International Nuclear Information System (INIS)

    Park, Yoon Shin; Lim, Goh-Woon; Cho, Kyung-Ah; Woo, So-Youn; Shin, Meeyoung; Yoo, Eun-Sun; Chan Ra, Jeong; Ryu, Kyung-Ha

    2012-01-01

    Highlights: ► Neutropenia is a principal complication of cancer treatment. ► Co-culture of neutrophils with AD-MSC retained cell survival and proliferation and inhibited neutrophil apoptosis under serum starved conditions. ► AD-MSC increased functions of neutrophil. ► AD-MSC promoted the viability of neutrophils by enhancing respiratory burst through the expression of IFN-α, G-CSF, and TGF-β. ► AD-MSC can be used to improve immunity for neutropenia treatment. -- Abstract: Neutropenia is a principal complication of cancer treatment. We investigated the supportive effect of adipose tissue-derived mesenchymal stem cells (AD-MSCs) on the viability and function of neutrophils. Neutrophils were derived from HL-60 cells by dimethylformamide stimulation and cultured with or without AD-MSCs under serum-starved conditions to evaluate neutrophil survival, proliferation, and function. Serum starvation resulted in the apoptosis of neutrophils and decreased cell survival. The co-culture of neutrophils and AD-MSCs resulted in cell survival and inhibited neutrophil apoptosis under serum-starved conditions. The survival rate of neutrophils was prolonged up to 72 h, and the expression levels of interferon (IFN)-α, granulocyte colony-stimulating factor (G-CSF), granulocyte–macrophage colony-stimulating factor, and transforming growth factor (TGF)-β in AD-MSCs were increased after co-culture with neutrophils. AD-MSCs promoted the viability of neutrophils by inhibiting apoptosis as well as enhancing respiratory burst, which could potentially be mediated by the increased expression of IFN-α, G-CSF, and TGF-β. Thus, we conclude that the use of AD-MSCs may be a promising cell-based therapy for increasing immunity by accelerating neutrophil function.

  19. Improved viability and activity of neutrophils differentiated from HL-60 cells by co-culture with adipose tissue-derived mesenchymal stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Park, Yoon Shin; Lim, Goh-Woon [Department of Pediatrics, Ewha Womans University, School of Medicine, Ewha Medical Research Center, Seoul (Korea, Republic of); Cho, Kyung-Ah; Woo, So-Youn; Shin, Meeyoung [Department of Microbiology, Ewha Womans University, School of Medicine, Ewha Medical Research Center, Seoul (Korea, Republic of); Yoo, Eun-Sun [Department of Pediatrics, Ewha Womans University, School of Medicine, Ewha Medical Research Center, Seoul (Korea, Republic of); Chan Ra, Jeong [Stem Cell Research Center, RNL BIO, Seoul 153-768 (Korea, Republic of); Ryu, Kyung-Ha, E-mail: ykh@ewha.ac.kr [Department of Pediatrics, Ewha Womans University, School of Medicine, Ewha Medical Research Center, Seoul (Korea, Republic of)

    2012-06-22

    Highlights: Black-Right-Pointing-Pointer Neutropenia is a principal complication of cancer treatment. Black-Right-Pointing-Pointer Co-culture of neutrophils with AD-MSC retained cell survival and proliferation and inhibited neutrophil apoptosis under serum starved conditions. Black-Right-Pointing-Pointer AD-MSC increased functions of neutrophil. Black-Right-Pointing-Pointer AD-MSC promoted the viability of neutrophils by enhancing respiratory burst through the expression of IFN-{alpha}, G-CSF, and TGF-{beta}. Black-Right-Pointing-Pointer AD-MSC can be used to improve immunity for neutropenia treatment. -- Abstract: Neutropenia is a principal complication of cancer treatment. We investigated the supportive effect of adipose tissue-derived mesenchymal stem cells (AD-MSCs) on the viability and function of neutrophils. Neutrophils were derived from HL-60 cells by dimethylformamide stimulation and cultured with or without AD-MSCs under serum-starved conditions to evaluate neutrophil survival, proliferation, and function. Serum starvation resulted in the apoptosis of neutrophils and decreased cell survival. The co-culture of neutrophils and AD-MSCs resulted in cell survival and inhibited neutrophil apoptosis under serum-starved conditions. The survival rate of neutrophils was prolonged up to 72 h, and the expression levels of interferon (IFN)-{alpha}, granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor, and transforming growth factor (TGF)-{beta} in AD-MSCs were increased after co-culture with neutrophils. AD-MSCs promoted the viability of neutrophils by inhibiting apoptosis as well as enhancing respiratory burst, which could potentially be mediated by the increased expression of IFN-{alpha}, G-CSF, and TGF-{beta}. Thus, we conclude that the use of AD-MSCs may be a promising cell-based therapy for increasing immunity by accelerating neutrophil function.

  20. Dietary n-3 PUFA affect TcR-mediated activation of purified murine T cells and accessory cell function in co-cultures

    Science.gov (United States)

    CHAPKIN, R S; ARRINGTON, J L; APANASOVICH, T V; CARROLL, R J; MCMURRAY, D N

    2002-01-01

    Diets enriched in n-3 polyunsaturated fatty acids (PUFA) suppress several functions of murine splenic T cells by acting directly on the T cells and/or indirectly on accessory cells. In this study, the relative contribution of highly purified populations of the two cell types to the dietary suppression of T cell function was examined. Mice were fed diets containing different levels of n-3 PUFA; safflower oil (SAF; control containing no n-3 PUFA), fish oil (FO) at 2% and 4%, or 1% purified docosahexaenoic acid (DHA) for 2 weeks. Purified (>90%) T cells were obtained from the spleen, and accessory cells (>95% adherent, esterase-positive) were obtained by peritoneal lavage. Purified T cells or accessory cells from each diet group were co-cultured with the alternative cell type from every other diet group, yielding a total of 16 different co-culture combinations. The T cells were stimulated with either concanavalin A (ConA) or antibodies to the T cell receptor (TcR)/CD3 complex and the costimulatory molecule CD28 (αCD3/αCD28), and proliferation was measured after four days. Suppression of T cell proliferation in the co-cultures was dependent upon the dose of dietary n-3 PUFA fed to mice from which the T cells were derived, irrespective of the dietary treatment of accessory cell donors. The greatest dietary effect was seen in mice consuming the DHA diet (P = 0·034 in the anova; P = 0·0053 in the Trend Test), and was observed with direct stimulation of the T cell receptor and CD28 costimulatory ligand, but not with ConA. A significant dietary effect was also contributed accessory cells (P = 0·033 in the Trend Test). We conclude that dietary n-3 PUFA affect TcR-mediated by T cell activation by both direct and indirect (accessory cell) mechanisms. PMID:12296847

  1. LFA-1 and ICAM-1 expression induced during melanoma-endothelial cell co-culture favors the transendothelial migration of melanoma cell lines in vitro

    International Nuclear Information System (INIS)

    Ghislin, Stephanie; Obino, Dorian; Middendorp, Sandrine; Boggetto, Nicole; Alcaide-Loridan, Catherine; Deshayes, Frederique

    2012-01-01

    Patients with metastatic melanoma have a poor median rate of survival. It is therefore necessary to increase our knowledge about melanoma cell dissemination which includes extravasation, where cancer cells cross the endothelial barrier. Extravasation is well understood during travelling of white blood cells, and involves integrins such as LFA-1 (composed of two chains, CD11a and CD18) expressed by T cells, while ICAM-1 is induced during inflammation by endothelial cells. Although melanoma cell lines cross endothelial cell barriers, they do not express LFA-1. We therefore hypothesized that melanoma-endothelial cell co-culture might induce the LFA-1/ICAM ligand/receptor couple during melanoma transmigration. A transwell approach has been used as well as blocking antibodies against CD11a, CD18 and ICAM-1. Data were analyzed with an epifluorescence microscope. Fluorescence intensity was quantified with the ImageJ software. We show here that HUVEC-conditioned medium induce cell-surface expression of LFA-1 on melanoma cell lines. Similarly melanoma-conditioned medium activates ICAM-1 expression in endothelial cells. Accordingly blocking antibodies of ICAM-1, CD11a or CD18 strongly decrease melanoma transmigration. We therefore demonstrate that melanoma cells can cross endothelial monolayers in vitro due to the induction of ICAM-1 and LFA-1 occurring during the co-culture of melanoma and endothelial cells. Our data further suggest a role of LFA-1 and ICAM-1 in the formation of melanoma cell clumps enhancing tumor cell transmigration. Melanoma-endothelial cell co-culture induces LFA-1 and ICAM-1 expression, thereby favoring in vitro melanoma trans-migration

  2. Fighting for territories: time-lapse analysis of dental pulp and dental follicle stem cells in co-culture reveals specific migratory capabilities

    Directory of Open Access Journals (Sweden)

    C Schiraldi

    2012-11-01

    Full Text Available Stem cell migration is a critical step during the repair of damaged tissues. In order to achieve appropriate cell-based therapies for tooth and periodontal ligament repair it is necessary first to understand the dynamics of tissue-specific stem cell populations such as dental pulp stem cells (DPSC and dental follicle stem cells (DFSC. Using time-lapse imaging, we analysed migratory and proliferative capabilities of these two human stem cell lines in vitro. When cultured alone, both DPSC and DFSC exhibited low and irregular migration profiles. In co-cultures, DFSC, but not DPSC, spectacularly increased their migration activity and velocity. DFSC rapidly surrounded the DPSC, thus resembling the in vivo developmental process, where follicle cells encircle both dental epithelium and pulp. Cell morphology was dependent on the culture conditions (mono-culture or co-culture and changed over time. Regulatory genes involved in dental cell migration and differentiation such as TWIST1, MSX1, RUNX2, SFRP1 and ADAM28, were also evaluated in co-cultures. MSX1 up-regulation indicates that DPSC and DFSC retain their odontogenic potential. However, DPSC lose their capacity to differentiate into odontoblasts in the presence of DFSC, as suggested by RUNX2 up-regulation and TWIST1 down-regulation. In contrast, the unchanged levels of SFRP1 expression suggest that DFSC retain their potential to form periodontal tissues even in the presence of DPSC. These findings demonstrate that stem cells behave differently according to their environment, retain their genetic memory, and compete with each other to acquire the appropriate territory. Understanding the mechanisms involved in stem cell migration may lead to new therapeutic approaches for tooth repair.

  3. LFA-1 and ICAM-1 expression induced during melanoma-endothelial cell co-culture favors the transendothelial migration of melanoma cell lines in vitro

    Directory of Open Access Journals (Sweden)

    Ghislin Stephanie

    2012-10-01

    Full Text Available Abstract Background Patients with metastatic melanoma have a poor median rate of survival. It is therefore necessary to increase our knowledge about melanoma cell dissemination which includes extravasation, where cancer cells cross the endothelial barrier. Extravasation is well understood during travelling of white blood cells, and involves integrins such as LFA-1 (composed of two chains, CD11a and CD18 expressed by T cells, while ICAM-1 is induced during inflammation by endothelial cells. Although melanoma cell lines cross endothelial cell barriers, they do not express LFA-1. We therefore hypothesized that melanoma-endothelial cell co-culture might induce the LFA-1/ICAM ligand/receptor couple during melanoma transmigration. Methods A transwell approach has been used as well as blocking antibodies against CD11a, CD18 and ICAM-1. Data were analyzed with an epifluorescence microscope. Fluorescence intensity was quantified with the ImageJ software. Results We show here that HUVEC-conditioned medium induce cell-surface expression of LFA-1 on melanoma cell lines. Similarly melanoma-conditioned medium activates ICAM-1 expression in endothelial cells. Accordingly blocking antibodies of ICAM-1, CD11a or CD18 strongly decrease melanoma transmigration. We therefore demonstrate that melanoma cells can cross endothelial monolayers in vitro due to the induction of ICAM-1 and LFA-1 occurring during the co-culture of melanoma and endothelial cells. Our data further suggest a role of LFA-1 and ICAM-1 in the formation of melanoma cell clumps enhancing tumor cell transmigration. Conclusion Melanoma-endothelial cell co-culture induces LFA-1 and ICAM-1 expression, thereby favoring in vitro melanoma trans-migration.

  4. Role of IL-1 beta and COX2 in silica-induced IL-6 release and loss of pneumocytes in co-cultures.

    Science.gov (United States)

    Herseth, Jan I; Refsnes, Magne; Låg, Marit; Schwarze, Per E

    2009-10-01

    The pro-inflammatory cytokines IL-1 beta, TNF-alpha and IL-6 are of great importance in the development of silica-induced lung damage and repair. In this study we investigated the role of IL-1 beta, TNF-alpha and COX2 in silica-induced regulation of IL-6 release and pneumocyte loss in various mono- and co-cultures of monocytes, pneumocytes and endothelial cells. All co-cultures with monocytes, and especially cultures including endothelial cells, showed an increase of silica-induced release of IL-6 compared to the respective monocultures. Treatment with the antagonist IL-1 ra strongly decreased IL-1 beta and IL-6 release in contact co-cultures of monocytes and pneumocytes. COX2 up-regulation by silica and IL-1 beta was eliminated by IL-1 ra. Inhibition of COX2 markedly reduced both IL-1 beta and IL-6 release. IL-1 ra was more effective than COX2-inhibition in reduction of IL-6, but not of IL-1 beta. Silica-induced pneumocyte loss was reduced by IL-1 beta, but this effect was not counteracted by the IL-1 receptor antagonist. Our findings suggest that silica-induced IL-6 release from pneumocytes is mainly mediated via IL-1 beta release from the monocytes, via both COX2-dependent and -independent pathways. Notably, COX2-derived mediators seem crucial for a positive feed-back regulation of IL-1 beta release from the monocytes. In contrast to silica-induced IL-6, the reduction in pneumocyte loss by IL-1 beta does not seem to be regulated through an IL-1R1-dependent mechanism.

  5. Palmitoylethanolamide Blunts Amyloid-β42-Induced Astrocyte Activation and Improves Neuronal Survival in Primary Mouse Cortical Astrocyte-Neuron Co-Cultures.

    Science.gov (United States)

    Beggiato, Sarah; Borelli, Andrea Celeste; Ferraro, Luca; Tanganelli, Sergio; Antonelli, Tiziana; Tomasini, Maria Cristina

    2018-01-01

    Based on the pivotal role of astrocytes in brain homeostasis and the strong metabolic cooperation existing between neurons and astrocytes, it has been suggested that astrocytic dysfunctions might cause and/or contribute to neuroinflammation and neurodegenerative processes. Therapeutic approaches aimed at both neuroprotection and neuroinflammation reduction may prove particularly effective in slowing the progression of these diseases. The endogenous lipid mediator palmitoylethanolamide (PEA) displayed neuroprotective and anti(neuro)inflammatory properties, and demonstrated interesting potential as a novel treatment for Alzheimer's disease. We firstly evaluated whether astrocytes could participate in regulating the Aβ42-induced neuronal damage, by using primary mouse astrocytes cell cultures and mixed astrocytes-neurons cultures. Furthermore, the possible protective effects of PEA against Aβ42-induced neuronal toxicity have also been investigated by evaluating neuronal viability, apoptosis, and morphometric parameters. The presence of astrocytes pre-exposed to Aβ42 (0.5μM; 24 h) induced a reduction of neuronal viability in primary mouse astrocytes-neurons co-cultures. Furthermore, under these experimental conditions, an increase in the number of neuronal apoptotic nuclei and a decrease in the number of MAP-2 positive neurons were observed. Finally, astrocytic Aβ42 pre-exposure induced an increase in the number of neurite aggregations/100μm as compared to control (i.e., untreated) astrocytes-neurons co-cultures. These effects were not observed in neurons cultured in the presence of astrocytes pre-exposed to PEA (0.1μM), applied 1 h before and maintained during Aβ42 treatment. Astrocytes contribute to Aβ42-induced neurotoxicity and PEA, by blunting Aβ42-induced astrocyte activation, improved neuronal survival in mouse astrocyte-neuron co-cultures.

  6. Protein hydrolysate from canned sardine and brewing by-products improves TNF-α-induced inflammation in an intestinal-endothelial co-culture cell model.

    Science.gov (United States)

    Vieira, Elsa F; Van Camp, John; Ferreira, Isabel M P L V O; Grootaert, Charlotte

    2017-07-17

    The anti-inflammatory activity of sardine protein hydrolysates (SPH) obtained by hydrolysis with proteases from brewing yeast surplus was ascertained. For this purpose, a digested and desalted SPH fraction with molecular weight lower than 10 kDa was investigated using an endothelial cell line (EA.hy926) as such and in a co-culture model with an intestinal cell line (Caco-2). Effects of SPH <10 kDa on nitric oxide (NO) production, reactive oxygen species (ROS) inhibition and secretion of monocyte chemoattractant protein 1 (MCP-1), vascular endothelial growth factor (VEGF), chemokine IL-8 (IL-8) and intercellular adhesion molecule-1 (ICAM-1) were evaluated in TNF-α-treated and untreated cells. Upon TNF-α treatment, levels of NO, MCP-1, VEGF, IL-8, ICAM-1 and endothelial ROS were significantly increased in both mono- and co-culture models. Treatment with SPH <10 kDa (2.0 mg peptides/mL) significantly decreased all the inflammation markers when compared to TNF-α-treated control. This protective effect was more pronounced in the co-culture model, suggesting that SPH <10 kDa Caco-2 cells metabolites produced in the course of intestinal absorption may provide a more relevant protective effect against endothelial dysfunction. Additionally, indirect cross-talk between two cell types was established, suggesting that SPH <10 kDa may also bind to receptors on the Caco-2 cells, thereby triggering a pathway to secrete the pro-inflammatory compounds. Overall, these in vitro screening results, in which intestinal digestion, absorption and endothelial bioactivity are simulated, show the potential of SPH to be used as a functional food with anti-inflammatory properties.

  7. Potassium 2-(1-hydroxypentyl-benzoate attenuates neuronal apoptosis in neuron–astrocyte co-culture system through neurotrophy and neuroinflammation pathway

    Directory of Open Access Journals (Sweden)

    Dongmei Liu

    2017-09-01

    Full Text Available Potassium 2-(1-hydroxypentyl-benzoate (d,l-PHPB, a new drug candidate for ischemic stroke at the phase II clinic trial, has been shown to protect neurons by inhibiting oxidative injury and reducing neuron apoptosis in previous studies. But the mechanisms of d,l-PHPB remain to be studied. In this study, a neuron–astrocytes co-culture system was used to elucidate the roles of astrocytes in neuroprotection of d,l-PHPB under oxygen-glucose deprivation/reoxygenation (OGD/R condition. Our data showed that d,l-PHPB reduced neuronal apoptosis in mono-culture system and this effect was enhanced in neuron–astrocyte co-culture system under the OGD/R condition. Meanwhile, d,l-PHPB obviously increased the levels of brain-derived neurotrophic factor (BDNF and nerve growth factor (NGF, which were mainly secreted from astrocytes, in the co-culture system after OGD/R. The PI3K/AKT and ERK signaling pathways as well as the p-TRKA/B receptors were involved in the process. In addition, the levels of TNF-α and IL-1β secreted from astrocytes after OGD/R were markedly reduced after d,l-PHPB treatment, which was mainly due to the suppression of phosphorylated p38. In conclusion, the present study demonstrates that the neuroprotective effects of d,l-PHPB were improved by astrocytes, mainly mediated by increasing the release of BDNF/NGF and attenuating inflammatory cytokines.

  8. The angiogenic behaviors of human umbilical vein endothelial cells (HUVEC) in co-culture with osteoblast-like cells (MG-63) on different titanium surfaces.

    Science.gov (United States)

    Shi, Bin; Andrukhov, Oleh; Berner, Simon; Schedle, Andreas; Rausch-Fan, Xiaohui

    2014-08-01

    Interaction between osteogenesis and angiogenesis plays an important role in implant osseointegration. In the present study we investigated the influence of titanium surface properties on the angiogenic behaviors of endothelial cells grown in direct contact co-culture with osteoblasts. Human umbilical vein endothelial cells (HUVECs) and osteoblast-like cells (MG-63 cells) were grown in direct co-culture on the following titanium surfaces: acid-etched (A), hydrophilic A (modA), coarse-gritblasted and acid-etched (SLA) and hydrophilic SLA (SLActive). Cell proliferation was evaluated by cell counting combined with flow cytometry. The expression of von Willebrand Factor (vWF), thrombomodulin (TM), endothelial cell protein C receptor (EPCR), E-Selectin, as well as vascular endothelial growth factor (VEGF) receptors Flt-1 and KDR in HUVECs and VEGF in MG-63 were measured by qPCR. The dynamic behavior of endothelial cells was recorded by time-lapse microscopy. Proliferation of HUVECs was highest on A, followed by SLA, modA and SLActive surfaces. The expression of vWF, TM, EPCR, E-Selectin and Flt-1 in HUVECs was significantly higher on A than on all other surfaces. The expression of KDR in HUVECs grown on A surface was below detection limit. VEGF expression in MG-63 cells was significantly higher on SLActive vs SLA and modA vs A surfaces. Time-lapse microscopy revealed that HUVECs moved quickest and formed cell clusters earlier on A surface, followed by SLA, modA and SLActive surface. In co-culture conditions, proliferation and expression of angiogenesis associated genes in HUVECs are promoted by smooth hydrophobic Ti surface, which is in contrast to previous mono-culture studies. Copyright © 2014 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.

  9. Immunoregulatory effects of human dental pulp-derived stem cells on T cells: comparison of transwell co-culture and mixed lymphocyte reaction systems.

    Science.gov (United States)

    Demircan, Pinar Cetinalp; Sariboyaci, Ayla Eker; Unal, Zehra Seda; Gacar, Gulcin; Subasi, Cansu; Karaoz, Erdal

    2011-11-01

    BACKGROUND AIMS. Studies performed using human and animal models have indicated the immunoregulatory capability of mesenchymal stromal cells in several lineages. We investigated whether human dental pulp-derived stem cells (hDP-SC) have regulatory effects on phytohemagglutinin (PHA)-activated CD3(+) T cells. We aimed to define the regulatory mechanisms associated with hDP-SC that occur in mixed lymphocyte reaction (MLR) and transwell systems with PHA-CD3(+) T cells and hDP-SC at a ratio of 1:1. METHODS. Proliferation, apoptosis and pro- and anti-inflammatory cytokines of PHA-CD3(+)T cells, the expression of Regulatory T cells (Treg) markers and some regulatory factors related to hDP-SC, were studied in Both transwell and MLR are co-cultures systems. RESULTS. Anti-proliferative and apoptotic effects of hDP-SC were determined in co-culture systems. Elevated expression levels of human leukocyte antigen (HLA)-G, hepatocyte growth factor (HGF)-β1, intracellular adhesion molecule (ICAM-1)-1, interleukin (IL)-6, IL-10, transforming growth factor (TGF)-β1, vascular adhesion molecule (VCAM)-1 and vascular endothelial growth factor (VEGF) by hDP-SC were detected in the co-culture systems. We observed decreased expression levels of pro-inflammatory cytokines [interferon (IFN)-γ, IL-2, IL-6 receptor (R), IL-12, Interleukin-17A (IL-17A), tumor necrosis factor (TNF)-α] and increased expression levels of anti-inflammatory cytokine [inducible protein (IP)-10] from PHA-CD3(+) T cells in the transwell system. Expression of Treg (CD4(+) CD25(+) Foxp3(+)) markers was significantly induced by hDP-SC in both co-culture systems. We observed apoptosis of PHA-CD3(+) T cells with 24 h using time-lapse camera photographs and active caspase labeling; it is likely that paracrine soluble factors and molecular signals secreted by hDP-SC led this apoptosis. CONCLUSIONS. We suggest that hDP-SC have potent immunoregulatory functions because of their soluble factors and cytokines via paracrine

  10. Metastases of Hepatocellular Carcinoma Misdiagnosed as Isolated Hypertrophic Cardiomyopathy.

    Science.gov (United States)

    Greco, Assunta; De Masi, Roberto; Orlando, Stefania; Metrangolo, Antonio; Zecca, Vittorio; Morciano, Giancarlo; De Donno, Antonella; Bagordo, Francesco; Piccinni, Giancarlo

    At present, cardiac metastasis of hepatocellular carcinoma is rarely mentioned in the literature. We report a hepatocellular carcinoma patient with cardiac metastasis misdiagnosed as hypertrophic cardiomyopathy in 2011. Two years later, on presentation of syncope, an abnormal ventricular septal size was recorded by ultrasound scan, and was subsequently shown by magnetic resonance imaging to be a tumour lesion. A myocardial biopsy confirmed infiltration of hepatocellular carcinoma. This observation underlines the risk of hepatocellular carcinoma cardiac metastasis, manifested in its infiltrative form as hypertrophic cardiomyopathy. In conclusion, we suggest that the ultrasound appearance of hypertrophic cardiomyopathy in hepatocellular carcinoma patients should be seen as a "red flag" and recommend the introduction of magnetic resonance imaging assessment of transplant candidates.

  11. The potential of cell sheet technique on the development of hepatocellular carcinoma in rat models.

    Directory of Open Access Journals (Sweden)

    Alaa T Alshareeda

    Full Text Available Hepatocellular carcinoma (HCC is considered the 3rd leading cause of death by cancer worldwide with the majority of patients were diagnosed in the late stages. Currently, there is no effective therapy. The selection of an animal model that mimics human cancer is essential for the identification of prognostic/predictive markers, candidate genes underlying cancer induction and the examination of factors that may influence the response of cancers to therapeutic agents and regimens. In this study, we developed a HCC nude rat models using cell sheet and examined the effect of human stromal cells (SCs on the development of the HCC model and on different liver parameters such as albumin and urea.Transplanted cell sheet for HCC rat models was fabricated using thermo-responsive culture dishes. The effect of human umbilical cord mesenchymal stromal cells (UC-MSCs and human bone marrow mesenchymal stromal cells (BM-MSCs on the developed tumour was tested. Furthermore, development of tumour and detection of the liver parameter was studied. Additionally, angiogenesis assay was performed using Matrigel.HepG2 cells requires five days to form a complete cell sheet while HepG2 co-cultured with UC-MSCs or BM-MSCs took only three days. The tumour developed within 4 weeks after transplantation of the HCC sheet on the liver of nude rats. Both UC-MSCs and BM-MSCs improved the secretion of liver parameters by increasing the secretion of albumin and urea. Comparatively, the UC-MSCs were more effective than BM-MSCs, but unlike BM-MSCs, UC-MSCs prevented liver tumour formation and the tube formation of HCC.Since this is a novel study to induce liver tumour in rats using hepatocellular carcinoma sheet and stromal cells, the data obtained suggest that cell sheet is a fast and easy technique to develop HCC models as well as UC-MSCs have therapeutic potential for liver diseases. Additionally, the data procured indicates that stromal cells enhanced the fabrication of HepG2

  12. Elevated serum levels of Chromogranin A in hepatocellular carcinoma.

    Science.gov (United States)

    Biondi, Antonio; Malaguarnera, Giulia; Vacante, Marco; Berretta, Massimiliano; D'Agata, Velia; Malaguarnera, Michele; Basile, Francesco; Drago, Filippo; Bertino, Gaetano

    2012-01-01

    During the past three decades, the incidence of hepatocellular carcinoma in the United States has tripled. The neuroendocrine character has been observed in some tumor cells within some hepatocellular carcinoma nodules and elevated serum chromogranin A also been reported in patients with hepatocellular carcinoma. The aim of this work was to investigate the role of serum concentration of chromogranin A in patients with hepatocellular carcinoma at different stages. The study population consisted of 96 patients (63 males and 33 females age range 52-84) at their first hospital admission for hepatocellular carcinoma. The control group consisted of 35 volunteers (20 males and 15 females age range 50-80). The hepatocellular carcinoma patients were stratified according the Barcelona-Clinic Liver Cancer classification. Venous blood samples were collected before treatment from each patients before surgery, centrifuged to obtain serum samples and stored at -80° C until assayed. The chromogranin A serum levels were elevated (> 100 ng/ml) in 72/96 patients with hepatocellular carcinoma. The serum levels of chromogranin A were significantly correlated (p<0.05) with alpha-fetoprotein. In comparison with controls, the hepatocellular carcinoma patients showed a significant increase (p<0.001) vs controls. The chromogranin A levels in the Barcelona staging of hepatocellular carcinoma was higher in stage D compared to stage C (p<0.01), to stage B (p<0.001), and to stage A (p<0.001). Molecular markers, such as chromogranin A, could be very useful tools for hepatocellular carcinoma diagnosis. However the molecular classification should be incorporated into a staging scheme, which effectively separated patients into groups with homogeneous prognosis and response to treatment, and thus serves to aid in the selection of appropriate therapy.

  13. CT diagnosis of rare histological variant of hepatocellular carcinoma

    International Nuclear Information System (INIS)

    Li Huaibo; Feng Zhipeng; Duan Shaoyin; Zhaugn Xiangrong

    2009-01-01

    Objective: To explore and understand the CT findings of 5 rare histological variants of hepatocellular carcinoma. Methods: CT findings of 31 cases of rare histological variants confirmed by surgery and pathology were analyzed retrospectively. Results: 13 cases were clear cell hepatocellular carcinoma. 3 cases of them showed patchy fat density in plain scans. Enhanced CT showed features of 'fast in fast out' which was similar to the common hepatocellular carcinoma. 4 cases belonged to sclerosis hepatocellular carcinoma. They appeared as heterogeneous, slowly enhancement on arterial phase images, and delay enhancement on portal venous phase and delay phase images. 9 cases belonged to mixed hepatocellular carcinoma. 5 cases of them showed inhomogeneous enhancement and 4 without enhancement during arterial phase, 3 cases showed delay enhancement and 4 without during portal venous and delay phase. 3 cases were fibrolamellar hepatocellular carcinoma. All showed obvious and fastly enhancement on arterial phase images, subsided slowly on the portal venous and delay phase images, showing features of 'fast in slow out', no enhancement was seen in the central scar. Shrinkage phenomenon on the surface of liver could be seen on the CT plain scans in sclerosis, mixed and fibrolamellar hepatocellular carcinoma. 2 cases were the type of dense hepatocellular carcinoma. The surrounding part in the 2 cases were slightly enhanced, while the most part of the center were not enhanced similar to necrosis. Conclusion: The CT findings of rate histological variant of hepatocellular carcinoma are characteristic. Analyzing the CT plain and enhancement finding is helpful to the diagnosis of these types of hepatocellular carcinoma. (authors)

  14. A qualitative signature for early diagnosis of hepatocellular carcinoma based on relative expression orderings.

    Science.gov (United States)

    Ao, Lu; Zhang, Zimei; Guan, Qingzhou; Guo, Yating; Guo, You; Zhang, Jiahui; Lv, Xingwei; Huang, Haiyan; Zhang, Huarong; Wang, Xianlong; Guo, Zheng

    2018-04-23

    Currently, using biopsy specimens to confirm suspicious liver lesions of early hepatocellular carcinoma are not entirely reliable because of insufficient sampling amount and inaccurate sampling location. It is necessary to develop a signature to aid early hepatocellular carcinoma diagnosis using biopsy specimens even when the sampling location is inaccurate. Based on the within-sample relative expression orderings of gene pairs, we identified a simple qualitative signature to distinguish both hepatocellular carcinoma and adjacent non-tumour tissues from cirrhosis tissues of non-hepatocellular carcinoma patients. A signature consisting of 19 gene pairs was identified in the training data sets and validated in 2 large collections of samples from biopsy and surgical resection specimens. For biopsy specimens, 95.7% of 141 hepatocellular carcinoma tissues and all (100%) of 108 cirrhosis tissues of non-hepatocellular carcinoma patients were correctly classified. Especially, all (100%) of 60 hepatocellular carcinoma adjacent normal tissues and 77.5% of 80 hepatocellular carcinoma adjacent cirrhosis tissues were classified to hepatocellular carcinoma. For surgical resection specimens, 99.7% of 733 hepatocellular carcinoma specimens were correctly classified to hepatocellular carcinoma, while 96.1% of 254 hepatocellular carcinoma adjacent cirrhosis tissues and 95.9% of 538 hepatocellular carcinoma adjacent normal tissues were classified to hepatocellular carcinoma. In contrast, 17.0% of 47 cirrhosis from non-hepatocellular carcinoma patients waiting for liver transplantation were classified to hepatocellular carcinoma, indicating that some patients with long-lasting cirrhosis could have already gained hepatocellular carcinoma characteristics. The signature can distinguish both hepatocellular carcinoma tissues and tumour-adjacent tissues from cirrhosis tissues of non-hepatocellular carcinoma patients even using inaccurately sampled biopsy specimens, which can aid early

  15. Water filtration rate and infiltration/accumulation of low density lipoproteins in 3 different modes of endothelial/smooth muscle cell co-cultures.

    Science.gov (United States)

    Ding, ZuFeng; Fan, YuBo; Deng, XiaoYan

    2009-11-01

    Using different endothelial/smooth muscle cell co-culture modes to simulate the intimal structure of blood vessels, the water filtration rate and the infiltration/accumulation of LDL of the cultured cell layers were studied. The three cell culture modes of the study were: (i) The endothelial cell monolayer (EC/Phi); (ii) endothelial cells directly co-cultured on the smooth muscle cell monolayer (EC-SMC); (iii) endothelial cells and smooth muscle cells cultured on different sides of a Millicell-CM membrane (EC/SMC). It was found that under the same condition, the water filtration rate was the lowest for the EC/SMC mode and the highest for the EC/Phi mode, while the infiltration/accumulation of DiI-LDLs was the lowest in the EC/Phi mode and the highest in the EC-SMC mode. It was also found that DiI-LDL infiltration/accumulation in the cultured cell layers increased with the increasing water filtration rate. The results from the in vitro model study therefore suggest that the infiltration/accumulation of the lipids within the arterial wall is positively correlated with concentration polarization of atherogenic lipids, and the integrity of the endothelium plays an important role in the penetration and accumulation of atherogenic lipids in blood vessel walls.

  16. Demonstration of neuron-glia transfer of precursors for GABA biosynthesis in a co-culture system of dissociated mouse cerebral cortex.

    Science.gov (United States)

    Leke, Renata; Bak, Lasse K; Schousboe, Arne; Waagepetersen, Helle S

    2008-12-01

    Co-cultures of neurons and astrocytes were prepared from dissociated embryonic mouse cerebral cortex and cultured for 7 days. To investigate if these cultures may serve as a functional model system to study neuron-glia interaction with regard to GABA biosynthesis, the cells were incubated either in media containing [U-(13)C]glutamine (0.1, 0.3 and 0.5 mM) or 1 mM acetate plus 2.5 mM glucose plus 1 mM lactate. In the latter case one of the 3 substrates was uniformly (13)C labeled. Cellular contents and (13)C labeling of glutamate, GABA, aspartate and glutamine were determined in the cells after an incubation period of 2.5 h. The GABA biosynthetic machinery exhibited the expected complexity with regard to metabolic compartmentation and involvement of TCA cycle activity as seen in other culture systems containing GABAergic neurons. Metabolism of acetate clearly demonstrated glial synthesis of glutamine and its transfer to the neuronal compartment. It is concluded that this co-culture system serves as a reliable model in which functional and pharmacological aspects of GABA biosynthesis can be investigated.

  17. Growth of Chlamydomonas reinhardtii in acetate-free medium when co-cultured with alginate-encapsulated, acetate-producing strains of Synechococcus sp. PCC 7002.

    Science.gov (United States)

    Therien, Jesse B; Zadvornyy, Oleg A; Posewitz, Matthew C; Bryant, Donald A; Peters, John W

    2014-01-01

    The model alga Chlamydomonas reinhardtii requires acetate as a co-substrate for optimal production of lipids, and the addition of acetate to culture media has practical and economic implications for algal biofuel production. Here we demonstrate the growth of C. reinhardtii on acetate provided by mutant strains of the cyanobacterium Synechococcus sp. PCC 7002. Optimal growth conditions for co-cultivation of C. reinhardtii with wild-type and mutant strains of Synechococcus sp. 7002 were established. In co-culture, acetate produced by a glycogen synthase knockout mutant of Synechococcus sp. PCC 7002 was able to support the growth of a lipid-accumulating mutant strain of C. reinhardtii defective in starch production. Encapsulation of Synechococcus sp. PCC 7002 using an alginate matrix was successfully employed in co-cultures to limit growth and maintain the stability. The ability of immobilized strains of the cyanobacterium Synechococcus sp. PCC 7002 to produce acetate at a level adequate to support the growth of lipid-accumulating strains of C. reinhartdii offers a potentially practical, photosynthetic alternative to providing exogenous acetate into growth media.

  18. 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.

  19. Release of neuropeptides from a neuro-cutaneous co-culture model: A novel in vitro model for studying sensory effects of ciguatoxins.

    Science.gov (United States)

    Le Garrec, Raphaele; L'herondelle, Killian; Le Gall-Ianotto, Christelle; Lebonvallet, Nicolas; Leschiera, Raphael; Buhe, Virginie; Talagas, Matthieu; Vetter, Irina; Lewis, Richard J; Misery, Laurent

    2016-06-15

    Ciguatoxins are the major toxins responsible for ciguatera fish poisoning, a disease dominated by muco-cutaneous sensory disorders including paresthesiae, cold dysesthesia and pruritus. While the ciguatoxins are well known to target voltage-gated sodium channels (VGSCs), the ensuing molecular mechanisms underlying these sensory disorders remain poorly understood. In this study, we propose a primary sensory neuron-keratinocyte co-culture as an appropriate model to study the neuro-cutaneous effects of ciguatoxins. Using this model, we show for the first time that nanomolar concentrations of Pacific ciguatoxin-2 (P-CTX-2) induced a VGSC-dependent release of substance P (SP) and calcitonin gene-related peptide (CGRP). As these neuropeptides are known mediators of pain and itch sensations, the ciguatoxin-induced sensory disturbances in ciguatera fish poisoning may involve the release of these neuropeptides. We further determined time- and P-CTX-2 concentration-dependence of the release of SP and CGRP from the co-culture model. Moreover, we highlighted the influence of extracellular calcium on the release of neuropeptides elicited by P-CTX-2. These findings underline the usefulness of this novel in vitro model for studying the cellular and molecular mechanisms of the neuro-cutaneous effects of ciguatoxins, which may assist with identifying potential therapeutics for ciguatera fish poisoning. Copyright © 2015 Elsevier Ltd. All rights reserved.

  20. Valorization of kitchen biowaste for ethanol production via simultaneous saccharification and fermentation using co-cultures of the yeasts Saccharomyces cerevisiae and Pichia stipitis.

    Science.gov (United States)

    Ntaikou, Ioanna; Menis, Nikolaos; Alexandropoulou, Maria; Antonopoulou, Georgia; Lyberatos, Gerasimos

    2018-04-30

    The biotransformation of the pre-dried and shredded organic fraction of kitchen waste to ethanol was investigated, via co-cultures of the yeasts Saccharomyces cerevisiae and Pichia stipitis (Scheffersomyces stipitis). Preliminary experiments with synthetic media were performed, in order to investigate the effect of different operational parameters on the ethanol production efficiency of the co-culture. The control of the pH and the supplementation with organic nitrogen were shown to be key factors for the optimization of the process. Subsequently, the ethanol production efficiency from the waste was assessed via simultaneous saccharification and fermentation experiments. Different loadings of cellulolytic enzymes and mixtures of cellulolytic with amylolytic enzymatic blends were tested in order to enhance the substrate conversion efficiency. It was further shown that for solids loading up to 40% waste on dry mass basis, corresponding to 170 g.L -1 initial concentration of carbohydrates, no substrate inhibition occurred, and ethanol concentration up to 45 g.L -1 was achieved. Copyright © 2018 Elsevier Ltd. All rights reserved.

  1. APOEε4 increases trauma induced early apoptosis via reducing delayed rectifier K(+) currents in neuronal/glial co-cultures model.

    Science.gov (United States)

    Chen, Ligang; Sun, Xiaochuan; Jiang, Yong; Kuai, Li

    2015-06-10

    Traumatic brain injury (TBI) is a commonly encountered emergency and severe neurosurgical injury. Previous studies have shown that the presence of the apolipoprotein E (APOE) ε4 allele has adverse outcomes across the spectrum of TBI severity. Our objective was to evaluate the effects of APOE alleles on trauma induced early apoptosis via modification of delayed rectifier K(+) current (Ik(DR)) in neuronal/glial co-cultures model. An ex vivo neuronal/glial co-cultures model carrying individual APOE alleles (ε2, ε3, ε4) of mechanical injury was developed. Flow cytometry and patch clamp recording were performed to analyze the correlations among APOE genotypes, early apoptosis and Ik(DR). We found that APOEε4 increased early apoptosis at 24h (p<0.05) compared to the ones transfected with APOEε3 and APOEε2. Noticeably, APOEε4 significantly reduced the amplitude of the Ik(DR) at 24h compared to the APOEε3 and APOEε2 (p<0.05) which exacerbate Ca(2+) influx. This indicates a possible effect of APOEε4 on early apoptosis via inhibiting Ik(DR) following injury which may adversely affect the outcome of TBI. Copyright © 2015 Elsevier Inc. All rights reserved.

  2. Study on O2 generation and CO2 absorption capability of four co-cultured salad plants in an enclosed system

    Science.gov (United States)

    Guo, Shuangsheng; Ai, Weidang; Tang, Yongkang; Cheng, Quanyong; Shen, Yunze; Qin, Lifeng; Ma, Jialu; Zhu, Jingtao; Ren, Jin

    2014-06-01

    The ability to generate O2 and absorb CO2 of several co-cultured vegetable plants in an enclosed system was studied to provide theoretical reference for the future man-plant integrated tests. Four kinds of salad plants (Lactuca sativa L. var. Dasusheng, Lactuca sativa L. var. Youmaicai, Gynura bicolor and Cichorium endivia L.) were grown in the CELSS Integration Test Platform (CITP). The environmental factors including O2 and CO2 concentration were continuously monitored on-line and the plant biomass was measured at the end of the test. The changing rules of O2 and CO2 concentration in the system were basically understood and it was found that the O2 generated by the plants could satisfy the respiratory needs of 1.75 persons by calculation. It was also found that the plants could absorb the CO2 breathed out by 2 persons when the light intensity was raised to 550 mmol m-2 s-1 PPF. The results showed that the co-cultured plants hold good compatibility and excellent O2-generating and CO2-absorbing capability. They could also supply some fresh edible vegetable for a 2-person crew.

  3. 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

  4. Effects of Treatment with Bone Morphogenetic Protein 4 and Co-culture on Expression of Piwil2 Gene in Mouse Differentiated Embryonic Stem Cells

    Directory of Open Access Journals (Sweden)

    Mehdi Forouzandeh-Moghadam

    2009-01-01

    Full Text Available Background: Specific growth factors and feeder layers seem to have important roles in in vitroembryonic stem cells (ESCs differentiation. In this study,the effects of bone morphogenetic protein4 (BMP4 and mouse embryonic fibroblasts (MEFs co-culture system on germ cell differentiationfrom mouse ESCs were studied.Materials and Methods: Cell suspension was prepared from one-day-old embryoid body (EBand cultured for four days in DMEM medium containing 20% fetal bovine serum (FBS in thefollowing groups: simple culture (SC, simple culture with BMP4 (SCB, co-culture (CO-C andco-culture with BMP4 (CO-CB. Expression of piwi-like homolog 2 (Piwil2, the germ cell-specificgene, was evaluated in the different study groups by using quantitative real time polymerase chainreaction (RT-PCR. Testis was used as a positive control.Results: The maximum and minimum Piwil2 expression was observed in SC and SCB groups,respectively. A significant difference was observed in Piwil2 expression between SCB and otherstudy groups (p<0.05.Conclusion: The findings of this study showed that neither the addition of BMP4 in culture mediumnor the use of MEFs as a feeder layer have a positive effect on late germ cell induction from mouseESCs.

  5. Co-Culture with Human Osteoblasts and Exposure to Extremely Low Frequency Pulsed Electromagnetic Fields Improve Osteogenic Differentiation of Human Adipose-Derived Mesenchymal Stem Cells

    Directory of Open Access Journals (Sweden)

    Sabrina Ehnert

    2018-03-01

    Full Text Available Human adipose-derived mesenchymal stem cells (Ad-MSCs have been proposed as suitable option for cell-based therapies to support bone regeneration. In the bone environment, Ad-MSCs will receive stimuli from resident cells that may favor their osteogenic differentiation. There is recent evidence that this process can be further improved by extremely low frequency pulsed electromagnetic fields (ELF-PEMFs. Thus, the project aimed at (i investigating whether co-culture conditions of human osteoblasts (OBs and Ad-MSCs have an impact on their proliferation and osteogenic differentiation; (ii whether this effect can be further improved by repetitive exposure to two specific ELF-PEMFs (16 and 26 Hz; (iii and the effect of these ELF-PEMFs on human osteoclasts (OCs. Osteogenic differentiation was improved by co-culturing OBs and Ad-MSCs when compared to the individual mono-cultures. An OB to Ad-MSC ratio of 3:1 had best effects on total protein content, alkaline phosphatase (AP activity, and matrix mineralization. Osteogenic differentiation was further improved by both ELF-PEMFs investigated. Interestingly, only repetitive exposure to 26 Hz ELF-PEMF increased Trap5B activity in OCs. Considering this result, a treatment with gradually increasing frequency might be of interest, as the lower frequency (16 Hz could enhance bone formation, while the higher frequency (26 Hz could enhance bone remodeling.

  6. Ecological effects of co-culturing the sea cucumber Apostichopus japonicus with the Chinese white shrimp Fenneropenaeus chinensis in an earthen pond

    Science.gov (United States)

    Zhou, Shun; Ren, Yichao; Pearce, Christopher M.; Dong, Shuanglin; Tian, Xiangli; Gao, Qinfeng; Wang, Fang

    2017-01-01

    Using net enclosures in an earthen pond, we established three culture treatments with the sea cucumber Apostichopus japonicus and the Chinese white shrimp Fenneropenaeus chinensis: monoculture of sea cucumbers (C), monoculture of shrimp (S), and co-culture of the two species (CS). We measured levels of suspended particulate matter in the water column; total organic matter, total organic carbon, total nitrogen, and carbon/nitrogen ratios in both settling particles and the sediment; and chlorophyll a levels in the sediment. We then compared these variables between the three treatments. We also examined growth, survival, and yield of the two species in the different treatments. From June to September, the mean monthly suspended particulate matter sedimentation rates in the CS and S treatments were significantly ( Pshrimp reared in the two systems. The bioturbation of the sediment and fecal production of the shrimp likely supplied natural food for the sea cucumbers. Co-culture of the two species is a viable option for increasing yield per unit area, maximizing use of the water body, and diversifying crop production.

  7. Small airway epithelial cells exposure to printer-emitted engineered nanoparticles induces cellular effects on human microvascular endothelial cells in an alveolar-capillary co-culture model.

    Science.gov (United States)

    Sisler, Jennifer D; Pirela, Sandra V; Friend, Sherri; Farcas, Mariana; Schwegler-Berry, Diane; Shvedova, Anna; Castranova, Vincent; Demokritou, Philip; Qian, Yong

    2015-01-01

    The printer is one of the most common office equipment. Recently, it was reported that toner formulations for printing equipment constitute nano-enabled products (NEPs) and contain engineered nanomaterials (ENMs) that become airborne during printing. To date, insufficient research has been performed to understand the potential toxicological properties of printer-emitted particles (PEPs) with several studies using bulk toner particles as test particles. These studies demonstrated the ability of toner particles to cause chronic inflammation and fibrosis in animal models. However, the toxicological implications of inhalation exposures to ENMs emitted from laser printing equipment remain largely unknown. The present study investigates the toxicological effects of PEPs using an in vitro alveolar-capillary co-culture model with Human Small Airway Epithelial Cells (SAEC) and Human Microvascular Endothelial Cells (HMVEC). Our data demonstrate that direct exposure of SAEC to low concentrations of PEPs (0.5 and 1.0 µg/mL) caused morphological changes of actin remodeling and gap formations within the endothelial monolayer. Furthermore, increased production of reactive oxygen species (ROS) and angiogenesis were observed in the HMVEC. Analysis of cytokine and chemokine levels demonstrates that interleukin (IL)-6 and MCP-1 may play a major role in the cellular communication observed between SAEC and HMVEC and the resultant responses in HMVEC. These data indicate that PEPs at low, non-cytotoxic exposure levels are bioactive and affect cellular responses in an alveolar-capillary co-culture model, which raises concerns for potential adverse health effects.

  8. 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

  9. Live Faecalibacterium prausnitzii Does Not Enhance Epithelial Barrier Integrity in an Apical Anaerobic Co-Culture Model of the Large Intestine

    Directory of Open Access Journals (Sweden)

    Eva Maier

    2017-12-01

    Full Text Available Appropriate intestinal barrier maturation during infancy largely depends on colonization with commensal bacteria. Faecalibacterium prausnitzii is an abundant obligate anaerobe that colonizes during weaning and is thought to maintain colonic health throughout life. We previously showed that F. prausnitzii induced Toll-like receptor 2 (TLR2 activation, which is linked to enhanced tight junction formation. Therefore, we hypothesized that F. prausnitzii enhances barrier integrity, an important factor in appropriate intestinal barrier maturation. In order to test metabolically active bacteria, we used a novel apical anaerobic co-culture system that allows the survival of both obligate anaerobic bacteria and oxygen-requiring intestinal epithelial cells (Caco-2. The first aim was to optimize the culture medium to enable growth and active metabolism of F. prausnitzii while maintaining the viability and barrier integrity, as measured by trans-epithelial electrical resistance (TEER, of the Caco-2 cells. This was achieved by supplementing the apical cell culture medium with bacterial culture medium. The second aim was to test the effect of F. prausnitzii on TEER across Caco-2 cell layers. Live F. prausn