A new level set model for cell image segmentation

International Nuclear Information System (INIS)

Ma Jing-Feng; Chen Chun; Hou Kai; Bao Shang-Lian

2011-01-01

In this paper we first determine three phases of cell images: background, cytoplasm and nucleolus according to the general physical characteristics of cell images, and then develop a variational model, based on these characteristics, to segment nucleolus and cytoplasm from their relatively complicated backgrounds. In the meantime, the preprocessing obtained information of cell images using the OTSU algorithm is used to initialize the level set function in the model, which can speed up the segmentation and present satisfactory results in cell image processing. (cross-disciplinary physics and related areas of science and technology)

Restoration of Mitochondrial NAD+ Levels Delays Stem Cell Senescence and Facilitates Reprogramming of Aged Somatic Cells.

Science.gov (United States)

Son, Myung Jin; Kwon, Youjeong; Son, Taekwon; Cho, Yee Sook

2016-12-01

The fundamental tenet that aging is irreversible has been challenged by the development of reprogramming technology that can restore molecular and cellular age by reversing the progression of aging. The use of cells from aged individuals as sources for reprogramming or transplantation creates a major barrier in stem cell therapy with respect to cell quality and quantity. Here, we investigated the molecular features underlying senescence and rejuvenation during aged cell reprogramming and identified novel factors that can overcome age-associated barriers. Enzymes, such as nicotinamide nucleotide transhydrogenase (NNT) and nicotinamide mononucleotide adenylyltransferase 3 (NMNAT3), that control mitochondrial NAD + levels appear to be susceptible to aging. In aged cells, mitochondrial NAD + levels decrease, accompanied by reduced SIRT3 activity; these changes severely impede cell fate transition. However, in cells collected from aged p16 knockout mice, which exhibit delayed cellular senescence, no changes in NNT or NMNAT3 expression were found. Importantly, restoring mitochondrial NAD + levels by overexpressing NNT and NMNAT3 enhanced reprogramming efficiency of aged somatic cells and extended the lifespan of human mesenchymal stem cells by delaying replicative senescence. These results demonstrate that maintenance of mitochondrial NAD + levels is critical for reversing the mechanisms of aging and ensuring that cells collected from aged individuals are of high quality. Stem Cells 2016;34:2840-2851. © 2016 AlphaMed Press.

Nicotinamidase modulation of NAD+ biosynthesis and nicotinamide levels separately affect reproductive development and cell survival in C. elegans.

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Vrablik, Tracy L; Huang, Li; Lange, Stephanie E; Hanna-Rose, Wendy

2009-11-01

Nicotinamide adenine dinucleotide (NAD(+)) is a central molecule in cellular metabolism and an obligate co-substrate for NAD(+)-consuming enzymes, which regulate key biological processes such as longevity and stress responses. Although NAD(+) biosynthesis has been intensely studied, little analysis has been done in developmental models. We have uncovered novel developmental roles for a nicotinamidase (PNC), the first enzyme in the NAD(+) salvage pathway of invertebrates. Mutations in the Caenorhabditis elegans nicotinamidase PNC-1 cause developmental and functional defects in the reproductive system; the development of the gonad is delayed, four uterine cells die by necrosis and the mutant animals are egg-laying defective. The temporal delay in gonad development results from depletion of the salvage pathway product NAD(+), whereas the uv1 cell necrosis and egg-laying defects result from accumulation of the substrate nicotinamide. Thus, regulation of both substrate and product level is key to the biological activity of PNC-1. We also find that diet probably affects the levels of these metabolites, as it affects phenotypes. Finally, we identified a secreted isoform of PNC-1 and confirmed its extracellular localization and functional activity in vivo. We demonstrate that nicotinamide phosphoribosyltransferase (Nampt), the equivalent enzyme in nicotinamide recycling to NAD(+) in vertebrates, can functionally substitute for PNC-1. As Nampt is also secreted, we postulate an evolutionarily conserved extracellular role for NAD(+) biosynthetic enzymes during development and physiology.

Apoptosis induction in Jurkat cells and sCD95 levels in women's sera are related with the risk of developing cervical cancer

International Nuclear Information System (INIS)

Aguilar-Lemarroy, Adriana; Jave-Suarez, Luis F; Romero-Ramos, Jose E; Olimon-Andalon, Vicente; Hernandez-Flores, Georgina; Lerma-Diaz, Jose M; Ortiz-Lazareno, Pablo C; Morgan-Villela, Gilberto; Toro-Arreola, Susana del; Bravo-Cuellar, Alejandro

2008-01-01

Currently, there is clear evidence that apoptosis plays an important role in the development and progression of tumors. One of the best characterized apoptosis triggering systems is the CD95/Fas/APO-1 pathway; previous reports have demonstrated high levels of soluble CD95 (sCD95) in serum of patients with some types of cancer. Cervical cancer is the second most common cancer among women worldwide. As a first step in an attempt to design a minimally invasive test to predict the risk of developing cervical cancer in patients with precancerous lesions, we used a simple assay based on the capacity of human serum to induce apoptosis in Jurkat cells. We evaluated the relationship between sCD95 levels and the ability to induce apoptosis in Jurkat cells in cervical cancer patients and controls. Jurkat cells were exposed to serum from 63 women (20 healthy volunteers, 21 with cervical intraepithelial neoplasia grade I [CIN 1] and 22 with cervical-uterine carcinoma). The apoptotic rate was measured by flow cytometry using Annexin-V-Fluos and Propidium Iodide as markers. Serum levels of sCD95 and soluble CD95 ligand (sCD95L) were measured by ELISA kits. We found that serum from almost all healthy women induced apoptosis in Jurkat cells, while only fifty percent of the sera from women with CIN 1 induced cell death in Jurkat cells. Interestingly, only one serum sample from a patient with cervical-uterine cancer was able to induce apoptosis, the rest of the sera protected Jurkat cells from this killing. We were able to demonstrate that elimination of Jurkat cells was mediated by the CD95/Fas/Apo-1 apoptotic pathway. Furthermore, the serum levels of sCD95 measured by ELISA were significantly higher in women with cervical cancer. Our results demonstrate that there is a strong correlation between low levels of sCD95 in serum of normal women and higher apoptosis induction in Jurkat cells. We suggest that an analysis of the apoptotic rate induced by serum in Jurkat cells and the

Development of a photon-cell interactive monte carlo simulation for non-invasive measurement of blood glucose level by Raman spectroscopy.

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Sakota, Daisuke; Kosaka, Ryo; Nishida, Masahiro; Maruyama, Osamu

2015-01-01

Turbidity variation is one of the major limitations in Raman spectroscopy for quantifying blood components, such as glucose, non-invasively. To overcome this limitation, we have developed a Raman scattering simulation using a photon-cell interactive Monte Carlo (pciMC) model that tracks photon migration in both the extra- and intracellular spaces without relying on the macroscopic scattering phase function and anisotropy factor. The interaction of photons at the plasma-cell boundary of randomly oriented three-dimensionally biconcave red blood cells (RBCs) is modeled using geometric optics. The validity of the developed pciMCRaman was investigated by comparing simulation and experimental results of Raman spectroscopy of glucose level in a bovine blood sample. The scattering of the excitation laser at a wavelength of 785 nm was simulated considering the changes in the refractive index of the extracellular solution. Based on the excitation laser photon distribution within the blood, the Raman photon derived from the hemoglobin and glucose molecule at the Raman shift of 1140 cm(-1) = 862 nm was generated, and the photons reaching the detection area were counted. The simulation and experimental results showed good correlation. It is speculated that pciMCRaman can provide information about the ability and limitations of the measurement of blood glucose level.

Plant Systems Biology at the Single-Cell Level.

Science.gov (United States)

Libault, Marc; Pingault, Lise; Zogli, Prince; Schiefelbein, John

2017-11-01

Our understanding of plant biology is increasingly being built upon studies using 'omics and system biology approaches performed at the level of the entire plant, organ, or tissue. Although these approaches open new avenues to better understand plant biology, they suffer from the cellular complexity of the analyzed sample. Recent methodological advances now allow plant scientists to overcome this limitation and enable biological analyses of single-cells or single-cell-types. Coupled with the development of bioinformatics and functional genomics resources, these studies provide opportunities for high-resolution systems analyses of plant phenomena. In this review, we describe the recent advances, current challenges, and future directions in exploring the biology of single-cells and single-cell-types to enhance our understanding of plant biology as a system. Copyright © 2017 Elsevier Ltd. All rights reserved.

Sorting of cells of the same size, shape, and cell cycle stage for a single cell level assay without staining

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Yomo Tetsuya

2006-06-01

Full Text Available Abstract Background Single-cell level studies are being used increasingly to measure cell properties not directly observable in a cell population. High-performance data acquisition systems for such studies have, by necessity, developed in synchrony. However, improvements in sample purification techniques are also required to reveal new phenomena. Here we assessed a cell sorter as a sample-pretreatment tool for a single-cell level assay. A cell sorter is routinely used for selecting one type of cells from a heterogeneous mixture of cells using specific fluorescence labels. In this case, we wanted to select cells of exactly the same size, shape, and cell-cycle stage from a population, without using a specific fluorescence label. Results We used four light scatter parameters: the peak height and area of the forward scatter (FSheight and FSarea and side scatter (SSheight and SSarea. The rat pheochromocytoma PC12 cell line, a neuronal cell line, was used for all experiments. The living cells concentrated in the high FSarea and middle SSheight/SSarea fractions. Single cells without cell clumps were concentrated in the low SS and middle FS fractions, and in the higher FSheight/FSarea and SSheight/SSarea fractions. The cell populations from these viable, single-cell-rich fractions were divided into twelve subfractions based on their FSarea-SSarea profiles, for more detailed analysis. We found that SSarea was proportional to the cell volume and the FSarea correlated with cell roundness and elongation, as well as with the level of DNA in the cell. To test the method and to characterize the basic properties of the isolated single cells, sorted cells were cultured in separate wells. The cells in all subfractions survived, proliferated and differentiated normally, suggesting that there was no serious damage. The smallest, roundest, and smoothest cells had the highest viability. There was no correlation between proliferation and differentiation. NGF increases

Cell surface N-glycans influence the level of functional E-cadherin at the cell–cell border

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M. Kristen Hall

2014-01-01

Full Text Available E-cadherin is crucial for adhesion of cells to each other and thereby development and maintenance of tissue. While it is has been established that N-glycans inside the cell impact the level of E-cadherin at the cell surface of epithelial-derived cells, it is unclear whether N-glycans outside the cell control the clustering of E-cadherin at the cell–cell border. Here, we demonstrate reduction of N-glycans at the cell surface weakened the recruitment and retention of E-cadherin at the cell–cell border, and consequently reduced the strength of cell–cell interactions. We conclude that N-glycans at the cell surface are tightly linked to the placement of E-cadherin at the cell–cell border and thereby control E-cadherin mediated cell–cell adhesion.

Evaluation of mRNA expression levels and electrophysiological function of neuron-like cells derived from canine bone marrow stromal cells.

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Nakano, Rei; Edamura, Kazuya; Sugiya, Hiroshi; Narita, Takanori; Okabayashi, Ken; Moritomo, Tadaaki; Teshima, Kenji; Asano, Kazushi; Nakayama, Tomohiro

2013-10-01

To investigate the in vitro differentiation of canine bone marrow stromal cells (BMSCs) into functional, mature neurons. Bone marrow from 6 adult dogs. BMSCs were isolated from bone marrow and chemically induced to develop into neurons. The morphology of the BMSCs during neuronal induction was monitored, and immunocytochemical analyses for neuron markers were performed after the induction. Real-time PCR methods were used to evaluate the mRNA expression levels of markers for neural stem or progenitor cells, neurons, and ion channels, and western blotting was used to assess the expression of neuronal proteins before and after neuronal induction. The electrophysiological properties of the neuron-like cells induced from canine BMSCs were evaluated with fluorescent dye to monitor Ca(2)+ influx. Canine BMSCs developed a neuron-like morphology after neuronal induction. Immunocytochemical analysis revealed that these neuron-like cells were positive for neuron markers. After induction, the cells' mRNA expression levels of almost all neuron and ion channel markers increased, and the protein expression levels of nestin and neurofilament-L increased significantly. However, the neuron-like cells derived from canine BMSCs did not have the Ca(2)+ influx characteristic of spiking neurons. Although canine BMSCs had neuron-like morphological and biochemical properties after induction, they did not develop the electrophysiological characteristics of neurons. Thus, these results have suggested that canine BMSCs could have the capacity to differentiate into a neuronal lineage, but the differentiation protocol used may have been insufficient to induce development into functional neurons.

Changes of serum prolactin level in patients with oral cavity squamous cell carcinoma

International Nuclear Information System (INIS)

Zheng Jian; Li Hairu; Chen Yaming; Tang Guihong; Xu Yalan

2004-01-01

To investigate the change of serum prolactin (PRL) level in patients with oral cavity squamous cell carcinoma, serum PRL level in 79 normal person and 68 cases of patient s was measured by RIA. The result showed that serum PRL level was significantly higher in 26 patients (38.2%, 26/68) than that in the control (P 0.05) between the sex and region of lesion. The above results indicated that proportion of patient with oral cavity squamous cell carcinoma was hyperprolactinaemia and the change of PRL was related to the development in oral cavity squamous cell carcinoma. (authors)

Macrocyclic peptides decrease c-Myc protein levels and reduce prostate cancer cell growth.

Science.gov (United States)

Mukhopadhyay, Archana; Hanold, Laura E; Thayele Purayil, Hamsa; Gisemba, Solomon A; Senadheera, Sanjeewa N; Aldrich, Jane V

2017-08-03

The oncoprotein c-Myc is often overexpressed in cancer cells, and the stability of this protein has major significance in deciding the fate of a cell. Thus, targeting c-Myc levels is an attractive approach for developing therapeutic agents for cancer treatment. In this study, we report the anti-cancer activity of the macrocyclic peptides [D-Trp]CJ-15,208 (cyclo[Phe-D-Pro-Phe-D-Trp]) and the natural product CJ-15,208 (cyclo[Phe-D-Pro-Phe-Trp]). [D-Trp]CJ-15,208 reduced c-Myc protein levels in prostate cancer cells and decreased cell proliferation with IC 50 values ranging from 2.0 to 16 µM in multiple PC cell lines. [D-Trp]CJ-15,208 induced early and late apoptosis in PC-3 cells following 48 hours treatment, and growth arrest in the G2 cell cycle phase following both 24 and 48 hours treatment. Down regulation of c-Myc in PC-3 cells resulted in loss of sensitivity to [D-Trp]CJ-15,208 treatment, while overexpression of c-Myc in HEK-293 cells imparted sensitivity of these cells to [D-Trp]CJ-15,208 treatment. This macrocyclic tetrapeptide also regulated PP2A by reducing the levels of its phosphorylated form which regulates the stability of cellular c-Myc protein. Thus [D-Trp]CJ-15,208 represents a new lead compound for the potential development of an effective treatment of prostate cancer.

Measurement of Heme Synthesis Levels in Mammalian Cells.

Science.gov (United States)

Hooda, Jagmohan; Alam, Maksudul; Zhang, Li

2015-07-09

Heme serves as the prosthetic group for a wide variety of proteins known as hemoproteins, such as hemoglobin, myoglobin and cytochromes. It is involved in various molecular and cellular processes such as gene transcription, translation, cell differentiation and cell proliferation. The biosynthesis levels of heme vary across different tissues and cell types and is altered in diseased conditions such as anemia, neuropathy and cancer. This technique uses [4-(14)C] 5-aminolevulinic acid ([(14)C] 5-ALA), one of the early precursors in the heme biosynthesis pathway to measure the levels of heme synthesis in mammalian cells. This assay involves incubation of cells with [(14)C] 5-ALA followed by extraction of heme and measurement of the radioactivity incorporated into heme. This procedure is accurate and quick. This method measures the relative levels of heme biosynthesis rather than the total heme content. To demonstrate the use of this technique the levels of heme biosynthesis were measured in several mammalian cell lines.

Development of a single ion micro-irradiation facility for experimental radiobiology at cell level

International Nuclear Information System (INIS)

Barberet, Ph.

2003-10-01

A micro-irradiation device has been developed for radiobiology applications at the scale of the cell. This device is based on an upgrade of an existing micro-beam line that was already able to deliver a 1 to 3 MeV proton or alpha beam of low intensity and whose space resolution is lower than 1 micrometer in vacuum. The important part of this work has been the development of an irradiation stage designed to fit on the micro-probe and able to deliver ions in the air with an absolute accuracy of a few micrometers. A program has been set up to monitor the complete irradiation line in testing and in automatic irradiation operating phases. Simulation tools based on Monte-Carlo calculations have been validated through comparisons with experimental data particularly in the field of spatial resolution and of the number of ions delivered. The promising results show the possibility in a near future to use this tool to study the response of cells to very low irradiation doses down to the extreme limit of one ion per cell

Development of molten carbonate fuel cells for power generation

Science.gov (United States)

1980-04-01

The broad and comprehensive program included elements of system definition, cell and system modeling, cell component development, cell testing in pure and contaminated environments, and the first stages of technology scale up. Single cells, with active areas of 45 sq cm and 582 sq cm, were operated at 650 C and improved to state of the art levels through the development of cell design concepts and improved electrolyte and electrode components. Performance was shown to degrade by the presence of fuel contaminants, such as sulfur and chlorine, and due to changes in electrode structure. Using conventional hot press fabrication techniques, electrolyte structures up to 20" x 20" were fabricated. Promising approaches were developed for nonhot pressed electrolyte structure fabrication and a promising electrolyte matrix material was identified. This program formed the basis for a long range effort to realize the benefits of molten carbonate fuel cell power plants.

LG Solid Oxide Fuel Cell (SOFC) Model Development

Energy Technology Data Exchange (ETDEWEB)

Haberman, Ben [LG Fuel Cell Systems Inc., North Canton, OH (United States); Martinez-Baca, Carlos [LG Fuel Cell Systems Inc., North Canton, OH (United States); Rush, Greg [LG Fuel Cell Systems Inc., North Canton, OH (United States)

2013-05-31

This report presents a summary of the work performed by LG Fuel Cell Systems Inc. during the project LG Solid Oxide Fuel Cell (SOFC) Model Development (DOE Award Number: DE-FE0000773) which commenced on October 1, 2009 and was completed on March 31, 2013. The aim of this project is for LG Fuel Cell Systems Inc. (formerly known as Rolls-Royce Fuel Cell Systems (US) Inc.) (LGFCS) to develop a multi-physics solid oxide fuel cell (SOFC) computer code (MPC) for performance calculations of the LGFCS fuel cell structure to support fuel cell product design and development. A summary of the initial stages of the project is provided which describes the MPC requirements that were developed and the selection of a candidate code, STAR-CCM+ (CD-adapco). This is followed by a detailed description of the subsequent work program including code enhancement and model verification and validation activities. Details of the code enhancements that were implemented to facilitate MPC SOFC simulations are provided along with a description of the models that were built using the MPC and validated against experimental data. The modeling work described in this report represents a level of calculation detail that has not been previously available within LGFCS.

Midkine inhibits inducible regulatory T cell differentiation by suppressing the development of tolerogenic dendritic cells.

Science.gov (United States)

Sonobe, Yoshifumi; Li, Hua; Jin, Shijie; Kishida, Satoshi; Kadomatsu, Kenji; Takeuchi, Hideyuki; Mizuno, Tetsuya; Suzumura, Akio

2012-03-15

Midkine (MK), a heparin-binding growth factor, reportedly contributes to inflammatory diseases, including Crohn's disease and rheumatoid arthritis. We previously showed that MK aggravates experimental autoimmune encephalomyelitis (EAE) by decreasing regulatory CD4(+)CD25(+)Foxp3(+) T cells (Tregs), a population that regulates the development of autoimmune responses, although the precise mechanism remains uncertain. In this article, we show that MK produced in inflammatory conditions suppresses the development of tolerogenic dendritic cells (DCregs), which drive the development of inducible Treg. MK suppressed DCreg-mediated expansion of the CD4(+)CD25(+)Foxp3(+) Treg population. DCregs expressed significantly higher levels of CD45RB and produced significantly less IL-12 compared with conventional dendritic cells. However, MK downregulated CD45RB expression and induced IL-12 production by reducing phosphorylated STAT3 levels via src homology region 2 domain-containing phosphatase-2 in DCreg. Inhibiting MK activity with anti-MK RNA aptamers, which bind to the targeted protein to suppress the function of the protein, increased the numbers of CD11c(low)CD45RB(+) dendritic cells and Tregs in the draining lymph nodes and suppressed the severity of EAE, an animal model of multiple sclerosis. Our results also demonstrated that MK was produced by inflammatory cells, in particular, CD4(+) T cells under inflammatory conditions. Taken together, these results suggest that MK aggravates EAE by suppressing DCreg development, thereby impairing the Treg population. Thus, MK is a promising therapeutic target for various autoimmune diseases.

Taurine Induces Proliferation of Neural Stem Cells and Synapse Development in the Developing Mouse Brain

Science.gov (United States)

Shivaraj, Mattu Chetana; Marcy, Guillaume; Low, Guoliang; Ryu, Jae Ryun; Zhao, Xianfeng; Rosales, Francisco J.; Goh, Eyleen L. K.

2012-01-01

Taurine is a sulfur-containing amino acid present in high concentrations in mammalian tissues. It has been implicated in several processes involving brain development and neurotransmission. However, the role of taurine in hippocampal neurogenesis during brain development is still unknown. Here we show that taurine regulates neural progenitor cell (NPC) proliferation in the dentate gyrus of the developing brain as well as in cultured early postnatal (P5) hippocampal progenitor cells and hippocampal slices derived from P5 mice brains. Taurine increased cell proliferation without having a significant effect on neural differentiation both in cultured P5 NPCs as well as cultured hippocampal slices and in vivo. Expression level analysis of synaptic proteins revealed that taurine increases the expression of Synapsin 1 and PSD 95. We also found that taurine stimulates the phosphorylation of ERK1/2 indicating a possible role of the ERK pathway in mediating the changes that we observed, especially in proliferation. Taken together, our results demonstrate a role for taurine in neural stem/progenitor cell proliferation in developing brain and suggest the involvement of the ERK1/2 pathways in mediating these actions. Our study also shows that taurine influences the levels of proteins associated with synapse development. This is the first evidence showing the effect of taurine on early postnatal neuronal development using a combination of in vitro, ex-vivo and in vivo systems. PMID:22916184

Dose levels in the hot cells area ININ

International Nuclear Information System (INIS)

Torre, J. De la; Ramirez, J.M.; Solis, M.L.

2004-01-01

The Laboratory of Hot Cells (LCC) located in the National Institute of Nuclear Research (ININ) is an institution, it is an area where radioactive material is managed with different activity values, in function of its original design for 10,000 curies of Co-60. Managing this materials in the installation, it implies to measure and to analyze the dose levels that the POE will receive as well as the implementation of appropriate measures of radiological protection and radiological safety, so that that is completed settled down by the concept ALARA. In this work they are carried out mensurations of the levels of the dose to receive for the POE when managing radionuclides with maximum activities that can be allowed in function of the current conditions of the cells and an evaluation of the obtained results is made comparing them with the effective international norms as well as the application of the program of surveillance and radiological protection implemented for the development of the works that are carry out in the installation. (Author)

Low-level lasers alter mRNA levels from traditional reference genes used in breast cancer cells

Science.gov (United States)

Teixeira, A. F.; Canuto, K. S.; Rodrigues, J. A.; Fonseca, A. S.; Mencalha, A. L.

2017-07-01

Cancer is among the leading causes of mortality worldwide, increasing the importance of treatment development. Low-level lasers are used in several diseases, but some concerns remains on cancers. Reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) is a technique used to understand cellular behavior through quantification of mRNA levels. Output data from target genes are commonly relative to a reference that cannot vary according to treatment. This study evaluated reference genes levels from MDA-MB-231 cells exposed to red or infrared lasers at different fluences. Cultures were exposed to red and infrared lasers, incubated (4 h, 37 °C), total RNA was extracted and cDNA synthesis was performed to evaluate mRNA levels from ACTB, GUSB and TRFC genes by RT-qPCR. Specific amplification was verified by melting curves and agarose gel electrophoresis. RefFinder enabled data analysis by geNorm, NormFinder and BestKeeper. Specific amplifications were obtained and, although mRNA levels from ACTB, GUSB or TRFC genes presented no significant variation through traditional statistical analysis, Excel-based tools revealed that the use of these reference genes are dependent of laser characteristics. Our data showed that exposure to low-level red and infrared lasers at different fluences alter the mRNA levels from ACTB, GUSB and TRFC in MDA-MB-231 cells.

Development of an ultralow-light-level luminescence image analysis system for dynamic measurements of transcriptional activity in living and migrating cells.

Science.gov (United States)

Maire, E; Lelièvre, E; Brau, D; Lyons, A; Woodward, M; Fafeur, V; Vandenbunder, B

2000-04-10

We have developed an approach to study in single living epithelial cells both cell migration and transcriptional activation, which was evidenced by the detection of luminescence emission from cells transfected with luciferase reporter vectors. The image acquisition chain consists of an epifluorescence inverted microscope, connected to an ultralow-light-level photon-counting camera and an image-acquisition card associated to specialized image analysis software running on a PC computer. Using a simple method based on a thin calibrated light source, the image acquisition chain has been optimized following comparisons of the performance of microscopy objectives and photon-counting cameras designed to observe luminescence. This setup allows us to measure by image analysis the luminescent light emitted by individual cells stably expressing a luciferase reporter vector. The sensitivity of the camera was adjusted to a high value, which required the use of a segmentation algorithm to eliminate the background noise. Following mathematical morphology treatments, kinetic changes of luminescent sources were analyzed and then correlated with the distance and speed of migration. Our results highlight the usefulness of our image acquisition chain and mathematical morphology software to quantify the kinetics of luminescence changes in migrating cells.

Joint level-set and spatio-temporal motion detection for cell segmentation.

Science.gov (United States)

Boukari, Fatima; Makrogiannis, Sokratis

2016-08-10

Cell segmentation is a critical step for quantification and monitoring of cell cycle progression, cell migration, and growth control to investigate cellular immune response, embryonic development, tumorigenesis, and drug effects on live cells in time-lapse microscopy images. In this study, we propose a joint spatio-temporal diffusion and region-based level-set optimization approach for moving cell segmentation. Moving regions are initially detected in each set of three consecutive sequence images by numerically solving a system of coupled spatio-temporal partial differential equations. In order to standardize intensities of each frame, we apply a histogram transformation approach to match the pixel intensities of each processed frame with an intensity distribution model learned from all frames of the sequence during the training stage. After the spatio-temporal diffusion stage is completed, we compute the edge map by nonparametric density estimation using Parzen kernels. This process is followed by watershed-based segmentation and moving cell detection. We use this result as an initial level-set function to evolve the cell boundaries, refine the delineation, and optimize the final segmentation result. We applied this method to several datasets of fluorescence microscopy images with varying levels of difficulty with respect to cell density, resolution, contrast, and signal-to-noise ratio. We compared the results with those produced by Chan and Vese segmentation, a temporally linked level-set technique, and nonlinear diffusion-based segmentation. We validated all segmentation techniques against reference masks provided by the international Cell Tracking Challenge consortium. The proposed approach delineated cells with an average Dice similarity coefficient of 89 % over a variety of simulated and real fluorescent image sequences. It yielded average improvements of 11 % in segmentation accuracy compared to both strictly spatial and temporally linked Chan

Evolutionary insights into postembryonic development of adult intestinal stem cells

Directory of Open Access Journals (Sweden)

Ishizuya-Oka Atsuko

2011-11-01

Full Text Available Abstract In the adult vertebrate intestine, multi-potent stem cells continuously generate all of the epithelial cells throughout the adulthood. While it has long been known that the frog intestine is formed via the development of adult intestinal stem cells during thyroid hormone (TH-dependent metamorphosis, the basic structure of the adult intestine is formed by birth in mammals and it is unclear if the subsequent maturation of the intestine involves any changes in the intestinal stem cells. Two recent papers showing that B lymphocyte-induced maturation protein 1 (Blimp1 regulates postnatal epithelial stem cell reprogramming during mouse intestinal maturation support the model that adult intestinal stem cells are developed during postembryonic development in mammals, in a TH-dependent process similar to intestinal remodeling during amphibian metamorphosis. Since the formation of the adult intestine in both mammals and amphibians is closely associated with the adaptation from aquatic to terrestrial life during the peak of endogenous TH levels, the molecular mechanisms by which the adult stem cells are developed are likely evolutionally conserved.

Reconstructing human pancreatic differentiation by mapping specific cell populations during development

DEFF Research Database (Denmark)

Ramond, Cyrille; Glaser, Nicolas; Berthault, Claire

2017-01-01

. Endocrine maturation progresses by up-regulating SUSD2 and lowering ECAD levels. Finally, in vitro differentiation of pancreatic endocrine cells derived from human pluripotent stem cells mimics key in vivo events. Our work paves the way to extend our understanding of the origin of mature human pancreatic......Information remains scarce on human development compared to animal models. Here, we reconstructed human fetal pancreatic differentiation using cell surface markers. We demonstrate that at 7weeks of development, the glycoprotein 2 (GP2) marks a multipotent cell population that will differentiate...... cell types and how such lineage decisions are regulated....

Influences of lamin A levels on induction of pluripotent stem cells

Directory of Open Access Journals (Sweden)

Bingfeng Zuo

2012-09-01

Lamin A is an inner nuclear membrane protein that maintains nuclear structure integrity, is involved in transcription, DNA damage response and genomic stability, and also links to cell differentiation, senescence, premature aging and associated diseases. Induced pluripotent stem (iPS cells have been successfully generated from various types of cells and used to model human diseases. It remains unclear whether levels of lamin A influence reprogramming of somatic cells to pluripotent states during iPS induction. Consistently, lamin A is expressed more in differentiated than in relatively undifferentiated somatic cells, and increases in expression levels with age. Somatic cells with various expression levels of lamin A differ in their dynamics and efficiency during iPS cell induction. Cells with higher levels of lamin A show slower reprogramming and decreased efficiency to iPS cells. Furthermore, depletion of lamin A by transient shRNA accelerates iPS cell induction from fibroblasts. Reduced levels of lamin A are associated with increased expression of pluripotent genes Oct4 and Nanog, and telomerase genes Tert and Terc. On the contrary, overexpression of lamin A retards somatic cell reprogramming to iPS-like colony formation. Our data suggest that levels of lamin A influence reprogramming of somatic cells to pluripotent stem cells and that artificial silencing of lamin A facilitates iPS cell induction. These findings may have implications in enhancing rejuvenation of senescent or older cells by iPS technology and manipulating lamin A levels.

Signaling profiling at the single-cell level identifies a distinct signaling signature in murine hematopoietic stem cells.

Science.gov (United States)

Du, Juan; Wang, Jinyong; Kong, Guangyao; Jiang, Jing; Zhang, Jingfang; Liu, Yangang; Tong, Wei; Zhang, Jing

2012-07-01

Hematopoietic stem cell (HSC) function is tightly regulated by cytokine signaling. Although phospho-flow cytometry allows us to study signaling in defined populations of cells, there has been tremendous hurdle to carry out this study in rare HSCs due to unrecoverable critical HSC markers, low HSC number, and poor cell recovery rate. Here, we overcame these difficulties and developed a "HSC phospho-flow" method to analyze cytokine signaling in murine HSCs at the single-cell level and compare HSC signaling profile to that of multipotent progenitors (MPPs), a cell type immediately downstream of HSCs, and commonly used Lin(-) cKit(+) cells (LK cells, enriched for myeloid progenitors). We chose to study signaling evoked from three representative cytokines, stem cell factor (SCF) and thrombopoietin (TPO) that are essential for HSC function and granulocyte macrophage-colony-stimulating factor (GM-CSF) that is dispensable for HSCs. HSCs display a distinct TPO and GM-CSF signaling signature from MPPs and LK cells, which highly correlates with receptor surface expression. In contrast, although majority of LK cells express lower levels of cKit than HSCs and MPPs, SCF-evoked ERK1/2 activation in LK cells shows a significantly increased magnitude for a prolonged period. These results suggest that specific cellular context plays a more important role than receptor surface expression in SCF signaling. Our study of HSC signaling at the homeostasis stage paves the way to investigate signaling changes in HSCs under conditions of stress, aging, and hematopoietic diseases. Copyright © 2012 AlphaMed Press.

Modular Approach for Continuous Cell-Level Balancing to Improve Performance of Large Battery Packs: Preprint

Energy Technology Data Exchange (ETDEWEB)

Muneed ur Rehman, M.; Evzelman, M.; Hathaway, K.; Zane, R.; Plett, G. L.; Smith, K.; Wood, E.; Maksimovic, D.

2014-10-01

Energy storage systems require battery cell balancing circuits to avoid divergence of cell state of charge (SOC). A modular approach based on distributed continuous cell-level control is presented that extends the balancing function to higher level pack performance objectives such as improving power capability and increasing pack lifetime. This is achieved by adding DC-DC converters in parallel with cells and using state estimation and control to autonomously bias individual cell SOC and SOC range, forcing healthier cells to be cycled deeper than weaker cells. The result is a pack with improved degradation characteristics and extended lifetime. The modular architecture and control concepts are developed and hardware results are demonstrated for a 91.2-Wh battery pack consisting of four series Li-ion battery cells and four dual active bridge (DAB) bypass DC-DC converters.

Assessing T cell differentiation at the single-cell level

NARCIS (Netherlands)

Gerlach, Carmen

2012-01-01

This thesis describes the development and use of a novel technology for single-cell fate mapping, called cellular barcoding. With this technology, unique and heritable genetic tags (barcodes) are introduced into naïve T cells. Using cellular barcoding, we investigated I) how different

Tools for Genomic and Transcriptomic Analysis of Microbes at Single-Cell Level

Directory of Open Access Journals (Sweden)

Zixi Chen

2017-09-01

Full Text Available Microbiologists traditionally study population rather than individual cells, as it is generally assumed that the status of individual cells will be similar to that observed in the population. However, the recent studies have shown that the individual behavior of each single cell could be quite different from that of the whole population, suggesting the importance of extending traditional microbiology studies to single-cell level. With recent technological advances, such as flow cytometry, next-generation sequencing (NGS, and microspectroscopy, single-cell microbiology has greatly enhanced the understanding of individuality and heterogeneity of microbes in many biological systems. Notably, the application of multiple ‘omics’ in single-cell analysis has shed light on how individual cells perceive, respond, and adapt to the environment, how heterogeneity arises under external stress and finally determines the fate of the whole population, and how microbes survive under natural conditions. As single-cell analysis involves no axenic cultivation of target microorganism, it has also been demonstrated as a valuable tool for dissecting the microbial ‘dark matter.’ In this review, current state-of-the-art tools and methods for genomic and transcriptomic analysis of microbes at single-cell level were critically summarized, including single-cell isolation methods and experimental strategies of single-cell analysis with NGS. In addition, perspectives on the future trends of technology development in the field of single-cell analysis was also presented.

Programmed Cell Death and Caspase Functions During Neural Development.

Science.gov (United States)

Yamaguchi, Yoshifumi; Miura, Masayuki

2015-01-01

Programmed cell death (PCD) is a fundamental component of nervous system development. PCD serves as the mechanism for quantitative matching of the number of projecting neurons and their target cells through direct competition for neurotrophic factors in the vertebrate peripheral nervous system. In addition, PCD plays roles in regulating neural cell numbers, canceling developmental errors or noise, and tissue remodeling processes. These findings are mainly derived from genetic studies that prevent cells from dying by apoptosis, which is a major form of PCD and is executed by activation of evolutionarily conserved cysteine protease caspases. Recent studies suggest that caspase activation can be coordinated in time and space at multiple levels, which might underlie nonapoptotic roles of caspases in neural development in addition to apoptotic roles. © 2015 Elsevier Inc. All rights reserved.

Development of nanosensors for studying intracellular phosphate levels

DEFF Research Database (Denmark)

Gu, Hong

-time monitoring of Pi metabolism in living cells, providing a new tool for fluxomics (measurement of metabolic flux), analysis of pathophysiology or changes of Pi during cell activity. Transformation of plants with FLIPs had resulted in only low expression levels. As an alternative a protein transduction domain......Abstract Inorganic phosphate (Pi) is an essential macronutrient that plays a central role in metabolism and signal transduction in plants. Uptake, compartmentation and transport are important players of cellular Pi homeostasis; however, methods to determine the cellular phosphate concentration...... of a substrate-binding protein linked to two fluorescent reporter proteins. Substrate binding changes the conformation of the nanosensor and, hence, the efficiency of fluorescence resonance energy transfer (FRET) between the reporter proteins. The aim of the present project was to develop nanosensors for Pi...

Mass Cytometry for Detection of Silver at the Bacterial Single Cell Level

Directory of Open Access Journals (Sweden)

Yuting Guo

2017-07-01

Full Text Available Background: Mass cytometry (Cytometry by Time of Flight, CyTOF allows single-cell characterization on the basis of specific metal-based cell markers. In addition, other metals in the mass range such as silver can be detected per cell. Bacteria are known to be sensible to silver and a protocol was developed to measure both the number of affected cells per population and the quantities of silver per cell.Methods: For mass cytometry ruthenium red was used as a marker for all cells of a population while parallel application of cisplatin discriminated live from dead cells. Silver quantities per cell and frequencies of silver containing cells in a population were measured by mass cytometry. In addition, live/dead subpopulations were analyzed by flow cytometry and distinguished by cell sorting based on ruthenium red and propidium iodide double staining. Verification of the cells’ silver load was performed on the bulk level by using ICP-MS in combination with cell sorting. The protocol was developed by conveying both, fast and non-growing Pseudomonas putida cells as test organisms.Results: A workflow for labeling bacteria in order to be analyzed by mass cytometry was developed. Three different parameters were tested: ruthenium red provided counts for all bacterial cells in a population while consecutively applied cisplatin marked the frequency of dead cells. Apparent population heterogeneity was detected by different frequencies of silver containing cells. Silver quantities per cell were also well measurable. Generally, AgNP-10 treatment caused higher frequencies of dead cells, higher frequencies of silver containing cells and higher per-cell silver quantities. Due to an assumed chemical equilibrium of free and bound silver ions live and dead cells were associated with silver in equal quantities and this preferably during exponential growth. With ICP-MS up to 1.5 fg silver per bacterial cell were detected.Conclusion: An effective mass cytometry

Multiweek cell culture project for use in upper-level biology laboratories.

Science.gov (United States)

Marion, Rebecca E; Gardner, Grant E; Parks, Lisa D

2012-06-01

This article describes a laboratory protocol for a multiweek project piloted in a new upper-level biology laboratory (BIO 426) using cell culture techniques. Human embryonic kidney-293 cells were used, and several culture media and supplements were identified for students to design their own experiments. Treatments included amino acids, EGF, caffeine, epinephrine, heavy metals, and FBS. Students researched primary literature to determine their experimental variables, made their own solutions, and treated their cells over a period of 2 wk. Before this, a sterile technique laboratory was developed to teach students how to work with the cells and minimize contamination. Students designed their experiments, mixed their solutions, seeded their cells, and treated them with their control and experimental media. Students had the choice of manipulating a number of variables, including incubation times, exposure to treatment media, and temperature. At the end of the experiment, students observed the effects of their treatment, harvested and dyed their cells, counted relative cell numbers in control and treatment flasks, and determined the ratio of living to dead cells using a hemocytometer. At the conclusion of the experiment, students presented their findings in a poster presentation. This laboratory can be expanded or adapted to include additional cell lines and treatments. The ability to design and implement their own experiments has been shown to increase student engagement in the biology-related laboratory activities as well as develop the critical thinking skills needed for independent research.

Control of cyclin C levels during development of Dictyostelium.

Directory of Open Access Journals (Sweden)

David M Greene

2010-05-01

Full Text Available Cdk8 and its partner cyclin C form part of the mediator complex which links the basal transcription machinery to regulatory proteins. The pair are required for correct regulation of a subset of genes and have been implicated in control of development in a number of organisms including the social amoeba Dictyostelium discoideum. When feeding, Dictyostelium amoebae are unicellular but upon starvation they aggregate to form a multicellular structure which develops into a fruiting body containing spores. Cells in which the gene encoding Cdk8 has been deleted fail to enter aggregates due to a failure of early gene expression.We have monitored the expression levels of cyclin C protein during development and find levels decrease after the multicellular mound is formed. This decrease is triggered by extracellular cAMP that, in turn, is working in part through an increase in intracellular cAMP. The loss of cyclin C is coincident with a reduction in the association of Cdk8 with a high molecular weight complex in the nucleus. Overexpression of cyclin C and Cdk8 lead to an increased rate of early development, consistent with the levels being rate limiting.Overall these results show that both cyclin C and Cdk8 are regulated during development in response to extracellular signals and the levels of these proteins are important in controlling the timing of developmental processes. These findings have important implications for the role of these proteins in controlling development, suggesting that they are targets for developmental signals to regulate gene expression.

Early postnatal docosahexaenoic acid levels and improved preterm brain development

OpenAIRE

Tam, Emily W.Y.; Chau, Vann; Barkovich, A. James; Ferriero, Donna M.; Miller, Steven P.; Rogers, Elizabeth E.; Grunau, Ruth E.; Synnes, Anne R.; Xu, Duan; Foong, Justin; Brant, Rollin; Innis, Sheila M.

2016-01-01

Background Preterm birth has a dramatic impact on polyunsaturated fatty acid exposures for the developing brain. This study examined the association between postnatal fatty acid levels and measures of brain injury and development, as well as outcomes. Methods A cohort of 60 preterm newborns (24?32 weeks GA) was assessed using early and near-term MRI studies. Red blood cell fatty acid composition was analyzed coordinated with each scan. Outcome at a mean of 33 months corrected age was assessed...

Photoacoustic bio-quantification of graphene based nanomaterials at a single cell level (Conference Presentation)

Science.gov (United States)

Nedosekin, Dmitry A.; Nolan, Jacqueline; Biris, Alexandru S.; Zharov, Vladimir P.

2017-03-01

Arkansas Nanomedicine Center at the University of Arkansas for Medical Sciences in collaboration with other Arkansas Universities and the FDA-based National Center of Toxicological Research in Jefferson, AR is developing novel techniques for rapid quantification of graphene-based nanomaterials (GBNs) in various biological samples. All-carbon GBNs have wide range of potential applications in industry, agriculture, food processing and medicine; however, quantification of GBNs is difficult in carbon reach biological tissues. The accurate quantification of GBNs is essential for research on material toxicity and the development of GBNs-based drug delivery platforms. We have developed microscopy and cytometry platforms for detection and quantification of GBNs in single cells, tissue and blood samples using photoacoustic contrast of GBNs. We demonstrated PA quantification of individual graphene uptake by single cells. High-resolution PA microscopy provided mapping of GBN distribution within live cells to establish correlation with intracellular toxic phenomena using apoptotic and necrotic assays. This new methodology and corresponding technical platform provide the insight on possible toxicological risks of GBNs at singe cells levels. In addition, in vivo PA image flow cytometry demonstrated the capability to monitor of GBNs pharmacokinetics in mouse model and to map the resulting biodistribution of GBNs in mouse tissues. The integrated PA platform provided an unprecedented sensitivity toward GBNs and allowed to enhance conventional toxicology research by providing a direct correlation between uptake of GBNs at a single cell level and cell viability status.

Hippocampal development in the rat: cytogenesis and morphogenesis examined with autoradiography and low-level x-irradiation

International Nuclear Information System (INIS)

Bayer, S.A.; Altman, J.

1974-01-01

The cytogenesis and morphogenesis of the rat hippocampus was examined with the techniques of 3 H-thymidine autoradiography, cell pyknosis produced by low-level x-irradiation, and quantitative histology. The procedure of progressively delayed cumulative labelling was used for autoradiography. Groups of rats were injected with four successive daily doses of 3 H-thymidine during non-overlapping periods ranging from birth to day 19. They were killed at 60 days of age, and the percentage of labelled cells was determined. Cell pyknosis in Ammon's horn reaches a maximal level prenatally and declines rapidly during the early postnatal period. Cell pyknosis in the dentate gyrus reaches its highest level during the second postnatal week and declines gradually with some radiosensitive cells still present in the adult. Immature granule cells are also at their highest level during the second postnatal week, while mature granule cells gradually accumulate to attain asymptotic levels at around two months of age. The alignment of the pyramidal cells to form the characteristic curvature of Ammon's horn occurs shortly after pyramidal cell cytogenesis is completed. Mechanisms for the morphological development of the dentate gyrus along with a consideration of the possible migratory route of granule cell precursors are discussed. (U.S.)

Classification of Normal and Apoptotic Cells from Fluorescence Microscopy Images Using Generalized Polynomial Chaos and Level Set Function.

Science.gov (United States)

Du, Yuncheng; Budman, Hector M; Duever, Thomas A

2016-06-01

Accurate automated quantitative analysis of living cells based on fluorescence microscopy images can be very useful for fast evaluation of experimental outcomes and cell culture protocols. In this work, an algorithm is developed for fast differentiation of normal and apoptotic viable Chinese hamster ovary (CHO) cells. For effective segmentation of cell images, a stochastic segmentation algorithm is developed by combining a generalized polynomial chaos expansion with a level set function-based segmentation algorithm. This approach provides a probabilistic description of the segmented cellular regions along the boundary, from which it is possible to calculate morphological changes related to apoptosis, i.e., the curvature and length of a cell's boundary. These features are then used as inputs to a support vector machine (SVM) classifier that is trained to distinguish between normal and apoptotic viable states of CHO cell images. The use of morphological features obtained from the stochastic level set segmentation of cell images in combination with the trained SVM classifier is more efficient in terms of differentiation accuracy as compared with the original deterministic level set method.

Single cell transcriptome profiling of developing chick retinal cells.

Science.gov (United States)

Laboissonniere, Lauren A; Martin, Gregory M; Goetz, Jillian J; Bi, Ran; Pope, Brock; Weinand, Kallie; Ellson, Laura; Fru, Diane; Lee, Miranda; Wester, Andrea K; Liu, Peng; Trimarchi, Jeffrey M

2017-08-15

The vertebrate retina is a specialized photosensitive tissue comprised of six neuronal and one glial cell types, each of which develops in prescribed proportions at overlapping timepoints from a common progenitor pool. While each of these cells has a specific function contributing to proper vision in the mature animal, their differential representation in the retina as well as the presence of distinctive cellular subtypes makes identifying the transcriptomic signatures that lead to each retinal cell's fate determination and development challenging. We have analyzed transcriptomes from individual cells isolated from the chick retina throughout retinogenesis. While we focused our efforts on the retinal ganglion cells, our transcriptomes of developing chick cells also contained representation from multiple retinal cell types, including photoreceptors and interneurons at different stages of development. Most interesting was the identification of transcriptomes from individual mixed lineage progenitor cells in the chick as these cells offer a window into the cell fate decision-making process. Taken together, these data sets will enable us to uncover the most critical genes acting in the steps of cell fate determination and early differentiation of various retinal cell types. © 2017 Wiley Periodicals, Inc.

Ezh2 represses the basal cell lineage during lung endoderm development.

Science.gov (United States)

Snitow, Melinda E; Li, Shanru; Morley, Michael P; Rathi, Komal; Lu, Min Min; Kadzik, Rachel S; Stewart, Kathleen M; Morrisey, Edward E

2015-01-01

The development of the lung epithelium is regulated in a stepwise fashion to generate numerous differentiated and stem cell lineages in the adult lung. How these different lineages are generated in a spatially and temporally restricted fashion remains poorly understood, although epigenetic regulation probably plays an important role. We show that the Polycomb repressive complex 2 component Ezh2 is highly expressed in early lung development but is gradually downregulated by late gestation. Deletion of Ezh2 in early lung endoderm progenitors leads to the ectopic and premature appearance of Trp63+ basal cells that extend the entire length of the airway. Loss of Ezh2 also leads to reduced secretory cell differentiation. In their place, morphologically similar cells develop that express a subset of basal cell genes, including keratin 5, but no longer express high levels of either Trp63 or of standard secretory cell markers. This suggests that Ezh2 regulates the phenotypic switch between basal cells and secretory cells. Together, these findings show that Ezh2 restricts the basal cell lineage during normal lung endoderm development to allow the proper patterning of epithelial lineages during lung formation. © 2015. Published by The Company of Biologists Ltd.

Cell cycle in egg cell and its progression during zygotic development in rice.

Science.gov (United States)

Sukawa, Yumiko; Okamoto, Takashi

2018-03-01

Rice egg is arrested at G1 phase probably by OsKRP2. After fusion with sperm, karyogamy, OsWEE1-mediated parental DNA integrity in zygote nucleus, zygote progresses cell cycle to produce two-celled embryo. In angiosperms, female and male gametes exist in gametophytes after the complementation of meiosis and the progression of nuclear/cell division of the haploid cell. Within the embryo sac, the egg cell is specially differentiated for fertilization and subsequent embryogenesis, and cellular programs for embryonic development, such as restarting the cell cycle and de novo gene expression, are halted. There is only limited knowledge about how the cell cycle in egg cells restarts toward zygotic division, although the conversion of the cell cycle from a quiescent and arrested state to an active state is the most evident transition of cell status from egg cell to zygote. This is partly due to the difficulty in direct access and analysis of egg cells, zygotes and early embryos, which are deeply embedded in ovaries. In this study, precise relative DNA amounts in the nuclei of egg cells, developing zygotes and cells of early embryos were measured, and the cell cycle of a rice egg cell was estimated as the G1 phase with a 1C DNA level. In addition, increases in DNA content in zygote nuclei via karyogamy and DNA replication were also detectable according to progression of the cell cycle. In addition, expression profiles for cell cycle-related genes in egg cells and zygotes were also addressed, and it was suggested that OsKRP2 and OsWEE1 function in the inhibition of cell cycle progression in egg cells and in checkpoint of parental DNA integrity in zygote nucleus, respectively.

'Fluorescent Cell Chip' for immunotoxicity testing: Development of the c-fos expression reporter cell lines

International Nuclear Information System (INIS)

Trzaska, Dominika; Zembek, Patrycja; Olszewski, Maciej; Adamczewska, Violetta; Ulleras, Erik; Dastych, JarosIaw

2005-01-01

The Fluorescent Cell Chip for in vitro immunotoxicity testing employs cell lines derived from lymphocytes, mast cells, and monocytes-macrophages transfected with various EGFP cytokine reporter gene constructs. While cytokine expression is a valid endpoint for in vitro immunotoxicity screening, additional marker for the immediate-early response gene expression level could be of interest for further development and refinement of the Fluorescent Cell Chip. We have used BW.5147.3 murine thymoma transfected with c-fos reporter constructs to obtain reporter cell lines expressing ECFP under the control of murine c-fos promoter. These cells upon serum withdrawal and readdition and incubation with heavy metal compounds showed paralleled induction of c-Fos expression as evidenced by Real-Time PCR and ECFP fluorescence as evidenced by computer-supported fluorescence microscopy. In conclusion, we developed fluorescent reporter cell lines that could be employed in a simple and time-efficient screening assay for possible action of chemicals on c-Fos expression in lymphocytes. The evaluation of usefulness of these cells for the Fluorescent Cell Chip-based detection of immunotoxicity will require additional testing with a larger number of chemicals

[Development of a Fluorescence Probe for Live Cell Imaging].

Science.gov (United States)

Shibata, Aya

2017-01-01

 Probes that detect specific biological materials are indispensable tools for deepening our understanding of various cellular phenomena. In live cell imaging, the probe must emit fluorescence only when a specific substance is detected. In this paper, we introduce a new probe we developed for live cell imaging. Glutathione S-transferase (GST) activity is higher in tumor cells than in normal cells and is involved in the development of resistance to various anticancer drugs. We previously reported the development of a general strategy for the synthesis of probes for detection of GST enzymes, including fluorogenic, bioluminogenic, and 19 F-NMR probes. Arylsulfonyl groups were used as caging groups during probe design. The fluorogenic probes were successfully used to quantitate very low levels of GST activity in cell extracts and were also successfully applied to the imaging of microsomal MGST1 activity in living cells. The bioluminogenic and 19 F-NMR probes were able to detect GST activity in Escherichia coli cells. Oligonucleotide-templated reactions are powerful tools for nucleic acid sensing. This strategy exploits the target strand as a template for two functionalized probes and provides a simple molecular mechanism for multiple turnover reactions. We developed a nucleophilic aromatic substitution reaction-triggered fluorescent probe. The probe completed its reaction within 30 s of initiation and amplified the fluorescence signal from 0.5 pM target oligonucleotide by 1500 fold under isothermal conditions. Additionally, we applied the oligonucleotide-templated reaction for molecular releasing and peptide detection.

A microfluidic-based lid device for conventional cell culture dishes to automatically control oxygen level.

Science.gov (United States)

Lee, Seung Yeob; Yang, Sung

2018-04-25

Most conventional hypoxic cell culture systems undergo reoxygenation during experimental manipulations, resulting in undesirable effects including the reduction of cell viability. A lid device was developed herein for conventional cell culture dishes to resolve this limitation. The integration of multilayered microfluidic channels inside a thin membrane was designed to prevent the reoxygenation caused by reagent infusion and automatically control the oxygen level. The experimental data clearly show the reducibility of the dissolved oxygen in the infusing reagent and the controllability of the oxygen level inside the dish. The feasibility of the device for hypoxia studies was confirmed by HIF-1α experiments. Therefore, the device could be used as a compact and convenient hypoxic cell culture system to prevent reoxygenation-related issues.

Acceptable contamination levels in solar grade silicon: From feedstock to solar cell

International Nuclear Information System (INIS)

Hofstetter, J.; Lelievre, J.F.; Canizo, C.; Luque, A. del

2009-01-01

Ultimately, alternative ways of silicon purification for photovoltaic applications are developed and applied. There is an ongoing debate about what are the acceptable contamination levels within the purified silicon feedstock to specify the material as solar grade silicon. Applying a simple model and making some additional assumptions, we calculate the acceptable contamination levels of different characteristic impurities for each fabrication step of a typical industrial mc-Si solar cell. The acceptable impurity concentrations within the finished solar cell are calculated for SRH recombination exclusively and under low injection conditions. It is assumed that during solar cell fabrication impurity concentrations are only altered by a gettering step. During the crystallization process, impurity segregation at the solid-liquid interface and at extended defects are taken into account. Finally, the initial contamination levels allowed within the feedstock are deduced. The acceptable concentration of iron in the finished solar cell is determined to be 9.7x10 -3 ppma whereas the concentration in the silicon feedstock can be as high as 12.5 ppma. In comparison, the titanium concentration admitted in the solar cell is calculated to be 2.7x10 -4 ppma and the allowed concentration of 2.2x10 -2 ppma in the feedstock is only two orders of magnitude higher. Finally, it is shown theoretically and experimentally that slow cooling rates can lead to a decrease of the interstitial Fe concentration and thus relax the purity requirements in the feedstock.

Chronically Elevated Levels of Short-Chain Fatty Acids Induce T Cell-Mediated Ureteritis and Hydronephrosis.

Science.gov (United States)

Park, Jeongho; Goergen, Craig J; HogenEsch, Harm; Kim, Chang H

2016-03-01

Short-chain fatty acids (SCFAs) are major products of gut microbial fermentation and profoundly affect host health and disease. SCFAs generate IL-10(+) regulatory T cells, which may promote immune tolerance. However, SCFAs can also induce Th1 and Th17 cells upon immunological challenges and, therefore, also have the potential to induce inflammatory responses. Because of the seemingly paradoxical SCFA activities in regulating T cells, we investigated, in depth, the impact of elevated SCFA levels on T cells and tissue inflammation in mice. Orally administered SCFAs induced effector (Th1 and Th17) and regulatory T cells in ureter and kidney tissues, and they induced T cell-mediated ureteritis, leading to kidney hydronephrosis (hereafter called acetate-induced renal disease, or C2RD). Kidney hydronephrosis in C2RD was caused by ureteral obstruction, which was, in turn, induced by SCFA-induced inflammation in the ureteropelvic junction and proximal ureter. Oral administration of all major SCFAs, such as acetate, propionate, and butyrate, induced the disease. We found that C2RD development is dependent on mammalian target of rapamycin activation, T cell-derived inflammatory cytokines such as IFN-γ and IL-17, and gut microbiota. Young or male animals were more susceptible than old or female animals, respectively. However, SCFA receptor (GPR41 or GPR43) deficiency did not affect C2RD development. Thus, SCFAs, when systemically administered at levels higher than physiological levels, cause dysregulated T cell responses and tissue inflammation in the renal system. The results provide insights into the immunological and pathological effects of chronically elevated SCFAs. Copyright © 2016 by The American Association of Immunologists, Inc.

Development of level 2 processor for the readout of TMC

International Nuclear Information System (INIS)

Arai, Y.; Ikeno, M.; Murata, T.; Sudo, F.; Emura, T.

1995-01-01

We have developed a prototype 8-bit processor for the level 2 data processing for the Time Memory Cell (TMC). The first prototype processor successfully runs with 18 MHz clock. The operation of same clock frequency as TMC (30 MHz) will be easily achieved with simple modifications. Although the processor is very primitive one but shows its powerful performance and flexibility. To realize the compact TMC/L2P (Level 2 Processor) system, it is better to include the microcode memory within the chip. Encoding logic of the microcode must be included to reduce the microcode memory in this case. (J.P.N.)

Low-level lasers on microRNA and uncoupling protein 2 mRNA levels in human breast cancer cells

Science.gov (United States)

Canuto, K. S.; Teixeira, A. F.; Rodrigues, J. A.; Paoli, F.; Nogueira, E. M.; Mencalha, A. L.; Fonseca, A. S.

2017-06-01

MicroRNA is short non-coding RNA and is a mediator of post-transcriptional regulation of gene expression. In addition, uncoupling proteins (UCPs) regulate thermogenesis, metabolic and energy balance, and decrease reactive oxygen species production. Both microRNA and UCP2 expression can be altered in cancer cells. At low power, laser wavelength, frequency, fluence and emission mode deternube photobiological responses, which are the basis of low-level laser therapy. There are few studies on miRNA and UCP mRNA levels after low-level laser exposure on cancer cells. In this work, we evaluate the micrRNA (mir-106b and mir-15a) and UCP2 mRNA levels in human breast cancer cells exposed to low-level lasers. MDA-MB-231 human breast cancer cells were exposed to low-level red and infrared lasers, total RNA was extracted for cDNA synthesis and mRNA levels by real time quantitative polymerase chain reaction were evaluated. Data show that mir-106b and mir-15a relative levels are not altered, but UCP2 mRNA relative levels are increased in MDA-MB-231 human breast cancer cells exposed to low-level red and infrared lasers at fluences used in therapeutic protocols.

Development of Thymic Epithelial Cells

DEFF Research Database (Denmark)

Ulyanchenko, Svetlana; Vaidya, Harsh J.; O'Neill, Kathy E.

2016-01-01

The thymus is the primary lymphoid organ in which the T cell repertoire is generated. The complex cellularity of this organ is uniquely designed to facilitate T cell development: defects in thymus development or function can cause immunodeficiencies ranging from the absence of T cell-mediated imm......The thymus is the primary lymphoid organ in which the T cell repertoire is generated. The complex cellularity of this organ is uniquely designed to facilitate T cell development: defects in thymus development or function can cause immunodeficiencies ranging from the absence of T cell......-mediated immunity to broad-spectrum autoimmune disease. Peak thymus size and output occurs early in life, after which the thymus undergoes a natural process of involution. This results in the progressive loss of functional thymus tissue and correspondingly in decreased production of new naïve T cells with age...... - contributing to the diminished capacity of the aged immune system to adequately respond to new antigenic challenge. Age-related thymic involutions, together with the thymic involutions associated with cytotoxic therapies (e.g., radio- or chemotherapy), have raised interest in development of clinically useful...

High fat diet triggers cell cycle arrest and excessive apoptosis of granulosa cells during the follicular development

Energy Technology Data Exchange (ETDEWEB)

Wu, Yanqing; Zhang, Zhenghong; Liao, Xinghui; Wang, Zhengchao, E-mail: zcwang@fjnu.edu.cn

2015-10-23

The regulatory mechanism of granulosa cells (GCs) proliferation during the follicular development is complicated and multifactorial, which is essential for the oocyte growth and normal ovarian functions. To investigate the role of high fat diet (HFD) on the proliferation of GCs, 4-week old female mice were fed with HFD or normal control diet (NC) for 15 weeks or 20 weeks and then detected the expression level of some regulatory molecules of cell cycle and apoptosis. The abnormal ovarian morphology was observed at 20 weeks. Further mechanistic studies indicated that HFD induced-obesity caused elevated apoptotic levels in GCs of the ovaries in a time-dependent manner. Moreover, cell cycle progress was also impacted after HFD fed. The cell cycle inhibitors, p27{sup Kip1} and p21{sup Cip1}, were significantly induced in the ovaries from the mice in HFD group when compared with that in the ovaries from the mice in NC group. Subsequently, the expression levels of Cyclin D1, D3 and CDK4 were also significantly influenced in the ovaries from the mice fed with HFD in a time-dependent manner. The present results suggested that HFD induced-obesity may trigger cell cycle arrest and excessive apoptosis of GCs, causing the abnormal follicular development and ovarian function failure. - Highlights: • HFD induced-obesity leads to abnormal ovarian morphology. • HFD induced-obesity triggers excessive apoptosis in the ovary. • HFD induced-obesity up-regulates cell cycle inhibitors p21{sup Cip1} and p27{sup Kip1} in the ovary. • HFD induced-obesity causes cell cycle arrest in the ovary.

Gene-specific correlation of RNA and protein levels in human cells and tissues

DEFF Research Database (Denmark)

Edfors, Fredrik; Danielsson, Frida; Hallström, Björn M.

2016-01-01

An important issue for molecular biology is to establish whether transcript levels of a given gene can be used as proxies for the corresponding protein levels. Here, we have developed a targeted proteomics approach for a set of human non-secreted proteins based on parallel reaction monitoring...... to measure, at steady-state conditions, absolute protein copy numbers across human tissues and cell lines and compared these levels with the corresponding mRNA levels using transcriptomics. The study shows that the transcript and protein levels do not correlate well unless a gene-specific RNA-to-protein (RTP...

Inverse associations between obesity indicators and thymic T-cell production levels in aging atomic-bomb survivors.

Directory of Open Access Journals (Sweden)

Kengo Yoshida

Full Text Available Reduction of the naive T-cell population represents a deteriorating state in the immune system that occurs with advancing age. In animal model studies, obesity compromises the T-cell immune system as a result of enhanced adipogenesis in primary lymphoid organs and systemic inflammation. In this study, to test the hypothesis that obesity may contribute to the aging of human T-cell immunity, a thousand atomic-bomb survivors were examined for obesity status and ability to produce naive T cells, i.e., T-cell receptor excision circle (TREC numbers in CD4 and CD8 T cells. The number of TRECs showed a strong positive correlation with naive T cell numbers, and lower TREC numbers were associated with higher age. We found that the TREC number was inversely associated with levels of obesity indicators (BMI, hemoglobin A1c and serum CRP levels. Development of type-2 diabetes and fatty liver was also associated with lower TREC numbers. This population study suggests that obesity with enhanced inflammation is involved in aging of the human T-cell immune system. Given the fact that obesity increases the risk of numerous age-related diseases, attenuated immune competence is a possible mechanistic link between obesity and disease development among the elderly.

Inverse associations between obesity indicators and thymic T-cell production levels in aging atomic-bomb survivors.

Science.gov (United States)

Yoshida, Kengo; Nakashima, Eiji; Kubo, Yoshiko; Yamaoka, Mika; Kajimura, Junko; Kyoizumi, Seishi; Hayashi, Tomonori; Ohishi, Waka; Kusunoki, Yoichiro

2014-01-01

Reduction of the naive T-cell population represents a deteriorating state in the immune system that occurs with advancing age. In animal model studies, obesity compromises the T-cell immune system as a result of enhanced adipogenesis in primary lymphoid organs and systemic inflammation. In this study, to test the hypothesis that obesity may contribute to the aging of human T-cell immunity, a thousand atomic-bomb survivors were examined for obesity status and ability to produce naive T cells, i.e., T-cell receptor excision circle (TREC) numbers in CD4 and CD8 T cells. The number of TRECs showed a strong positive correlation with naive T cell numbers, and lower TREC numbers were associated with higher age. We found that the TREC number was inversely associated with levels of obesity indicators (BMI, hemoglobin A1c) and serum CRP levels. Development of type-2 diabetes and fatty liver was also associated with lower TREC numbers. This population study suggests that obesity with enhanced inflammation is involved in aging of the human T-cell immune system. Given the fact that obesity increases the risk of numerous age-related diseases, attenuated immune competence is a possible mechanistic link between obesity and disease development among the elderly.

Hydrogen Fuel Cell development in Columbia (SC)

Energy Technology Data Exchange (ETDEWEB)

Reifsnider, Kenneth [Univ. of South Carolina, Columbia, SC (United States); Chen, Fanglin [Univ. of South Carolina, Columbia, SC (United States); Popov, Branko [Univ. of South Carolina, Columbia, SC (United States); Chao, Yuh [Univ. of South Carolina, Columbia, SC (United States); Xue, Xingjian [Univ. of South Carolina, Columbia, SC (United States)

2012-09-15

This is an update to the final report filed after the extension of this program to May of 2011. The activities of the present program contributed to the goals and objectives of the Fuel Cell element of the Hydrogen, Fuel Cells and Infrastructure Technologies Program of the Department of Energy through five sub-projects. Three of these projects have focused on PEM cells, addressing the creation of carbon-based metal-free catalysts, the development of durable seals, and an effort to understand contaminant adsorption/reaction/transport/performance relationships at low contaminant levels in PEM cells. Two programs addressed barriers in SOFCs; an effort to create a new symmetrical and direct hydrocarbon fuel SOFC designs with greatly increased durability, efficiency, and ease of manufacturing, and an effort to create a multiphysics engineering durability model based on electrochemical impedance spectroscopy interpretations that associate the micro-details of how a fuel cell is made and their history of (individual) use with specific prognosis for long term performance, resulting in attendant reductions in design, manufacturing, and maintenance costs and increases in reliability and durability.

Fuel Cell Manufacturing Research and Development | Hydrogen and Fuel Cells

Science.gov (United States)

| NREL Fuel Cell Manufacturing Research and Development Fuel Cell Manufacturing Research and Development NREL's fuel cell manufacturing R&D focuses on improving quality-inspection practices for high costs. A researcher monitoring web-line equipment in the Manufacturing Laboratory Many fuel cell

Paradoxical binding levels of vasoactive amines to cultured cerebral microvessel derived endothelial cells

International Nuclear Information System (INIS)

Robinson, R.A.; TenEyck, C.J.; Linthicum, D.S.; Hart, M.N.

1986-01-01

Vascular sensitization to vasoactive amines (VAA) may be critical for the development of experimental autoimmune encephalitis as well as other autoimmune diseases. Some inbred stains of mice such as SJL/J are particularly sensitive to the effects of VAA while others (BALB/c) are not. This study was performed to determine if the differing response to VAA in vivo is due to differing levels of binding of VAA to cultured brain endothelial (En) cells in vitro. Cells were isolated, grown to confluence, washed twice with binding buffer and incubated with either 3 H-histamine, 3 H-mepyramine or 3 H-5 hydroxytryptamine (5HT) for 1 hour at 37 0 C. Results showed that the BALB derived En cells specifically bound approximately twice as much mepyramine and three times as much 5-HT as the SJL derived En cells. The relative low binding of VAA to SJL En cells may reflect the extreme in vivo sensitivity that this mouse strain displays toward VAA. These seemingly paradoxical levels of VAA binding in the cultured cerebral endothelium may be due to genetic factors and may give insight into diseases that affect the blood brain barrier

Detection of the Level of Reactive Oxygen Species Induced by Ionizing Radiation in Cells

Energy Technology Data Exchange (ETDEWEB)

Kim, Jin Kyu; Chung, Dong Min; Kim, Jin-Hong [Korea Atomic Energy Research Institute, Daejeon (Korea, Republic of)

2015-05-15

By definition, the direct effect is referred to interaction between photon and DNA molecule, whereas the indirect effect is mediated by the reactive oxygen species (ROS) generated by radiolysis and subsequent reaction. It has been reported that ROS produced after exposure to IR can react with cellular materials such as DNA, proteins, carbohydrates and lipids. ROS is free radicals such as the superoxide anion, hydroxyl radicals and the non-radical hydrogen peroxide. Cells generate ROS during aerobic metabolism. Excessive production of ROS can lead to oxidative stress, genetic alteration and even cell death. It has been reported that ROS plays a critical role in radiation-induced cell injury. Thus, it is of great interest to determine the radiation-induced ROS level. Many kinds of methods to detect the level of ROS have been developed so far. There were random changes of fluorescence intensity in the treatment after irradiation. This result meant that this protocol was not appropriate for determination of radiation-induced ROS. On the other hand, the fluorescence intensity was increased in a dose-dependent manner when the cells were treated with the DCFH-DA solution before irradiation. Conclusions can be drawn from the experimental results of this study. In order to properly measure the ROS level in the cells exposed to ionizing radiation, the cells should be treated with the DCFH-DA solution before irradiation.

Connexin 43 Communication Channels in Follicular Dendritic Cell Development and in Follicular Lymphomas

Directory of Open Access Journals (Sweden)

Hajnalka Rajnai

2015-01-01

Full Text Available Follicular dendritic cells (FDC show homo- and heterocellular metabolic coupling through connexin 43 (Cx43 gap junctions and support B cell selection and maturation in germinal centers. In follicular lymphomas B cells escape apoptosis while FDC develop abnormally. Here we tested Cx43 channels in reactive FDC development and follicular lymphomas. In culture, the treatment of FDC-B cell clusters (resembling to “ex vivo” germinal centers with Gap27 peptide, mimicking the 2nd extracellular loop of Cx43 protein, significantly impaired FDC-B cell cluster formation and cell survival. In untreated cultures of intact clusters, cell proliferation showed a moderate reduction. In tissues, Cx43 protein levels run parallel with the density of FDC both in reactive germinal centers and in malformed follicles of follicular lymphomas and showed strong upregulation in newly generated and/or degrading bi-/multinuclear FDC of rudimentary processes. However, the inverse correlation between Cx43 expression and B cell proliferation seen in reactive germinal centers was not detected in follicular lymphomas. Furthermore, Cx43 levels were not associated with either lymphoma grade or bone marrow involvement. Our results suggest that Cx43 channels are critical in FDC and “ex vivo” germinal center development and in the persistence of FDC in follicular lymphomas but do not affect tumor progression.

System-level modeling and simulation of the cell culture microfluidic biochip ProCell

DEFF Research Database (Denmark)

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

2010-01-01

Microfluidic biochips offer a promising alternative to a conventional biochemical laboratory. There are two technologies for the microfluidic biochips: droplet-based and flow-based. In this paper we are interested in flow-based microfluidic biochips, where the liquid flows continuously through pre......-defined micro-channels using valves and pumps. We present an approach to the system-level modeling and simulation of a cell culture microfluidic biochip called ProCell, Programmable Cell Culture Chip. ProCell contains a cell culture chamber, which is envisioned to run 256 simultaneous experiments (viewed...

Development of Large-Format Lithium-Ion Cells with Silicon Anode and Low Flammable Electrolyte

Science.gov (United States)

Wu, James J.; Hernandez-Lugo, D. M.; Smart, M. C.; Ratnakumar, B. V.; Miller, T. B.; Lvovich, V. F.; Lytle, J. K.

2014-01-01

NASA is developing safe, high energy and high capacity lithium-ion cell designs and batteries for future missions under NASAs Advanced Space Power System (ASPS) project. Advanced cell components, such as high specific capacity silicon anodes and low-flammable electrolytes have been developed for improving the cell specific energy and enhancing safety. To advance the technology readiness level, we have developed large-format flight-type hermetically sealed battery cells by incorporating high capacity silicon anodes, commercially available lithium nickel, cobalt, aluminum oxide (NCA) cathodes, and low-flammable electrolytes. In this report, we will present the performance results of these various battery cells. In addition, we will also discuss the post-test cell analysis results as well.

Asymmetric cell division during T cell development controls downstream fate

Science.gov (United States)

Pham, Kim; Shimoni, Raz; Charnley, Mirren; Ludford-Menting, Mandy J.; Hawkins, Edwin D.; Ramsbottom, Kelly; Oliaro, Jane; Izon, David; Ting, Stephen B.; Reynolds, Joseph; Lythe, Grant; Molina-Paris, Carmen; Melichar, Heather; Robey, Ellen; Humbert, Patrick O.; Gu, Min

2015-01-01

During mammalian T cell development, the requirement for expansion of many individual T cell clones, rather than merely expansion of the entire T cell population, suggests a possible role for asymmetric cell division (ACD). We show that ACD of developing T cells controls cell fate through differential inheritance of cell fate determinants Numb and α-Adaptin. ACD occurs specifically during the β-selection stage of T cell development, and subsequent divisions are predominantly symmetric. ACD is controlled by interaction with stromal cells and chemokine receptor signaling and uses a conserved network of polarity regulators. The disruption of polarity by deletion of the polarity regulator, Scribble, or the altered inheritance of fate determinants impacts subsequent fate decisions to influence the numbers of DN4 cells arising after the β-selection checkpoint. These findings indicate that ACD enables the thymic microenvironment to orchestrate fate decisions related to differentiation and self-renewal. PMID:26370500

Age-dependent transition from cell-level to population-level control in murine intestinal homeostasis revealed by coalescence analysis.

Directory of Open Access Journals (Sweden)

Zheng Hu

Full Text Available In multi-cellular organisms, tissue homeostasis is maintained by an exquisite balance between stem cell proliferation and differentiation. This equilibrium can be achieved either at the single cell level (a.k.a. cell asymmetry, where stem cells follow strict asymmetric divisions, or the population level (a.k.a. population asymmetry, where gains and losses in individual stem cell lineages are randomly distributed, but the net effect is homeostasis. In the mature mouse intestinal crypt, previous evidence has revealed a pattern of population asymmetry through predominantly symmetric divisions of stem cells. In this work, using population genetic theory together with previously published crypt single-cell data obtained at different mouse life stages, we reveal a strikingly dynamic pattern of stem cell homeostatic control. We find that single-cell asymmetric divisions are gradually replaced by stochastic population-level asymmetry as the mouse matures to adulthood. This lifelong process has important developmental and evolutionary implications in understanding how adult tissues maintain their homeostasis integrating the trade-off between intrinsic and extrinsic regulations.

[The level of superoxide dismutase expression in primary and metastatic colorectal cancer cells in hypoxia and tissue normoxia].

Science.gov (United States)

Skrzycki, Michał; Czeczot, Hanna; Chrzanowska, Alicja; Otto-Ślusarczyk, Dagmara

2015-11-01

Superoxide oxidase (SOD) is a key antioxidant enzyme protecting cells against oxidative stress, which might induce cancerogenesis. In tumor cells SOD influences the level of the reactive oxygen species (ROS) allowing for survival and proliferation. High rate of cells proliferation in tumor leads to their temporary hypoxia due to lower rate of angiogenesis. Therefore during tumor development, cancer cells function in conditions of hypoxia or tissue normoxia. The aim of study was to evaluate of SOD isoenzymes (SOD1 and SOD2) expression level in cell lines of primary (SW 480) and metastatic (SW 620) colorectal cancer, cultured in hypoxia (1% oxygen), tissue normoxia (10% oxygen), and atmospheric normoxia (21% oxygen). Cells were cultured in MEM medium in different oxygen concentrations (1%, 10%, 21%) in hypoxic chamber with oxygenation regulator. The number of living cells in lines SW 480 and 620 was determined by trypan blue method. Expression of SOD1 and SOD2 at the mRNA level was determined by RT-PCR and PCR. In both studied cell lines (SW 480 and SW 620), the number of living cells (viability) was increased in hypoxia and atmospheric normoxia. The expression level of SOD1 and SOD2 in studied cell lines was different. The lowest level of expression of both SOD isoenzymes was observed in hypoxia. In conditions of atmospheric normoxia the expression level of SOD1 in SW480 cell line was increased, and similar in SW620 cell line comparing to tissue normoxia. Whereas the SOD2 expression level in atmospheric normoxia conditions in both cell lines was significantly increased. Observed differences were statistically significant (p ≤ 0,05). The profile of expression of SOD1 and SOD2 in cell lines SW480 and SW620 indicates differentiated response of tumor cells depending on access to oxygen. Low level of SOD isoenzymes expression in SW480 and SW620 cells in hypoxia indicates decreased production of ROS. Differences of SOD isoenzymes expression level in tissue normoxia

Mapping organism expression levels at cellular resolution in developing Drosophila

Science.gov (United States)

Knowles, David W.; Keranen, Soile; Biggin, Mark D.; Sudar, Damir

2002-05-01

The development of an animal embryo is orchestrated by a network of genetically determined, temporal and spatial gene expression patterns that determine the animals final form. To understand such networks, we are developing novel quantitative optical imaging techniques to map gene expression levels at cellular and sub-cellular resolution within pregastrula Drosophila. Embryos at different stages of development are labeled for total DNA and specific gene products using different fluorophors and imaged in 3D with confocal microscopy. Innovative steps have been made which allow the DNA-image to be automatically segmented to produce a morphological mask of the individual nuclear boundaries. For each stage of development an average morphology is chosen to which images from different embryo are compared. The morphological mask is then used to quantify gene-product on a per nuclei basis. What results is an atlas of the relative amount of the specific gene product expressed within the nucleus of every cell in the embryo at the various stages of development. We are creating a quantitative database of transcription factor and target gene expression patterns in wild-type and factor mutant embryos with single cell resolution. Our goal is to uncover the rules determining how patterns of gene expression are generated.

Elevated level of polysaccharides in a high level UV-B tolerant cell ...

African Journals Online (AJOL)

Jane

2011-04-26

Apr 26, 2011 ... A cell line of Bupleurum scorzonerifolium Willd with high level ... mechanisms to repair UV-induced damages via repairing ... for treatment or prevention of solar radiation. ..... working as both UV-B absorbing compounds and.

Hydrogen peroxide production is affected by oxygen levels in mammalian cell culture.

Science.gov (United States)

Maddalena, Lucas A; Selim, Shehab M; Fonseca, Joao; Messner, Holt; McGowan, Shannon; Stuart, Jeffrey A

2017-11-04

Although oxygen levels in the extracellular space of most mammalian tissues are just a few percent, under standard cell culture conditions they are not regulated and are often substantially higher. Some cellular sources of reactive oxygen species, like NADPH oxidase 4, are sensitive to oxygen levels in the range between 'normal' physiological (typically 1-5%) and standard cell culture (up to 18%). Hydrogen peroxide in particular participates in signal transduction pathways via protein redox modifications, so the potential increase in its production under standard cell culture conditions is important to understand. We measured the rates of cellular hydrogen peroxide production in some common cell lines, including C2C12, PC-3, HeLa, SH-SY5Y, MCF-7, and mouse embryonic fibroblasts (MEFs) maintained at 18% or 5% oxygen. In all instances the rate of hydrogen peroxide production by these cells was significantly greater at 18% oxygen than at 5%. The increase in hydrogen peroxide production at higher oxygen levels was either abolished or substantially reduced by treatment with GKT 137831, a selective inhibitor of NADPH oxidase subunits 1 and 4. These data indicate that oxygen levels experienced by cells in culture influence hydrogen peroxide production via NADPH oxidase 1/4, highlighting the importance of regulating oxygen levels in culture near physiological values. However, we measured pericellular oxygen levels adjacent to cell monolayers under a variety of conditions and with different cell lines and found that, particularly when growing at 5% incubator oxygen levels, pericellular oxygen was often lower and variable. Together, these observations indicate the importance, and difficulty, of regulating oxygen levels experienced by cells in culture. Copyright © 2017 Elsevier Inc. All rights reserved.

Effect of 60 Co gamma radiation on trehalose level of Saccharomyces boulardii cells

International Nuclear Information System (INIS)

Santos, Raquel Gouvea dos; Neves, Maria Jose; Duarte, Rinaldo; Nicoli, Jacques R.

1996-01-01

The medicine Floratil used for diarrhea treatment is composed by lyophilized yeast cells of Saccharomyces boulardii. The action mechanism of this product is unknown. The efficacy of S. boulardii depends on its viability. The increase in the trehalose level in S. cerevisiae cells submitted to a previous sub-lethal level heat shock results in resistance to a lethal shock. The aim of this work was to study weather the cell trehalose level is involved also in the resistance of gamma irradiation. It was noted that the yeast cells surviving to gamma irradiation had more trehalose level than control cells. Apparently there is a positive correlation between of trehalose level and gamma irradiation resistance of S.boulardii cells. (author)

Design and development of microbioreactors for long-term cell culture in controlled oxygen microenvironments.

Science.gov (United States)

Abaci, Hasan E; Devendra, Raghavendra; Smith, Quinton; Gerecht, Sharon; Drazer, German

2012-02-01

The ability to control the oxygen level to which cells are exposed in tissue culture experiments is crucial for many applications. Here, we design, develop and test a microbioreactor (MBR) for long-term cell culture studies with the capability to accurately control and continuously monitor the dissolved oxygen (DO) level in the cell microenvironment. In addition, the DO level can be controlled independently from other cues, such as the viscous shear-stress acting on the cells. We first analyze the transport of oxygen in the proposed device and determine the materials and dimensions that are compatible with uniform oxygen tension and low shear-stress at the cell level. The device is also designed to culture a statistically significant number of cells. We use fully transparent materials and the overall design of the device is compatible with live-cell imaging. The proposed system includes real-time read-out of actual DO levels, is simple to fabricate at low cost, and can be easily expanded to control the concentration of other microenvironmental solutes. We performed control experiments in the absence of cells to demonstrate that the MBR can be used to accurately modulate DO levels ranging from atmospheric level to 1%, both under no flow and perfusion conditions. We also demonstrate cancer cell attachment and viability within the MBR. The proposed MBR offers the unprecedented capability to perform on-line measurement and analysis of DO levels in the microenvironment of adherent cultures and to correlate them with various cellular responses.

High levels of circulating triiodothyronine induce plasma cell differentiation.

Science.gov (United States)

Bloise, Flavia Fonseca; Oliveira, Felipe Leite de; Nobrega, Alberto Félix; Vasconcellos, Rita; Cordeiro, Aline; Paiva, Luciana Souza de; Taub, Dennis D; Borojevic, Radovan; Pazos-Moura, Carmen Cabanelas; Mello-Coelho, Valéria de

2014-03-01

The effects of hyperthyroidism on B-cell physiology are still poorly known. In this study, we evaluated the influence of high-circulating levels of 3,5,3'-triiodothyronine (T3) on bone marrow, blood, and spleen B-cell subsets, more specifically on B-cell differentiation into plasma cells, in C57BL/6 mice receiving daily injections of T3 for 14 days. As analyzed by flow cytometry, T3-treated mice exhibited increased frequencies of pre-B and immature B-cells and decreased percentages of mature B-cells in the bone marrow, accompanied by an increased frequency of blood B-cells, splenic newly formed B-cells, and total CD19(+)B-cells. T3 administration also promoted an increase in the size and cellularity of the spleen as well as in the white pulp areas of the organ, as evidenced by histological analyses. In addition, a decreased frequency of splenic B220(+) cells correlating with an increased percentage of CD138(+) plasma cells was observed in the spleen and bone marrow of T3-treated mice. Using enzyme-linked immunospot assay, an increased number of splenic immunoglobulin-secreting B-cells from T3-treated mice was detected ex vivo. Similar results were observed in mice immunized with hen egg lysozyme and aluminum adjuvant alone or together with treatment with T3. In conclusion, we provide evidence that high-circulating levels of T3 stimulate plasma cytogenesis favoring an increase in plasma cells in the bone marrow, a long-lived plasma cell survival niche. These findings indicate that a stimulatory effect on plasma cell differentiation could occur in untreated patients with Graves' disease.

Fuel Cell Development for NASA's Human Exploration Program: Benchmarking with "The Hydrogen Economy"

Science.gov (United States)

Scott, John H.

2007-01-01

The theoretically high efficiency and low temperature operation of hydrogen-oxygen fuel cells has motivated them to be the subject of much study since their invention in the 19th Century, but their relatively high life cycle costs kept them as a "solution in search of a problem" for many years. The first problem for which fuel cells presented a truly cost effective solution was that of providing a power source for NASA's human spaceflight vehicles in the 1960 s. NASA thus invested, and continues to invest, in the development of fuel cell power plants for this application. This development program continues to place its highest priorities on requirements for minimum system mass and maximum durability and reliability. These priorities drive fuel cell power plant design decisions at all levels, even that of catalyst support. However, since the mid-1990's, prospective environmental regulations have driven increased governmental and industrial interest in "green power" and the "Hydrogen Economy." This has in turn stimulated greatly increased investment in fuel cell development for a variety of commercial applications. This investment is bringing about notable advances in fuel cell technology, but, as these development efforts place their highest priority on requirements for minimum life cycle cost and field safety, these advances are yielding design solutions quite different at almost every level from those needed for spacecraft applications. This environment thus presents both opportunities and challenges for NASA's Human Exploration Program

Levels of small molecules and enzymes in the mother cell compartment and the forespore of sporulating Bacillus megaterium.

Science.gov (United States)

Singh, R P; Setlow, B; Setlow, P

1977-06-01

We have determined the amounts of a number of small molecules and enzymes in the mother cell compartment and the developing forespore during sporulation of Bacillus megaterium. Significant amounts of adenosine 5'-triphosphate and reduced nicotinamide adenine dinucleotide were present in the forespore compartment before accumulation of dipicolinic acid (DPA), but these compounds disappeared as DPA was accumulated. 3-Phosphoglyceric acid (3-PGA) accumulated only within the developing forespore, beginning 1 to 2 h before DPA accumulation. Throughout its development the forespore contained constant levels of enzymes of both 3-PGA synthesis (phosphoglycerate kinase and glyceraldehyde-3-phosphate dehydrogenase) and 3-PGA utilization (phosphoglycerate mutase, enolase, and pyruvate kinase) at levels similar to those in the mother cell and the dormant spore. Despite the presence of enzymes for 3-PGA utilization, this compound was stable within isolated forespores. Two acid-soluble proteins (A and B proteins) also accumulated only in the forespore, beginning 1 to 2 h before DPA accumulation. At this time the specific protease involved in degradation of the A and B proteins during germination also appeared, but only in the forespore compartment. Nevertheless, the A and B proteins were stable within isolated forespores. Arginine and glutamic acid accumulated within the forespore in parallel with DPA accumulation. The forespore also contained the enzyme arginase at a level similar to that in the mother cell and a level of glutamic acid decarboxylase 2- to 25-fold higher than that in the mother cell, depending on when in sporulation the forespores were isolated. The specific activities of several other enzymes (protease active on hemoglobin, ornithine transcarbamylase, malate dehydrogenase, aconitase, and isocitrate dehydrogenase) in forespores were about 10% or less of the values in the mother cell. Aminopeptidase was present at similar levels in both compartments; threonine

Dysfunction of irradiated thymus for the development of helper T cells

International Nuclear Information System (INIS)

Amagai, T.; Kina, T.; Hirokawa, K.; Nishikawa, S.; Imanishi, J.; Katsura, Y.

1987-01-01

The development of cytotoxic T cells and helper T cells in an intact or irradiated thymus was investigated. C57BL/6 (H-2b, Thy-1.2) mice were whole body-irradiated, or were irradiated with shielding over either the thymus or right leg and tail, and were transferred with 1.5 X 10(7) bone marrow cells from B10.Thy-1.1 mice (H-2b, Thy-1.1). At various days after reconstitution, thymus cells from the recipient mice were harvested and a peanut agglutinin low-binding population was isolated. This population was further treated with anti-Thy-1.2 plus complement to remove host-derived cells and was assayed for the frequency of cytotoxic T cell precursors (CTLp) and for the activity of helper T cells (Th). In the thymus of thymus-shielded and irradiated mice, Th activity reached normal control level by day 25, whereas CTLp frequency remained at a very low level during these days. In the thymus of whole body-irradiated mice, generation of CTLp was highly accelerated while that of Th was retarded, the period required for reconstitution being 25 days and more than 42 days for CTLp and Th, respectively. Preferential development of CTLp was also seen in right leg- and tail-shielded (L-T-shielded) and irradiated recipients. Histological observation indicated that Ia+ nonlymphoid cells were well preserved in the thymus of thymus-shielded and irradiated recipients, whereas in L-T-shielded and irradiated recipients, such cells in the medulla were markedly reduced in number. These results suggest strongly that the generation of Th but not CTLp is dependent on radiosensitive thymic component(s), and that such components may represent Ia+ cells themselves in the medulla or some microenvironment related to Ia+ cells

Cell to cell signalling during vertebrate limb bud development

NARCIS (Netherlands)

Panman, Lia

2004-01-01

Communication between cells is essential during embryonic development. The vertebrate limb bud provides us a model to study signalling interactions between cells during patterning of embryonic tissues and organogenesis. In chapter 1 I give an introduction about limb bud development that is focussed

Differences in the mechanical properties of the developing cerebral cortical proliferative zone between mice and ferrets at both the tissue and single-cell levels

Directory of Open Access Journals (Sweden)

Arata Nagasaka

2016-11-01

Full Text Available Cell-producing events in developing tissues are mechanically dynamic throughout the cell cycle. In many epithelial systems, cells are apicobasally tall, with nuclei and somata that adopt different apicobasal positions because nuclei and somata move in a cell cycle–dependent manner. This movement is apical during G2 phase and basal during G1 phase, whereas mitosis occurs at the apical surface. These movements are collectively referred to as interkinetic nuclear migration, and such epithelia are called pseudostratified. The embryonic mammalian cerebral cortical neuroepithelium is a good model for highly pseudostratified epithelia, and we previously found differences between mice and ferrets in both horizontal cellular density (greater in ferrets and nuclear/somal movements (slower during G2 and faster during G1 in ferrets. These differences suggest that neuroepithelial cells alter their nucleokinetic behavior in response to physical factors that they encounter, which may form the basis for evolutionary transitions towards more abundant brain-cell production from mice to ferrets and primates. To address how mouse and ferret neuroepithelia may differ physically in a quantitative manner, we used atomic force microscopy to determine that the vertical stiffness of their apical surface is greater in ferrets (Young’s modulus = 1700 Pa than in mice (1400 Pa. We systematically analyzed factors underlying the apical-surface stiffness through experiments to pharmacologically inhibit actomyosin or microtubules and to examine recoiling behaviors of the apical surface upon laser ablation and also through electron microscopy to observe adherens junction. We found that although both actomyosin and microtubules are partly responsible for the apical-surface stiffness, the mousecells is

Human and great ape red blood cells differ in plasmalogen levels and composition.

Science.gov (United States)

Moser, Ann B; Steinberg, Steven J; Watkins, Paul A; Moser, Hugo W; Ramaswamy, Krishna; Siegmund, Kimberly D; Lee, D Rick; Ely, John J; Ryder, Oliver A; Hacia, Joseph G

2011-06-17

Plasmalogens are ether phospholipids required for normal mammalian developmental, physiological, and cognitive functions. They have been proposed to act as membrane antioxidants and reservoirs of polyunsaturated fatty acids as well as influence intracellular signaling and membrane dynamics. Plasmalogens are particularly enriched in cells and tissues of the human nervous, immune, and cardiovascular systems. Humans with severely reduced plasmalogen levels have reduced life spans, abnormal neurological development, skeletal dysplasia, impaired respiration, and cataracts. Plasmalogen deficiency is also found in the brain tissue of individuals with Alzheimer disease. In a human and great ape cohort, we measured the red blood cell (RBC) levels of the most abundant types of plasmalogens. Total RBC plasmalogen levels were lower in humans than bonobos, chimpanzees, and gorillas, but higher than orangutans. There were especially pronounced cross-species differences in the levels of plasmalogens with a C16:0 moiety at the sn-1 position. Humans on Western or vegan diets had comparable total RBC plasmalogen levels, but the latter group showed moderately higher levels of plasmalogens with a C18:1 moiety at the sn-1 position. We did not find robust sex-specific differences in human or chimpanzee RBC plasmalogen levels or composition. Furthermore, human and great ape skin fibroblasts showed only modest differences in peroxisomal plasmalogen biosynthetic activity. Human and chimpanzee microarray data indicated that genes involved in plasmalogen biosynthesis show cross-species differential expression in multiple tissues. We propose that the observed differences in human and great ape RBC plasmalogens are primarily caused by their rates of biosynthesis and/or turnover. Gene expression data raise the possibility that other human and great ape cells and tissues differ in plasmalogen levels. Based on the phenotypes of humans and rodents with plasmalogen disorders, we propose that cross

Reactive glia promote development of CD103+ CD69+ CD8+ T-cells through programmed cell death-ligand 1 (PD-L1).

Science.gov (United States)

Prasad, Sujata; Hu, Shuxian; Sheng, Wen S; Chauhan, Priyanka; Lokensgard, James R

2018-06-01

Previous work from our laboratory has demonstrated in vivo persistence of CD103 + CD69 + brain resident memory CD8 + T-cells (bT RM ) following viral infection, and that the PD-1: PD-L1 pathway promotes development of these T RM cells within the brain. Although glial cells express low basal levels of PD-L1, its expression is upregulated upon IFN-γ-treatment, and they have been shown to modulate antiviral T-cell effector responses through the PD-1: PD-L1 pathway. We performed flow cytometric analysis of cells from co-cultures of mixed glia and CD8 + T-cells obtained from wild type mice to investigate the role of glial cells in the development of bT RM . In this study, we show that interactions between reactive glia and anti-CD3 Ab-stimulated CD8 + T-cells promote development of CD103 + CD69 + CD8 + T-cells through engagement of the PD-1: PD-L1 pathway. These studies used co-cultures of primary murine glial cells obtained from WT animals along with CD8 + T-cells obtained from either WT or PD-1 KO mice. We found that αCD3 Ab-stimulated CD8 + T-cells from WT animals increased expression of CD103 and CD69 when co-cultured with primary murine glial cells. In contrast, significantly reduced expression of CD103 and CD69 was observed using CD8 + T-cells from PD-1 KO mice. We also observed that reactive glia promoted high levels of CD127, a marker of memory precursor effector cells (MPEC), on CD69 + CD8 + T-cells, which promotes development of T RM cells. Interestingly, results obtained using T-cells from PD-1 KO animals showed significantly reduced expression of CD127 on CD69 + CD8 + cells. Additionally, blocking of glial PD-L1 resulted in decreased expression of CD103, along with reduced CD127 on CD69 + CD8 + T-cells. Taken together, these results demonstrate a role for activated glia in promoting development of bT RM through the PD-1: PD-L1 pathway. © 2018 The Authors. Immunity, Inflammation and Disease Published by John Wiley & Sons Ltd.

Early dynamic fate changes in haemogenic endothelium characterized at the single-cell level

Science.gov (United States)

Swiers, Gemma; Baumann, Claudia; O'Rourke, John; Giannoulatou, Eleni; Taylor, Stephen; Joshi, Anagha; Moignard, Victoria; Pina, Cristina; Bee, Thomas; Kokkaliaris, Konstantinos D.; Yoshimoto, Momoko; Yoder, Mervin C.; Frampton, Jon; Schroeder, Timm; Enver, Tariq; Göttgens, Berthold; de Bruijn, Marella F. T. R.

2013-12-01

Haematopoietic stem cells (HSCs) are the founding cells of the adult haematopoietic system, born during ontogeny from a specialized subset of endothelium, the haemogenic endothelium (HE) via an endothelial-to-haematopoietic transition (EHT). Although recently imaged in real time, the underlying mechanism of EHT is still poorly understood. We have generated a Runx1 +23 enhancer-reporter transgenic mouse (23GFP) for the prospective isolation of HE throughout embryonic development. Here we perform functional analysis of over 1,800 and transcriptional analysis of 268 single 23GFP+ HE cells to explore the onset of EHT at the single-cell level. We show that initiation of the haematopoietic programme occurs in cells still embedded in the endothelial layer, and is accompanied by a previously unrecognized early loss of endothelial potential before HSCs emerge. Our data therefore provide important insights on the timeline of early haematopoietic commitment.

Performance analysis of three-dimensional-triple-level cell and two-dimensional-multi-level cell NAND flash hybrid solid-state drives

Science.gov (United States)

Sakaki, Yukiya; Yamada, Tomoaki; Matsui, Chihiro; Yamaga, Yusuke; Takeuchi, Ken

2018-04-01

In order to improve performance of solid-state drives (SSDs), hybrid SSDs have been proposed. Hybrid SSDs consist of more than two types of NAND flash memories or NAND flash memories and storage-class memories (SCMs). However, the cost of hybrid SSDs adopting SCMs is more expensive than that of NAND flash only SSDs because of the high bit cost of SCMs. This paper proposes unique hybrid SSDs with two-dimensional (2D) horizontal multi-level cell (MLC)/three-dimensional (3D) vertical triple-level cell (TLC) NAND flash memories to achieve higher cost-performance. The 2D-MLC/3D-TLC hybrid SSD achieves up to 31% higher performance than the conventional 2D-MLC/2D-TLC hybrid SSD. The factors of different performance between the proposed hybrid SSD and the conventional hybrid SSD are analyzed by changing its block size, read/write/erase latencies, and write unit of 3D-TLC NAND flash memory, by means of a transaction-level modeling simulator.

Strategic alliances for the development of fuel cell vehicles

Energy Technology Data Exchange (ETDEWEB)

Maruo, Kanehira [Goeteborg Univ. (Sweden). Section of Science and Technology Studies

1998-12-01

The aim of this paper is to explore and describe the current stage of fuel cell vehicle development in the world. One can write three possible future scenarios - an optimistic, a realistic, and a pessimistic scenario: - The optimistic scenario -- The Daimler/Ballard/Ford alliance continues to develop fuel cell stacks and fuel cell vehicle systems as eagerly as they have been doing in recent years. Daimler(/Chrysler)-Benz continues to present its Necar 4, Necar 5, and so on, as planned, and thus keeps Toyota and Honda under severe pressure. Toyota`s and Honda`s real motivation seems to be not to allow Daimler-Benz to be the first to market. Their investment in fuel cell technology will be very large. At the same time, governments and other stake-holders will quickly and in a timely fashion build up infrastructures. We will then see many fuel cell vehicles by 2004. A paradigm shift in automotive technology will have taken place. - The realistic scenario -- Fuel cell vehicles will reach the same level of development by 2004/2005 as pure electric vehicles were at in 1997/1998. This means that fuel cell vehicles will be produced at the rate of several hundred vehicles per year per manufacturer and cost about $40,000 or more, which is still considerably more expensive than ordinary gasoline cars. These fuel cell vehicles will have a performance similar to today`s advanced electric vehicles, e.g., Toyota`s RAV4/EV and Honda`s EV Plus. To go further from this stage to the mass-production stage strong government incentives will be needed. - The pessimistic scenario -- It turns out that fuel cells are not as pure or efficient as in theory and in laboratory experiments. Prices of gasoline and diesel gas continue to be very low. The Californian 10% ZEV Requirement that has been meant to be valid at least ten years from 2003 through 2012 will be suspended or greatly modified. Daimler-Benz, Toyota, and Honda slow down their fuel cell vehicle development activities. No one is

Differing levels of excision repair in human fetal dermis and brain cells

International Nuclear Information System (INIS)

Gibson, R.E.; D'Ambrosio, S.M.; Ohio State Univ., Columbus

1982-01-01

The levels of DNA excision repair, as measured by unscheduled DNA synthesis (UDS) and the UV-endonuclease sensitive site assay, were compared in cells derived from human fetal brain and dermal tissues. The level of UDS induced following ultraviolet (UV) irradiation was found to be lower (approx. 60%) in the fetal brain cells than in fetal dermal cells. It was determined, using the UV-endonuclease sensitive site assay to confirm the UDS observation, that 50% of the dimers induced by UV in fetal dermal cells were repaired in 8 h. while only 15% were removed in the fetal brain cells during the same period of time. Even after 24 h. only 44% of the dimers induced by UV in the fetal brain cells were repaired, while 65% were removed in the dermal cells. These data suggest that cultured human fetal brain cells exhibit lower levels of excision repair compared to cultured human fetal dermal cells. (author)

Technological development and prospect of alkaline fuel cells

International Nuclear Information System (INIS)

Meng Ni; Michael KH Leung; Dennis YC Leung

2006-01-01

This paper reviewed the technological development of alkaline fuel cell (AFC). Although the technology was popular in 1970's and 1980's, there has been a decline in AFC research over the past decade, mainly due to the poisoning of CO 2 . Continuous efforts have demonstrated that CO 2 concentration could be reduced to an acceptable level by a number of viable methods such as absorption, adsorption, electrochemical process, electrolyte circulation, use of liquid hydrogen, and use of solid anionic exchange membranes. Literature survey showed that AFC lifetime could achieve up to 5000 hours. In addition, the use of ammonia as a fuel for AFC was identified as a promising technology. Comparison between AFC and proton exchange membrane fuel cell (PEMFC) was presented to evaluate the AFC technology and its economics. The present review and assessment showed the promise of AFC for the coming hydrogen economy and sustainable development. (authors)

Cell chirality: its origin and roles in left-right asymmetric development.

Science.gov (United States)

Inaki, Mikiko; Liu, Jingyang; Matsuno, Kenji

2016-12-19

An item is chiral if it cannot be superimposed on its mirror image. Most biological molecules are chiral. The homochirality of amino acids ensures that proteins are chiral, which is essential for their functions. Chirality also occurs at the whole-cell level, which was first studied mostly in ciliates, single-celled protozoans. Ciliates show chirality in their cortical structures, which is not determined by genetics, but by 'cortical inheritance'. These studies suggested that molecular chirality directs whole-cell chirality. Intriguingly, chirality in cellular structures and functions is also found in metazoans. In Drosophila, intrinsic cell chirality is observed in various left-right (LR) asymmetric tissues, and appears to be responsible for their LR asymmetric morphogenesis. In other invertebrates, such as snails and Caenorhabditis elegans, blastomere chirality is responsible for subsequent LR asymmetric development. Various cultured cells of vertebrates also show intrinsic chirality in their cellular behaviours and intracellular structural dynamics. Thus, cell chirality may be a general property of eukaryotic cells. In Drosophila, cell chirality drives the LR asymmetric development of individual organs, without establishing the LR axis of the whole embryo. Considering that organ-intrinsic LR asymmetry is also reported in vertebrates, this mechanism may contribute to LR asymmetric development across phyla.This article is part of the themed issue 'Provocative questions in left-right asymmetry'. © 2016 The Authors.

Cell chirality: its origin and roles in left–right asymmetric development

Science.gov (United States)

Inaki, Mikiko; Liu, Jingyang

2016-01-01

An item is chiral if it cannot be superimposed on its mirror image. Most biological molecules are chiral. The homochirality of amino acids ensures that proteins are chiral, which is essential for their functions. Chirality also occurs at the whole-cell level, which was first studied mostly in ciliates, single-celled protozoans. Ciliates show chirality in their cortical structures, which is not determined by genetics, but by ‘cortical inheritance’. These studies suggested that molecular chirality directs whole-cell chirality. Intriguingly, chirality in cellular structures and functions is also found in metazoans. In Drosophila, intrinsic cell chirality is observed in various left–right (LR) asymmetric tissues, and appears to be responsible for their LR asymmetric morphogenesis. In other invertebrates, such as snails and Caenorhabditis elegans, blastomere chirality is responsible for subsequent LR asymmetric development. Various cultured cells of vertebrates also show intrinsic chirality in their cellular behaviours and intracellular structural dynamics. Thus, cell chirality may be a general property of eukaryotic cells. In Drosophila, cell chirality drives the LR asymmetric development of individual organs, without establishing the LR axis of the whole embryo. Considering that organ-intrinsic LR asymmetry is also reported in vertebrates, this mechanism may contribute to LR asymmetric development across phyla. This article is part of the themed issue ‘Provocative questions in left–right asymmetry’. PMID:27821533

Cell death in Pseudomonas aeruginosa biofilm development

DEFF Research Database (Denmark)

Webb, J.S.; Thompson, L.S.; James, S.

2003-01-01

Bacteria growing in biofilms often develop multicellular, three-dimensional structures known as microcolonies. Complex differentiation within biofilms of Pseudomonas aeruginosa occurs, leading to the creation of voids inside microcolonies and to the dispersal of cells from within these voids....... However, key developmental processes regulating these events are poorly understood. A normal component of multicellular development is cell death. Here we report that a repeatable pattern of cell death and lysis occurs in biofilms of P. aeruginosa during the normal course of development. Cell death...... occurred with temporal and spatial organization within biofilms, inside microcolonies, when the biofilms were allowed to develop in continuous-culture flow cells. A subpopulation of viable cells was always observed in these regions. During the onset of biofilm killing and during biofilm development...

Fuel cell serves as oxygen level detector

Science.gov (United States)

1965-01-01

Monitoring the oxygen level in the air is accomplished by a fuel cell detector whose voltage output is proportional to the partial pressure of oxygen in the sampled gas. The relationship between output voltage and partial pressure of oxygen can be calibrated.

Visualization and measurement of ATP levels in living cells replicating hepatitis C virus genome RNA.

Directory of Open Access Journals (Sweden)

Tomomi Ando

Full Text Available Adenosine 5'-triphosphate (ATP is the primary energy currency of all living organisms and participates in a variety of cellular processes. Although ATP requirements during viral lifecycles have been examined in a number of studies, a method by which ATP production can be monitored in real-time, and by which ATP can be quantified in individual cells and subcellular compartments, is lacking, thereby hindering studies aimed at elucidating the precise mechanisms by which viral replication energized by ATP is controlled. In this study, we investigated the fluctuation and distribution of ATP in cells during RNA replication of the hepatitis C virus (HCV, a member of the Flaviviridae family. We demonstrated that cells involved in viral RNA replication actively consumed ATP, thereby reducing cytoplasmic ATP levels. Subsequently, a method to measure ATP levels at putative subcellular sites of HCV RNA replication in living cells was developed by introducing a recently-established Förster resonance energy transfer (FRET-based ATP indicator, called ATeam, into the NS5A coding region of the HCV replicon. Using this method, we were able to observe the formation of ATP-enriched dot-like structures, which co-localize with non-structural viral proteins, within the cytoplasm of HCV-replicating cells but not in non-replicating cells. The obtained FRET signals allowed us to estimate ATP concentrations within HCV replicating cells as ∼5 mM at possible replicating sites and ∼1 mM at peripheral sites that did not appear to be involved in HCV replication. In contrast, cytoplasmic ATP levels in non-replicating Huh-7 cells were estimated as ∼2 mM. To our knowledge, this is the first study to demonstrate changes in ATP concentration within cells during replication of the HCV genome and increased ATP levels at distinct sites within replicating cells. ATeam may be a powerful tool for the study of energy metabolism during replication of the viral genome.

The change of intracellular NAD level at the process of fusarium sambucinum growth and development

International Nuclear Information System (INIS)

Gulyamova, T.G.; Ehshtukhtarova, M.Kh.; Umarova, G.D.; Kerbalaeva, A.M.; Khalmuradov, A.G.

1996-01-01

Alterations of intracellular NAD(Nicotinamide-Adenine Dinucleotide) level have been studied in the process of growth and development of Fusanium sambucinum, selected earlier as a potential NAD producer. It was established that essential fluctuations of NAD concentration are dependent on growth phase, morphological cell type and DNA biosynthesis, that allowed to propose a real linkage between coenzyme pool and replicative activity of cells. (author). 7 refs., 2 figs

Identifying long-term memory B-cells in vaccinated children despite waning antibody levels specific for Bordetella pertussis proteins.

Science.gov (United States)

Hendrikx, Lotte H; Oztürk, Kemal; de Rond, Lia G H; Veenhoven, Reinier H; Sanders, Elisabeth A M; Berbers, Guy A M; Buisman, Anne-Marie

2011-02-04

Whooping cough is a respiratory disease caused by Bordetella pertussis. Since the 1950s in developed countries pertussis vaccinations are included in the national immunization program. However, antibody levels rapidly wane after both whole cell and acellular pertussis vaccination. Therefore protection against pertussis may depend largely on long-term B- and T-cell immunities. We investigated long-term pertussis-specific memory B-cell responses in children who were primed at infant age with the Dutch wP-vaccine (ISRCTN65428640). Purified B-cells were characterized by FACS-analysis and after polyclonal stimulation memory B-cells were detected by ELISPOT-assays specific for pertussis toxin, filamentous haemagglutinin, pertactin and tetanus. In addition, plasma IgG levels directed to the same antigens were measured by a fluorescent bead-based multiplex immunoassay. Two and 3 years after wP priming as well as 2 and 5 years after the aP booster at the age of 4, low plasma IgG levels to the pertussis proteins were found. At the same time, however pertussis protein-specific memory B-cells could be detected and their number increased with age. The number of tetanus-specific memory B-cells was similar in all age groups, whereas IgG-tetanus levels were high 2 years after tetanus booster compared to pre- and 5 years post-booster levels. This study shows the presence of long-term pertussis protein-specific memory B-cells in children despite waning antibody levels after vaccination, which suggests that memory B-cells in addition to antibodies may contribute to protection against pertussis. Copyright © 2010 Elsevier Ltd. All rights reserved.

Distinct gene expression signatures in human embryonic stem cells differentiated towards definitive endoderm at single-cell level

DEFF Research Database (Denmark)

Norrman, Karin; Strömbeck, Anna; Semb, Henrik

2013-01-01

for the three activin A based protocols applied. Our data provide novel insights in DE gene expression at the cellular level of in vitro differentiated human embryonic stem cells, and illustrate the power of using single-cell gene expression profiling to study differentiation heterogeneity and to characterize...... of anterior definitive endoderm (DE). Here, we differentiated human embryonic stem cells towards DE using three different activin A based treatments. Differentiation efficiencies were evaluated by gene expression profiling over time at cell population level. A panel of key markers was used to study DE...... formation. Final DE differentiation was also analyzed with immunocytochemistry and single-cell gene expression profiling. We found that cells treated with activin A in combination with sodium butyrate and B27 serum-free supplement medium generated the most mature DE cells. Cell population studies were...

Research, development and pilot production of high output thin silicon solar cells

Science.gov (United States)

Iles, P. A.

1976-01-01

Work was performed to define and apply processes which could lead to high output from thin (2-8 mils) silicon solar cells. The overall problems are outlined, and two satisfactory process sequences were developed. These sequences led to good output cells in the thickness range to just below 4 mils; although the initial contract scope was reduced, one of these sequences proved capable of operating beyond a pilot line level, to yield good quality 4-6 mil cells of high output.

Extended Temperature Solar Cell Technology Development

Science.gov (United States)

Landis, Geoffrey A.; Jenkins, Phillip; Scheiman, David; Rafaelle, Ryne

2004-01-01

Future NASA missions will require solar cells to operate both in regimes closer to the sun, and farther from the sun, where the operating temperatures will be higher and lower than standard operational conditions. NASA Glenn is engaged in testing solar cells under extended temperature ranges, developing theoretical models of cell operation as a function of temperature, and in developing technology for improving the performance of solar cells for both high and low temperature operation.

Reduced levels of SCD1 accentuate palmitate-induced stress in insulin-producing β-cells

Directory of Open Access Journals (Sweden)

Hovsepyan Meri

2010-09-01

Full Text Available Abstract Background Stearoyl-CoA desaturase 1 (SCD1 is an ER resident enzyme introducing a double-bond in saturated fatty acids. Global knockout of SCD1 in mouse increases fatty acid oxidation and insulin sensitivity which makes the animal resistant to diet-induced obesity. Inhibition of SCD1 has therefore been proposed as a potential therapy of the metabolic syndrome. Much of the work has focused on insulin target tissue and very little is known about how reduced levels of SCD1 would affect the insulin-producing β-cell, however. The aim of the present study was therefore to investigate how reduced levels of SCD1 affect the β-cell. Results Insulin-secreting MIN6 cells with reduced levels of SCD1 were established by siRNA mediated knockdown. When fatty acid oxidation was measured, no difference between cells with reduced levels of SCD1 and mock-transfected cells were found. Also, reducing levels of SCD1 did not affect insulin secretion in response to glucose. To investigate how SCD1 knockdown affected cellular mechanisms, differentially regulated proteins were identified by a proteomic approach. Cells with reduced levels of SCD1 had higher levels of ER chaperones and components of the proteasome. The higher amounts did not protect the β-cell from palmitate-induced ER stress and apoptosis. Instead, rise in levels of p-eIF2α and CHOP after palmitate exposure was 2-fold higher in cells with reduced levels of SCD1 compared to mock-transfected cells. Accordingly, apoptosis rose to higher levels after exposure to palmitate in cells with reduced levels of SCD1 compared to mock-transfected cells. Conclusions In conclusion, reduced levels of SCD1 augment palmitate-induced ER stress and apoptosis in the β-cell, which is an important caveat when considering targeting this enzyme as a treatment of the metabolic syndrome.

In vitro development rate of preimplantation rabbit embryos cultured with different levels of melatonin.

Science.gov (United States)

Mehaisen, Gamal Mohamed Kamel; Saeed, Ayman Moustafa

2015-02-01

This study aimed to investigate the effect of melatonin supplementation at different levels in culture medium on embryo development in rabbits. Embryos of 2-4 cells, 8-16 cells and morula stages were recovered from nulliparous Red Baladi rabbit does by laparotomy technique 24, 48 and 72 h post-insemination, respectively. Normal embryos from each stage were cultured to hatched blastocyst stages in either control culture medium (TCM-199 + 20% fetal bovine serum) or control supplemented with melatonin at 10(-3) M, 10(-6) M or 10(-9) M. No effect of melatonin was found on development of embryos recovered at 24 h post-insemination. The high level of melatonin at 10(-3) M adversely affected the in vitro development rates of embryos recovered at 48 h post-insemination (52 versus 86, 87 and 80% blastocyst rate; 28 versus 66, 78 and 59% hatchability rate for 10(-3) M versus 10(-9) M, 10(-6) M and control, respectively, P< 0.05). At the morula stage, melatonin at 10-3 M significantly increased the in vitro development of embryos (92% for 10(-3) M versus 76% for control, P < 0.05), while the hatchability rate of these embryos was not improved by melatonin (16-30% versus 52% for melatonin groups versus control, P < 0.05). Results show that a moderate level of melatonin (10(-6) M) may improve the development and hatchability rates of preimplantation rabbit embryos. The addition of melatonin at a 10-3 M concentration enhances the development of rabbit morulae but may negatively affect the development of earlier embryos. More studies are needed to optimize the use of melatonin in in vitro embryo culture in rabbits.

Development of one body α-γ type manipulator for hot cell facility

International Nuclear Information System (INIS)

Jung, S. K.; Lee, S. B.; Lee, E. P.

2004-01-01

To handle the high level radioactive materials in a sealed type hot cell, our company has developed the one body alpha-gamma type manipulator and this is an improved model compared with the previously developed beta-gamma and separated alpha-gamma type manipulators. The successful development of one body alpha-gamma type manipulator means our company has a whole capacity to design and fabricate all kinds of manipulators using in hot cells. Until now most of the manipulators in Korea were imported from other countries. The development of Korean manipulators gives us the easier maintenance and lower price compared to the foreign products. It is also possible to export the Korean manipulators to overseas

The level of circulating endothelial progenitor cells may be associated with the occurrence and recurrence of chronic subdural hematoma

Directory of Open Access Journals (Sweden)

Yan Song

2013-01-01

Full Text Available OBJECTIVES: The onset of chronic subdural hematoma may be associated with direct or indirect minor injuries to the head or a poorly repaired vascular injury. Endothelial progenitor cells happen to be one of the key factors involved in hemostasis and vascular repair. This study was designed to observe the levels of endothelial progenitor cells, white blood cells, platelets, and other indicators in the peripheral blood of patients diagnosed with chronic subdural hematoma to determine the possible relationship between the endothelial progenitor cells and the occurrence, development, and outcomes of chronic subdural hematoma. METHOD: We enrolled 30 patients with diagnosed chronic subdural hematoma by computer tomography scanning and operating procedure at Tianjin Medical University General Hospital from July 2009 to July 2011. Meanwhile, we collected 30 cases of peripheral blood samples from healthy volunteers over the age of 50. Approximately 2 ml of blood was taken from veins of the elbow to test the peripheral blood routine and coagulation function. The content of endothelial progenitor cells in peripheral blood mononuclear cells was determined by flow cytometry. RESULTS: The level of endothelial progenitor cells in peripheral blood was significantly lower in preoperational patients with chronic subdural hematomas than in controls. There were no significant differences between the two groups regarding the blood routine and coagulation function. However, the levels of circulating endothelial progenitor cells were significantly different between the recurrent group and the non-recurrent group. CONCLUSIONS: The level of circulating endothelial progenitor cells in chronic subdural hematoma patients was significantly lower than the level in healthy controls. Meanwhile, the level of endothelial progenitor cells in recurrent patients was significantly lower than the level in patients without recurrence. Endothelial progenitor cells may be related to the

Effect of low-level laser-treated mesenchymal stem cells on myocardial infarction.

Science.gov (United States)

El Gammal, Zaynab H; Zaher, Amr M; El-Badri, Nagwa

2017-09-01

Cardiovascular disease is the leading cause of death worldwide. Although cardiac transplantation is considered the most effective therapy for end-stage cardiac diseases, it is limited by the availability of matching donors and the complications of the immune suppressive regimen used to prevent graft rejection. Application of stem cell therapy in experimental animal models was shown to reverse cardiac remodeling, attenuate cardiac fibrosis, improve heart functions, and stimulate angiogenesis. The efficacy of stem cell therapy can be amplified by low-level laser radiation. It is well established that the bio-stimulatory effect of low-level laser is influenced by the following parameters: wavelength, power density, duration, energy density, delivery time, and the type of irradiated target. In this review, we evaluate the available experimental data on treatment of myocardial infarction using low-level laser. Eligible papers were characterized as in vivo experimental studies that evaluated the use of low-level laser therapy on stem cells in order to attenuate myocardial infarction. The following descriptors were used separately and in combination: laser therapy, low-level laser, low-power laser, stem cell, and myocardial infarction. The assessed low-level laser parameters were wavelength (635-804 nm), power density (6-50 mW/cm 2 ), duration (20-150 s), energy density (0.96-1 J/cm 2 ), delivery time (20 min-3 weeks after myocardial infarction), and the type of irradiated target (bone marrow or in vitro-cultured bone marrow mesenchymal stem cells). The analysis focused on the cardioprotective effect of this form of therapy, the attenuation of scar tissue, and the enhancement of angiogenesis as primary targets. Other effects such as cell survival, cell differentiation, and homing are also included. Among the evaluated protocols using different parameters, the best outcome for treating myocardial infarction was achieved by treating the bone marrow by one dose of low-level

Diverse phosphorylation patterns of B cell receptor-associated signaling in naïve and memory human B cells revealed by phosphoflow, a powerful technique to study signaling at the single cell level

Directory of Open Access Journals (Sweden)

Franklin R Toapanta

2012-10-01

Full Text Available Following interaction with cognate antigens, B cells undergo cell activation, proliferation and differentiation. Ligation of the B cell receptor (BCR leads to the phosphorylation of BCR-associated signaling proteins within minutes of antigen binding, a process with profound consequences for the fate of the cells and development of effector immunity. Phosphoflow allows a rapid evaluation of various signaling pathways in complex heterogenous cell subsets. This novel technique was used in combination with multi-chromatic flow cytometry and fluorescent-cell barcoding to study phosphorylation of BCR-associated signaling pathways in naïve and memory human B cell subsets. Proteins of the initiation (Syk, propagation (Btk, Akt and integration (p38MAPK and Erk1/2 signaling units were studied. Switched memory (Sm CD27+ and Sm CD27- phosphorylation patterns were similar when stimulated with anti-IgA or -IgG. In contrast, naïve and unswitched memory (Um cells showed significant differences following IgM stimulation. Enhanced phosphorylation of Syk was observed in Um cells, suggesting a lower activation threshold. This is likely the result of higher amounts of IgM on the cell surface, higher pan-Syk levels and enhanced susceptibility to phosphatase inhibition. All other signaling proteins evaluated also showed some degree of enhanced phosphorylation in Um cells. Furthermore, both the PLC-γ2 and PI3K pathways were activated in Um cells, while only the PI3K pathway was activated on naïve cells. Um cells were the only ones that activated signaling pathways when stimulated with fluorescently-labeled S. Typhi and S. pneumoniae. Finally, simultaneous evaluation of signaling proteins at the single cell level (multi-phosphorylated cells revealed that interaction with gram positive and negative bacteria resulted in complex and diverse signaling patterns. Phosphoflow holds great potential to accelerate vaccine development by identifying signaling profiles in good

Effect of oxygen levels on the physiology of dendritic cells: implications for adoptive cell therapy.

Science.gov (United States)

Futalan, Diahnn; Huang, Chien-Tze; Schmidt-Wolf, Ingo G H; Larsson, Marie; Messmer, Davorka

2011-01-01

Dendritic cell (DC)-based adoptive tumor immunotherapy approaches have shown promising results, but the incidence of tumor regression is low and there is an evident call for identifying culture conditions that produce DCs with a more potent Th1 potential. Routinely, DCs are differentiated in CO(2) incubators under atmospheric oxygen conditions (21% O(2)), which differ from physiological oxygen levels of only 3-5% in tissue, where most DCs reside. We investigated whether differentiation and maturation of DCs under physiological oxygen levels could produce more potent T-cell stimulatory DCs for use in adoptive immunotherapy. We found that immature DCs differentiated under physiological oxygen levels showed a small but significant reduction in their endocytic capacity. The different oxygen levels did not influence their stimuli-induced upregulation of cluster of differentiation 54 (CD54), CD40, CD83, CD86, C-C chemokine receptor type 7 (CCR7), C-X-C chemokine receptor type 4 (CXCR4) and human leukocyte antigen (HLA)-DR or the secretion of interleukin (IL)-6, tumor necrosis factor (TNF)-α and IL-10 in response to lipopolysaccharide (LPS) or a cytokine cocktail. However, DCs differentiated under physiological oxygen level secreted higher levels of IL-12(p70) after exposure to LPS or CD40 ligand. Immature DCs differentiated at physiological oxygen levels caused increased T-cell proliferation, but no differences were observed for mature DCs with regard to T-cell activation. In conclusion, we show that although DCs generated under atmospheric or physiological oxygen conditions are mostly similar in function and phenotype, DCs differentiated under physiological oxygen secrete larger amounts of IL-12(p70). This result could have implications for the use of ex vivo-generated DCs for clinical studies, since DCs differentiated at physiological oxygen could induce increased Th1 responses in vivo.

Energetic consumption levels and human development indexes

International Nuclear Information System (INIS)

Boa Nova, Antonio Carlos

1999-01-01

The article overviews the energetic consumption levels and human development indexes. The human development indexes are described based on the United Nations Development Programme. A comparison between the energetic consumption levels and human development indexes is also presented

Quantitative intracellular flux modeling and applications in biotherapeutic development and production using CHO cell cultures.

Science.gov (United States)

Huang, Zhuangrong; Lee, Dong-Yup; Yoon, Seongkyu

2017-12-01

Chinese hamster ovary (CHO) cells have been widely used for producing many recombinant therapeutic proteins. Constraint-based modeling, such as flux balance analysis (FBA) and metabolic flux analysis (MFA), has been developing rapidly for the quantification of intracellular metabolic flux distribution at a systematic level. Such methods would produce detailed maps of flows through metabolic networks, which contribute significantly to better understanding of metabolism in cells. Although these approaches have been extensively established in microbial systems, their application to mammalian cells is sparse. This review brings together the recent development of constraint-based models and their applications in CHO cells. The further development of constraint-based modeling approaches driven by multi-omics datasets is discussed, and a framework of potential modeling application in cell culture engineering is proposed. Improved cell culture system understanding will enable robust developments in cell line and bioprocess engineering thus accelerating consistent process quality control in biopharmaceutical manufacturing. © 2017 Wiley Periodicals, Inc.

Identifying EGFR-Expressed Cells and Detecting EGFR Multi-Mutations at Single-Cell Level by Microfluidic Chip

Science.gov (United States)

Li, Ren; Zhou, Mingxing; Li, Jine; Wang, Zihua; Zhang, Weikai; Yue, Chunyan; Ma, Yan; Peng, Hailin; Wei, Zewen; Hu, Zhiyuan

2018-03-01

EGFR mutations companion diagnostics have been proved to be crucial for the efficacy of tyrosine kinase inhibitor targeted cancer therapies. To uncover multiple mutations occurred in minority of EGFR-mutated cells, which may be covered by the noises from majority of un-mutated cells, is currently becoming an urgent clinical requirement. Here we present the validation of a microfluidic-chip-based method for detecting EGFR multi-mutations at single-cell level. By trapping and immunofluorescently imaging single cells in specifically designed silicon microwells, the EGFR-expressed cells were easily identified. By in situ lysing single cells, the cell lysates of EGFR-expressed cells were retrieved without cross-contamination. Benefited from excluding the noise from cells without EGFR expression, the simple and cost-effective Sanger's sequencing, but not the expensive deep sequencing of the whole cell population, was used to discover multi-mutations. We verified the new method with precisely discovering three most important EGFR drug-related mutations from a sample in which EGFR-mutated cells only account for a small percentage of whole cell population. The microfluidic chip is capable of discovering not only the existence of specific EGFR multi-mutations, but also other valuable single-cell-level information: on which specific cells the mutations occurred, or whether different mutations coexist on the same cells. This microfluidic chip constitutes a promising method to promote simple and cost-effective Sanger's sequencing to be a routine test before performing targeted cancer therapy.[Figure not available: see fulltext.

Radiosensitizing Effects of Ectopic miR-101 on Non–Small-Cell Lung Cancer Cells Depend on the Endogenous miR-101 Level

International Nuclear Information System (INIS)

Chen, Susie; Wang Hongyan; Ng, Wooi Loon; Curran, Walter J.; Wang Ya

2011-01-01

Purpose: Previously, we showed that ectopic miR-101 could sensitize human tumor cells to radiation by targeting ATM and DNA-PK catalytic subunit (DNA-PKcs) to inhibit DNA repair, as the endogenous miR-101 levels are low in tumors in general. However, the heterogeneity of human cancers may result in an exception. The purpose of this study was to test the hypothesis that a few tumor cell lines with a high level of endogenous miR-101 would prove less response to ectopic miR-101. Methods and Materials: Fourteeen non–small-cell lung cancer (NSCLC) cell lines and one immortalized non-malignant lung epithelial cell line (NL20) were used for comparing endogenous miR-101 levels by real-time reverse transcription–polymerase chain reaction. Based on the different miR-101 levels, four cell lines with different miR-101 levels were chosen for transfection with a green fluorescent protein–lentiviral plasmid encoding miR-101. The target protein levels were measured by using Western blotting. The radiosensitizing effects of ectopic miR-101 on these NSCLC cell lines were determined by a clonogenic assay and xenograft mouse model. Results: The endogenous miR-101 level was similar or lower in 13 NSCLC cell lines but was 11-fold higher in one cell line (H157) than in NL20 cells. Although ectopic miR-101 efficiently decreased the ATM and DNA-PKcs levels and increased the radiosensitization level in H1299, H1975, and A549 cells, it did not change the levels of the miR-101 targets or radiosensitivity in H157 cells. Similar results were observed in xenograft mice. Conclusions: A small number of NSCLC cell lines could have a high level of endogenous miR-101. The ectopic miR-101 was able to radiosensitize most NSCLC cells, except for the NSCLC cell lines that had a much higher endogenous miR-101 level. These results suggest that when we choose one miRNA as a therapeutic tool, the endogenous level of the miRNA in each tumor should be considered.

Radiosensitizing Effects of Ectopic miR-101 on Non-Small-Cell Lung Cancer Cells Depend on the Endogenous miR-101 Level

Energy Technology Data Exchange (ETDEWEB)

Chen, Susie; Wang Hongyan; Ng, Wooi Loon; Curran, Walter J. [Department of Radiation Oncology, School of Medicine and the Winship Cancer Institute, Emory University, Atlanta, GA (United States); Wang Ya, E-mail: ywang94@emory.edu [Department of Radiation Oncology, School of Medicine and the Winship Cancer Institute, Emory University, Atlanta, GA (United States)

2011-12-01

Purpose: Previously, we showed that ectopic miR-101 could sensitize human tumor cells to radiation by targeting ATM and DNA-PK catalytic subunit (DNA-PKcs) to inhibit DNA repair, as the endogenous miR-101 levels are low in tumors in general. However, the heterogeneity of human cancers may result in an exception. The purpose of this study was to test the hypothesis that a few tumor cell lines with a high level of endogenous miR-101 would prove less response to ectopic miR-101. Methods and Materials: Fourteeen non-small-cell lung cancer (NSCLC) cell lines and one immortalized non-malignant lung epithelial cell line (NL20) were used for comparing endogenous miR-101 levels by real-time reverse transcription-polymerase chain reaction. Based on the different miR-101 levels, four cell lines with different miR-101 levels were chosen for transfection with a green fluorescent protein-lentiviral plasmid encoding miR-101. The target protein levels were measured by using Western blotting. The radiosensitizing effects of ectopic miR-101 on these NSCLC cell lines were determined by a clonogenic assay and xenograft mouse model. Results: The endogenous miR-101 level was similar or lower in 13 NSCLC cell lines but was 11-fold higher in one cell line (H157) than in NL20 cells. Although ectopic miR-101 efficiently decreased the ATM and DNA-PKcs levels and increased the radiosensitization level in H1299, H1975, and A549 cells, it did not change the levels of the miR-101 targets or radiosensitivity in H157 cells. Similar results were observed in xenograft mice. Conclusions: A small number of NSCLC cell lines could have a high level of endogenous miR-101. The ectopic miR-101 was able to radiosensitize most NSCLC cells, except for the NSCLC cell lines that had a much higher endogenous miR-101 level. These results suggest that when we choose one miRNA as a therapeutic tool, the endogenous level of the miRNA in each tumor should be considered.

Plasmacytoid dendritic cells: Development, functions, and role in atherosclerotic inflammation

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Dimitry A Chistiakov

2014-07-01

Full Text Available Plasmacytoid dendritic cells (pDCs are a specialized subset of DCs that links innate and adaptive immunity. They sense viral and bacterial pathogens and release high levels of Type I interferons (IFN-I in response to infection. pDCs were shown to contribute to inflammatory responses in the steady state and in pathology. In atherosclerosis, pDCs are involved in priming vascular inflammation and atherogenesis through production of IFN-I and chemokines that attract inflammatory cells to inflamed sites. pDCs also contribute to the proinflammatory activation of effector T cells, cytotoxic T cells, and conventional DCs. However, tolerogenic populations of pDCs are found that suppress atherosclerosis-associated inflammation through down-regulation of function and proliferation of proinflammatory T cell subsets and induction of regulatory T cells with potent immunomodulatory properties. Notably, atheroprotective tolerogenic DCs could be induced by certain self-antigens or bacterial antigens that suggests for great therapeutic potential of these DCs for development of DC-based anti-atherogenic vaccines.

Development of galvanic high energy cells with molten salt electrolytes

Energy Technology Data Exchange (ETDEWEB)

Borger, W.; Ely, G.; Kunze, D.; Laig-Hoerstebrock, H.; Panesar, H.; Sterr, G.; Wunderlich, A.

1985-01-01

The development work during the period 1980-1983 was mainly directed towards the development of technical LiAl/FeS cells, the development of separators, tests of cells and modules, and more basic work. An important objective was the improvement of cycle life at constant specific energy. Technical cells with 140 Ah nominal capacity at the five hour rate and 100 Wh.kg/sup -1/ specific energy performed up to 400 full cycles (30 A discharge), while in 10 Ah test cells more than 2000 full cycles have been demonstrated. The improvement of cycle life of technical cells was achieved by the use of improved separators fabricated from MgO-powder and by a vacuum-tight electrical feedthrough. A design concept of a 10 cell module has been developed based upon 200 Ah cell with two positive and three negative plates. A detailed investigation of safety aspects showed that there is no specific risk related to the LiAl/molten salt/FeS system. Thermal management of a 24 kWh battery was investigated and the Ohmic heat generated in the leads seems to be the critical factor. A range of total materials cost between 60 and 130 DM/kWh has been estimated. The price of LiAl/FeS batteries will most probably also be in the range of conventional secondary batteries. The cost/benefit analysis shows a considerable potential of energy conservation by the use of light-weight high energy batteries. Compared with a expected technical life of 7 years a pay-back period between 2 and 6 years seems attractive. However, the economy of the electric vehicle is strongly influenced by the higher purchase price of an electric vehicle and the present energy level.

Human and great ape red blood cells differ in plasmalogen levels and composition

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Ely John J

2011-06-01

Full Text Available Abstract Background Plasmalogens are ether phospholipids required for normal mammalian developmental, physiological, and cognitive functions. They have been proposed to act as membrane antioxidants and reservoirs of polyunsaturated fatty acids as well as influence intracellular signaling and membrane dynamics. Plasmalogens are particularly enriched in cells and tissues of the human nervous, immune, and cardiovascular systems. Humans with severely reduced plasmalogen levels have reduced life spans, abnormal neurological development, skeletal dysplasia, impaired respiration, and cataracts. Plasmalogen deficiency is also found in the brain tissue of individuals with Alzheimer disease. Results In a human and great ape cohort, we measured the red blood cell (RBC levels of the most abundant types of plasmalogens. Total RBC plasmalogen levels were lower in humans than bonobos, chimpanzees, and gorillas, but higher than orangutans. There were especially pronounced cross-species differences in the levels of plasmalogens with a C16:0 moiety at the sn-1 position. Humans on Western or vegan diets had comparable total RBC plasmalogen levels, but the latter group showed moderately higher levels of plasmalogens with a C18:1 moiety at the sn-1 position. We did not find robust sex-specific differences in human or chimpanzee RBC plasmalogen levels or composition. Furthermore, human and great ape skin fibroblasts showed only modest differences in peroxisomal plasmalogen biosynthetic activity. Human and chimpanzee microarray data indicated that genes involved in plasmalogen biosynthesis show cross-species differential expression in multiple tissues. Conclusion We propose that the observed differences in human and great ape RBC plasmalogens are primarily caused by their rates of biosynthesis and/or turnover. Gene expression data raise the possibility that other human and great ape cells and tissues differ in plasmalogen levels. Based on the phenotypes of humans and

Stomach development, stem cells and disease

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Kim, Tae-Hee; Shivdasani, Ramesh A.

2016-01-01

The stomach, an organ derived from foregut endoderm, secretes acid and enzymes and plays a key role in digestion. During development, mesenchymal-epithelial interactions drive stomach specification, patterning, differentiation and growth through selected signaling pathways and transcription factors. After birth, the gastric epithelium is maintained by the activity of stem cells. Developmental signals are aberrantly activated and stem cell functions are disrupted in gastric cancer and other disorders. Therefore, a better understanding of stomach development and stem cells can inform approaches to treating these conditions. This Review highlights the molecular mechanisms of stomach development and discusses recent findings regarding stomach stem cells and organoid cultures, and their roles in investigating disease mechanisms. PMID:26884394

Breast cancer instructs dendritic cells to prime interleukin 13–secreting CD4+ T cells that facilitate tumor development

Science.gov (United States)

Aspord, Caroline; Pedroza-Gonzalez, Alexander; Gallegos, Mike; Tindle, Sasha; Burton, Elizabeth C.; Su, Dan; Marches, Florentina; Banchereau, Jacques; Palucka, A. Karolina

2007-01-01

We previously reported (Bell, D., P. Chomarat, D. Broyles, G. Netto, G.M. Harb, S. Lebecque, J. Valladeau, J. Davoust, K.A. Palucka, and J. Banchereau. 1999. J. Exp. Med. 190: 1417–1426) that breast cancer tumors are infiltrated with mature dendritic cells (DCs), which cluster with CD4+ T cells. We now show that CD4+ T cells infiltrating breast cancer tumors secrete type 1 (interferon γ) as well as high levels of type 2 (interleukin [IL] 4 and IL-13) cytokines. Immunofluorescence staining of tissue sections revealed intense IL-13 staining on breast cancer cells. The expression of phosphorylated signal transducer and activator of transcription 6 in breast cancer cells suggests that IL-13 actually delivers signals to cancer cells. To determine the link between breast cancer, DCs, and CD4+ T cells, we implanted human breast cancer cell lines in nonobese diabetic/LtSz-scid/scid β2 microglobulin–deficient mice engrafted with human CD34+ hematopoietic progenitor cells and autologous T cells. There, CD4+ T cells promote early tumor development. This is dependent on DCs and can be partially prevented by administration of IL-13 antagonists. Thus, breast cancer targets DCs to facilitate its development. PMID:17438063

Non-Linear Relationships between Aflatoxin B1 Levels and the Biological Response of Monkey Kidney Vero Cells

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Mendel Friedman

2013-08-01

Full Text Available Aflatoxin-producing fungi contaminate food and feed during pre-harvest, storage and processing periods. Once consumed, aflatoxins (AFs accumulate in tissues, causing illnesses in animals and humans. Most human exposure to AF seems to be a result of consumption of contaminated plant and animal products. The policy of blending and dilution of grain containing higher levels of aflatoxins with uncontaminated grains for use in animal feed implicitly assumes that the deleterious effects of low levels of the toxins are linearly correlated to concentration. This assumption may not be justified, since it involves extrapolation of these nontoxic levels in feed, which are not of further concern. To develop a better understanding of the significance of low dose effects, in the present study, we developed quantitative methods for the detection of biologically active aflatoxin B1 (AFB1 in Vero cells by two independent assays: the green fluorescent protein (GFP assay, as a measure of protein synthesis by the cells, and the microculture tetrazolium (MTT assay, as a measure of cell viability. The results demonstrate a non-linear dose-response relationship at the cellular level. AFB1 at low concentrations has an opposite biological effect to higher doses that inhibit protein synthesis. Additional studies showed that heat does not affect the stability of AFB1 in milk and that the Vero cell model can be used to determine the presence of bioactive AFB1 in spiked beef, lamb and turkey meat. The implication of the results for the cumulative effects of low amounts of AFB1 in numerous foods is discussed.

The relationship of metabolic burden to productivity levels in CHO cell lines.

Science.gov (United States)

Zou, Wu; Edros, Raihana; Al-Rubeai, Mohamed

2018-03-01

The growing demand for recombinant therapeutics has driven biotechnologists to develop new production strategies. One such strategy for increasing the expression of heterologous proteins has focused on enhancing cell-specific productivity through environmental perturbations. In this work, the effects of hypothermia, hyperosmolarity, high shear stress, and sodium butyrate treatment on growth and productivity were studied using three (low, medium, and high producing) CHO cell lines that differed in their specific productivities of monoclonal antibody. In all three cell lines, the inhibitory effect of these parameters on proliferation was demonstrated. Additionally, compared to the control, specific productivity was enhanced under all conditions and exhibited a consistent cell line specific pattern, with maximum increases (50-290%) in the low producer, and minimum increases (7-20%) in the high producer. Thus, the high-producing cell line was less responsive to environmental perturbations than the low-producing cell line. We hypothesize that this difference is most likely due to the bottleneck associated with a higher metabolic burden caused by higher antibody expression. Increased recombinant mRNA levels and pyruvate carboxylase activities due to low temperature and hyperosmotic stress were found to be positively associated with the metabolic burden. © 2017 International Union of Biochemistry and Molecular Biology, Inc.

Application of low level laser on skin cell lines

CSIR Research Space (South Africa)

Ndhundhuma, IM

2010-01-01

Full Text Available Lasers have emerged as powerful tools for tissue engineering. To examine cellular growth, and cell to cell interactions, in vitro skin models have been developed combining two major cell types of skin, keratinocytes and fibroblasts. The main...

Model-based development of low-level control strategies for transient operation of solid oxide fuel cell systems

Science.gov (United States)

Sorrentino, Marco; Pianese, Cesare

The exploitation of an SOFC-system model to define and test control and energy management strategies is presented. Such a work is motivated by the increasing interest paid to SOFC technology by industries and governments due to its highly appealing potentialities in terms of energy savings, fuel flexibility, cogeneration, low-pollution and low-noise operation. The core part of the model is the SOFC stack, surrounded by a number of auxiliary devices, i.e. air compressor, regulating pressure valves, heat exchangers, pre-reformer and post-burner. Due to the slow thermal dynamics of SOFCs, a set of three lumped-capacity models describes the dynamic response of fuel cell and heat exchangers to any operation change. The dynamic model was used to develop low-level control strategies aimed at guaranteeing targeted performance while keeping stack temperature derivative within safe limits to reduce stack degradation due to thermal stresses. Control strategies for both cold-start and warmed-up operations were implemented by combining feedforward and feedback approaches. Particularly, the main cold-start control action relies on the precise regulation of methane flow towards anode and post-burner via by-pass valves; this strategy is combined with a cathode air-flow adjustment to have a tight control of both stack temperature gradient and warm-up time. Results are presented to show the potentialities of the proposed model-based approach to: (i) serve as a support to control strategies development and (ii) solve the trade-off between fast SOFC cold-start and avoidance of thermal-stress caused damages.

Modeling Human Natural Killer Cell Development in the Era of Innate Lymphoid Cells.

Science.gov (United States)

Scoville, Steven D; Freud, Aharon G; Caligiuri, Michael A

2017-01-01

Decades after the discovery of natural killer (NK) cells, their developmental pathways in mice and humans have not yet been completely deciphered. Accumulating evidence indicates that NK cells can develop in multiple tissues throughout the body. Moreover, detailed and comprehensive models of NK cell development were proposed soon after the turn of the century. However, with the recent identification and characterization of other subtypes of innate lymphoid cells (ILCs), which show some overlapping functional and phenotypic features with NK cell developmental intermediates, the distinct stages through which human NK cells develop from early hematopoietic progenitor cells remain unclear. Thus, there is a need to reassess and refine older models of NK cell development in the context of new data and in the era of ILCs. Our group has focused on elucidating the developmental pathway of human NK cells in secondary lymphoid tissues (SLTs), including tonsils and lymph nodes. Here, we provide an update of recent progress that has been made with regard to human NK cell development in SLTs, and we discuss these new findings in the context of contemporary models of ILC development.

High normal fasting glucose level in obese youth: a marker for insulin resistance and beta cell dysregulation.

LENUS (Irish Health Repository)

O'Malley, G

2010-06-01

A high but normal fasting plasma glucose level in adults is a risk factor for future development of type 2 diabetes mellitus and cardiovascular disease. We investigated whether normal fasting plasma glucose levels (<5.60 mmol\\/l) are associated with decreases in insulin sensitivity and beta cell function, as well as an adverse cardiovascular profile in obese youth.

Development of model for analysing respective collections of intended hematopoietic stem cells and harvests of unintended mature cells in apheresis for autologous hematopoietic stem cell collection.

Science.gov (United States)

Hequet, O; Le, Q H; Rodriguez, J; Dubost, P; Revesz, D; Clerc, A; Rigal, D; Salles, G; Coiffier, B

2014-04-01

Hematopoietic stem cells (HSCs) required to perform peripheral hematopoietic autologous stem cell transplantation (APBSCT) can be collected by processing several blood volumes (BVs) in leukapheresis sessions. However, this may cause granulocyte harvest in graft and decrease in patient's platelet blood level. Both consequences may induce disturbances in patient. One apheresis team's current purpose is to improve HSC collection by increasing HSC collection and prevent increase in granulocyte and platelet harvests. Before improving HSC collection it seemed important to know more about the way to harvest these types of cells. The purpose of our study was to develop a simple model for analysing respective collections of intended CD34+ cells among HSC (designated here as HSC) and harvests of unintended platelets or granulocytes among mature cells (designated here as mature cells) considering the number of BVs processed and factors likely to influence cell collection or harvest. For this, we processed 1, 2 and 3 BVs in 59 leukapheresis sessions and analysed corresponding collections and harvests with a referent device (COBE Spectra). First we analysed the amounts of HSC collected and mature cells harvested and second the evolution of the respective shares of HSC and mature cells collected or harvested throughout the BV processes. HSC collections and mature cell harvests increased globally (pcollections and harvests, which showed that only pre-leukapheresis blood levels (CD34+cells and platelets) influenced both cell collections and harvests (CD34+cells and platelets) (pcollections and mature unintended cells harvests (pcollections or unintended mature cell harvests were pre-leukapheresis blood cell levels. Our model was meant to assist apheresis teams in analysing shares of HSC collected and mature cells harvested with new devices or with new types of HSC mobilization. Copyright © 2014 Elsevier Ltd. All rights reserved.

In Vitro Modeling of Human Germ Cell Development Using Pluripotent Stem Cells

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Yuncheng Zhao

2018-02-01

Full Text Available Summary: Due to differences across species, the mechanisms of cell fate decisions determined in mice cannot be readily extrapolated to humans. In this study, we developed a feeder- and xeno-free culture protocol that efficiently induced human pluripotent stem cells (iPSCs into PLZF+/GPR125+/CD90+ spermatogonium-like cells (SLCs. These SLCs were enriched with key genes in germ cell development such as MVH, DAZL, GFRα1, NANOS3, and DMRT1. In addition, a small fraction of SLCs went through meiosis in vitro to develop into haploid cells. We further demonstrated that this chemically defined induction protocol faithfully recapitulated the features of compromised germ cell development of PSCs with NANOS3 deficiency or iPSC lines established from patients with non-obstructive azoospermia. Taken together, we established a powerful experimental platform to investigate human germ cell development and pathology related to male infertility. : In this article, Wang and colleagues established a feeder- and xeno-free system to robustly induce human pluripotent stem cells (PSCs into spermatogonia-like cells. This chemically defined induction protocol faithfully recapitulated the features of compromised germ cell development of PSCs with NANOS3 deficiency or iPSC lines established from patients with non-obstructive azoospermia. Keywords: pluripotent stem cells, spermatogonia, infertility, non-obstructive azoospermia

High-level expression of human insulin receptor cDNA in mouse NIH 3T3 cells

International Nuclear Information System (INIS)

Whittaker, J.; Okamoto, A.K.; Thys, R.; Bell, G.I.; Steiner, D.F.; Hofmann, C.A.

1987-01-01

In order to develop a simple, efficient system for the high-level expression of human insulin receptors in eukaryotic cells, a full-length human kidney insulin receptor cDNA was inserted into a bovine papilloma virus vector under the control of the mouse metallothionein promoter. After transfection of mouse NIH 3T3 cells with this construct, seven cell lines expressing insulin receptors were isolated; two cell lines had more than 10 6 receptors per cell. The cell line with the highest 125 I-insulin binding (NIH 3T3 HIR3.5) had 6 x 10 6 receptors with a K/sub d/ of 10 -9 M. This level was not dependent on exposure to metals but could be increased further to 2 x 10 7 receptors per cell by addition of sodium butyrate to the culture medium. The α and β subunits had apparent molecular weights of 147,000 and 105,000, respectively (compared to 135,000 and 95,000 in IM-9 human lymphocytes), values identical to those of the α and β subunits of the insulin receptors of nontransformed NIH 3T3 cells. This size difference was due to altered carbohydrate composition, as N-glycanase digestion reduced the apparent receptor subunit size of the transfected cells and IM-9 lymphocytes to identical values. The alteration in N-linked oligosaccharide composition could not be ascribed to differences in the kinetics of posttranslational processing of the insulin receptors, which was comparable to that of other cells studied. The basal rate of glycogen synthesis in the cells overexpressing insulin receptors was increased 4- to 5-fold compared with controls. Low levels of added insulin (0.1 nM) caused a 50% increase in the rate of glycogen synthesis

Development of hematopoietic stem and progenitor cells from human pluripotent stem cells.

Science.gov (United States)

Chen, Tong; Wang, Fen; Wu, Mengyao; Wang, Zack Z

2015-07-01

Human pluripotent stem cells (hPSCs), including human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs), provide a new cell source for regenerative medicine, disease modeling, drug discovery, and preclinical toxicity screening. Understanding of the onset and the sequential process of hematopoietic cells from differentiated hPSCs will enable the achievement of personalized medicine and provide an in vitro platform for studying of human hematopoietic development and disease. During embryogenesis, hemogenic endothelial cells, a specified subset of endothelial cells in embryonic endothelium, are the primary source of multipotent hematopoietic stem cells. In this review, we discuss current status in the generation of multipotent hematopoietic stem and progenitor cells from hPSCs via hemogenic endothelial cells. We also review the achievements in direct reprogramming from non-hematopoietic cells to hematopoietic stem and progenitor cells. Further characterization of hematopoietic differentiation in hPSCs will improve our understanding of blood development and expedite the development of hPSC-derived blood products for therapeutic purpose. © 2015 Wiley Periodicals, Inc.

Exposure to ultrafine particles, intracellular production of reactive oxygen species in leukocytes and altered levels of endothelial progenitor cells

DEFF Research Database (Denmark)

Jantzen, Kim; Møller, Peter Horn; Karottki, Dorina Gabriela

2016-01-01

. Additionally, the early endothelial progenitor cell levels were positively associated with a personalised measure of ultrafine particle exposure and negatively associated with both basal and capacity for reactive oxygen species production in lymphocytes and granulocytes, respectively. Our results indicate......Exposure to particles in the fine and ultrafine size range has been linked to induction of low-grade systemic inflammation, oxidative stress and development of cardiovascular diseases. Declining levels of endothelial progenitor cells within systemic circulation have likewise been linked...... to progression of cardiovascular diseases. The objective was to determine if exposure to fine and ultrafine particles from indoor and outdoor sources, assessed by personal and residential indoor monitoring, is associated with altered levels of endothelial progenitor cells, and whether such effects are related...

Low-level laser irradiation induces in vitro proliferation of mesenchymal stem cells

International Nuclear Information System (INIS)

Barboza, Carlos Augusto Galvão; Ginani, Fernanda; Soares, Diego Moura; Henriques, Águida Cristina Gomes; Freitas, Roseana de Almeida

2014-01-01

To evaluate the effect of low-level laser irradiation on the proliferation and possible nuclear morphological changes of mouse mesenchymal stem cells. Mesenchymal stem cells derived from bone marrow and adipose tissue were submitted to two applications (T0 and T48 hours) of low-level laser irradiation (660nm; doses of 0.5 and 1.0J/cm"2). The trypan blue assay was used to evaluate cell viability, and growth curves were used to analyze proliferation at zero, 24, 48, and 72 hours. Nuclear alterations were evaluated by staining with DAPI (4'-6-diamidino-2-phenylindole) at 72 hours. Bone marrow-derived mesenchymal stem cells responded to laser therapy in a dose-dependent manner. Higher cell growth was observed when the cells were irradiated with a dose of 1.0J/cm"2, especially after 24 hours (p<0.01). Adipose-derived mesenchymal stem cells responded better to a dose of 1.0J/cm"2, but higher cell proliferation was observed after 48 hours (p<0.05) and 72 hours (p<0.01). Neither nuclear alterations nor a significant change in cell viability was detected in the studied groups. Low-level laser irradiation stimulated the proliferation of mouse mesenchymal stem cells without causing nuclear alterations. The biostimulation of mesenchymal stem cells using laser therapy might be an important tool for regenerative therapy and tissue engineering

Signaling hierarchy regulating human endothelial cell development.

Science.gov (United States)

Kelly, Melissa A; Hirschi, Karen K

2009-05-01

Our present knowledge of the regulation of mammalian endothelial cell differentiation has been largely derived from studies of mouse embryonic development. However, unique mechanisms and hierarchy of signals that govern human endothelial cell development are unknown and, thus, explored in these studies. Using human embryonic stem cells as a model system, we were able to reproducibly and robustly generate differentiated endothelial cells via coculture on OP9 marrow stromal cells. We found that, in contrast to studies in the mouse, bFGF and VEGF had no specific effects on the initiation of human vasculogenesis. However, exogenous Ihh promoted endothelial cell differentiation, as evidenced by increased production of cells with cobblestone morphology that coexpress multiple endothelial-specific genes and proteins, form lumens, and exhibit DiI-AcLDL uptake. Inhibition of BMP signaling using Noggin or BMP4, specifically, using neutralizing antibodies suppressed endothelial cell formation; whereas, addition of rhBMP4 to cells treated with the hedgehog inhibitor cyclopamine rescued endothelial cell development. Our studies revealed that Ihh promoted human endothelial cell differentiation from pluripotent hES cells via BMP signaling, providing novel insights applicable to modulating human endothelial cell formation and vascular regeneration for human clinical therapies.

SCA-1 Expression Level Identifies Quiescent Hematopoietic Stem and Progenitor Cells

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Mina N.F. Morcos

2017-06-01

Full Text Available Blood cell generation depends on continuous cellular output by the sequential hierarchy of hematopoietic stem cell (HSC and progenitor populations that all contain quiescent and actively cycling cells. Hematopoietic stem and progenitor cells (HSPCs express the surface molecule Stem cell antigen 1 (SCA-1/LY6A. Using histone 2B-red fluorescent fusion protein label retention and cell-cycle reporter mice, we demonstrate that high SCA-1 expression (SCA-1hi identifies not only quiescent HSCs but quiescent cells on all hierarchical levels within the lineage−SCA-1+KIT+ (LSK population. Each transplanted SCA-1hi HSPC population also displayed self-renewal potential superior to that of the respective SCA-1lo population. SCA-1 expression is inducible by type I interferon (IFN. We show, however, that quiescence and high self-renewal capacity of cells with brighter SCA-1 expression at steady state were independent of type I IFN signaling. We conclude that SCA-1 expression levels can be used to prospectively isolate functionally heterogeneous HSPC subpopulations.

Load leveling total system. Part 2. Development of load leveling logic for residential customer; Fuka heijunka total system. 2. Kateiyo juyoka wo taisho to shita heijunka ronri no kaihatsu

Energy Technology Data Exchange (ETDEWEB)

Asari, M.; Nanahara, T. [Central Research Institute of Electric Power Industry, Tokyo (Japan)

1995-05-01

It is essential, in order to meet steadily increasing demands for electrical power, to develop techniques for effective utilization of energy and load leveling. Described herein is development, by the aid of linear programming, of logic for daily management of charge/discharge of load conditioners and reverse power flow, for predicted loads and patterns of power generated by photovoltaic cells installed at individual customers. It is aimed at minimizing power rate and leveling of load at distribution systems. Predicted loads, outputs by photovoltaic cell units and different power rates by time zone for the next day are inputted, to determine the charge/discharge schedules and power supply/reverse flow patterns for that day, in order to minimize power rates and level loads at higher hierarchical levels. The logic-aided daily simulation for various districts confirms the operational patterns that realize improved utilization of pole-mounted transformers while reducing costs at customers, and effects of prediction errors. 4 refs., 14 figs.

Fuel Cell Development and Test Laboratory | Energy Systems Integration

Science.gov (United States)

Facility | NREL Fuel Cell Development and Test Laboratory Fuel Cell Development and Test Laboratory The Energy System Integration Facility's Fuel Cell Development and Test Laboratory supports fuel cell research and development projects through in-situ fuel cell testing. Photo of a researcher running

BolA inhibits cell elongation and regulates MreB expression levels.

Science.gov (United States)

Freire, Patrick; Moreira, Ricardo Neves; Arraiano, Cecília Maria

2009-02-06

The morphogene bolA is a general stress response gene in Escherichia coli that induces a round morphology when overexpressed. Results presented in this report show that increased BolA levels can inhibit cell elongation mechanisms. MreB polymerization is crucial for the bacterial cell cytoskeleton, and this protein is essential for the maintenance of a cellular rod shape. In this report, we demonstrate that bolA overexpression affects the architecture of MreB filaments. An increase in BolA leads to a significant reduction in MreB protein levels and mreB transcripts. BolA affects the mreBCD operon in vivo at the level of transcription. Furthermore, our results show that BolA is a new transcriptional repressor of MreB. The alterations in cell morphology induced by bolA seem to be mediated by a complex pathway that integrates PBP5, PBP6, MreB, and probably other regulators of cell morphology/elongation.

Genome-Wide Analyses Suggest Mechanisms Involving Early B-Cell Development in Canine IgA Deficiency.

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Mia Olsson

Full Text Available Immunoglobulin A deficiency (IgAD is the most common primary immune deficiency disorder in both humans and dogs, characterized by recurrent mucosal tract infections and a predisposition for allergic and other immune mediated diseases. In several dog breeds, low IgA levels have been observed at a high frequency and with a clinical resemblance to human IgAD. In this study, we used genome-wide association studies (GWAS to identify genomic regions associated with low IgA levels in dogs as a comparative model for human IgAD. We used a novel percentile groups-approach to establish breed-specific cut-offs and to perform analyses in a close to continuous manner. GWAS performed in four breeds prone to low IgA levels (German shepherd, Golden retriever, Labrador retriever and Shar-Pei identified 35 genomic loci suggestively associated (p <0.0005 to IgA levels. In German shepherd, three genomic regions (candidate genes include KIRREL3 and SERPINA9 were genome-wide significantly associated (p <0.0002 with IgA levels. A ~20kb long haplotype on CFA28, significantly associated (p = 0.0005 to IgA levels in Shar-Pei, was positioned within the first intron of the gene SLIT1. Both KIRREL3 and SLIT1 are highly expressed in the central nervous system and in bone marrow and are potentially important during B-cell development. SERPINA9 expression is restricted to B-cells and peaks at the time-point when B-cells proliferate into antibody-producing plasma cells. The suggestively associated regions were enriched for genes in Gene Ontology gene sets involving inflammation and early immune cell development.

Transcript levels of several epigenome regulatory genes in bovine somatic donor cells are not correlated with their cloning efficiency.

Science.gov (United States)

Zhou, Wenli; Sadeghieh, Sanaz; Abruzzese, Ronald; Uppada, Subhadra; Meredith, Justin; Ohlrichs, Charletta; Broek, Diane; Polejaeva, Irina

2009-09-01

Among many factors that potentially affect somatic cell nuclear transfer (SCNT) embryo development is the donor cell itself. Cloning potentials of somatic donor cells vary greatly, possibly because the cells have different capacities to be reprogrammed by ooplasma. It is therefore intriguing to identify factors that regulate the reprogrammability of somatic donor cells. Gene expression analysis is a widely used tool to investigate underlying mechanisms of various phenotypes. In this study, we conducted a retrospective analysis investigating whether donor cell lines with distinct cloning efficiencies express different levels of genes involved in epigenetic reprogramming including histone deacetylase-1 (HDAC1), -2 (HDAC2); DNA methyltransferase-1 (DNMT1), -3a (DNMT3a),-3b (DNMT3b), and the bovine homolog of yeast sucrose nonfermenting-2 (SNF2L), a SWI/SNF family of ATPases. Cell samples from 12 bovine donor cell lines were collected at the time of nuclear transfer experiments and expression levels of the genes were measured using quantitative polymerase chain reaction (PCR). Our results show that there are no significant differences in expression levels of these genes between donor cell lines of high and low cloning efficiency defined as live calving rates, although inverse correlations are observed between in vitro embryo developmental rates and expression levels of HDAC2 and SNF2L. We also show that selection of stable reference genes is important for relative quantification, and different batches of cells can have different gene expression patterns. In summary, we demonstrate that expression levels of these epigenome regulatory genes in bovine donor cells are not correlated with cloning potential. The experimental design and data analysis method reported here can be applied to study any genes expressed in donor cells.

Effect of cryopreservation and in vitro culture of bovine fibroblasts on histone acetylation levels and in vitro development of hand-made cloned embryos

Science.gov (United States)

Chacon, L.; Gomez, M.C.; Jenkins, J.A.; Leibo, S.P.; Wirtu, G.; Dresser, B.L.; Pope, C.E.

2011-01-01

In this study, the relative acetylation levels of histone 3 in lysine 9 (H3K9ac) in cultured and cryopreserved bovine fibroblasts was measured and we determined the influence of the epigenetic status of three cultured (C1, C2 and C3) donor cell lines on the in vitro development of reconstructed bovine embryos. Results showed that cryopreservation did not alter the overall acetylation levels of H3K9 in bovine fibroblasts analysed immediately after thawing (frozen/thawed) compared with fibroblasts cultured for a period of time after thawing. However, reduced cleavage rates were noted in embryos reconstructed with fibroblasts used immediately after thawing. Cell passage affects the levels of H3K9ac in bovine fibroblasts, decreasing after P1 and donor cells with lower H3K9ac produced a greater frequency of embryo development to the blastocyst stage. Cryopreservation did not influence the total cell and ICM numbers, or the ICM/TPD ratios of reconstructed embryos. However, the genetic source of donor cells did influence the total number of cells and the trophectoderm cell numbers, and the cell passage influenced the total ICM cell numbers. ?? Copyright Cambridge University Press 2010.

High-level production of human interleukin-10 fusions in tobacco cell suspension cultures

Science.gov (United States)

Kaldis, Angelo; Ahmad, Adil; Reid, Alexandra; McGarvey, Brian; Brandle, Jim; Ma, Shengwu; Jevnikar, Anthony; Kohalmi, Susanne E; Menassa, Rima

2013-01-01

The production of pharmaceutical proteins in plants has made much progress in recent years with the development of transient expression systems, transplastomic technology and humanizing glycosylation patterns in plants. However, the first therapeutic proteins approved for administration to humans and animals were made in plant cell suspensions for reasons of containment, rapid scale-up and lack of toxic contaminants. In this study, we have investigated the production of human interleukin-10 (IL-10) in tobacco BY-2 cell suspension and evaluated the effect of an elastin-like polypeptide tag (ELP) and a green fluorescent protein (GFP) tag on IL-10 accumulation. We report the highest accumulation levels of hIL-10 obtained with any stable plant expression system using the ELP fusion strategy. Although IL-10-ELP has cytokine activity, its activity is reduced compared to unfused IL-10, likely caused by interference of ELP with folding of IL-10. Green fluorescent protein has no effect on IL-10 accumulation, but examining the trafficking of IL-10-GFP over the cell culture cycle revealed fluorescence in the vacuole during the stationary phase of the culture growth cycle. Analysis of isolated vacuoles indicated that GFP alone is found in vacuoles, while the full-size fusion remains in the whole-cell extract. This indicates that GFP is cleaved off prior to its trafficking to the vacuole. On the other hand, IL-10-GFP-ELP remains mostly in the ER and accumulates to high levels. Protein bodies were observed at the end of the culture cycle and are thought to arise as a consequence of high levels of accumulation in the ER. PMID:23297698

Low-level laser irradiation induces in vitro proliferation of mesenchymal stem cells

Energy Technology Data Exchange (ETDEWEB)

Barboza, Carlos Augusto Galvão; Ginani, Fernanda [Universidade Federal do Rio Grande do Norte, Natal, RN (Brazil); Soares, Diego Moura [Universidade Federal de Pernambuco, Recife, PE (Brazil); Henriques, Águida Cristina Gomes; Freitas, Roseana de Almeida [Universidade Federal do Rio Grande do Norte, Natal, RN (Brazil)

2014-07-01

To evaluate the effect of low-level laser irradiation on the proliferation and possible nuclear morphological changes of mouse mesenchymal stem cells. Mesenchymal stem cells derived from bone marrow and adipose tissue were submitted to two applications (T0 and T48 hours) of low-level laser irradiation (660nm; doses of 0.5 and 1.0J/cm{sup 2}). The trypan blue assay was used to evaluate cell viability, and growth curves were used to analyze proliferation at zero, 24, 48, and 72 hours. Nuclear alterations were evaluated by staining with DAPI (4'-6-diamidino-2-phenylindole) at 72 hours. Bone marrow-derived mesenchymal stem cells responded to laser therapy in a dose-dependent manner. Higher cell growth was observed when the cells were irradiated with a dose of 1.0J/cm{sup 2}, especially after 24 hours (p<0.01). Adipose-derived mesenchymal stem cells responded better to a dose of 1.0J/cm{sup 2}, but higher cell proliferation was observed after 48 hours (p<0.05) and 72 hours (p<0.01). Neither nuclear alterations nor a significant change in cell viability was detected in the studied groups. Low-level laser irradiation stimulated the proliferation of mouse mesenchymal stem cells without causing nuclear alterations. The biostimulation of mesenchymal stem cells using laser therapy might be an important tool for regenerative therapy and tissue engineering.

Concentrator-solar-cell development

Science.gov (United States)

Grenon, L.

1982-07-01

A program is described which is a continuation of earlier programs for the development of high-efficiency, low-cost, silicon concentrator solar cells. The base-line process steps and process sequences identified in these earlier contracts were evaluated and specific processes reviewed. In particular, emphasis on the use of Czochralski-grown silicon wafers rather than float-zone wafers were examined. Additionally, a study of the trade-offs between textured and nontextured cells was initiated, and the limits within which the low-cost plated nickel copper metallization can be used in concentrator solar cell applications was identified.

Development of a Micro-Fiber Nickel Electrode for Nickel-Hydrogen Cell

Science.gov (United States)

Britton, Doris L.

1996-01-01

The development of a high specific energy battery is one of the objectives of the lightweight nickel-hydrogen (NiH2) program at the NASA Lewis Research Center. The approach has been to improve the nickel electrode by continuing combined in-house and contract efforts to develop a more efficient and lighter weight electrode for the nickel-hydrogen fuel cell. Small fiber diameter nickel plaques are used as conductive supports for the nickel hydroxide active material. These plaques are commercial products and have an advantage of increased surface area available for the deposition of active materials. Initial tests include activation and capacity measurements at different discharge levels followed by half-cell cycle testing at 80 percent depth-of-discharge in a low Earth orbit regime. The electrodes that pass the initial tests are life cycle tested in a boiler plate nickel-hydrogen cell before flightweight designs are built and tested.

Silicon Web Process Development. [for solar cell fabrication

Science.gov (United States)

Duncan, C. S.; Seidensticker, R. G.; Hopkins, R. H.; Mchugh, J. P.; Hill, F. E.; Heimlich, M. E.; Driggers, J. M.

1979-01-01

Silicon dendritic web, ribbon form of silicon and capable of fabrication into solar cells with greater than 15% AMl conversion efficiency, was produced from the melt without die shaping. Improvements were made both in the width of the web ribbons grown and in the techniques to replenish the liquid silicon as it is transformed to web. Through means of improved thermal shielding stress was reduced sufficiently so that web crystals nearly 4.5 cm wide were grown. The development of two subsystems, a silicon feeder and a melt level sensor, necessary to achieve an operational melt replenishment system, is described. A gas flow management technique is discussed and a laser reflection method to sense and control the melt level as silicon is replenished is examined.

How to Draw Energy Level Diagrams in Excitonic Solar Cells.

Science.gov (United States)

Zhu, X-Y

2014-07-03

Emerging photovoltaic devices based on molecular and nanomaterials are mostly excitonic in nature. The initial absorption of a photon in these materials creates an exciton that can subsequently dissociate in each material or at their interfaces to give charge carriers. Any attempt at mechanistic understanding of excitonic solar cells must start with drawing energy level diagrams. This seemingly elementary exercise, which is described in textbooks for inorganic solar cells, has turned out to be a difficult subject in the literature. The problem stems from conceptual confusion of single-particle energy with quasi-particle energy and the misleading practice of mixing the two on the same energy level diagram. Here, I discuss how to draw physically accurate energy diagrams in excitonic solar cells using only single-particle energies (ionization potentials and electron affinities) of both ground and optically excited states. I will briefly discuss current understanding on the electronic energy landscape responsible for efficient charge separation in excitonic solar cells.

Relationship of visfatin level to pancreatic endocrine hormone level, HOMA-IR index, and HOMA β-cell index in overweight women who performed hydraulic resistance exercise.

Science.gov (United States)

Ha, Chang Ho; Swearingin, Brenda; Jeon, Yong Kyun

2015-09-01

[Purpose] This study aimed to examine the correlation of visfatin level to pancreatic endocrine hormone level, homeostasis model assessment of insulin resistance (HOMA-IR) index, and HOMA β-cell index in hydraulic resistance exercise. Furthermore, it investigated the relationship between visfatin level and other variables affected by exercise in overweight women. [Subjects and Methods] The exercise group trained for 12 weeks, 70 minutes/day, 5 days/week. Visfatin level, pancreatic endocrine hormone level, HOMA-IR index, and HOMA β-cell index were measured before and after the intervention. Based on the blood insulin and glucose concentrations, HOMA-IR index, the indicator of insulin resistance, and HOMA β-cell index, the indicator of insulin secretion level, were assessed. [Results] Interaction effects on visfatin level, insulin level, HOMA-IR index, and HOMA β-cell index were observed. Interaction effects on glucagon and glucose levels were not observed between the intervention groups. The correlations of visfatin level to insulin, glucagon, and glucose levels, and HOMA-IR and HOMA β-cell indexes were not significant for any of the subjects. [Conclusion] Therefore, the 12-week resistance exercise affected body composition, visfatin level, insulin level, HOMA-IR index, and HOMA β-cell index. Finally, visfatin was not related to insulin, glucagon, and glucose levels, and HOMA-IR and HOMA β-cell indexes.

Development of planar SOE/SOFC reversible cell

International Nuclear Information System (INIS)

Kusunoki, A.; Matsubara, H.; Kikuoka, Y.; Yanagi, C.; Kugimiya, K.; Yoshino, M.; Tokura, M.; Watanabe, K.; Ueda, S.; Sumi, M.; Miyamoto, H.; Tokunaga, S.

1993-01-01

A new energy storage system using SOE/SOFC (solid oxide electrolysis-solid oxide fuel cells) reversible cells is presented, where a unit cell works as a fuel cell during a period of high electric power demand and alternately works as an electrolysis cell during a period of low power demand. A planar cell configuration is used which allows for a compact and low cost energy storage and load leveling system for power stations. Tests were performed to verify the reversibility of the planar cell, at 1000 deg C, with YSZ (Yttria stabilized zirconia) as the solid electrolyte, to improve the cell performance by reducing the overvoltage in electrolysis, and to obtain fundamental characteristics of a reversible cell. 3 figs

Layout level design for testability strategy applied to a CMOS cell library

NARCIS (Netherlands)

Blom, F.C.; Oliver, J.; Rullan, M.; Ferrer, C.

1993-01-01

The layout level design for testability (LLDFT) rules used here allow to avoid some hard to detect faults or even undetectable faults on a cell library by modifying the cell layout without changing their behavior and achieving a good level of reliability. These rules avoid some open faults or reduce

High Concentration of Melatonin Regulates Leaf Development by Suppressing Cell Proliferation and Endoreduplication in Arabidopsis.

Science.gov (United States)

Wang, Qiannan; An, Bang; Shi, Haitao; Luo, Hongli; He, Chaozu

2017-05-05

N -acetyl-5-methoxytryptamine (Melatonin), as a crucial messenger in plants, functions in adjusting biological rhythms, stress tolerance, plant growth and development. Several studies have shown the retardation effect of exogenous melatonin treatment on plant growth and development. However, the in vivo role of melatonin in regulating plant leaf growth and the underlying mechanism are still unclear. In this study, we found that high concentration of melatonin suppressed leaf growth in Arabidopsis by reducing both cell size and cell number. Further kinetic analysis of the fifth leaves showed that melatonin remarkably inhibited cell division rate. Additionally, flow cytometic analysis indicated that melatonin negatively regulated endoreduplication during leaf development. Consistently, the expression analysis revealed that melatonin regulated the transcriptional levels of key genes of cell cycle and ribosome. Taken together, this study suggests that high concentration of melatonin negatively regulated the leaf growth and development in Arabidopsis , through modulation of endoreduplication and the transcripts of cell cycle and ribosomal key genes.

Nano-scale microfluidics to study 3D chemotaxis at the single cell level.

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Corina Frick

Full Text Available Directed migration of cells relies on their ability to sense directional guidance cues and to interact with pericellular structures in order to transduce contractile cytoskeletal- into mechanical forces. These biomechanical processes depend highly on microenvironmental factors such as exposure to 2D surfaces or 3D matrices. In vivo, the majority of cells are exposed to 3D environments. Data on 3D cell migration are mostly derived from intravital microscopy or collagen-based in vitro assays. Both approaches offer only limited controllability of experimental conditions. Here, we developed an automated microfluidic system that allows positioning of cells in 3D microenvironments containing highly controlled diffusion-based chemokine gradients. Tracking migration in such gradients was feasible in real time at the single cell level. Moreover, the setup allowed on-chip immunocytochemistry and thus linking of functional with phenotypical properties in individual cells. Spatially defined retrieval of cells from the device allows down-stream off-chip analysis. Using dendritic cells as a model, our setup specifically allowed us for the first time to quantitate key migration characteristics of cells exposed to identical gradients of the chemokine CCL19 yet placed on 2D vs in 3D environments. Migration properties between 2D and 3D migration were distinct. Morphological features of cells migrating in an in vitro 3D environment were similar to those of cells migrating in animal tissues, but different from cells migrating on a surface. Our system thus offers a highly controllable in vitro-mimic of a 3D environment that cells traffic in vivo.

Cyclic adenosine monophosphate levels and the function of skin microvascular endothelial cells.

Science.gov (United States)

Tuder, R M; Karasek, M A; Bensch, K G

1990-02-01

The maintenance of the normal epithelioid morphology of human dermal microvascular endothelial cells (MEC) grown in vitro depends strongly on the presence of factors that increase intracellular levels of cyclic AMP. Complete removal of dibutyryl cAMP and isobutylmethylxanthine (IMX) from the growth medium results in a progressive transition from an epithelioid to a spindle-shaped cell line. This transition cannot be reversed by the readdition of dibutyryl cAMP and IMX to the growth medium or by addition of agonists that increase cAMP levels. Spindle-shaped MEC lose the ability to express Factor VIII rAG and DR antigens and to bind peripheral blood mononuclear leukocyte (PBML). Ultrastructural analyses of transitional cells and spindle-shaped cells show decreased numbers of Weibel-Palade bodies in transitional cells and their complete absence in spindle-shaped cells. Interferon-gamma alters several functional properties of both epithelioid and spindle-shaped cells. In the absence of dibutyryl cAMP it accelerates the transition from epithelial to spindle-shaped cells, whereas in the presence of cyclic AMP interferon-gamma increases the binding of PBMLs to both epithelioid and spindle-shaped MEC and the endocytic activity of the endothelial cells. These results suggest that cyclic AMP is an important second messenger in the maintenance of several key functions of microvascular endothelial cells. Factors that influence the levels of this messenger in vivo can be expected to influence the angiogenic and immunologic functions of the microvasculature.

Protein Expression Analyses at the Single Cell Level

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Masae Ohno

2014-09-01

Full Text Available The central dogma of molecular biology explains how genetic information is converted into its end product, proteins, which are responsible for the phenotypic state of the cell. Along with the protein type, the phenotypic state depends on the protein copy number. Therefore, quantification of the protein expression in a single cell is critical for quantitative characterization of the phenotypic states. Protein expression is typically a dynamic and stochastic phenomenon that cannot be well described by standard experimental methods. As an alternative, fluorescence imaging is being explored for the study of protein expression, because of its high sensitivity and high throughput. Here we review key recent progresses in fluorescence imaging-based methods and discuss their application to proteome analysis at the single cell level.

Molecular Imaging: A Useful Tool for the Development of Natural Killer Cell-Based Immunotherapies

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Prakash Gangadaran

2017-09-01

Full Text Available Molecular imaging is a relatively new discipline that allows visualization, characterization, and measurement of the biological processes in living subjects, including humans, at a cellular and molecular level. The interaction between cancer cells and natural killer (NK cells is complex and incompletely understood. Despite our limited knowledge, progress in the search for immune cell therapies against cancer could be significantly improved by dynamic and non-invasive visualization and tracking of immune cells and by visualization of the response of cancer cells to therapies in preclinical and clinical studies. Molecular imaging is an essential tool for these studies, and a multimodal molecular imaging approach can be applied to monitor immune cells in vivo, for instance, to visualize therapeutic effects. In this review, we discuss the usefulness of NK cells in cancer therapies and the preclinical and clinical usefulness of molecular imaging in NK cell-based therapies. Furthermore, we discuss different molecular imaging modalities for use with NK cell-based therapies, and their preclinical and clinical applications in animal and human subjects. Molecular imaging has contributed to the development of NK cell-based therapies against cancers in animal models and to the refinement of current cell-based cancer immunotherapies. Developing sensitive and reproducible non-invasive molecular imaging technologies for in vivo NK cell monitoring and for real-time assessment of therapeutic effects will accelerate the development of NK cell therapies.

Does Ploidy Level Directly Control Cell Size? Counterevidence from Arabidopsis Genetics

OpenAIRE

Tsukaya, Hirokazu

2013-01-01

Ploidy level affects cell size in many organisms, and ploidy-dependent cell enlargement has been used to breed many useful organisms. However, how polyploidy affects cell size remains unknown. Previous studies have explored changes in transcriptome data caused by polyploidy, but have not been successful. The most naïve theory explaining ploidy-dependent cell enlargement is that increases in gene copy number increase the amount of protein, which in turn increases the cell volume. This hypothes...

Iκb Kinase α Is Essential for Mature B Cell Development and Function

Science.gov (United States)

Kaisho, Tsuneyasu; Takeda, Kiyoshi; Tsujimura, Tohru; Kawai, Taro; Nomura, Fumiko; Terada, Nobuyuki; Akira, Shizuo

2001-01-01

IκB kinase (IKK) α and β phosphorylate IκB proteins and activate the transcription factor, nuclear factor (NF)-κB. Although both are highly homologous kinases, gene targeting experiments revealed their differential roles in vivo. IKKα is involved in skin and limb morphogenesis, whereas IKKβ is essential for cytokine signaling. To elucidate in vivo roles of IKKα in hematopoietic cells, we have generated bone marrow chimeras by transferring control and IKKα-deficient fetal liver cells. The mature B cell population was decreased in IKKα−/− chimeras. IKKα−/− chimeras also exhibited a decrease of serum immunoglobulin basal level and impaired antigen-specific immune responses. Histologically, they also manifested marked disruption of germinal center formation and splenic microarchitectures that depend on mature B cells. IKKα−/− B cells not only showed impairment of survival and mitogenic responses in vitro, accompanied by decreased, although inducible, NF-κB activity, but also increased turnover rate in vivo. In addition, transgene expression of bcl-2 could only partially rescue impaired B cell development in IKKα−/− chimeras. Taken together, these results demonstrate that IKKα is critically involved in the prevention of cell death and functional development of mature B cells. PMID:11181694

Lower adiponectin levels at first trimester of pregnancy are associated with increased insulin resistance and higher risk of developing gestational diabetes mellitus.

Science.gov (United States)

Lacroix, Marilyn; Battista, Marie-Claude; Doyon, Myriam; Ménard, Julie; Ardilouze, Jean-Luc; Perron, Patrice; Hivert, Marie-France

2013-06-01

To evaluate the associations between adiponectin levels and 1) the risk of developing gestational diabetes mellitus (GDM), and 2) insulin resistance/sensitivity, β-cell function, and compensation indices in a prospective cohort representative of the general population of pregnant women. We performed anthropometric measurements and collected blood samples at 1st (6-13 weeks) and 2nd (24-28 weeks) trimesters. Diagnosis of GDM was made at 2nd trimester based on a 75-g oral glucose tolerance test (International Association of the Diabetes and Pregnancy Study Groups criteria). Insulin was measured (ELISA; Luminex) to estimate homeostasis model assessment of insulin resistance (HOMA-IR), β-cell function (HOMA-B), insulin sensitivity (Matsuda index), insulin secretion (AUC(insulin/glucose)), and β-cell compensation (insulin secretion sensitivity index-2). Adiponectin was measured by radioimmunoassay. Among the 445 participants included in this study, 38 women developed GDM. Women who developed GDM had lower 1st-trimester adiponectin levels (9.67 ± 3.84 vs. 11.92 ± 4.59 µg/mL in women with normal glucose tolerance). Lower adiponectin levels were associated with higher risk of developing GDM (OR, 1.12 per 1 µg/mL decrease of adiponectin levels; P = 0.02, adjusted for BMI and HbA1c at 1st trimester). Adiponectin levels at 1st and 2nd trimesters were associated with HOMA-IR (both: r = -0.22, P insulin/glucose). Pregnant women with lower adiponectin levels at 1st trimester have higher levels of insulin resistance and are more likely to develop GDM independently of adiposity or glycemic measurements.

Progress and challenges in the development of a cell-based therapy for hemophilia A.

Science.gov (United States)

Fomin, M E; Togarrati, P P; Muench, M O

2014-12-01

Hemophilia A results from an insufficiency of factor VIII (FVIII). Although replacement therapy with plasma-derived or recombinant FVIII is a life-saving therapy for hemophilia A patients, such therapy is a life-long treatment rather than a cure for the disease. In this review, we discuss the possibilities, progress, and challenges that remain in the development of a cell-based cure for hemophilia A. The success of cell therapy depends on the type and availability of donor cells, the age of the host and method of transplantation, and the levels of engraftment and production of FVIII by the graft. Early therapy, possibly even prenatal transplantation, may yield the highest levels of engraftment by avoiding immunological rejection of the graft. Potential cell sources of FVIII include a specialized subset of endothelial cells known as liver sinusoidal endothelial cells (LSECs) present in the adult and fetal liver, or patient-specific endothelial cells derived from induced pluripotent stem cells that have undergone gene editing to produce FVIII. Achieving sufficient engraftment of transplanted LSECs is one of the obstacles to successful cell therapy for hemophilia A. We discuss recent results from transplants performed in animals that show production of functional and clinically relevant levels of FVIII obtained from donor LSECs. Hence, the possibility of treating hemophilia A can be envisioned through persistent production of FVIII from transplanted donor cells derived from a number of potential cell sources or through creation of donor endothelial cells from patient-specific induced pluripotent stem cells. © 2014 International Society on Thrombosis and Haemostasis.

Involvement of host stroma cells and tissue fibrosis in pancreatic tumor development in transgenic mice.

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Itai Spector

Full Text Available INTRODUCTION: Stroma cells and extracellular matrix (ECM components provide the pivotal microenvironment for tumor development. The study aimed to evaluate the importance of the pancreatic stroma for tumor development. METHODS: Pancreatic tumor cells were implanted subcutaneously into green fluorescent protein transgenic mice, and stroma cells invading the tumors were identified through immunohistochemistry. Inhibition of tumor invasion by stroma cells was achieved with halofuginone, an inhibitor of TGFβ/Smad3 signaling, alone or in combination with chemotherapy. The origin of tumor ECM was evaluated with species-specific collagen I antibodies and in situ hybridization of collagen α1(I gene. Pancreatic fibrosis was induced by cerulean injection and tumors by spleen injection of pancreatic tumor cells. RESULTS: Inhibition of stroma cell infiltration and reduction of tumor ECM levels by halofuginone inhibited development of tumors derived from mouse and human pancreatic cancer cells. Halofuginone reduced the number only of stroma myofibroblasts expressing both contractile and collagen biosynthesis markers. Both stroma myofibroblasts and tumor cells generated ECM that contributes to tumor growth. Combination of treatments that inhibit stroma cell infiltration, cause apoptosis of myofibroblasts and inhibit Smad3 phosphorylation, with chemotherapy that increases tumor-cell apoptosis without affecting Smad3 phosphorylation was more efficacious than either treatment alone. More tumors developed in fibrotic than in normal pancreas, and prevention of tissue fibrosis greatly reduced tumor development. CONCLUSIONS: The utmost importance of tissue fibrosis and of stroma cells for tumor development presents potential new therapy targets, suggesting combination therapy against stroma and neoplastic cells as a treatment of choice.

324 Radiochemical engineering cells and high level vault tanks mixed waste compliance status

International Nuclear Information System (INIS)

1994-01-01

The 324 Building in the Hanford 300 Area contains Radiochemical Engineering Cells and High Level Vault tanks (the open-quotes REC/HLVclose quotes) for research and development activities involving radioactive materials. Radioactive mixed waste within this research installation, found primarily in B-Cell and three of the high level vault tanks, is subject to RCRA/DWR (open-quotes RCRAclose quotes) regulations for storage. This white paper provides a baseline RCRA compliance summary of MW management in the REC/HLV, based on best available knowledge. The REC/HLV compliance project, of which this paper is a part, is intended to achieve the highest degree of compliance practicable given the special technical difficulties of managing high activity radioactive materials, and to assure protection of human health and safety and the environment. The REC/HLV was constructed in 1965 to strict standards for the safe management of highly radioactive materials. Mixed waste in the REC/HLV consists of discarded tools and equipment, dried feed stock from nuclear waste melting experiments, contaminated particulate matter, and liquid feed stock from various experimental programs in the vault tanks. B-Cell contains most of these materials. Total radiological inventory in B-Cell is estimated at 3 MCi, about half of which is potentially open-quotes dispersibleclose quotes, that is, it is in small pieces or mobile particles. Most of the mixed waste currently in the REC/HLV was generated or introduced before mixed wastes were subjected to RCRA in 1987

A low-level diode laser therapy reduces the lipopolysaccharide (LPS)-induced periodontal ligament cell inflammation

International Nuclear Information System (INIS)

Huang, T H; Chen, C C; Liu, S L; Lu, Y C; Kao, C T

2014-01-01

The purpose of this study was to investigate the cytologic effects of inflammatory periodontal ligament cells in vitro after low-level laser therapy. Human periodontal ligament cells were cultured, exposed to lipopolysaccharide and subjected to low-level laser treatment of 5 J cm −2 or 10 J cm −2 using a 920 nm diode laser. A periodontal ligament cell attachment was observed under a microscope, and the cell viability was quantified by a mitochondrial colorimetric assay. Lipopolysaccharide-treated periodontal ligament cells were irradiated with the low-level laser, and the expression levels of several inflammatory markers, iNOS, TNF-α and IL-1, and pErk kinase, were analyzed by reverse transcription polymerase chain reaction and western blot. The data were collected and analyzed by one-way analysis of variance; p < 0.05 indicated a statistically significant difference. The low-level laser treatment of periodontal ligament cells increased their ability to attach and survive. After irradiation, the expression levels of iNOS, TNF-α and IL-1 in lipopolysaccharide-exposed periodontal ligament cells decreased over time (p < 0.05). In periodontal ligament cells, low-level diode laser treatment increased the cells’ proliferative ability and decreased the expression of the examined inflammatory mediators. (letters)

Resolving tumor heterogeneity: genes involved in chordoma cell development identified by low-template analysis of morphologically distinct cells.

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Amin El-Heliebi

Full Text Available The classical sacrococcygeal chordoma tumor presents with a typical morphology of lobulated myxoid tumor tissue with cords, strands and nests of tumor cells. The population of cells consists of small non-vacuolated cells, intermediate cells with a wide range of vacuolization and large heavily vacuolated (physaliferous cells. To date analysis was only performed on bulk tumor mass because of its rare incidence, lack of suited model systems and technical limitations thereby neglecting its heterogeneous composition. We intended to clarify whether the observed cell types are derived from genetically distinct clones or represent different phenotypes. Furthermore, we aimed at elucidating the differences between small non-vacuolated and large physaliferous cells on the genomic and transcriptomic level. Phenotype-specific analyses of small non-vacuolated and large physaliferous cells in two independent chordoma cell lines yielded four candidate genes involved in chordoma cell development. UCHL3, coding for an ubiquitin hydrolase, was found to be over-expressed in the large physaliferous cell phenotype of MUG-Chor1 (18.7-fold and U-CH1 (3.7-fold cells. The mannosyltransferase ALG11 (695-fold and the phosphatase subunit PPP2CB (18.6-fold were found to be up-regulated in large physaliferous MUG-Chor1 cells showing a similar trend in U-CH1 cells. TMEM144, an orphan 10-transmembrane family receptor, yielded contradictory data as cDNA microarray analysis showed up- but RT-qPCR data down-regulation in large physaliferous MUG-Chor1 cells. Isolation of few but morphologically identical cells allowed us to overcome the limitations of bulk analysis in chordoma research. We identified the different chordoma cell phenotypes to be part of a developmental process and discovered new genes linked to chordoma cell development representing potential targets for further research in chordoma tumor biology.

Acquired tolerance in cadmium-adapted lung epithelial cells: Roles of the c-Jun N-terminal kinase signaling pathway and basal level of metallothionein

International Nuclear Information System (INIS)

Lau, Andy T.Y.; Zhang Jian; Chiu, J.-F.

2006-01-01

Cadmium-resistant cells were developed in our laboratory with rat lung epithelial cells (LECs) by stepwise exposure of LECs to cadmium chloride from 1 μM to 20 μM after 20 passages. To investigate the Cd-resistant phenotype in a long-term perspective, cadmium-resistant cells adapted to 20 μM cadmium (Cd R ) were then cultured in the absence of cadmium for various passages [Cd R (-n)]. All these adapted cells were significantly protected from cadmium toxicity as compared to parental cadmium-sensitive LECs (Cd S ). The cadmium-resistant phenotype of adapted cells was relatively stable in the absence of cadmium for as long as 40 passages. Basal mRNA level of metallothionein-1 (MT-1) was dramatically higher in Cd R than in Cd R (-), which may account for the higher Cd-resistance of Cd R than Cd R (-). MT-1 mRNA level decreased drastically in Cd R after cadmium removal, suggesting that the high basal level of MT-1 in Cd R may be only partially responsible for cadmium-resistance. Treatment of cells with high levels of cadmium resulted in decreased phosphorylation of c-Jun N-terminal kinase (JNK1/2) in adapted cells than in sensitive cells and this cadmium-induced JNK activity was blocked by JNK inhibitor II, SP600125. Ro318220, a strong activator of JNK, reverted cadmium-sensitive phenotype in adapted cells. Taken together, our results suggest that during cadmium adaptation, cells develop tolerance to cell death, generally due to perturbation of the JNK signaling pathway and the nonresponsiveness of JNK phosphorylation is critical for the Cd-tolerance in these cells

A novel Atoh1 "self-terminating" mouse model reveals the necessity of proper Atoh1 level and duration for hair cell differentiation and viability.

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Ning Pan

Full Text Available Atonal homolog1 (Atoh1 is a bHLH transcription factor essential for inner ear hair cell differentiation. Targeted expression of Atoh1 at various stages in development can result in hair cell differentiation in the ear. However, the level and duration of Atoh1 expression required for proper hair cell differentiation and maintenance remain unknown. We generated an Atoh1 conditional knockout (CKO mouse line using Tg(Atoh1-cre, in which the cre expression is driven by an Atoh1 enhancer element that is regulated by Atoh1 protein to "self-terminate" its expression. The mutant mice show transient, limited expression of Atoh1 in all hair cells in the ear. In the organ of Corti, reduction and delayed deletion of Atoh1 result in progressive loss of almost all the inner hair cells and the majority of the outer hair cells within three weeks after birth. The remaining cells express hair cell marker Myo7a and attract nerve fibers, but do not differentiate normal stereocilia bundles. Some Myo7a-positive cells persist in the cochlea into adult stages in the position of outer hair cells, flanked by a single row of pillar cells and two to three rows of disorganized Deiters cells. Gene expression analyses of Atoh1, Barhl1 and Pou4f3, genes required for survival and maturation of hair cells, reveal earlier and higher expression levels in the inner compared to the outer hair cells. Our data show that Atoh1 is crucial for hair cell mechanotransduction development, viability, and maintenance and also suggest that Atoh1 expression level and duration may play a role in inner vs. outer hair cell development. These genetically engineered Atoh1 CKO mice provide a novel model for establishing critical conditions needed to regenerate viable and functional hair cells with Atoh1 therapy.

Progress and challenges in the development of a cell-based therapy for hemophilia A

Science.gov (United States)

Fomin, Marina E.; Togarrati, Padma Priya; Muench, Marcus O.

2015-01-01

Hemophilia A results from an insufficiency of factor VIII (FVIII). Although replacement therapy with plasma-derived or recombinant FVIII is a life-saving therapy for hemophilia A patients, such therapy is a life-long treatment rather than a cure for the disease. In this review we discuss the possibilities, progress and challenges that remain in the development of a cell-based cure for hemophilia A. The success of cell therapy depends on the type and availability of donor cells, the age of the host and method of transplantation, and the levels of engraftment and production of FVIII by the graft. Early therapy, possibly even prenatal transplantation, may yield the highest levels of engraftment by avoiding immunological rejection of the graft. Potential cell sources of FVIII include a specialized subset of endothelial cells known as liver sinusoidal endothelial cells (LSECs) present in the adult and fetal liver, or patient-specific endothelial cells derived from induced pluripotent stem cells (iPSCs) that have undergone gene editing to produce FVIII. Achieving sufficient engraftment of transplanted LSECs is one of the obstacles to successful cell therapy for hemophilia A. We discuss recent results from transplants performed in animals that show production of functional and clinically relevant levels of FVIII obtained from donor LSECs. Hence, the possibility of treating hemophilia A can be envisioned through persistent production of FVIII from transplanted donor cells derived from a number of potential cell sources or through creation of donor endothelial cells from patient-specific iPSCs. PMID:25297648

Levels of immune cells in transcendental meditation practitioners

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Jose R Infante

2014-01-01

Conclusions: The technique of meditation studied seems to have a significant effect on immune cells, manifesting in the different circulating levels of lymphocyte subsets analyzed. The significant effect of TM on the neuroendocrine axis and its relationship with the immune system may partly explain our results.

Upregulation of mitochondrial NAD+ levels impairs the clonogenicity of SSEA1+ glioblastoma tumor-initiating cells.

Science.gov (United States)

Son, Myung Jin; Ryu, Jae-Sung; Kim, Jae Yun; Kwon, Youjeong; Chung, Kyung-Sook; Mun, Seon Ju; Cho, Yee Sook

2017-06-09

Emerging evidence has emphasized the importance of cancer therapies targeting an abnormal metabolic state of tumor-initiating cells (TICs) in which they retain stem cell-like phenotypes and nicotinamide adenine dinucleotide (NAD + ) metabolism. However, the functional role of NAD + metabolism in regulating the characteristics of TICs is not known. In this study, we provide evidence that the mitochondrial NAD + levels affect the characteristics of glioma-driven SSEA1 + TICs, including clonogenic growth potential. An increase in the mitochondrial NAD + levels by the overexpression of the mitochondrial enzyme nicotinamide nucleotide transhydrogenase (NNT) significantly suppressed the sphere-forming ability and induced differentiation of TICs, suggesting a loss of the characteristics of TICs. In addition, increased SIRT3 activity and reduced lactate production, which are mainly observed in healthy and young cells, appeared following NNT-overexpressed TICs. Moreover, in vivo tumorigenic potential was substantially abolished by NNT overexpression. Conversely, the short interfering RNA-mediated knockdown of NNT facilitated the maintenance of TIC characteristics, as evidenced by the increased numbers of large tumor spheres and in vivo tumorigenic potential. Our results demonstrated that targeting the maintenance of healthy mitochondria with increased mitochondrial NAD + levels and SIRT3 activity could be a promising strategy for abolishing the development of TICs as a new therapeutic approach to treating aging-associated tumors.

Functional importance of GLP-1 receptor species and expression levels in cell lines.

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Knudsen, Lotte Bjerre; Hastrup, Sven; Underwood, Christina Rye; Wulff, Birgitte Schjellerup; Fleckner, Jan

2012-04-10

Of the mammalian species, only the GLP-1 receptors of rat and human origin have been described and characterized. Here, we report the cloning of the homologous GLP-1 receptors from mouse, rabbit, pig, cynomolgus monkey and chimp. The GLP-1 receptor is highly conserved across species, thus underlining the physiological importance of the peptide hormone and its receptor across a wide range of mammals. We expressed the receptors by stable transfection of BHK cells, both in cell lines with high expression levels of the cloned receptors, as well as in cell lines with lower expression levels, more comparable to endogenous expression of these receptors. High expression levels of cloned GLP-1 receptors markedly increased the potency of GLP-1 and other high affinity ligands, whereas the K(d) values were not affected. For a low affinity ligand like the ago-allosteric modulator Compound 2, expression levels of the human GLP-1 receptor were important for maximal efficacy as well as potency. The two natural metabolites of GLP-1, GLP-1(9-37) and GLP-1(9-36)amide were agonists when tested on a cell line with high expression of the recombinant human GLP-1 receptor, whereas they behaved as (low potent) antagonists on a cell line that expressed the receptor endogenously, as well as cells expressing a moderate level of the recombinant human GLP-1 receptor. The amide form was a more potent agonist than the free acid from. In conclusion, receptor expression level is an important parametre for selecting cell lines with cloned GLP-1 receptors for functional characterization of physiological and pharmaceutical ligands. Copyright © 2011 Elsevier B.V. All rights reserved.

Assessment of the Current Level of Automation in the Manufacture of Fuel Cell Systems for Combined Heat and Power Applications

Energy Technology Data Exchange (ETDEWEB)

Ulsh, M.; Wheeler, D.; Protopappas, P.

2011-08-01

The U.S. Department of Energy (DOE) is interested in supporting manufacturing research and development (R&D) for fuel cell systems in the 10-1,000 kilowatt (kW) power range relevant to stationary and distributed combined heat and power applications, with the intent to reduce manufacturing costs and increase production throughput. To assist in future decision-making, DOE requested that the National Renewable Energy Laboratory (NREL) provide a baseline understanding of the current levels of adoption of automation in manufacturing processes and flow, as well as of continuous processes. NREL identified and visited or interviewed key manufacturers, universities, and laboratories relevant to the study using a standard questionnaire. The questionnaire covered the current level of vertical integration, the importance of quality control developments for automation, the current level of automation and source of automation design, critical balance of plant issues, potential for continuous cell manufacturing, key manufacturing steps or processes that would benefit from DOE support for manufacturing R&D, the potential for cell or stack design changes to support automation, and the relationship between production volume and decisions on automation.

Sustained levels of FGF2 maintain undifferentiated stem cell cultures with biweekly feeding.

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Steven Lotz

Full Text Available An essential aspect of stem cell culture is the successful maintenance of the undifferentiated state. Many types of stem cells are FGF2 dependent, and pluripotent stem cells are maintained by replacing FGF2-containing media daily, while tissue-specific stem cells are typically fed every 3rd day. Frequent feeding, however, results in significant variation in growth factor levels due to FGF2 instability, which limits effective maintenance due to spontaneous differentiation. We report that stabilization of FGF2 levels using controlled release PLGA microspheres improves expression of stem cell markers, increases stem cell numbers and decreases spontaneous differentiation. The controlled release FGF2 additive reduces the frequency of media changes needed to maintain stem cell cultures, so that human embryonic stem cells and induced pluripotent stem cells can be maintained successfully with biweekly feedings.

Development and characterization of cell culture systems from Puntius (Tor) chelynoides (McClelland).

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Goswami, M; Sharma, B S; Tripathi, A K; Yadav, Kamalendra; Bahuguna, S N; Nagpure, N S; Lakra, W S; Jena, J K

2012-05-25

Puntius (Tor) chelynoides, commonly known as dark mahseer, is a commercially important coldwater fish species which inhabits fast-flowing hill-streams of India and Nepal. Cell culture systems were developed from eye, fin, heart and swim bladder tissues of P. chelynoides using explant method. The cell culture system developed from eye has been maintained towards a continuous cell line designated as PCE. The cells were grown in 25cm(2) tissue culture flasks with Leibovitz' L-15 media supplemented with 20 % fetal bovine serum (FBS) at 24°C. The PCE cell line consists of predominantly fibroblast-like cells and showed high plating efficiency. The monolayer formed from the fin and heart explants were comprised of epithelial as well as fibroblast-like cells, a prominent and rhythmic heartbeat was also observed in heart explants. Monolayer formed from swim bladder explants showed the morphology of fibroblast-like cells. All the cells from different tissues are able to grow at an optimum temperature of 24°C and growth rate increased as the FBS concentration increased. The PCE cell line was characterized using amplification of mitochondrial cytochrome oxidase subunit I (COI) & 16S rRNA genes which confirmed that the cell line originated from P. chelynoides. Cytogenetic analysis of PCE cell line and cells from fin revealed a diploid count of 100 chromosomes. Upon transfection with pEGFP-C1 plasmid, bright fluorescent signals were observed, suggesting that this cell line can be used for transgenic and genetic manipulation studies. Further, genotoxicity assessment of PCE cells illustrated the utility of this cell line as an in vitro model for aquatic toxicological studies. The PCE cell line was successfully cryopreserved and revived at different passage levels. The cell line and culture systems are being maintained to develop continuous cell lines for further studies. Copyright © 2012 Elsevier B.V. All rights reserved.

Unbiased analysis of TCRα/β chains at the single-cell level in human CD8+ T-cell subsets.

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Sun, Xiaoming; Saito, Masumichi; Sato, Yoshinori; Chikata, Takayuki; Naruto, Takuya; Ozawa, Tatsuhiko; Kobayashi, Eiji; Kishi, Hiroyuki; Muraguchi, Atsushi; Takiguchi, Masafumi

2012-01-01

T-cell receptor (TCR) α/β chains are expressed on the surface of CD8(+) T-cells and have been implicated in antigen recognition, activation, and proliferation. However, the methods for characterization of human TCRα/β chains have not been well established largely because of the complexity of their structures owing to the extensive genetic rearrangements that they undergo. Here we report the development of an integrated 5'-RACE and multiplex PCR method to amplify the full-length transcripts of TCRα/β at the single-cell level in human CD8(+) subsets, including naive, central memory, early effector memory, late effector memory, and effector phenotypic cells. Using this method, with an approximately 47% and 62% of PCR success rate for TCRα and for TCRβ chains, respectively, we were able to analyze more than 1,000 reads of transcripts of each TCR chain. Our comprehensive analysis revealed the following: (1) chimeric rearrangements of TCRδ-α, (2) control of TCRα/β transcription with multiple transcriptional initiation sites, (3) altered utilization of TCRα/β chains in CD8(+) subsets, and (4) strong association between the clonal size of TCRα/β chains and the effector phenotype of CD8(+) T-cells. Based on these findings, we conclude that our method is a useful tool to identify the dynamics of the TCRα/β repertoire, and provides new insights into the study of human TCRα/β chains.

Tubular solid oxide fuel cell development program

Energy Technology Data Exchange (ETDEWEB)

Ray, E.R.; Cracraft, C.

1995-12-31

This paper presents an overview of the Westinghouse Solid Oxide Fuel Cell (SOFC) development activities and current program status. The Westinghouse goal is to develop a cost effective cell that can operate for 50,000 to 100,000 hours. Progress toward this goal will be discussed and test results presented for multiple single cell tests which have now successfully exceeded 56,000 hours of continuous power operation at temperature. Results of development efforts to reduce cost and increase power output of tubular SOFCs are described.

Vitamin E levels in buccal cells of arsenicosis patients following vitamin E supplementation

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Mir Misbahuddin

2013-08-01

Full Text Available To understand the role of vitamin E in the treatment of arsenical melanosis and keratosis, the buccal cells of 19 patients, 14 arsenic exposed controls and 13 healthy volunteers were collected for the estimation of vitamin E both before and after supplementation with vitamin E (200 IU, caplet daily orally for 20 weeks. The vitamin E levels in the buccal cells of patients were significantly low in comparison to healthy volunteers (healthy vs patients: 17.2 ± 4.4 vs 12.3 ± 6.1 mg/mg of protein; p=0.012. These low level of vitamin E in patients returned toward normal levels following supplementation with vitamin E for 20 weeks (p=0.044. The vitamin E levels in serum of patients were also low (healthy vs patients: 18.9 ± 4.4 vs 10.2 ± 2.6 mg/mL; p=0.000. Supplementation with vitamin E overcomed the low levels of vitamin E in serum. The cholesterol levels in buccal cells and serum of patients were significantly low in comparison to healthy volunteers (buccal cells of healthy vs patients: 24.5 ± 14.1 vs 10.3 ± 9.8 mg/mg of protein; p=0.005; serum of healthy vs patients: 153.5 ± 22.8 vs 125.3 ± 37.0 mg/dL; p=0.012. After supplementation of vitamin E, there was no significant change in cholesterol levels in both buccal cells and serum of patients.

Transgenic cells with increased plastoquinone levels and methods of use

Energy Technology Data Exchange (ETDEWEB)

Sayre, Richard T.; Subramanian, Sowmya; Cahoon, Edgar

2016-12-27

Disclosed herein are transgenic cells expressing a heterologous nucleic acid encoding a prephenate dehydrogenase (PDH) protein, a heterologous nucleic acid encoding a homogentisate solanesyl transferase (HST) protein, a heterologous nucleic acid encoding a deoxyxylulose phosphate synthase (DXS) protein, or a combination of two or more thereof. In particular examples, the disclosed transgenic cells have increased plastoquinone levels. Also disclosed are methods of increasing cell growth rates or production of biomass by cultivating transgenic cells expressing a heterologous nucleic acid encoding a PDH protein, a heterologous nucleic acid encoding an HST protein, a heterologous nucleic acid encoding a DXS protein, or a combination of two or more thereof under conditions sufficient to produce cell growth or biomass.

Accurate control of oxygen level in cells during culture on silicone rubber membranes with application to stem cell differentiation.

Science.gov (United States)

Powers, Daryl E; Millman, Jeffrey R; Bonner-Weir, Susan; Rappel, Michael J; Colton, Clark K

2010-01-01

Oxygen level in mammalian cell culture is often controlled by placing culture vessels in humidified incubators with a defined gas phase partial pressure of oxygen (pO(2gas)). Because the cells are consuming oxygen supplied by diffusion, a difference between pO(2gas) and that experienced by the cells (pO(2cell)) arises, which is maximal when cells are cultured in vessels with little or no oxygen permeability. Here, we demonstrate theoretically that highly oxygen-permeable silicone rubber membranes can be used to control pO(2cell) during culture of cells in monolayers and aggregates much more accurately and can achieve more rapid transient response following a disturbance than on polystyrene and fluorinated ethylene-propylene copolymer membranes. Cell attachment on silicone rubber was achieved by physical adsorption of fibronectin or Matrigel. We use these membranes for the differentiation of mouse embryonic stem cells to cardiomyocytes and compare the results with culture on polystyrene or on silicone rubber on top of polystyrene. The fraction of cells that are cardiomyocyte-like increases with decreasing pO(2) only when using oxygen-permeable silicone membrane-based dishs, which contract on silicone rubber but not polystyrene. The high permeability of silicone rubber results in pO(2cell) being equal to pO(2gas) at the tissue-membrane interface. This, together with geometric information from histological sections, facilitates development of a model from which the pO(2) distribution within the resulting aggregates is computed. Silicone rubber membranes have significant advantages over polystyrene in controlling pO(2cell), and these results suggest they are a valuable tool for investigating pO(2) effects in many applications, such as stem cell differentiation. Copyright 2009 American Institute of Chemical Engineers

Regulation of hTERT by BCR-ABL at multiple levels in K562 cells

International Nuclear Information System (INIS)

Chai, Juin Hsien; Zhang, Yong; Tan, Wei Han; Chng, Wee Joo; Li, Baojie; Wang, Xueying

2011-01-01

The cytogenetic characteristic of Chronic Myeloid Leukemia (CML) is the formation of the Philadelphia chromosome gene product, BCR-ABL. Given that BCR-ABL is the specific target of Gleevec in CML treatment, we investigated the regulation of the catalytic component of telomerase, hTERT, by BCR-ABL at multiple levels in K562 cells. Molecular techniques such as over expression, knockdown, real-time PCR, immunoprecipitation, western blotting, reporter assay, confocal microscopy, telomerase assays and microarray were used to suggest that hTERT expression and activity is modulated by BCR-ABL at multiple levels. Our results suggest that BCR-ABL plays an important role in regulating hTERT in K562 (BCR-ABL positive human leukemia) cells. When Gleevec inhibited the tyrosine kinase activity of BCR-ABL, phosphorylation of hTERT was downregulated, therefore suggesting a positive correlation between BCR-ABL and hTERT. Gleevec treatment inhibited hTERT at mRNA level and significantly reduced telomerase activity (TA) in K562 cells, but not in HL60 or Jurkat cells (BCR-ABL negative cells). We also demonstrated that the transcription factor STAT5a plays a critical role in hTERT gene regulation in K562 cells. Knockdown of STAT5a, but not STAT5b, resulted in a marked downregulation of hTERT mRNA level, TA and hTERT protein level in K562 cells. Furthermore, translocation of hTERT from nucleoli to nucleoplasm was observed in K562 cells induced by Gleevec. Our data reveal that BCR-ABL can regulate TA at multiple levels, including transcription, post-translational level, and proper localization. Thus, suppression of cell growth and induction of apoptosis by Gleevec treatment may be partially due to TA inhibition. Additionally, we have identified STAT5a as critical mediator of the hTERT gene expression in BCR-ABL positive CML cells, suggesting that targeting STAT5a may be a promising therapeutic strategy for BCR-ABL positive CML patients

High-Level Development of Multiserver Online Games

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Frank Glinka

2008-01-01

Full Text Available Multiplayer online games with support for high user numbers must provide mechanisms to support an increasing amount of players by using additional resources. This paper provides a comprehensive analysis of the practically proven multiserver distribution mechanisms, zoning, instancing, and replication, and the tasks for the game developer implied by them. We propose a novel, high-level development approach which integrates the three distribution mechanisms seamlessly in today's online games. As a possible base for this high-level approach, we describe the real-time framework (RTF middleware system which liberates the developer from low-level tasks and allows him to stay at high level of design abstraction. We explain how RTF supports the implementation of single-server online games and how RTF allows to incorporate the three multiserver distribution mechanisms during the development process. Finally, we describe briefly how RTF provides manageability and maintenance functionality for online games in a grid context with dynamic resource allocation scenarios.

Reconstruction of incomplete cell paths through a 3D-2D level set segmentation

Science.gov (United States)

Hariri, Maia; Wan, Justin W. L.

2012-02-01

Segmentation of fluorescent cell images has been a popular technique for tracking live cells. One challenge of segmenting cells from fluorescence microscopy is that cells in fluorescent images frequently disappear. When the images are stacked together to form a 3D image volume, the disappearance of the cells leads to broken cell paths. In this paper, we present a segmentation method that can reconstruct incomplete cell paths. The key idea of this model is to perform 2D segmentation in a 3D framework. The 2D segmentation captures the cells that appear in the image slices while the 3D segmentation connects the broken cell paths. The formulation is similar to the Chan-Vese level set segmentation which detects edges by comparing the intensity value at each voxel with the mean intensity values inside and outside of the level set surface. Our model, however, performs the comparison on each 2D slice with the means calculated by the 2D projected contour. The resulting effect is to segment the cells on each image slice. Unlike segmentation on each image frame individually, these 2D contours together form the 3D level set function. By enforcing minimum mean curvature on the level set surface, our segmentation model is able to extend the cell contours right before (and after) the cell disappears (and reappears) into the gaps, eventually connecting the broken paths. We will present segmentation results of C2C12 cells in fluorescent images to illustrate the effectiveness of our model qualitatively and quantitatively by different numerical examples.

Emergence of cytotoxic resistance in cancer cell populations: Single-cell mechanisms and population-level consequences

International Nuclear Information System (INIS)

Lorenzi, Tommaso; Chisholm, Rebecca H.; Lorz, Alexander; Neves de Almeida, Luís; Clairambault, Jean; Larsen, Annette K.; Escargueil, Alexandre

2016-01-01

We formulate an individual-based model and a population model of phenotypic evolution, under cytotoxic drugs, in a cancer cell population structured by the expression levels of survival-potential and proliferation-potential. We apply these models to a recently studied experimental system. Our results suggest that mechanisms based on fundamental laws of biology can reversibly push an actively-proliferating, and drug-sensitive, cell population to transition into a weakly-proliferative and drug-tolerant state, which will eventually facilitate the emergence of more potent, proliferating and drug-tolerant cells.

Emergence of cytotoxic resistance in cancer cell populations: Single-cell mechanisms and population-level consequences

Energy Technology Data Exchange (ETDEWEB)

Lorenzi, Tommaso [Centre de Mathématiques et de Leurs Applications, ENS Cachan, CNRS, Cachan 94230 Cedex, France & INRIA-Paris-Rocquencourt, MAMBA Team, Domaine de Voluceau, BP105, 78153 Le Chesnay Cedex (France); Chisholm, Rebecca H. [School of Biotechnology and Biomolecular Sciences, University of New South Wales, Sydney NSW 2052 (Australia); Lorz, Alexander; Neves de Almeida, Luís; Clairambault, Jean [Sorbonne Universités, UPMC Univ Paris 06, UMR 7598, Laboratoire Jacques-Louis Lions, F-75005, Paris (France); CNRS, UMR 7598, Laboratoire Jacques-Louis Lions, F-75005, Paris (France); INRIA-Paris-Rocquencourt, MAMBA Team, Domaine de Voluceau, BP105, 78153 Le Chesnay Cedex (France); Larsen, Annette K.; Escargueil, Alexandre [Sorbonne Universités, UPMC Univ Paris 06, F-75005, Paris (France); INSERM, UMR-S 938, Laboratory of “Cancer Biology and Therapeutics”, F-75012, Paris (France)

2016-06-08

We formulate an individual-based model and a population model of phenotypic evolution, under cytotoxic drugs, in a cancer cell population structured by the expression levels of survival-potential and proliferation-potential. We apply these models to a recently studied experimental system. Our results suggest that mechanisms based on fundamental laws of biology can reversibly push an actively-proliferating, and drug-sensitive, cell population to transition into a weakly-proliferative and drug-tolerant state, which will eventually facilitate the emergence of more potent, proliferating and drug-tolerant cells.

A mouse model for monitoring islet cell genesis and developing therapies for diabetes

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Yoshinori Shimajiri

2011-03-01

Transient expression of the transcription factor neurogenin-3 marks progenitor cells in the pancreas as they differentiate into islet cells. We developed a transgenic mouse line in which the surrogate markers secreted alkaline phosphatase (SeAP and enhanced green florescent protein (EGFP can be used to monitor neurogenin-3 expression, and thus islet cell genesis. In transgenic embryos, cells expressing EGFP lined the pancreatic ducts. SeAP was readily detectable in embryos, in the media of cultured embryonic pancreases and in the serum of adult animals. Treatment with the γ-secretase inhibitor DAPT, which blocks Notch signaling, enhanced SeAP secretion rates and increased the number of EGFP-expressing cells as assayed by fluorescence-activated cell sorting (FACS and immunohistochemistry in cultured pancreases from embryos at embryonic day 11.5, but not in pancreases harvested 1 day later. By contrast, treatment with growth differentiation factor 11 (GDF11 reduced SeAP secretion rates. In adult mice, partial pancreatectomy decreased, whereas duct ligation increased, circulating SeAP levels. This model will be useful for studying signals involved in islet cell genesis in vivo and developing therapies that induce this process.

Do rice suspension-cultured cells treated with abscisic acid mimic developing seeds?

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Matsuno, Koya; Fujimura, Tatsuhito

2015-08-01

Starch synthesis is activated in the endosperm during seed development and also in rice suspension cells cultured with abscisic acid. In the anticipation that the mechanisms of starch synthesis are similar between the endosperm and the suspension cells cultured with abscisic acid, expression of genes involved in starch synthesis was evaluated in the suspension cells after abscisic acid treatment. However, it was found that the regulatory mechanism of starch synthesis in the suspension cells cultured with abscisic acid was different from that in developing seeds. Expression analyses of genes involved in oil bodies, which accumulate in the embryo and aleurone layer, and seed storage proteins, which accumulate mainly in the endosperm, showed that the former were activated in the suspension cells cultured with abscisic acid, but the latter were not. Master regulators for embryogenesis, OsVP1 (homologue of AtABI3) and OsLFL1 (homologue of AtFUS3 or AtLFL2), were expressed in the suspension cells at levels comparable to those in the embryo. From these results, it is suggested that interactions between regulators and abscisic acid control the synthesis of phytic acid and oil bodies in the cultured cells and embryo. We suggest that the system of suspension cells cultured with abscisic acid helps to reveal the mechanisms of phytic acid and oil body synthesis in embryo.

Hydrogen-bromine fuel cell advance component development

Science.gov (United States)

Charleston, Joann; Reed, James

1988-01-01

Advanced cell component development is performed by NASA Lewis to achieve improved performance and longer life for the hydrogen-bromine fuel cells system. The state-of-the-art hydrogen-bromine system utilizes the solid polymer electrolyte (SPE) technology, similar to the SPE technology developed for the hydrogen-oxygen fuel cell system. These studies are directed at exploring the potential for this system by assessing and evaluating various types of materials for cell parts and electrode materials for Bromine-hydrogen bromine environment and fabricating experimental membrane/electrode-catalysts by chemical deposition.

Development of a qPCR method to rapidly assess the function of NKT cells.

Science.gov (United States)

Sohn, Silke; Tiper, Irina; Japp, Emily; Sun, Wenji; Tkaczuk, Katherine; Webb, Tonya J

2014-05-01

NKT cells comprise a rare, but important subset of T cells which account for ~0.2% of the total circulating T cell population. NKT cells are known to have anti-tumor functions and rapidly produce high levels of cytokines following activation. Several clinical trials have sought to exploit the effector functions of NKT cells. While some studies have shown promise, NKT cells are approximately 50% lower in cancer patients compared to healthy donors of the same age and gender, thus limiting their therapeutic efficacy. These studies indicate that baseline levels of activation should be assessed before initiating an NKT cell based immunotherapeutic strategy. The goal of this study was to develop a sensitive method to rapidly assess NKT cell function. We utilized artificial antigen presenting cells in combination with qPCR in order to determine NKT cell function in peripheral blood mononuclear cells from healthy donors and breast cancer patients. We found that NKT cell activation can be detected by qPCR, but not by ELISA, in healthy donors as well as in breast cancer patients following four hour stimulation. This method utilizing CD1d-expressing aAPCs will enhance our knowledge of NKT cell biology and could potentially be used as a novel tool in adoptive immunotherapeutic strategies. Copyright © 2014 Elsevier B.V. All rights reserved.

Exposure to ultrafine particles, intracellular production of reactive oxygen species in leukocytes and altered levels of endothelial progenitor cells

International Nuclear Information System (INIS)

Jantzen, Kim; Møller, Peter; Karottki, Dorina Gabriela; Olsen, Yulia; Bekö, Gabriel; Clausen, Geo; Hersoug, Lars-Georg; Loft, Steffen

2016-01-01

Exposure to particles in the fine and ultrafine size range has been linked to induction of low-grade systemic inflammation, oxidative stress and development of cardiovascular diseases. Declining levels of endothelial progenitor cells within systemic circulation have likewise been linked to progression of cardiovascular diseases. The objective was to determine if exposure to fine and ultrafine particles from indoor and outdoor sources, assessed by personal and residential indoor monitoring, is associated with altered levels of endothelial progenitor cells, and whether such effects are related to leukocyte-mediated oxidative stress. The study utilized a cross sectional design performed in 58 study participants from a larger cohort. Levels of circulating endothelial progenitor cells, defined as either late (CD34 + KDR + cells) or early (CD34 + CD133 + KDR + cells) subsets were measured using polychromatic flow cytometry. We additionally measured production of reactive oxygen species in leukocyte subsets (lymphocytes, monocytes and granulocytes) by flow cytometry using intracellular 2′,7′-dichlorofluoroscein. The measurements encompassed both basal levels of reactive oxygen species production and capacity for reactive oxygen species production for each leukocyte subset. We found that the late endothelial progenitor subset was negatively associated with levels of ultrafine particles measured within the participant residences and with reactive oxygen species production capacity in lymphocytes. Additionally, the early endothelial progenitor cell levels were positively associated with a personalised measure of ultrafine particle exposure and negatively associated with both basal and capacity for reactive oxygen species production in lymphocytes and granulocytes, respectively. Our results indicate that exposure to fine and ultrafine particles derived from indoor sources may have adverse effects on human vascular health.

Identifying long-term memory B-cells in vaccinated children despite waning antibody levels specific for Bordetella pertussis proteins

NARCIS (Netherlands)

Hendrikx, Lotte H.; Ozturk, Kemal; de Rond, Lia G. H.; Veenhoven, Reinier H.; Sanders, Elisabeth A. M.; Berbers, Guy A. M.; Buisman, Anne-Marie

2011-01-01

Whooping cough is a respiratory disease caused by Bordetella pertussis. Since the 1950s in developed countries pertussis vaccinations are included in the national immunization program. However, antibody levels rapidly wane after both whole cell and acellular pertussis vaccination. Therefore

Serum Copper Level Significantly Influences Platelet Count, Lymphocyte Count and Mean Cell Hemoglobin in Sickle Cell Anemia

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Okocha Chide

2015-12-01

Full Text Available Background Changes in serum micro nutrients levels affect a number of critically important metabolic processes; these could potentially influence blood counts and ultimately disease presentation in patients with sickle cell anemia (SCA. Objectives To evaluate the influence of serum micro-nutrients levels; zinc, copper, selenium and magnesium on blood counts in steady state SCA patients. Methods A cross sectional study that involved 28 steady state adult SCA subjects. Seven milliliters (mls of blood was collected; 3 mls was for hemoglobin electrophoresis and full blood count determination while 4 mls was for measurement of serum micro nutrients levels, by the atomic absorption spectrophotometry. Correlation between serum micro-nutrient levels and blood counts was done by the Pearson’s linear regression. Ethical approval was obtained from the institutional review board and each participant gave informed consent. All data was analyzed by SPSS software version 20. Results There was a significant correlation between serum copper levels and mean cell hemoglobin (MCH, platelet and lymphocyte counts (r = 0.418; P = 0.02, r = -0.376; P = 0.04 and r = -0.383; P = 0.04, respectively. There were no significant correlations between serum levels of other micro nutrients (selenium, zinc and magnesium and blood counts. Conclusions Copper influences blood count in SCA patients probably by inducing red cell haemolysis, oxidant tissue damage and stimulating the immune system.

Multi-level Strategy for Identifying Proteasome-Catalyzed Spliced Epitopes Targeted by CD8+ T Cells during Bacterial Infection

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Anouk C.M. Platteel

2017-08-01

Full Text Available Proteasome-catalyzed peptide splicing (PCPS generates peptides that are presented by MHC class I molecules, but because their identification is challenging, the immunological relevance of spliced peptides remains unclear. Here, we developed a reverse immunology-based multi-level approach to identify proteasome-generated spliced epitopes. Applying this strategy to a murine Listeria monocytogenes infection model, we identified two spliced epitopes within the secreted bacterial phospholipase PlcB that primed antigen-specific CD8+ T cells in L. monocytogenes-infected mice. While reacting to the spliced epitopes, these CD8+ T cells failed to recognize the non-spliced peptide parts in the context of their natural flanking sequences. Thus, we here show that PCPS expands the CD8+ T cell response against L. monocytogenes by exposing spliced epitopes on the cell surface. Moreover, our multi-level strategy opens up opportunities to systematically investigate proteins for spliced epitope candidates and thus strategies for immunotherapies or vaccine design.

Signaling hierarchy regulating human endothelial cell development

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Our present knowledge of the regulation of mammalian endothelial cell differentiation has been largely derived from studies of mouse embryonic development. However, unique mechanisms and hierarchy of signals that govern human endothelial cell development are unknown and, thus, explored in these stud...

Puerarin Facilitates T-Tubule Development of Murine Embryonic Stem Cell-Derived Cardiomyocytes

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Lu Wang

2014-07-01

Full Text Available Aims: The embryonic stem cell-derived cardiomyocytes (ES-CM is one of the promising cell sources for repopulation of damaged myocardium. However, ES-CMs present immature structure, which impairs their integration with host tissue and functional regeneration. This study used murine ES-CMs as an in vitro model of cardiomyogenesis to elucidate the effect of puerarin, the main compound found in the traditional Chinese medicine the herb Radix puerariae, on t-tubule development of murine ES-CMs. Methods: Electron microscope was employed to examine the ultrastructure. The investigation of transverse-tubules (t-tubules was performed by Di-8-ANEPPS staining. Quantitative real-time PCR was utilized to study the transcript level of genes related to t-tubule development. Results: We found that long-term application of puerarin throughout cardiac differentiation improved myofibril array and sarcomeres formation, and significantly facilitated t-tubules development of ES-CMs. The transcript levels of caveolin-3, amphiphysin-2 and junctophinlin-2, which are crucial for the formation and development of t-tubules, were significantly upregulated by puerarin treatment. Furthermore, puerarin repressed the expression of miR-22, which targets to caveolin-3. Conclusion: Our data showed that puerarin facilitates t-tubule development of murine ES-CMs. This might be related to the repression of miR-22 by puerarin and upregulation of Cav3, Bin1 and JP2 transcripts.

Lithium-Ion Cell Charge-Control Unit Developed

Science.gov (United States)

Reid, Concha M.; Manzo, Michelle A.; Buton, Robert M.; Gemeiner, Russel

2005-01-01

A lithium-ion (Li-ion) cell charge-control unit was developed as part of a Li-ion cell verification program. This unit manages the complex charging scheme that is required when Li-ion cells are charged in series. It enables researchers to test cells together as a pack, while allowing each cell to charge individually. This allows the inherent cell-to-cell variations to be addressed on a series string of cells and reduces test costs substantially in comparison to individual cell testing.

Cell module and fuel conditioner development

Science.gov (United States)

Hoover, D. Q., Jr.

1980-01-01

Components for the first 5 cell stack (no cooling plates) of the MK-2 design were fabricated. Preliminary specfications and designs for the components of a 23 cell MK-1 stack with four DIGAS cooling plates were developed. The MK-2 was selected as a bench mark design and a preliminary design of the facilities required for high rate manufacture of fuel cell modules was developed. Two stands for testing 5 cell stacks were built and design work for modifying existing stands and building new stands for 23 and 80 cell stacks was initiated. Design and procurement of components and materials for the catalyst test stand were completed and construction initiated. Work on the specifications of pipeline gas, tap water and recovered water and definition of equipment required for treatment was initiated. An innovative geometry for the reformer was conceived and modifications of the computer program to be used in its design were stated.

Analysis of gene expression levels in individual bacterial cells without image segmentation.

Science.gov (United States)

Kwak, In Hae; Son, Minjun; Hagen, Stephen J

2012-05-11

Studies of stochasticity in gene expression typically make use of fluorescent protein reporters, which permit the measurement of expression levels within individual cells by fluorescence microscopy. Analysis of such microscopy images is almost invariably based on a segmentation algorithm, where the image of a cell or cluster is analyzed mathematically to delineate individual cell boundaries. However segmentation can be ineffective for studying bacterial cells or clusters, especially at lower magnification, where outlines of individual cells are poorly resolved. Here we demonstrate an alternative method for analyzing such images without segmentation. The method employs a comparison between the pixel brightness in phase contrast vs fluorescence microscopy images. By fitting the correlation between phase contrast and fluorescence intensity to a physical model, we obtain well-defined estimates for the different levels of gene expression that are present in the cell or cluster. The method reveals the boundaries of the individual cells, even if the source images lack the resolution to show these boundaries clearly. Copyright © 2012 Elsevier Inc. All rights reserved.

Tumor-promoting phorbol ester transiently down-modulates the p53 level and blocks the cell cycle

DEFF Research Database (Denmark)

Skouv, J.; Jensen, P O; Forchhammer, J

1994-01-01

Activation of the protein kinase C signaling pathway by tumor-promoting phorbol esters, such as 4 beta-phorbol 12-myristate 13-acetate (PMA), induced a decrease in the level of p53 mRNA in several serum-starved human cell lines. Also, the tumor-promoting phosphatase inhibitor okadaic acid induced...... a decrease in the p53 mRNA level in the cell lines. Normal diploid as well as various tumor cell lines were tested. Two tumor cell lines, HeLa and A549, both containing the wild-type p53 gene, but very different levels of p53 protein, were studied in detail. In both cell lines, the level of p53 m......RNA was minimal after 9 h of exposure to PMA. After approximately 120 h, the p53 mRNA level was similar to the pretreatment level. PMA induced a similar transient decrease in the level of p53 protein in the A549 cell line. The decrease in the p53 mRNA level could not be explained by changes in the transcriptional...

CD44-positive cells are candidates for astrocyte precursor cells in developing mouse cerebellum.

Science.gov (United States)

Cai, Na; Kurachi, Masashi; Shibasaki, Koji; Okano-Uchida, Takayuki; Ishizaki, Yasuki

2012-03-01

Neural stem cells are generally considered to be committed to becoming precursor cells before terminally differentiating into either neurons or glial cells during neural development. Neuronal and oligodendrocyte precursor cells have been identified in several areas in the murine central nervous system. The presence of astrocyte precursor cells (APCs) is not so well understood. The present study provides several lines of evidence that CD44-positive cells are APCs in the early postnatal mouse cerebellum. In developing mouse cerebellum, CD44-positive cells, mostly located in the white matter, were positive for the markers of the astrocyte lineage, but negative for the markers of mature astrocytes. CD44-positive cells were purified from postnatal cerebellum by fluorescence-activated cell sorting and characterized in vitro. In the absence of any signaling molecule, many cells died by apoptosis. The surviving cells gradually expressed glial fibrillary acidic protein, a marker for mature astrocytes, indicating that differentiation into mature astrocytes is the default program for these cells. The cells produced no neurospheres nor neurons nor oligodendrocytes under any condition examined, indicating these cells are not neural stem cells. Leukemia inhibitory factor greatly promoted astrocytic differentiation of CD44-positive cells, whereas bone morphogenetic protein 4 (BMP4) did not. Fibroblast growth factor-2 was a potent mitogen for these cells, but was insufficient for survival. BMP4 inhibited activation of caspase-3 and greatly promoted survival, suggesting a novel role for BMP4 in the control of development of astrocytes in cerebellum. We isolated and characterized only CD44 strongly positive large cells and discarded small and/or CD44 weakly positive cells in this study. Further studies are necessary to characterize these cells to help determine whether CD44 is a selective and specific marker for APCs in the developing mouse cerebellum. In conclusion, we succeeded in

Development and Prospect of Nanoarchitectured Solar Cells

OpenAIRE

Zhang, Bo; Xie, Wenxu; Xiang, Yong

2015-01-01

This paper gives an overview of the development and prospect of nanotechnologies utilized in the solar cell applications. Even though it is not clearly pointed out, nanostructures indeed have been used in the fabrication of conventional solar cells for a long time. However, in those circumstances, only very limited benefits of nanostructures have been used to improve cell performance. During the last decade, the development of the photovoltaic device theory and nanofabrication technology enab...

Pituitary gland development and disease: from stem cell to hormone production.

Science.gov (United States)

Davis, Shannon W; Ellsworth, Buffy S; Peréz Millan, María Inés; Gergics, Peter; Schade, Vanessa; Foyouzi, Nastaran; Brinkmeier, Michelle L; Mortensen, Amanda H; Camper, Sally A

2013-01-01

Many aspects of pituitary development have become better understood in the past two decades. The signaling pathways regulating pituitary growth and shape have emerged, and the balancing interactions between the pathways are now appreciated. Markers for multipotent progenitor cells are being identified, and signature transcription factors have been discovered for most hormone-producing cell types. We now realize that pulsatile hormone secretion involves a 3D integration of cellular networks. About a dozen genes are known to cause pituitary hypoplasia when mutated due to their essential roles in pituitary development. Similarly, a few genes are known that predispose to familial endocrine neoplasia, and several genes mutated in sporadic pituitary adenomas are documented. In the next decade, we anticipate gleaning a deeper appreciation of these processes at the molecular level, insight into the development of the hypophyseal portal blood system, and evolution of better therapeutics for congenital and acquired hormone deficiencies and for common craniopharyngiomas and pituitary adenomas. © 2013 Elsevier Inc. All rights reserved.

Differential cellular responses by oncogenic levels of c-Myc expression in long-term confluent retinal pigment epithelial cells.

Science.gov (United States)

Wang, Yiping; Cheng, Xiangdong; Samma, Muhammad Kaleem; Kung, Sam K P; Lee, Clement M; Chiu, Sung Kay

2018-06-01

c-Myc is a highly pleiotropic transcription factor known to control cell cycle progression, apoptosis, and cellular transformation. Normally, ectopic expression of c-Myc is associated with promoting cell proliferation or triggering cell death via activating p53. However, it is not clear how the levels of c-Myc lead to different cellular responses. Here, we generated a series of stable RPE cell clones expressing c-Myc at different levels, and found that consistent low level of c-Myc induced cellular senescence by activating AP4 in post-confluent RPE cells, while the cells underwent cell death at high level of c-Myc. In addition, high level of c-Myc could override the effect of AP4 on cellular senescence. Further knockdown of AP4 abrogated senescence-like phenotype in cells expressing low level of c-Myc, and accelerated cell death in cells with medium level of c-Myc, indicating that AP4 was required for cellular senescence induced by low level of c-Myc.

Cytochrome P450 levels are altered in patients with esophageal squamous-cell carcinoma

DEFF Research Database (Denmark)

Bergheim, I.; Wolfgarten, E.; Bollschweiler, E.

2007-01-01

AIM: To investigate the role of cytochrome P450 (CYP) in the carcinogenesis of squamous-cell carcinoma (SCC) in human esophagus by determining expression patterns and protein levels of representative CYPs in esophageal tissue of patients with SCC and controls. METHODS: mRNA expression of CYP2E1...... tissue (e.g. CYP2C8, CYP3A4, CYP3A5, and CYP2E1) between SCC patients and healthy subjects and may contribute to the development of SCC in the esophagus....

Human induced pluripotent cells resemble embryonic stem cells demonstrating enhanced levels of DNA repair and efficacy of nonhomologous end-joining

Energy Technology Data Exchange (ETDEWEB)

Fan Jinshui; Robert, Carine [Department of Radiation Oncology, University of Maryland School of Medicine, 655 West Baltimore Street, BRB 7-023A, Baltimore, MD 21201 (United States); Jang, Yoon-Young; Liu Hua; Sharkis, Saul; Baylin, Stephen Bruce [Johns Hopkins University School of Medicine, Department of Oncology, Baltimore, MD 21231-1000 (United States); Rassool, Feyruz Virgilia, E-mail: frassool@som.umaryland.edu [Department of Radiation Oncology, University of Maryland School of Medicine, 655 West Baltimore Street, BRB 7-023A, Baltimore, MD 21201 (United States)

2011-08-01

Highlights: {yields} iPSC and hESC demonstrate a similar cell cycle profile, with increased S phase cells and decreased G0/G1. {yields} iPSC and hESC increased ROS and decreased DSBs, compared with differentiated parental cells. {yields} iPSC and hESC demonstrate elevated DSB repair activity, including nonhomologous end-joining, compared with differentiated parental cells. {yields} iPSC however show a partial apoptotic response to DNA damage, compared to hESC. {yields} DNA damage responses may constitute important markers for the efficacy of iPSC reprogramming. - Abstract: To maintain the integrity of the organism, embryonic stem cells (ESC) need to maintain their genomic integrity in response to DNA damage. DNA double strand breaks (DSBs) are one of the most lethal forms of DNA damage and can have disastrous consequences if not repaired correctly, leading to cell death, genomic instability and cancer. How human ESC (hESC) maintain genomic integrity in response to agents that cause DSBs is relatively unclear. Adult somatic cells can be induced to 'dedifferentiate' into induced pluripotent stem cells (iPSC) and reprogram into cells of all three germ layers. Whether iPSC have reprogrammed the DNA damage response is a critical question in regenerative medicine. Here, we show that hESC demonstrate high levels of endogenous reactive oxygen species (ROS) which can contribute to DNA damage and may arise from high levels of metabolic activity. To potentially counter genomic instability caused by DNA damage, we find that hESC employ two strategies: First, these cells have enhanced levels of DNA repair proteins, including those involved in repair of DSBs, and they demonstrate elevated nonhomologous end-joining (NHEJ) activity and repair efficacy, one of the main pathways for repairing DSBs. Second, they are hypersensitive to DNA damaging agents, as evidenced by a high level of apoptosis upon irradiation. Importantly, iPSC, unlike the parent cells they are derived

Human induced pluripotent cells resemble embryonic stem cells demonstrating enhanced levels of DNA repair and efficacy of nonhomologous end-joining

International Nuclear Information System (INIS)

Fan Jinshui; Robert, Carine; Jang, Yoon-Young; Liu Hua; Sharkis, Saul; Baylin, Stephen Bruce; Rassool, Feyruz Virgilia

2011-01-01

Highlights: → iPSC and hESC demonstrate a similar cell cycle profile, with increased S phase cells and decreased G0/G1. → iPSC and hESC increased ROS and decreased DSBs, compared with differentiated parental cells. → iPSC and hESC demonstrate elevated DSB repair activity, including nonhomologous end-joining, compared with differentiated parental cells. → iPSC however show a partial apoptotic response to DNA damage, compared to hESC. → DNA damage responses may constitute important markers for the efficacy of iPSC reprogramming. - Abstract: To maintain the integrity of the organism, embryonic stem cells (ESC) need to maintain their genomic integrity in response to DNA damage. DNA double strand breaks (DSBs) are one of the most lethal forms of DNA damage and can have disastrous consequences if not repaired correctly, leading to cell death, genomic instability and cancer. How human ESC (hESC) maintain genomic integrity in response to agents that cause DSBs is relatively unclear. Adult somatic cells can be induced to 'dedifferentiate' into induced pluripotent stem cells (iPSC) and reprogram into cells of all three germ layers. Whether iPSC have reprogrammed the DNA damage response is a critical question in regenerative medicine. Here, we show that hESC demonstrate high levels of endogenous reactive oxygen species (ROS) which can contribute to DNA damage and may arise from high levels of metabolic activity. To potentially counter genomic instability caused by DNA damage, we find that hESC employ two strategies: First, these cells have enhanced levels of DNA repair proteins, including those involved in repair of DSBs, and they demonstrate elevated nonhomologous end-joining (NHEJ) activity and repair efficacy, one of the main pathways for repairing DSBs. Second, they are hypersensitive to DNA damaging agents, as evidenced by a high level of apoptosis upon irradiation. Importantly, iPSC, unlike the parent cells they are derived from, mimic hESC in their ROS levels

Keeping the Rhythm : Cardiac Pacemaker Cell Development

NARCIS (Netherlands)

Burkhard, S.B.

2017-01-01

The heart is the first organ to form and function in the developing vertebrate embryo. Its proper morphogenesis and function is crucial for survival. Here we focus on the development and characterization of a highly specialized subset of cardiac cells, the pacemaker cells. In the mammalian heart,

System-level Reliability Assessment of Power Stage in Fuel Cell Application

DEFF Research Database (Denmark)

Zhou, Dao; Wang, Huai; Blaabjerg, Frede

2016-01-01

reliability. In a case study of a 5 kW fuel cell power stage, the parameter variations of the lifetime model prove that the exponential factor of the junction temperature fluctuation is the most sensitive parameter. Besides, if a 5-out-of-6 redundancy is used, it is concluded both the B10 and the B1 system......High efficient and less pollutant fuel cell stacks are emerging and strong candidates of the power solution used for mobile base stations. In the application of the backup power, the availability and reliability hold the highest priority. This paper considers the reliability metrics from...... the component-level to the system-level for the power stage used in a fuel cell application. It starts with an estimation of the annual accumulated damage for the key power electronic components according to the real mission profile of the fuel cell system. Then, considering the parameter variations in both...

Increased level and interferon-γ production of circulating natural killer cells in patients with scrub typhus.

Science.gov (United States)

Kang, Seung-Ji; Jin, Hye-Mi; Cho, Young-Nan; Kim, Seong Eun; Kim, Uh Jin; Park, Kyung-Hwa; Jang, Hee-Chang; Jung, Sook-In; Kee, Seung-Jung; Park, Yong-Wook

2017-07-01

Natural killer (NK) cells are essential immune cells against several pathogens. Not much is known regarding the roll of NK cells in Orientia tsutsugamushi infection. Thus, this study aims to determine the level, function, and clinical relevance of NK cells in patients with scrub typhus. This study enrolled fifty-six scrub typhus patients and 56 health controls (HCs). The patients were divided into subgroups according to their disease severity. A flow cytometry measured NK cell level and function in peripheral blood. Circulating NK cell levels and CD69 expressions were significantly increased in scrub typhus patients. Increased NK cell levels reflected disease severity. In scrub typhus patients, tests showed their NK cells produced higher amounts of interferon (IFN)-γ after stimulation with interleukin (IL)-12 and IL-18 relative to those of HCs. Meanwhile, between scrub typhus patients and HCs, the cytotoxicity and degranulation of NK cells against K562 were comparable. CD69 expressions were recovered to the normal levels in the remission phase. This study shows that circulating NK cells are activated and numerically increased, and they produced more IFN-γ in scrub typhus patients.

Increased level and interferon-γ production of circulating natural killer cells in patients with scrub typhus.

Directory of Open Access Journals (Sweden)

Seung-Ji Kang

2017-07-01

Full Text Available Natural killer (NK cells are essential immune cells against several pathogens. Not much is known regarding the roll of NK cells in Orientia tsutsugamushi infection. Thus, this study aims to determine the level, function, and clinical relevance of NK cells in patients with scrub typhus.This study enrolled fifty-six scrub typhus patients and 56 health controls (HCs. The patients were divided into subgroups according to their disease severity. A flow cytometry measured NK cell level and function in peripheral blood. Circulating NK cell levels and CD69 expressions were significantly increased in scrub typhus patients. Increased NK cell levels reflected disease severity. In scrub typhus patients, tests showed their NK cells produced higher amounts of interferon (IFN-γ after stimulation with interleukin (IL-12 and IL-18 relative to those of HCs. Meanwhile, between scrub typhus patients and HCs, the cytotoxicity and degranulation of NK cells against K562 were comparable. CD69 expressions were recovered to the normal levels in the remission phase.This study shows that circulating NK cells are activated and numerically increased, and they produced more IFN-γ in scrub typhus patients.

Systems Analysis Reveals High Genetic and Antigen-Driven Predetermination of Antibody Repertoires throughout B Cell Development

Directory of Open Access Journals (Sweden)

Victor Greiff

2017-05-01

Full Text Available Antibody repertoire diversity and plasticity is crucial for broad protective immunity. Repertoires change in size and diversity across multiple B cell developmental stages and in response to antigen exposure. However, we still lack fundamental quantitative understanding of the extent to which repertoire diversity is predetermined. Therefore, we implemented a systems immunology framework for quantifying repertoire predetermination on three distinct levels: (1 B cell development (pre-B cell, naive B cell, plasma cell, (2 antigen exposure (three structurally different proteins, and (3 four antibody repertoire components (V-gene usage, clonal expansion, clonal diversity, repertoire size extracted from antibody repertoire sequencing data (400 million reads. Across all three levels, we detected a dynamic balance of high genetic (e.g., >90% for V-gene usage and clonal expansion in naive B cells and antigen-driven (e.g., 40% for clonal diversity in plasma cells predetermination and stochastic variation. Our study has implications for the prediction and manipulation of humoral immunity.

Unbiased analysis of TCRα/β chains at the single-cell level in human CD8+ T-cell subsets.

Directory of Open Access Journals (Sweden)

Xiaoming Sun

Full Text Available T-cell receptor (TCR α/β chains are expressed on the surface of CD8(+ T-cells and have been implicated in antigen recognition, activation, and proliferation. However, the methods for characterization of human TCRα/β chains have not been well established largely because of the complexity of their structures owing to the extensive genetic rearrangements that they undergo. Here we report the development of an integrated 5'-RACE and multiplex PCR method to amplify the full-length transcripts of TCRα/β at the single-cell level in human CD8(+ subsets, including naive, central memory, early effector memory, late effector memory, and effector phenotypic cells. Using this method, with an approximately 47% and 62% of PCR success rate for TCRα and for TCRβ chains, respectively, we were able to analyze more than 1,000 reads of transcripts of each TCR chain. Our comprehensive analysis revealed the following: (1 chimeric rearrangements of TCRδ-α, (2 control of TCRα/β transcription with multiple transcriptional initiation sites, (3 altered utilization of TCRα/β chains in CD8(+ subsets, and (4 strong association between the clonal size of TCRα/β chains and the effector phenotype of CD8(+ T-cells. Based on these findings, we conclude that our method is a useful tool to identify the dynamics of the TCRα/β repertoire, and provides new insights into the study of human TCRα/β chains.

Serum Endostatin Levels in Oral Squamous Cell Carcinoma

Directory of Open Access Journals (Sweden)

Maryam Mardani

2018-05-01

Full Text Available Introduction: Endostatin is a C-­terminal proteolytic fragment of collagen XVIII and, as with angiostatin and thrombospondin, is known as an anti­angiogenic agent. The aim of this study was to assess the level of serum endostatin in patients with oral squamous cell carcinoma (SCC, and its association with the clinicopathological characteristics of the tumor.   Materials and Methods:  Using an enzyme-linked immunosorbent assay (ELISA kit, we investigated the circulating levels of endostatin in the blood serum of 45 patients with oral SCC and 45 healthy controls.   Results: The mean level of serum endostatin in patients was significantly lower (68.8±85 ng/ml than in healthy controls (175.6±73 ng/ml (P

CD38 gene-modified dendritic cells inhibit murine asthma development by increasing IL-12 production and promoting Th1 cell differentiation.

Science.gov (United States)

Wang, Jiaoli; Zhu, Weiguo; Chen, Yinghu; Lin, Zhendong; Ma, Shenglin

2016-11-01

Predominant T helper (Th)2 and impaired Th1 cell polarization has a crucial role in the development of asthma. Cluster of differentiation (CD)38 is associated with the increased release of interleukin (IL)‑12 from dendritic cells (DCs) and DC‑induced Th1 cell polarization. However, whether CD38 expression affects DC function in asthma development remains unknown. In the current study, adenoviruses were constructed containing the murine CD38 gene. Overexpression of CD38 protein level in DCs induced from bone‑marrow derived DCs (BMDCs) by recombinant mouse granulocyte macrophage colony‑stimulating factor and IL‑4 was achieved through 24 h adenovirus infection. The results demonstrated that BMDCs with CD38 overexpression exhibited no phenotypic change; however, following stimulation with lipopolysaccharide (LPS), maturation and IL‑12 secretion were increased. In addition, CD38‑overexpressing BMDCs stimulated with LPS exhibited more effective Th1 cell differentiation. Mice that were administered CD38‑overexpressing BMDCs exhibited milder symptoms of asthma. Furthermore, decreased IL‑4, IL‑5 and IL‑13 levels were detected in bronchoalveolar lavage fluid (BALF), reduced immunoglobulin E levels were measured in the sera, and increased interferon‑γ was detected in BALF from the recipients of CD38‑overexpressing BMDCs. Increased phosphorylated‑p38 expression was also detected in LPS-stimulated CD38-overexpressing BMDCs, whereas pretreatment with a p38‑specific inhibitor was able to abolish the effects of LPS stimulation and CD38 overexpression on IL‑12 release and Th1 cell differentiation in BMDCs. These results suggested that CD38 may be involved in the DC function of alleviating asthma via restoration of the Th1/Th2 balance, thus providing a novel strategy for asthma therapy.

Epalrestat increases intracellular glutathione levels in Schwann cells through transcription regulation

Directory of Open Access Journals (Sweden)

Keisuke Sato

2014-01-01

Full Text Available Epalrestat (EPS, approved in Japan, is the only aldose reductase inhibitor that is currently available for the treatment of diabetic neuropathy. Here we report that EPS at near-plasma concentration increases the intracellular levels of glutathione (GSH, which is important for protection against oxidative injury, through transcription regulation. Treatment of Schwann cells with EPS caused a dramatic increase in intracellular GSH levels. EPS increased the mRNA levels of γ-glutamylcysteine synthetase (γ-GCS, the enzyme catalyzing the first and rate-limiting step in de novo GSH synthesis. Nuclear factor erythroid 2-related factor 2 (Nrf2 is a key transcription factor that plays a central role in regulating the expression of γ-GCS. ELISA revealed that EPS increased nuclear Nrf2 levels. Knockdown of Nrf2 by siRNA suppressed the EPS-induced GSH biosynthesis. Furthermore, pretreatment with EPS reduced the cytotoxicity induced by H2O2, tert-butylhydroperoxide, 2,2'-azobis (2-amidinopropane dihydrochloride, and menadione, indicating that EPS plays a role in protecting against oxidative stress. This is the first study to show that EPS induces GSH biosynthesis via the activation of Nrf2. We suggest that EPS has new beneficial properties that may prevent the development and progression of disorders caused by oxidative stress.

Manganese superoxide dismutase level in blood cells of patients with breast cancer

International Nuclear Information System (INIS)

Adzic, M.; Niciforovic, A.; Filipovic, D.; Vucic, V.; Neskovic-Konstantinovic, Z.; Radojcic, M.B. . E-mail address of corresponding author: marija@vin.bg.ac.yu; Radojcic, M.B.)

2005-01-01

The intracellular MnSOD levels were determined in peripheral blood cells obtained from two age groups of patients with breast cancer (BC): 30-45 year old patients (premenopausal, n=7, clinical stage 1 to 3) and 46-60 year old patients (peri- and postmenopausal women, n=12, clinical stage 3 or 4), at diagnosis, prior to any clinical treatment. The respective healthy women groups were used as controls. Blood cells were also irradiated in vitro with 2- and 9- Gy of gamma-ray radiation from 60 Co source. The MnSOD levels were determined by the specific immunostaining and quantified by the laser densitometry. The MANOVA analysis and Tukey post-hoc test indicated significantly higher MnSOD levels in both groups of BC patients in relation to the respective controls ( F=25.166, p<0.001). The data indicated that the increased initial MnSOD levels in peripheral blood cells may be related to the presence of BC i.e. they may reflect the system response to the presence of malignant tumour. In addition to that, in vitro radiation challenge of blood cells indicate that MnSOD overexpression may be a good indicator for selection of BC patients that would express increased resistance to oxidative stress imposed by the clinical treatment. (author)

The expression of SLAMF7 levels in malignant B cells: a novel ...

African Journals Online (AJOL)

Signalling lymphocyte activation molecule (SLAM) F7 is found on the surface of some immune cells including B-lymphocytes. Its activation leads to the proliferation or differentiation of immune cells. The objectives of the study were to measure SLAMF7 expression levels on B-CLL cells, and to upregulate the expression of ...

Reduction of nitric oxide level enhances the radiosensitivity of hypoxic non-small cell lung cancer

International Nuclear Information System (INIS)

Saleem, Wael; Suzuki, Yoshiyuki; Mobaraki, Abdulelah; Yoshida, Yukari; Noda, Shinei; Saitoh, Jun-ichi; Nakano, Takashi

2011-01-01

The epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (E-TKI) resistance has emerged as an important clinical issue. To overcome this resistance, researchers have examined different modalities, either for use as a monotherapy or in combination with E-TKI therapy. In the present study, we investigated whether a decrease in nitric oxide (NO) levels affects the radiosensitization of non-small cell lung cancer (NSCLC) cell lines. A549 and H3255 NSCLC cells were examined. They were subjected to hypoxic conditions and monotherapy, or combined therapy using radiation and N G -monomethyl- L -arginine, monoacetate (LNMMA). Reductions in nitric oxide levels enhanced the radiosensitivity of both cell lines and significantly reduced the expression of both hypoxia-inducible factor-1α (HIF-1α) and EGFR in H3255 cells compared to A549 cells. Since NO is significantly associated with cell metabolism, we measured the levels of pyruvate dehydrogenase kinase-1 (PDK-1), reactive oxygen species, and oxygen and observed that the expression of PDK-1 was significantly reduced. This reduction was seen simultaneously after the silencing of HIF-1α; however, not following LNMMA treatment. The oxygen concentration was significantly increased in the treated cells, and their viability decreased in parallel. Reactive oxygen species were decreased after LNMMA and radiation treatment. Adding EGFR-TKI to cells with reduced NO levels further suppressed cell viability when combined with radiation. This study suggests that a reduction in the NO level might substantially overcome the radioresistance of mutant NSCLC cells. (author)

Restricted cell elongation in Arabidopsis hypocotyls is associated with a reduced average pectin esterification level.

Science.gov (United States)

Derbyshire, Paul; McCann, Maureen C; Roberts, Keith

2007-06-17

Cell elongation is mainly limited by the extensibility of the cell wall. Dicotyledonous primary (growing) cell walls contain cellulose, xyloglucan, pectin and proteins, but little is known about how each polymer class contributes to the cell wall mechanical properties that control extensibility. We present evidence that the degree of pectin methyl-esterification (DE%) limits cell growth, and that a minimum level of about 60% DE is required for normal cell elongation in Arabidopsis hypocotyls. When the average DE% falls below this level, as in two gibberellic acid (GA) mutants ga1-3 and gai, and plants expressing pectin methyl-esterase (PME1) from Aspergillus aculeatus, then hypocotyl elongation is reduced. Low average levels of pectin DE% are associated with reduced cell elongation, implicating PMEs, the enzymes that regulate DE%, in the cell elongation process and in responses to GA. At high average DE% other components of the cell wall limit GA-induced growth.

Restricted cell elongation in Arabidopsis hypocotyls is associated with a reduced average pectin esterification level

Directory of Open Access Journals (Sweden)

Derbyshire Paul

2007-06-01

Full Text Available Abstract Background Cell elongation is mainly limited by the extensibility of the cell wall. Dicotyledonous primary (growing cell walls contain cellulose, xyloglucan, pectin and proteins, but little is known about how each polymer class contributes to the cell wall mechanical properties that control extensibility. Results We present evidence that the degree of pectin methyl-esterification (DE% limits cell growth, and that a minimum level of about 60% DE is required for normal cell elongation in Arabidopsis hypocotyls. When the average DE% falls below this level, as in two gibberellic acid (GA mutants ga1-3 and gai, and plants expressing pectin methyl-esterase (PME1 from Aspergillus aculeatus, then hypocotyl elongation is reduced. Conclusion Low average levels of pectin DE% are associated with reduced cell elongation, implicating PMEs, the enzymes that regulate DE%, in the cell elongation process and in responses to GA. At high average DE% other components of the cell wall limit GA-induced growth.

Development of analytical cell support for vitrification at the West Valley Demonstration Project. Topical report

Energy Technology Data Exchange (ETDEWEB)

Barber, F.H.; Borek, T.T.; Christopher, J.Z. [and others

1997-12-01

Analytical and Process Chemistry (A&PC) support is essential to the high-level waste vitrification campaign at the West Valley Demonstration Project (WVDP). A&PC characterizes the waste, providing information necessary to formulate the recipe for the target radioactive glass product. High-level waste (HLW) samples are prepared and analyzed in the analytical cells (ACs) and Sample Storage Cell (SSC) on the third floor of the main plant. The high levels of radioactivity in the samples require handling them in the shielded cells with remote manipulators. The analytical hot cells and third floor laboratories were refurbished to ensure optimal uninterrupted operation during the vitrification campaign. New and modified instrumentation, tools, sample preparation and analysis techniques, and equipment and training were required for A&PC to support vitrification. Analytical Cell Mockup Units (ACMUs) were designed to facilitate method development, scientist and technician training, and planning for analytical process flow. The ACMUs were fabricated and installed to simulate the analytical cell environment and dimensions. New techniques, equipment, and tools could be evaluated m in the ACMUs without the consequences of generating or handling radioactive waste. Tools were fabricated, handling and disposal of wastes was addressed, and spatial arrangements for equipment were refined. As a result of the work at the ACMUs the remote preparation and analysis methods and the equipment and tools were ready for installation into the ACs and SSC m in July 1995. Before use m in the hot cells, all remote methods had been validated and four to eight technicians were trained on each. Fine tuning of the procedures has been ongoing at the ACs based on input from A&PC technicians. Working at the ACs presents greater challenges than had development at the ACMUs. The ACMU work and further refinements m in the ACs have resulted m in a reduction m in analysis turnaround time (TAT).

Analysis of gene expression levels in individual bacterial cells without image segmentation

International Nuclear Information System (INIS)

Kwak, In Hae; Son, Minjun; Hagen, Stephen J.

2012-01-01

Highlights: ► We present a method for extracting gene expression data from images of bacterial cells. ► The method does not employ cell segmentation and does not require high magnification. ► Fluorescence and phase contrast images of the cells are correlated through the physics of phase contrast. ► We demonstrate the method by characterizing noisy expression of comX in Streptococcus mutans. -- Abstract: Studies of stochasticity in gene expression typically make use of fluorescent protein reporters, which permit the measurement of expression levels within individual cells by fluorescence microscopy. Analysis of such microscopy images is almost invariably based on a segmentation algorithm, where the image of a cell or cluster is analyzed mathematically to delineate individual cell boundaries. However segmentation can be ineffective for studying bacterial cells or clusters, especially at lower magnification, where outlines of individual cells are poorly resolved. Here we demonstrate an alternative method for analyzing such images without segmentation. The method employs a comparison between the pixel brightness in phase contrast vs fluorescence microscopy images. By fitting the correlation between phase contrast and fluorescence intensity to a physical model, we obtain well-defined estimates for the different levels of gene expression that are present in the cell or cluster. The method reveals the boundaries of the individual cells, even if the source images lack the resolution to show these boundaries clearly.

Assessment of flhDC mRNA levels in Serratia liquefaciens swarm cells

DEFF Research Database (Denmark)

Tolker-Nielsen, Tim; Christensen, Allan Beck; Holmstrøm, K.

2000-01-01

We reported previously that artificial overexpression of the flhDC operon in liquid-grown Serratia liquefaciens resulted in the formation of filamentous, multinucleated, and hyperflagellated cells that were indistinguishable from surface-induced swarm cells (L. Eberl, G. Christiansen, S. Molin, a......, vegetative cells. This suggests that surface-induced S. liquefaciens swarm cell differentiation, although dependent on flhDC gene expression, does not occur through elevated flhDC mRNA levels....

Study on serum TNF-α level, B-cell count and T-cell subsets distribution in peripheral blood in patients with rheumatoid arthritis

International Nuclear Information System (INIS)

Shi Buqing

2006-01-01

Objective: To study the changes of serum TNF-α levels, B-cell count and T-cell subsets distribution in peripheral blood in patients with rheumatoid arthritis. Methods: Serum TNF-α levels (with RIA), B cell as well as T cell subsets distribution type (with monoclonal antibody technique) were examined in 37 patients with rheumatoid arthritis and 30 controls. Results Serum TNF-α levels and B lymphocytes count were significantly higher in the patients than those in controls (P 3 , CD 4 and CD 4 /CD 8 were obviously lower (P<0.01). Conclusion: Rheumatoid arthritis is an autoimmune disease with abnormal immunoregulation. (authors)

The Development of Adult Innate Lymphoid Cells

Science.gov (United States)

Yang, Qi; Bhandoola, Avinash

2016-01-01

Innate lymphoid cells (ILC) are a specialized family of effector lymphocytes that transcriptionally and functionally mirror effector subsets of T cells, but differ from T cells in that they lack clonally-distributed adaptive antigen receptors. Our understanding of this family of lymphocytes is still in its infancy. In this review, we summarize current understanding and discuss recent insights into the cellular and molecular events that occur during early ILC development in adult mice. We discuss how these events overlap and diverge with the early development of adaptive T cells, and how they may influence the molecular and functional properties of mature ILC. PMID:26871595

Auxins differentially regulate root system architecture and cell cycle protein levels in maize seedlings.

Science.gov (United States)

Martínez-de la Cruz, Enrique; García-Ramírez, Elpidio; Vázquez-Ramos, Jorge M; Reyes de la Cruz, Homero; López-Bucio, José

2015-03-15

Maize (Zea mays) root system architecture has a complex organization, with adventitious and lateral roots determining its overall absorptive capacity. To generate basic information about the earlier stages of root development, we compared the post-embryonic growth of maize seedlings germinated in water-embedded cotton beds with that of plants obtained from embryonic axes cultivated in liquid medium. In addition, the effect of four different auxins, namely indole-3-acetic acid (IAA), 1-naphthaleneacetic acid (NAA), indole-3-butyric acid (IBA) and 2,4-dichlorophenoxyacetic acid (2,4-D) on root architecture and levels of the heat shock protein HSP101 and the cell cycle proteins CKS1, CYCA1 and CDKA1 were analyzed. Our data show that during the first days after germination, maize seedlings develop several root types with a simultaneous and/or continuous growth. The post-embryonic root development started with the formation of the primary root (PR) and seminal scutellar roots (SSR) and then continued with the formation of adventitious crown roots (CR), brace roots (BR) and lateral roots (LR). Auxins affected root architecture in a dose-response fashion; whereas NAA and IBA mostly stimulated crown root formation, 2,4-D showed a strong repressing effect on growth. The levels of HSP101, CKS1, CYCA1 and CDKA in root and leaf tissues were differentially affected by auxins and interestingly, HSP101 registered an auxin-inducible and root specific expression pattern. Taken together, our results show the timing of early branching patterns of maize and indicate that auxins regulate root development likely through modulation of the HSP101 and cell cycle proteins. Copyright © 2014 Elsevier GmbH. All rights reserved.

HIV-1 Tat protein induces glial cell autophagy through enhancement of BAG3 protein levels.

Science.gov (United States)

Bruno, Anna Paola; De Simone, Francesca Isabella; Iorio, Vittoria; De Marco, Margot; Khalili, Kamel; Sariyer, Ilker Kudret; Capunzo, Mario; Nori, Stefania Lucia; Rosati, Alessandra

2014-01-01

BAG3 protein has been described as an anti-apoptotic and pro-autophagic factor in several neoplastic and normal cells. We previously demonstrated that BAG3 expression is elevated upon HIV-1 infection of glial and T lymphocyte cells. Among HIV-1 proteins, Tat is highly involved in regulating host cell response to viral infection. Therefore, we investigated the possible role of Tat protein in modulating BAG3 protein levels and the autophagic process itself. In this report, we show that transfection with Tat raises BAG3 levels in glioblastoma cells. Moreover, BAG3 silencing results in highly reducing Tat- induced levels of LC3-II and increasing the appearance of sub G0/G1 apoptotic cells, in keeping with the reported role of BAG3 in modulating the autophagy/apoptosis balance. These results demonstrate for the first time that Tat protein is able to stimulate autophagy through increasing BAG3 levels in human glial cells.

Down-Regulation of Neuropathy Target Esterase in Preeclampsia Placenta Inhibits Human Trophoblast Cell Invasion via Modulating MMP-9 Levels

Directory of Open Access Journals (Sweden)

Ting Zhong

2018-02-01

Full Text Available Background/Aims: Neuropathy target esterase (NTE, also known as neurotoxic esterase is proven to deacylate phosphatidylcholine (PC to glycerophosphocholine as a phospholipase B. Recently; studies showed that artificial phosphatidylserine/PC microvesicles can induce preeclampsia (PE-like changes in pregnant mice. However, it is unclear whether NTE plays a key role in the pathology of PE, a pregnancy-related disease, which was characterized by deficient trophoblast invasion and reduced trophoblast-mediated remodeling of spiral arteries. The aim of this study was to investigate the expression pattern of NTE in the placenta from women with PE and normal pregnancy, and the molecular mechanism of NTE involved in the development of PE. Methods: NTE expression levels in placentas from 20 pregnant women with PE and 20 healthy pregnant women were detected using quantitative PCR and immunohistochemistry staining. The effect of NTE on trophoblast migration and invasion and the underlying mechanisms were examined in HTR-8/SVneo cell lines by transfection method. Results: NTE mRNA and protein expression levels were significantly decreased in preeclamptic placentas than normal control. Over-expression of NTE in HTR-8/SVneo cells significantly promoted trophoblast cells migration and invasion and was associated with increased MMP-9 levels. Conversely, shRNA-mediated down-regulation of NTE markedly inhibited the cell migration and invasion. In addition, silencing NTE reduced the MMP-9 activity and phosphorylated Erk1/2 and AKT levels. Conclusions: Our results suggest that the decreased NTE may contribute to the development of PE through impairing trophoblast invasion by down-regulating MMP-9 via the Erk1/2 and AKT signaling pathway.

Low baseline levels of NK cells may predict a positive response to ipilimumab in melanoma therapy.

Science.gov (United States)

Tietze, Julia K; Angelova, Daniela; Heppt, Markus V; Ruzicka, Thomas; Berking, Carola

2017-07-01

The introduction of immune checkpoint blockade (ICB) has been a breakthrough in the therapy of metastatic melanoma. The influence of ICB on T-cell populations has been studied extensively, but little is known about the effect on NK cells. In this study, we analysed the relative and absolute amounts of NK cells and of the subpopulations of CD56 dim and CD56 bright NK cells among the peripheral blood mononuclear cells (PBMCs) of 32 patients with metastatic melanoma before and under treatment with ipilimumab or pembrolizumab by flow cytometry. In 15 (47%) patients, an abnormal low amount of NK cells was found at baseline. Analysis of the subpopulations showed also low or normal baseline levels for CD56 dim NK cells, whereas the baseline levels of CD56 bright NK cells were either normal or abnormally high. The relative and absolute amounts of NK cells and of CD56 dim and CD56 bright NK cell subpopulations in patients with a normal baseline did not change under treatment. However, patients with a low baseline of NK cells and CD56 dim NK cells showed a significant increase in these immune cell subsets, but the amounts remained to be lower than the normal baseline. The amount of CD56 bright NK cells was unaffected by treatment. The baseline levels of NK cells were correlated with the number of metastatic organs. Their proportion increased, whereas the expression of NKG2D decreased significantly when more than one organ was affected by metastases. Low baseline levels of NK cells and CD56 dim NK cells as well as normal baseline levels of CD56 bright NK cells correlated significantly with a positive response to ipilimumab but not to pembrolizumab. Survival curves of patients with low amounts of CD56 dim NK cells treated with ipilimumab showed a trend to longer survival. Normal baseline levels of CD56 bright NK cells were significantly correlated with longer survival as compared to patients with high baseline levels. In conclusion, analysis of the amounts of total NK cells

Sodium arsenite alters cell cycle and MTHFR, MT1/2, and c-Myc protein levels in MCF-7 cells

International Nuclear Information System (INIS)

Ruiz-Ramos, Ruben; Lopez-Carrillo, Lizbeth; Albores, Arnulfo; Hernandez-Ramirez, Raul U.; Cebrian, Mariano E.

2009-01-01

There is limited available information on the effects of arsenic on enzymes participating in the folate cycle. Therefore, our aim was to evaluate the effects of sodium arsenite on the protein levels of methylenetetrahydrofolate reductase (MTHFR) and dihydrofolate reductase (DHFR) and its further relationship with the expression MT1/2 and c-myc in MCF-7 cells. Arsenite treatment (0-10 μM) for 4 h decreased MTHFR levels in a concentration-dependent fashion without significant effects on DHFR. The effects on MTHFR were observed at arsenite concentrations not significantly affecting cell viability. We also observed an increase in S-phase recruitment at all concentrations probed. Lower concentrations (< 5 μM) induced cell proliferation, showing a high proportion of BrdU-stained cells, indicating a higher DNA synthesis rate. However, higher concentrations (≥ 5 μM) or longer treatment periods induced apoptosis. Arsenite also induced dose-dependent increases in MT1/2 and c-Myc protein levels. The levels of MTHFR were inversely correlated to MT1/2 and c-Myc overexpression and increased S-phase recruitment. Our findings indicate that breast epithelial cells are responsive to arsenite and suggest that exposure may pose a risk for breast cancer. The reductions in MTHFR protein levels contribute to understand the mechanisms underlying the induction of genes influencing growth regulation, such as c-myc and MT1/2. However, further research is needed to ascertain if the effects here reported following short-time and high-dose exposure are relevant for human populations chronically exposed to low arsenic concentrations.

MG132 as a proteasome inhibitor induces cell growth inhibition and cell death in A549 lung cancer cells via influencing reactive oxygen species and GSH level.

Science.gov (United States)

Han, Yong Hwan; Park, Woo Hyun

2010-07-01

Carbobenzoxy-Leu-Leu-leucinal (MG132) as a proteasome inhibitor has been shown to induce apoptotic cell death through formation of reactive oxygen species (ROS). In the present study, we evaluated the effects of MG132 on the growth of A549 lung cancer cells in relation to cell growth, ROS and glutathione (GSH) levels. Treatment with MG132 inhibited the growth of A549 cells with an IC(50) of approximately 20 microM at 24 hours. DNA flow cytometric analysis indicated that 0.5 approximately 30 microM MG132 induced a G1 phase arrest of the cell cycle in A549 cells. Treatment with 10 or 30 microM MG132 also induced apoptosis, as evidenced by sub-G1 cells and annexin V staining cells. This was accompanied by the loss of mitochondrial membrane potential (MMP; Delta psi m). The intracellular ROS levels including O(2) (*-) were strongly increased in 10 or 30 microM MG132-treated A549 cells but were down-regulated in 0.1, 0.5 or 1 microM MG132-treated cells. Furthermore, 10 or 30 microM MG132 increased mitochondrial O(2) (*- ) level but 0.1, 0.5 or 1 microM MG132 decreased that. In addition, 10 or 30 microM MG132 induced GSH depletion in A549 cells. In conclusion, MG132 inhibited the growth of human A549 cells via inducing the cell cycle arrest as well as triggering apoptosis, which was in part correlated with the changes of ROS and GSH levels. Our present data provide important information on the anti-growth mechanisms of MG132 in A549 lung cancer cells in relation to ROS and GSH.

Selective ablation of the androgen receptor in mouse sertoli cells affects sertoli cell maturation, barrier formation and cytoskeletal development.

Directory of Open Access Journals (Sweden)

Ariane Willems

2010-11-01

Full Text Available The observation that mice with a selective ablation of the androgen receptor (AR in Sertoli cells (SC (SCARKO mice display a complete block in meiosis supports the contention that SC play a pivotal role in the control of germ cell development by androgens. To delineate the physiological and molecular mechanism responsible for this control, we compared tubular development in pubertal SCARKO mice and littermate controls. Particular attention was paid to differences in SC maturation, SC barrier formation and cytoskeletal organization and to the molecular mediators potentially involved. Functional analysis of SC barrier development by hypertonic perfusion and lanthanum permeation techniques and immunohistochemical analysis of junction formation showed that SCARKO mice still attempt to produce a barrier separating basal and adluminal compartment but that barrier formation is delayed and defective. Defective barrier formation was accompanied by disturbances in SC nuclear maturation (immature shape, absence of prominent, tripartite nucleoli and SC polarization (aberrant positioning of SC nuclei and cytoskeletal elements such as vimentin. Quantitative RT-PCR was used to study the transcript levels of genes potentially related to the described phenomena between day 8 and 35. Differences in the expression of SC genes known to play a role in junction formation could be shown from day 8 for Cldn11, from day 15 for Cldn3 and Espn, from day 20 for Cdh2 and Jam3 and from day 35 for ZO-1. Marked differences were also noted in the transcript levels of several genes that are also related to cell adhesion and cytoskeletal dynamics but that have not yet been studied in SC (Actn3, Ank3, Anxa9, Scin, Emb, Mpzl2. It is concluded that absence of a functional AR in SC impedes the remodeling of testicular tubules expected at the onset of spermatogenesis and interferes with the creation of the specific environment needed for germ cell development.

Radiation level analysis for the port cell of the ITER electron cyclotron-heating upper launcher

Energy Technology Data Exchange (ETDEWEB)

Weinhorst, Bastian, E-mail: bastian.weinhorst@kit.edu [KIT, Institute for Neutron Physics and Reactor Technology, Hermann-von-Helmholtz-Platz 1, 76344 Eggenstein-Leopoldshafen (Germany); Fischer, Ulrich; Lu, Lei [KIT, Institute for Neutron Physics and Reactor Technology, Hermann-von-Helmholtz-Platz 1, 76344 Eggenstein-Leopoldshafen (Germany); Strauss, Dirk; Spaeh, Peter; Scherer, Theo [KIT, Institute for Applied Materials, Hermann-von-Helmholtz-Platz 1, 76344 Eggenstein-Leopoldshafen (Germany); Leichtle, Dieter [F4E, Analysis & Codes/Technical Support Services, Josep Pla 2, Torres Diagonal Litoral B3, 08019 Barcelona (Spain)

2016-11-01

Highlights: • First detailed neutronic modelling of the ECHUL port cell with ECHUL equipment (including beam lines with diamond windows, the beam lines mounting box, conduit boxes and rails). • Three different bioshield port plug configurations and two different neutron source configurations are investigated. • Radiation Levels are calculated in the port cell, focusing on the position of the diamond window. • The dose rate in the port cell is below the limit for maintenance in the port cell. • The radiation level at the diamond window is very low and should not influence its performance. - Abstract: The electron cyclotron-heating upper launcher (ECHUL) will be installed in four upper ports of the ITER tokamak thermonuclear fusion reactor. Each ECHUL is able to deposit 8 MW power into the plasma for plasma mode stabilization via microwave beam lines. An essential part of these beam lines are the diamond windows. They are located in the upper port cell behind the bioshield to reduce the radiation levels to a minimum. The paper describes the first detailed neutronic modelling of the ECHUL port cell with ECHUL equipment. The bioshield plug is modelled including passageways for the microwave beam lines, piping and cables looms as well as rails and openings for ventilation. The port cell is equipped with the beam lines including the diamond windows, the beam lines mounting box, conduit boxes and rails. The neutrons are transported into the port cell starting from a surface source in front of the bioshield. Neutronic results are obtained for radiation levels in the port cell at different positions, mainly focusing on the diamond windows position. It is shown that the radiation level is below the limit for maintenance in the port cell. The radiation level at the diamond window is very low and should not influence its performance.

Multiweek Cell Culture Project for Use in Upper-Level Biology Laboratories

Science.gov (United States)

Marion, Rebecca E.; Gardner, Grant E.; Parks, Lisa D.

2012-01-01

This article describes a laboratory protocol for a multiweek project piloted in a new upper-level biology laboratory (BIO 426) using cell culture techniques. Human embryonic kidney-293 cells were used, and several culture media and supplements were identified for students to design their own experiments. Treatments included amino acids, EGF,…

The development of human mast cells. An historical reappraisal

Energy Technology Data Exchange (ETDEWEB)

Ribatti, Domenico, E-mail: domenico.ribatti@uniba.it

2016-03-15

The understanding of mast cell (MC) differentiation is derived mainly from in vitro studies of different stages of stem and progenitor cells. The hematopoietic lineage development of human MCs is unique compared to other myeloid-derived cells. Human MCs originate from CD34{sup +}/CD117{sup +}/CD13{sup +}multipotent hematopoietic progenitors, which undergo transendothelial recruitment into peripheral tissues, where they complete differentiation. Stem cell factor (SCF) is a major chemotactic factor for MCs and their progenitors. SCF also elicits cell-cell and cell-substratum adhesion, facilitates the proliferation, and sustains the survival, differentiation, and maturation, of MCs. Because MC maturation is influenced by local microenvironmental factors, different MC phenotypes can develop in different tissues and organs. - Highlights: • Human mast cells originate from CD34/CD117/CD13 positive multipotent hematopoietic progenitors. • Stem cell factor is a major chemotactic factor for mast cells and their progenitors. • Different mast cell phenotypes can develop in different tissues and organs.

The development of human mast cells. An historical reappraisal

International Nuclear Information System (INIS)

Ribatti, Domenico

2016-01-01

The understanding of mast cell (MC) differentiation is derived mainly from in vitro studies of different stages of stem and progenitor cells. The hematopoietic lineage development of human MCs is unique compared to other myeloid-derived cells. Human MCs originate from CD34"+/CD117"+/CD13"+multipotent hematopoietic progenitors, which undergo transendothelial recruitment into peripheral tissues, where they complete differentiation. Stem cell factor (SCF) is a major chemotactic factor for MCs and their progenitors. SCF also elicits cell-cell and cell-substratum adhesion, facilitates the proliferation, and sustains the survival, differentiation, and maturation, of MCs. Because MC maturation is influenced by local microenvironmental factors, different MC phenotypes can develop in different tissues and organs. - Highlights: • Human mast cells originate from CD34/CD117/CD13 positive multipotent hematopoietic progenitors. • Stem cell factor is a major chemotactic factor for mast cells and their progenitors. • Different mast cell phenotypes can develop in different tissues and organs.

Porcine pluripotency cell signaling develops from the inner cell mass to the epiblast during early development

DEFF Research Database (Denmark)

Hall, Vanessa Jane; Christensen, Josef; Gao, Yu

2009-01-01

  The signaling mechanisms regulating pluripotency in porcine embryonic stem cells and embryos are unknown. In this study, we characterize cell signaling in the in-vivo porcine inner cell mass and later-stage epiblast. We evaluate expression of OCT4, NANOG, SOX2, genes within the JAK/STAT pathway...... pluripotency in human embryonic stem cells is detectable in the porcine epiblast, but not in the inner cell mass. Copyright (c) 2009 Wiley-Liss, Inc.......  The signaling mechanisms regulating pluripotency in porcine embryonic stem cells and embryos are unknown. In this study, we characterize cell signaling in the in-vivo porcine inner cell mass and later-stage epiblast. We evaluate expression of OCT4, NANOG, SOX2, genes within the JAK/STAT pathway...... (LIF, LIFR, GP130), FGF pathway (bFGF, FGFR1, FGFR2), BMP pathway (BMP4), and downstream-activated genes (STAT3, c-Myc, c-Fos, and SMAD4). We discovered two different expression profiles exist in the developing porcine embryo. The D6 porcine blastocyst (inner cell mass stage) is devoid...

Correlation of Cell Surface Biomarker Expression Levels with Adhesion Contact Angle Measured by Lateral Microscopy.

Science.gov (United States)

Walz, Jenna A; Mace, Charles R

2018-06-05

Immunophenotyping is typically achieved using flow cytometry, but any influence a biomarker may have on adhesion or surface recognition cannot be determined concurrently. In this manuscript, we demonstrate the utility of lateral microscopy for correlating cell surface biomarker expression levels with quantitative descriptions of cell morphology. With our imaging system, we observed single cells from two T cell lines and two B cell lines adhere to antibody-coated substrates and quantified this adhesion using contact angle measurements. We found that SUP-T1 and CEM CD4+ cells, both of which express similar levels of CD4, experienced average changes in contact angle that were not statistically different from one another on surfaces coated in anti-CD4. However, MAVER-1 and BJAB K20 cells, both of which express different levels of CD20, underwent average changes in contact angle that were significantly different from one another on surfaces coated in anti-CD20. Our results indicate that changes in cell contact angles on antibody-coated substrates reflect the expression levels of corresponding antigens on the surfaces of cells as determined by flow cytometry. Our lateral microscopy approach offers a more reproducible and quantitative alternative to evaluate adhesion compared to commonly used wash assays and can be extended to many additional immunophenotyping applications to identify cells of interest within heterogeneous populations.

Stepwise development of MAIT cells in mouse and human.

Directory of Open Access Journals (Sweden)

Emmanuel Martin

2009-03-01

Full Text Available Mucosal-associated invariant T (MAIT cells display two evolutionarily conserved features: an invariant T cell receptor (TCRalpha (iTCRalpha chain and restriction by the nonpolymorphic class Ib major histocompatibility complex (MHC molecule, MHC-related molecule 1 (MR1. MR1 expression on thymus epithelial cells is not necessary for MAIT cell development but their accumulation in the gut requires MR1 expressing B cells and commensal flora. MAIT cell development is poorly known, as these cells have not been found in the thymus so far. Herein, complementary human and mouse experiments using an anti-humanValpha7.2 antibody and MAIT cell-specific iTCRalpha and TCRbeta transgenic mice in different genetic backgrounds show that MAIT cell development is a stepwise process, with an intra-thymic selection followed by peripheral expansion. Mouse MAIT cells are selected in an MR1-dependent manner both in fetal thymic organ culture and in double iTCRalpha and TCRbeta transgenic RAG knockout mice. In the latter mice, MAIT cells do not expand in the periphery unless B cells are added back by adoptive transfer, showing that B cells are not required for the initial thymic selection step but for the peripheral accumulation. In humans, contrary to natural killer T (NKT cells, MAIT cells display a naïve phenotype in the thymus as well as in cord blood where they are in low numbers. After birth, MAIT cells acquire a memory phenotype and expand dramatically, up to 1%-4% of blood T cells. Finally, in contrast with NKT cells, human MAIT cell development is independent of the molecular adaptor SAP. Interestingly, mouse MAIT cells display a naïve phenotype and do not express the ZBTB16 transcription factor, which, in contrast, is expressed by NKT cells and the memory human MAIT cells found in the periphery after birth. In conclusion, MAIT cells are selected by MR1 in the thymus on a non-B non-T hematopoietic cell, and acquire a memory phenotype and expand in the

Dissecting stem cell differentiation using single cell expression profiling

OpenAIRE

Moignard, Victoria Rachel; Göttgens, Berthold

2016-01-01

Many assumptions about the way cells behave are based on analyses of populations. However, it is now widely recognized that even apparently pure populations can display a remarkable level of heterogeneity. This is particularly true in stem cell biology where it hinders our understanding of normal development and the development of strategies for regenerative medicine. Over the past decade technologies facilitating gene expression analysis at the single cell level have become widespread, provi...

Development Track of Different County-Level Towns

OpenAIRE

ZHANG, Haijiao; ZHANG, Zhenghe

2015-01-01

County-level town is important space carrier of China’s urbanization and the emphasis and key of strategic distribution for urbanization. Backwardness of county-level towns limits nearby transfer of surplus labor. To promote development of county-level towns, on the basis of classifying county-level towns, this paper analyzed motive and resistance factors of evolution of different county-level towns using historical data collection and statistical method, comparative approach, typical case ...

The development of microfabricated biocatalytic fuel cells

Energy Technology Data Exchange (ETDEWEB)

Sasaki, Satoshi; Karube, Isao [University of Tokyo (Japan). Research Center for Advanced Science and Technology

1999-02-01

The production of electricity by biocatalytic fuel cells has been feasible for almost two decades and can produce electric power at a practical level. These fuel cells use immobilized microorganisms or enzymes as catalysts, and glucose as a fuel. A microfabricated enzyme battery has recently been made that is designed to function as a power supply for microsurgery robots or artificial organs. (author)

The effects of oxygen level and glucose concentration on the metabolism of porcine TMJ disc cells.

Science.gov (United States)

Cisewski, S E; Zhang, L; Kuo, J; Wright, G J; Wu, Y; Kern, M J; Yao, H

2015-10-01

To determine the combined effect of oxygen level and glucose concentration on cell viability, ATP production, and matrix synthesis of temporomandibular joint (TMJ) disc cells. TMJ disc cells were isolated from pigs aged 6-8 months and cultured in a monolayer. Cell cultures were preconditioned for 48 h with 0, 1.5, 5, or 25 mM glucose DMEM under 1%, 5%, 10%, or 21% O2 level, respectively. The cell viability was measured using the WST-1 assay. ATP production was determined using the Luciferin-Luciferase assay. Collagen and proteoglycan synthesis were determined by measuring the incorporation of [2, 3-(3)H] proline and [(35)S] sulfate into the cells, respectively. TMJ disc cell viability significantly decreased (P oxygen levels significantly increased viability (P oxygen levels significantly reduced ATP production (P oxygen was significant in regards to cell viability (P oxygen are important, glucose is the limiting nutrient for TMJ disc cell survival. At low oxygen levels, the production of ATP, collagen, and proteoglycan are severely inhibited. These results suggest that steeper nutrient gradients may exist in the TMJ disc and it may be vulnerable to pathological events that impede nutrient supply. Copyright © 2015 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.

A role for Lin28 in primordial germ cell development and germ cell malignancy

Science.gov (United States)

West, Jason A.; Viswanathan, Srinivas R.; Yabuuchi, Akiko; Cunniff, Kerianne; Takeuchi, Ayumu; Park, In-Hyun; Sero, Julia E.; Zhu, Hao; Perez-Atayde, Antonio; Frazier, A. Lindsay; Surani, M. Azim; Daley, George Q.

2009-01-01

The rarity and inaccessibility of the earliest primordial germ cells (PGCs) in the mouse embryo thwarts efforts to investigate molecular mechanisms of germ cell specification. Stella marks the minute founder population of the germ lineage1,2. Here we differentiate mouse embryonic stem cells (ESCs) carrying a Stella transgenic reporter into putative PGCs in vitro. The Stella+ cells possess a transcriptional profile similar to embryo-derived PGCs, and like their counterparts in vivo, lose imprints in a time-dependent manner. Using inhibitory RNAs to screen candidate genes for effects on the development of Stella+ cells in vitro, we discovered that Lin28, a negative regulator of let-7 microRNA processing3-6, is essential for proper PGC development. We further show that Blimp1, a let-7 target and a master regulator of PGC specification7-9, can rescue the effect of Lin28-deficiency during PGC development, thereby establishing a mechanism of action for Lin28 during PGC specification. Over-expression of Lin28 promotes formation of Stella+ cells in vitro and PGCs in chimeric embryos, and is associated with human germ cell tumours. The differentiation of putative PGCs from ESCs in vitro recapitulates the early stages of gamete development in vivo, and provides an accessible system for discovering novel genes involved in germ cell development and malignancy. PMID:19578360

Analysis of gene expression levels in individual bacterial cells without image segmentation

Energy Technology Data Exchange (ETDEWEB)

Kwak, In Hae; Son, Minjun [Physics Department, University of Florida, P.O. Box 118440, Gainesville, FL 32611-8440 (United States); Hagen, Stephen J., E-mail: sjhagen@ufl.edu [Physics Department, University of Florida, P.O. Box 118440, Gainesville, FL 32611-8440 (United States)

2012-05-11

Highlights: Black-Right-Pointing-Pointer We present a method for extracting gene expression data from images of bacterial cells. Black-Right-Pointing-Pointer The method does not employ cell segmentation and does not require high magnification. Black-Right-Pointing-Pointer Fluorescence and phase contrast images of the cells are correlated through the physics of phase contrast. Black-Right-Pointing-Pointer We demonstrate the method by characterizing noisy expression of comX in Streptococcus mutans. -- Abstract: Studies of stochasticity in gene expression typically make use of fluorescent protein reporters, which permit the measurement of expression levels within individual cells by fluorescence microscopy. Analysis of such microscopy images is almost invariably based on a segmentation algorithm, where the image of a cell or cluster is analyzed mathematically to delineate individual cell boundaries. However segmentation can be ineffective for studying bacterial cells or clusters, especially at lower magnification, where outlines of individual cells are poorly resolved. Here we demonstrate an alternative method for analyzing such images without segmentation. The method employs a comparison between the pixel brightness in phase contrast vs fluorescence microscopy images. By fitting the correlation between phase contrast and fluorescence intensity to a physical model, we obtain well-defined estimates for the different levels of gene expression that are present in the cell or cluster. The method reveals the boundaries of the individual cells, even if the source images lack the resolution to show these boundaries clearly.

Development of a Functional Schwann Cell Phenotype from Autologous Porcine Bone Marrow Mononuclear Cells for Nerve Repair

Directory of Open Access Journals (Sweden)

Michael J. Rutten

2012-01-01

Full Text Available Adult bone marrow mononuclear cells (BM-MNCs are a potential resource for making Schwann cells to repair damaged peripheral nerves. However, many methods of producing Schwann-like cells can be laborious with the cells lacking a functional phenotype. The objective of this study was to develop a simple and rapid method using autologous BM-MNCs to produce a phenotypic and functional Schwann-like cell. Adult porcine bone marrow was collected and enriched for BM-MNCs using a SEPAX device, then cells cultured in Neurobasal media, 4 mM L-glutamine and 20% serum. After 6–8 days, the cultures expressed Schwann cell markers, S-100, O4, GFAP, were FluoroMyelin positive, but had low p75(NGF expression. Addition of neuregulin (1–25 nM increased p75(NGF levels at 24–48 hrs. We found ATP dose-dependently increased intracellular calcium [Ca2+]i, with nucleotide potency being UTP=ATP>ADP>AMP>adenosine. Suramin blocked the ATP-induced [Ca2+]i but α, β,-methylene-ATP had little effect suggesting an ATP purinergic P2Y2 G-protein-coupled receptor is present. Both the Schwann cell markers and ATP-induced [Ca2+]i sensitivity decreased in cells passaged >20 times. Our studies indicate that autologous BM-MNCs can be induced to form a phenotypic and functional Schwann-like cell which could be used for peripheral nerve repair.

Intracellular cholesterol level regulates sensitivity of glioblastoma cells against temozolomide-induced cell death by modulation of caspase-8 activation via death receptor 5-accumulation and activation in the plasma membrane lipid raft.

Science.gov (United States)

Yamamoto, Yutaro; Tomiyama, Arata; Sasaki, Nobuyoshi; Yamaguchi, Hideki; Shirakihara, Takuya; Nakashima, Katsuhiko; Kumagai, Kosuke; Takeuchi, Satoru; Toyooka, Terushige; Otani, Naoki; Wada, Kojiro; Narita, Yoshitaka; Ichimura, Koichi; Sakai, Ryuichi; Namba, Hiroki; Mori, Kentaro

2018-01-01

Development of resistance against temozolomide (TMZ) in glioblastoma (GBM) after continuous treatment with TMZ is one of the critical problems in clinical GBM therapy. Intracellular cholesterol regulates cancer cell biology, but whether intracellular cholesterol is involved in TMZ resistance of GBM cells remains unclear. The involvement of intracellular cholesterol in acquired resistance against TMZ in GBM cells was investigated. Intracellular cholesterol levels were measured in human U251 MG cells with acquired TMZ resistance (U251-R cells) and TMZ-sensitive control U251 MG cells (U251-Con cells), and found that the intracellular cholesterol level was significantly lower in U251-R cells than in U251-Con cells. In addition, treatment by intracellular cholesterol remover, methyl-beta cyclodextrin (MβCD), or intracellular cholesterol inducer, soluble cholesterol (Chol), regulated TMZ-induced U251-Con cell death in line with changes in intracellular cholesterol level. Involvement of death receptor 5 (DR5), a death receptor localized in the plasma membrane, was evaluated. TMZ without or with MβCD and/or Chol caused accumulation of DR5 into the plasma membrane lipid raft and formed a complex with caspase-8, an extrinsic caspase cascade inducer, reflected in the induction of cell death. In addition, treatment with caspase-8 inhibitor or knockdown of DR5 dramatically suppressed U251-Con cell death induced by combination treatment with TMZ, MβCD, and Chol. Combined treatment of Chol with TMZ reversed the TMZ resistance of U251-R cells and another GBM cell model with acquired TMZ resistance, whereas clinical antihypercholesterolemia agents at physiological concentrations suppressed TMZ-induced cell death of U251-Con cells. These findings suggest that intracellular cholesterol level affects TMZ treatment of GBM mediated via a DR5-caspase-8 mechanism. Copyright © 2017 Elsevier Inc. All rights reserved.

324 Building radiochemical engineering cells, high-level vault, low-level vault, and associated areas closure plan

International Nuclear Information System (INIS)

Barnett, J.M.

1998-01-01

The Hanford Site, located adjacent to and north of Richland, Washington, is operated by the US Department of Energy, Richland Operations Office (RL). The 324 Building is located in the 300 Area of the Hanford Site. The 324 Building was constructed in the 1960s to support materials and chemical process research and development activities ranging from laboratory/bench-scale studies to full engineering-scale pilot plant demonstrations. In the mid-1990s, it was determined that dangerous waste and waste residues were being stored for greater than 90 days in the 324 Building Radiochemical Engineering Cells (REC) and in the High-Level Vault/Low-Level Vault (HLV/LLV) tanks. [These areas are not Resource Conservation and Recovery Act of 1976 (RCRA) permitted portions of the 324 Building.] Through the Hanford Federal Facility Agreement and Consent Order (Tri-Party Agreement) Milestone M-89, agreement was reached to close the nonpermitted RCRA unit in the 324 Building. This closure plan, managed under TPA Milestone M-20-55, addresses the identified building areas targeted by the Tri-Party Agreement and provides commitments to achieve the highest degree of compliance practicable, given the special technical difficulties of managing mixed waste that contains high-activity radioactive materials, and the physical limitations of working remotely in the areas within the subject closure unit. This closure plan is divided into nine chapters. Chapter 1.0 provides the introduction, historical perspective, 324 Building history and current mission, and the regulatory basis and strategy for managing the closure unit. Chapters 2.0, 3.0, 4.0, and 5.0 discuss the detailed facility description, process information, waste characteristics, and groundwater monitoring respectively. Chapter 6.0 deals with the closure strategy and performance standard, including the closure activities for the B-Cell, D-Cell, HLV, LLV; piping and miscellaneous associated building areas. Chapter 7.0 addresses the

Physicochemical properties of newly developed bioactive glass cement and its effects on various cells.

Science.gov (United States)

Washio, Ayako; Nakagawa, Aika; Nishihara, Tatsuji; Maeda, Hidefumi; Kitamura, Chiaki

2015-02-01

Biomaterials used in dental treatments are expected to have favorable properties such as biocompatibility and an ability to induce tissue formation in dental pulp and periapical tissue, as well as sealing to block external stimuli. Bioactive glasses have been applied in bone engineering, but rarely applied in the field of dentistry. In the present study, bioactive glass cement for dental treatment was developed, and then its physicochemical properties and effects on cell responses were analyzed. To clarify the physicochemical attributes of the cement, field emission scanning electron microscopy, X-ray diffraction, and pH measurement were carried out. Cell attachment, morphology, and viability to the cement were also examined to clarify the effects of the cement on odontoblast-like cells (KN-3 cells), osteoblastic cells (MC3T3-E1 cells), human periodontal ligament stem/progenitor cells and neuro-differentiative cells (PC-12 cells). Hydroxyapatite-like precipitation was formed on the surface of the hardened cement and the pH level changed from pH10 to pH9, then stabilized in simulate body fluid. The cement had no cytotxic effects on these cells, and particulary induced process elongation of PC-12 cells. Our results suggest that the newly developed bioactive glass cement have capability of the application in dental procedures as bioactive cement. © 2014 Wiley Periodicals, Inc.

Classroom-Level Teacher Professional Development and Satisfaction: Teachers Learn in the Context of Classroom-Level Curriculum Development

Science.gov (United States)

Shawer, Saad

2010-01-01

This qualitative study examined the impact of classroom-level teacher professional development (CTPD) and curriculum transmission on teacher professional development and satisfaction. Based on work with English-as-a-foreign-language college teachers and students, data analysis showed that CTPD significantly improved student-teacher subject,…

Vitamin C enhances in vitro and in vivo development of porcine somatic cell nuclear transfer embryos

Energy Technology Data Exchange (ETDEWEB)

Huang, Yongye; Tang, Xiaochun; Xie, Wanhua; Zhou, Yan; Li, Dong; Zhou, Yang; Zhu, Jianguo; Yuan, Ting; Lai, Liangxue [Jilin Province Key Laboratory of Animal Embryo Engineering, College of Animal Science and Veterinary Medicine, Jilin University, 5333 Xi An DaLu, Changchun 130062 (China); Pang, Daxin, E-mail: pdx@jlu.edu.cn [Jilin Province Key Laboratory of Animal Embryo Engineering, College of Animal Science and Veterinary Medicine, Jilin University, 5333 Xi An DaLu, Changchun 130062 (China); Ouyang, Hongsheng, E-mail: ouyh@jlu.edu.cn [Jilin Province Key Laboratory of Animal Embryo Engineering, College of Animal Science and Veterinary Medicine, Jilin University, 5333 Xi An DaLu, Changchun 130062 (China)

2011-07-29

Highlights: {yields} Report for the first time that vitamin C has a beneficial effect on the development of porcine SCNT embryos. {yields} The level of acH4K5 and Oct4 expression at blastocyst-stage was up-regulated after treatment. {yields} A higher rate of gestation and increased number of piglets born were harvested in the treated group. -- Abstract: The reprogramming of differentiated cells into a totipotent embryonic state through somatic cell nuclear transfer (SCNT) is still an inefficient process. Previous studies revealed that the generation of induced pluripotent stem (iPS) cells from mouse and human fibroblasts could be significantly enhanced with vitamin C treatment. Here, we investigated the effects of vitamin C, to our knowledge for the first time, on the in vitro and in vivo development of porcine SCNT embryos. The rate of blastocyst development in SCNT embryos treated with 50 {mu}g/mL vitamin C 15 h after activation (36.0%) was significantly higher than that of untreated SCNT embryos (11.5%). The enhanced in vitro development rate of vitamin C-treated embryos was associated with an increased acetylation level of histone H4 lysine 5 and higher Oct4, Sox2 and Klf4 expression levels in blastocysts, as determined by real-time PCR. In addition, treatment with vitamin C resulted in an increased pregnancy rate in pigs. These findings suggest that treatment with vitamin C is beneficial for enhancement of the in vitro and in vivo development of porcine SCNT embryos.

Vitamin C enhances in vitro and in vivo development of porcine somatic cell nuclear transfer embryos

International Nuclear Information System (INIS)

Huang, Yongye; Tang, Xiaochun; Xie, Wanhua; Zhou, Yan; Li, Dong; Zhou, Yang; Zhu, Jianguo; Yuan, Ting; Lai, Liangxue; Pang, Daxin; Ouyang, Hongsheng

2011-01-01

Highlights: → Report for the first time that vitamin C has a beneficial effect on the development of porcine SCNT embryos. → The level of acH4K5 and Oct4 expression at blastocyst-stage was up-regulated after treatment. → A higher rate of gestation and increased number of piglets born were harvested in the treated group. -- Abstract: The reprogramming of differentiated cells into a totipotent embryonic state through somatic cell nuclear transfer (SCNT) is still an inefficient process. Previous studies revealed that the generation of induced pluripotent stem (iPS) cells from mouse and human fibroblasts could be significantly enhanced with vitamin C treatment. Here, we investigated the effects of vitamin C, to our knowledge for the first time, on the in vitro and in vivo development of porcine SCNT embryos. The rate of blastocyst development in SCNT embryos treated with 50 μg/mL vitamin C 15 h after activation (36.0%) was significantly higher than that of untreated SCNT embryos (11.5%). The enhanced in vitro development rate of vitamin C-treated embryos was associated with an increased acetylation level of histone H4 lysine 5 and higher Oct4, Sox2 and Klf4 expression levels in blastocysts, as determined by real-time PCR. In addition, treatment with vitamin C resulted in an increased pregnancy rate in pigs. These findings suggest that treatment with vitamin C is beneficial for enhancement of the in vitro and in vivo development of porcine SCNT embryos.

Low-level radioactive waste management technology development

International Nuclear Information System (INIS)

Coleman, J.A.

1985-01-01

Although reviews of disposal practices and site performance indicated that there were no releases to the environment that would affect public health and safety, it became clear that: (a) several burial grounds were not performing as expected; (b) long-term maintenance of closed trenches could be a costly problem, and (c) more cost-effective methods could be developed for the treatment, packing, and disposal of low-level waste. As a result of these reviews, the Department of Energy developed the Low-level Waste Management Program to seek improvements in existing practices, correct obvious deficiencies, and develop site closure techniques that would avoid expensive long-term maintenance and monitoring. Such technology developments provide a better understanding of the physical and technical mechanisms governing low-level waste treatment and disposal and lead to improvement in the performance of disposal sites. The primary means of disposal of low-level waste has been the accepted and regulated practice of shallow land disposal, i.e., placement of low-level waste in trenches 5 to 10 meters deep with several meters of special soil cover. Department of Energy waste is primarily disposed at six major shallow land disposal sites. Commercial waste is currently disposed of at three major sites in the nation - Barnwell, South Carolina; Richland, Washington; and Beatty, Nevada. In the late 1970's public concern arose regarding the management practices of sites operated by the civilian sector and by the Department of Energy

Development and Prospect of Nanoarchitectured Solar Cells

Directory of Open Access Journals (Sweden)

Bo Zhang

2015-01-01

Full Text Available This paper gives an overview of the development and prospect of nanotechnologies utilized in the solar cell applications. Even though it is not clearly pointed out, nanostructures indeed have been used in the fabrication of conventional solar cells for a long time. However, in those circumstances, only very limited benefits of nanostructures have been used to improve cell performance. During the last decade, the development of the photovoltaic device theory and nanofabrication technology enables studies of more complex nanostructured solar cells with higher conversion efficiency and lower production cost. The fundamental principles and important features of these advanced solar cell designs are systematically reviewed and summarized in this paper, with a focus on the function and role of nanostructures and the key factors affecting device performance. Among various nanostructures, special attention is given to those relying on quantum effect.

Erythropoetin treatment can increase 2,3-diphosphoglycerate levels in red blood cells.

Science.gov (United States)

Birgegård, G; Sandhagen, B

2001-01-01

Some patients experience an improved well-being during treatment with recombinant human erythropoietin even with an unchanged Hb level. We have hypothesized that this may not be only a placebo effect. 2,3-diphosphoglycerate (2,3-DPG) in red blood cells increases in response to anaemia/hypoxia and causes a shift of the oxygen dissociation curve, allowing a more effective oxygen delivery. We have investigated red cell 2,3-DPG concentrations during erythropoietin treatment in healthy volunteers as a mediator of a possible physiological explanation. Thirteen healthy subjects with no iron deficiency were recruited and randomly assigned to a treatment group comprising five males and three females and a control group including three males and two females. The treatment group was treated with erythropoietin (Recormon), 20 IE/kg subcutaneously three times/week for 4 weeks. Blood samples were collected at each injection day and 10 days after the last injection and at corresponding times in the control group. B-Hb, red cell 2,3-DPG and P50 were measured by standard techniques and oxygen-releasing capacity was calculated. due to the sampling (26 ml each time, three times/week) the mean Hb level was lowered from 140.5 +/- 5.9 to 128.6 +/- 10.4 g/L in the control group whereas the erythropoietin treatment group maintained a mean Hb level of about 142 g/L (plevel curve as well as that for oxygen releasing capacity also differed significantly between the two groups (p levels. treatment with erythropoietin causes an increase in red cell 2,3-DPG levels.

Location and cellular stages of NK cell development

Science.gov (United States)

Yu, Jianhua; Freud, Aharon G.; Caligiuri, Michael A

2013-01-01

The identification of distinct tissue-specific natural killer (NK) cell populations that apparently mature from local precursor populations has brought new insight into the diversity and developmental regulation of this important lymphoid subset. NK cells provide a necessary link between the early (innate) and late (adaptive) immune responses to infection. Gaining a better understanding of the processes that govern NK cell development should allow us to better harness NK cell functions in multiple clinical settings as well as to gain further insight into how these cells undergo malignant transformation. In this review, we summarize recent advances in understanding sites and cellular stages of NK cell development in humans and mice. PMID:24055329

Red cell 2,3-diphosphoglycerate levels in children with hereditary haemolytic anaemias.

Science.gov (United States)

Haidas, S; Zannos-Mariolea, L; Matsaniotis, N

1975-12-01

The role of red cell 2,3-diphosphoglycerate (2,3-DPG) in increasing the availability of haemoglobin oxygen in neonatal jaundice and hereditary haemolytic anaemias was investigated. Measurements of 2,3-DPG were carried out on 58 normal children and six normal adults, 18 full-term newborns with neonatal jaundice and 57 cases (51 children and six adults) with hereditary haemolytic anaemias. In normal children and adults, with a mean haemoglobin of 12.69 g/dl, mean 2,3-DPG was 14.90 mumol/g Hb. In jaundiced newborns with a mean haemoglobin of 16.04 g/dl mean 2,3-DPG levels were 14.51 mumol/g Hb, i.e. normal. 2,3-DPG levels were increased in patients with beta-thalassaemia major, alpha-thalassaemia, sickle-cell disease, favism, hereditary spherocytosis and in heterozygotes for beta-thalassaemia with increased haemoglobin F. In heterozygotes for beta-thalassaemia with increased haemoglobin A2 only and in sickle cell trait 2,3-DPG levels were normal.

CD28 in thymocyte development and peripheral T cell activation in mice exposed to suspended particulate matter

International Nuclear Information System (INIS)

Drela, Nadzieja; Zesko, Izabela; Jakubowska, Martyna; Biernacka, Marzena

2006-01-01

The CD28:B7 signaling pathway is very important for the activity of mature peripheral T lymphocytes and thymocyte development. The proper development of thymocytes into mature single positive CD4 + and CD8 + T cells is crucial for almost all immune functions. In naturally occurring conditions, T cells maturation in the thymus is influenced by environmental agents. The expression of CD28 and the distribution of CD28 low/high thymocytes have been examined at various stages of thymocyte development in BALB/c mice exposed to air-suspended particulate matter (ASM). Acute exposure to ASM resulted in the decrease of CD28 expression in the total thymocyte population. The increase of the percentage of CD28 low and the decrease of CD28 high thymocytes were observed, which may account for the acceleration of thymocyte development under the conditions of elevated risk resulting from the exposure of animals to environmental xenobiotics. ASM exposure resulted in the increase of the level of proliferation of lymph node T cells induced by anti-CD3 and anti-CD28 monoclonal antibodies activation despite normal expression of CD28 molecule. In contrast, the level of proliferation of spleen T cells was lowered or normal dependently of the concentration of stimuli used for activation. Results of these studies demonstrate that acute exposure of mice to ASM can result in the progression of two contrasting processes in the immune system: upregulation of thymocyte development, which contributes to the maintenance of peripheral T cell pool, and over-activation of lymph node lymphocytes, which may lead to uncontrolled immunostimulation

Toward Development of Pluripotent Porcine Stem Cells by Road Mapping Early Embryonic Development

DEFF Research Database (Denmark)

Petkov, Stoyan; Freude, Kristine; Mashayekhi-Nezamabadi, Kaveh

2017-01-01

The lack in production of bona fide porcine pluripotent stem cells has definitely been hampered by a lack of research into porcine embryo development. Embryonic development in mammals is the extraordinary transition of a single-celled fertilized zygote into a complex fetus, which occurs...... in the uterus of the maternal adult during the early stages of gestation. Biomedical pig models could serve as genetic backgrounds for establishment of embryonic stem cells (ESCs) or other pluripotent stem cells (such as iPSC), which may be used to model and study diseases in vitro. This chapter provides...... insight into the current knowledge of pluripotent states in the developing pig embryo and the current status in establishment of bona fide porcine ESC (pESC) and piPSCs. It reflects the potential causes underlying the difficulty in establishing pluripotent stem cells and reviews recent data on global...

Development of a population of cancer cells: Observation and modeling by a Mixed Spatial Evolutionary Games approach.

Science.gov (United States)

Świerniak, Andrzej; Krześlak, Michał; Student, Sebastian; Rzeszowska-Wolny, Joanna

2016-09-21

Living cells, like whole living organisms during evolution, communicate with their neighbors, interact with the environment, divide, change their phenotypes, and eventually die. The development of specific ways of communication (through signaling molecules and receptors) allows some cellular subpopulations to survive better, to coordinate their physiological status, and during embryonal development to create tissues and organs or in some conditions to become tumors. Populations of cells cultured in vitro interact similarly, also competing for space and nutrients and stimulating each other to better survive or to die. The results of these intercellular interactions of different types seem to be good examples of biological evolutionary games, and have been the subjects of simulations by the methods of evolutionary game theory where individual cells are treated as players. Here we present examples of intercellular contacts in a population of living human cancer HeLa cells cultured in vitro and propose an evolutionary game theory approach to model the development of such populations. We propose a new technique termed Mixed Spatial Evolutionary Games (MSEG) which are played on multiple lattices corresponding to the possible cellular phenotypes which gives the possibility of simulating and investigating the effects of heterogeneity at the cellular level in addition to the population level. Analyses performed with MSEG suggested different ways in which cellular populations develop in the case of cells communicating directly and through factors released to the environment. Copyright © 2016 Elsevier Ltd. All rights reserved.

Novel, Moon and Mars, partial gravity simulation paradigms and their effects on the balance between cell growth and cell proliferation during early plant development.

Science.gov (United States)

Manzano, Aránzazu; Herranz, Raúl; den Toom, Leonardus A; Te Slaa, Sjoerd; Borst, Guus; Visser, Martijn; Medina, F Javier; van Loon, Jack J W A

2018-01-01

Clinostats and Random Positioning Machine (RPM) are used to simulate microgravity, but, for space exploration, we need to know the response of living systems to fractional levels of gravity (partial gravity) as they exist on Moon and Mars. We have developed and compared two different paradigms to simulate partial gravity using the RPM, one by implementing a centrifuge on the RPM (RPM HW ), the other by applying specific software protocols to driving the RPM motors (RPM SW ). The effects of the simulated partial gravity were tested in plant root meristematic cells, a system with known response to real and simulated microgravity. Seeds of Arabidopsis thaliana were germinated under simulated Moon (0.17  g ) and Mars (0.38  g ) gravity. In parallel, seeds germinated under simulated microgravity (RPM), or at 1  g control conditions. Fixed root meristematic cells from 4-day grown seedlings were analyzed for cell proliferation rate and rate of ribosome biogenesis using morphometrical methods and molecular markers of the regulation of cell cycle and nucleolar activity. Cell proliferation appeared increased and cell growth was depleted under Moon gravity, compared with the 1  g control. The effects were even higher at the Moon level than at simulated microgravity, indicating that meristematic competence (balance between cell growth and proliferation) is also affected at this gravity level. However, the results at the simulated Mars level were close to the 1  g static control. This suggests that the threshold for sensing and responding to gravity alteration in the root would be at a level intermediate between Moon and Mars gravity. Both partial g simulation strategies seem valid and show similar results at Moon g -levels, but further research is needed, in spaceflight and simulation facilities, especially around and beyond Mars g levels to better understand more precisely the differences and constrains in the use of these facilities for the space biology community.

Program-level and contextual-level determinants of low-median CD4+ cell count in cohorts of persons initiating ART in eight sub-Saharan African countries.

Science.gov (United States)

Nash, Denis; Wu, Yingfeng; Elul, Batya; Hoos, David; El Sadr, Wafaa

2011-07-31

In sub-Saharan Africa, many patients initiate antiretroviral therapy (ART) at CD4 cell counts much lower than those recommended in national guidelines. We examined program-level and contextual-level factors associated with low median CD4 cell count at ART initiation in populations initiating ART. Multilevel analysis of aggregate and program-level service delivery data. We examined data on 1690 cohorts of patients initiating ART during 2004-2008 in eight sub-Saharan African countries. Cohorts with median CD4 less than 111 cells/μl (the lowest quartile) were classified as having low median CD4 cell count at ART initiation. Cohort information was combined with time-updated program-level data and subnational contextual-level data, and analyzed using multilevel models. The 1690 cohorts had median CD4 cell count of 136 cells/μl and included 121,504 patients initiating ART at 267 clinics. Program-level factors associated with low cohort median CD4 cell count included urban setting [adjusted odds ratio (AOR) 2.1; 95% confidence interval (CI) 1.3-3.3], lower provider-to-patient ratio (AOR 2.2; 95% CI 1.3-4.0), no PMTCT program (AOR 3.6; 95% CI 1.0-12.8), outreach services for ART patients only vs. both pre-ART and ART patients (AOR 2.4; 95% CI 1.5-3.9), fewer vs. more adherence support services (AOR 1.6; 95% CI 1.0-2.5), and smaller cohort size (AOR 2.5; 95% CI 1.4-4.5). Contextual-level factors associated with low cohort median CD4 cell count included initiating ART in areas where a lower proportion of the population heard of AIDS, tested for HIV recently, and a higher proportion believed 'limiting themselves to one HIV-uninfected sexual partner reduces HIV risk'. Determinants of CD4 cell count at ART initiation in populations initiating ART operate at multiple levels. Structural interventions targeting points upstream from ART initiation along the continuum from infection to diagnosis to care engagement are needed.

Shikonin increases glucose uptake in skeletal muscle cells and improves plasma glucose levels in diabetic Goto-Kakizaki rats.

Directory of Open Access Journals (Sweden)

Anette I Öberg

Full Text Available BACKGROUND: There is considerable interest in identifying compounds that can improve glucose homeostasis. Skeletal muscle, due to its large mass, is the principal organ for glucose disposal in the body and we have investigated here if shikonin, a naphthoquinone derived from the Chinese plant Lithospermum erythrorhizon, increases glucose uptake in skeletal muscle cells. METHODOLOGY/PRINCIPAL FINDINGS: Shikonin increases glucose uptake in L6 skeletal muscle myotubes, but does not phosphorylate Akt, indicating that in skeletal muscle cells its effect is medaited via a pathway distinct from that used for insulin-stimulated uptake. Furthermore we find no evidence for the involvement of AMP-activated protein kinase in shikonin induced glucose uptake. Shikonin increases the intracellular levels of calcium in these cells and this increase is necessary for shikonin-mediated glucose uptake. Furthermore, we found that shikonin stimulated the translocation of GLUT4 from intracellular vesicles to the cell surface in L6 myoblasts. The beneficial effect of shikonin on glucose uptake was investigated in vivo by measuring plasma glucose levels and insulin sensitivity in spontaneously diabetic Goto-Kakizaki rats. Treatment with shikonin (10 mg/kg intraperitoneally once daily for 4 days significantly decreased plasma glucose levels. In an insulin sensitivity test (s.c. injection of 0.5 U/kg insulin, plasma glucose levels were significantly lower in the shikonin-treated rats. In conclusion, shikonin increases glucose uptake in muscle cells via an insulin-independent pathway dependent on calcium. CONCLUSIONS/SIGNIFICANCE: Shikonin increases glucose uptake in skeletal muscle cells via an insulin-independent pathway dependent on calcium. The beneficial effects of shikonin on glucose metabolism, both in vitro and in vivo, show that the compound possesses properties that make it of considerable interest for developing novel treatment of type 2 diabetes.

Geminivirus vectors for high-level expression of foreign proteins in plant cells.

Science.gov (United States)

Mor, Tsafrir S; Moon, Yong-Sun; Palmer, Kenneth E; Mason, Hugh S

2003-02-20

Bean yellow dwarf virus (BeYDV) is a monopartite geminivirus that can infect dicotyledonous plants. We have developed a high-level expression system that utilizes elements of the replication machinery of this single-stranded DNA virus. The replication initiator protein (Rep) mediates release and replication of a replicon from a DNA construct ("LSL vector") that contains an expression cassette for a gene of interest flanked by cis-acting elements of the virus. We used tobacco NT1 cells and biolistic delivery of plasmid DNA for evaluation of replication and expression of reporter genes contained within an LSL vector. By codelivery of a GUS reporter-LSL vector and a Rep-supplying vector, we obtained up to 40-fold increase in expression levels compared to delivery of the reporter-LSL vectors alone. High-copy replication of the LSL vector was correlated with enhanced expression of GUS. Rep expression using a whole BeYDV clone, a cauliflower mosaic virus 35S promoter driving either genomic rep or an intron-deleted rep gene, or 35S-rep contained in the LSL vector all achieved efficient replication and enhancement of GUS expression. We anticipate that this system can be adapted for use in transgenic plants or plant cell cultures with appropriately regulated expression of Rep, with the potential to greatly increase yield of recombinant proteins. Copyright 2003 Wiley Periodicals, Inc. Biotechnol Bioeng 81: 430-437, 2003.

Development of Cell Analysis Software for Cultivated Corneal Endothelial Cells.

Science.gov (United States)

Okumura, Naoki; Ishida, Naoya; Kakutani, Kazuya; Hongo, Akane; Hiwa, Satoru; Hiroyasu, Tomoyuki; Koizumi, Noriko

2017-11-01

To develop analysis software for cultured human corneal endothelial cells (HCECs). Software was designed to recognize cell borders and to provide parameters such as cell density, coefficient of variation, and polygonality of cultured HCECs based on phase contrast images. Cultured HCECs with high or low cell density were incubated with Ca-free and Mg-free phosphate-buffered saline for 10 minutes to reveal the cell borders and were then analyzed with software (n = 50). Phase contrast images showed that cell borders were not distinctly outlined, but these borders became more distinctly outlined after phosphate-buffered saline treatment and were recognized by cell analysis software. The cell density value provided by software was similar to that obtained using manual cell counting by an experienced researcher. Morphometric parameters, such as the coefficient of variation and polygonality, were also produced by software, and these values were significantly correlated with cell density (Pearson correlation coefficients -0.62 and 0.63, respectively). The software described here provides morphometric information from phase contrast images, and it enables subjective and noninvasive quality assessment for tissue engineering therapy of the corneal endothelium.

Strategic Partnerships in Fuel Cell Development

Science.gov (United States)

Diab, Dorey

2006-01-01

This article describes how forming strategic alliances with universities, emerging technology companies, the state of Ohio, the federal government, and the National Science Foundation, has enabled Stark State College to develop a $5.5 million Fuel Cell Prototyping Center and establish a Fuel Cell Technology program to promote economic development…

Notch pathway signaling in the skin antagonizes Merkel cell development.

Science.gov (United States)

Logan, Gregory J; Wright, Margaret C; Kubicki, Adam C; Maricich, Stephen M

2018-02-15

Merkel cells are mechanosensitive skin cells derived from the epidermal lineage whose development requires expression of the basic helix-loop-helix transcription factor Atoh1. The genes and pathways involved in regulating Merkel cell development during embryogenesis are poorly understood. Notch pathway signaling antagonizes Atoh1 expression in many developing body regions, so we hypothesized that Notch signaling might inhibit Merkel cell development. We found that conditional, constitutive overexpression of the Notch intracellular domain (NICD) in mouse epidermis significantly decreased Merkel cell numbers in whisker follicles and touch domes of hairy skin. Conversely, conditional deletion of the obligate NICD binding partner RBPj in the epidermis significantly increased Merkel cell numbers in whisker follicles, led to the development of ectopic Merkel cells outside of touch domes in hairy skin epidermis, and altered the distribution of Merkel cells in touch domes. Deletion of the downstream Notch effector gene Hes1 also significantly increased Merkel cell numbers in whisker follicles. Together, these data demonstrate that Notch signaling regulates Merkel cell production and patterning. Copyright © 2018 Elsevier Inc. All rights reserved.

Low-level red laser therapy alters effects of ultraviolet C radiation on Escherichia coli cells

International Nuclear Information System (INIS)

Canuto, K.S.; Guimaraes, O.R.; Geller, M.; Sergio, L.P.S.; Paoli, F.; Fonseca, A.S.

2015-01-01

Low-level lasers are used at low power densities and doses according to clinical protocols supplied with laser devices or based on professional practice. Although use of these lasers is increasing in many countries, the molecular mechanisms involved in effects of low-level lasers, mainly on DNA, are controversial. In this study, we evaluated the effects of low-level red lasers on survival, filamentation, and morphology of Escherichia coli cells that were exposed to ultraviolet C (UVC) radiation. Exponential and stationary wild-type and uvrA-deficient E. coli cells were exposed to a low-level red laser and in sequence to UVC radiation. Bacterial survival was evaluated to determine the laser protection factor (ratio between the number of viable cells after exposure to the red laser and UVC and the number of viable cells after exposure to UVC). Bacterial filaments were counted to obtain the percentage of filamentation. Area-perimeter ratios were calculated for evaluation of cellular morphology. Experiments were carried out in duplicate and the results are reported as the means of three independent assays. Pre-exposure to a red laser protected wild-type and uvrA-deficient E. coli cells against the lethal effect of UVC radiation, and increased the percentage of filamentation and the area-perimeter ratio, depending on UVC fluence and physiological conditions in the cells. Therapeutic, low-level red laser radiation can induce DNA lesions at a sub-lethal level. Consequences to cells and tissues should be considered when clinical protocols based on this laser are carried out. (author)

Low-level red laser therapy alters effects of ultraviolet C radiation on Escherichia coli cells

Energy Technology Data Exchange (ETDEWEB)

Canuto, K.S.; Guimaraes, O.R.; Geller, M. [Centro Universitario Serra dos Orgaos, Teresopolis, RJ (Brazil). Centro de Ciencias da Saude; Sergio, L.P.S. [Instituto de Biologia Roberto Alcantara Gomes, Rio de Janeiro, RJ (Brazil). Departamento de Biofisica e Biometria; Paoli, F. [Universidade Federal de Juiz de Fora (UFJF), Juiz de Fora, MG (Brazil). Departamento de Morfologia; Fonseca, A.S., E-mail: adnfonseca@ig.com.br [Universidade Federal do Estado do Rio de Janeiro (UFRJ), Rio de Janeiro, RJ (Brazil). Departamento de Ciencias Fisiologicas

2015-10-15

Low-level lasers are used at low power densities and doses according to clinical protocols supplied with laser devices or based on professional practice. Although use of these lasers is increasing in many countries, the molecular mechanisms involved in effects of low-level lasers, mainly on DNA, are controversial. In this study, we evaluated the effects of low-level red lasers on survival, filamentation, and morphology of Escherichia coli cells that were exposed to ultraviolet C (UVC) radiation. Exponential and stationary wild-type and uvrA-deficient E. coli cells were exposed to a low-level red laser and in sequence to UVC radiation. Bacterial survival was evaluated to determine the laser protection factor (ratio between the number of viable cells after exposure to the red laser and UVC and the number of viable cells after exposure to UVC). Bacterial filaments were counted to obtain the percentage of filamentation. Area-perimeter ratios were calculated for evaluation of cellular morphology. Experiments were carried out in duplicate and the results are reported as the means of three independent assays. Pre-exposure to a red laser protected wild-type and uvrA-deficient E. coli cells against the lethal effect of UVC radiation, and increased the percentage of filamentation and the area-perimeter ratio, depending on UVC fluence and physiological conditions in the cells. Therapeutic, low-level red laser radiation can induce DNA lesions at a sub-lethal level. Consequences to cells and tissues should be considered when clinical protocols based on this laser are carried out. (author)

Low Level Laser Therapy Reduces the Development of Lung Inflammation Induced by Formaldehyde Exposure.

Directory of Open Access Journals (Sweden)

Cristiane Miranda da Silva

Full Text Available Lung diseases constitute an important public health problem and its growing level of concern has led to efforts for the development of new therapies, particularly for the control of lung inflammation. Low Level Laser Therapy (LLLT has been highlighted as a non-invasive therapy with few side effects, but its mechanisms need to be better understood and explored. Considering that pollution causes several harmful effects on human health, including lung inflammation, in this study, we have used formaldehyde (FA, an environmental and occupational pollutant, for the induction of neutrophilic lung inflammation. Our objective was to investigate the local and systemic effects of LLLT after FA exposure. Male Wistar rats were exposed to FA (1% or vehicle (distillated water during 3 consecutive days and treated or not with LLLT (1 and 5 hours after each FA exposure. Non-manipulated rats were used as control. 24 h after the last FA exposure, we analyzed the local and systemic effects of LLLT. The treatment with LLLT reduced the development of neutrophilic lung inflammation induced by FA, as observed by the reduced number of leukocytes, mast cells degranulated, and a decreased myeloperoxidase activity in the lung. Moreover, LLLT also reduced the microvascular lung permeability in the parenchyma and the intrapulmonary bronchi. Alterations on the profile of inflammatory cytokines were evidenced by the reduced levels of IL-6 and TNF-α and the elevated levels of IL-10 in the lung. Together, our results showed that LLLT abolishes FA-induced neutrophilic lung inflammation by a reduction of the inflammatory cytokines and mast cell degranulation. This study may provide important information about the mechanisms of LLLT in lung inflammation induced by a pollutant.

High levels of γ-H2AX foci and cell membrane oxidation in adolescents with type 1 diabetes

Energy Technology Data Exchange (ETDEWEB)

Giovannini, Caterina [Unità di Genetica, Dipartimento di Biologia, Pisa University, Pisa (Italy); Piaggi, Simona [Sezione di Patologia Sperimentale, Dipartimento di Ricerca Traslazionale e delle Nuove Tecnologie in Medicina e Chirurgia, Pisa University, Pisa (Italy); Federico, Giovanni [Unità di Endocrinologia Pediatrica e Diabete, Dipartimento di Medicina Clinica e Sperimentale Pisa University, Pisa (Italy); Scarpato, Roberto, E-mail: roberto.scarpato@unipi.it [Unità di Genetica, Dipartimento di Biologia, Pisa University, Pisa (Italy)

2014-12-15

Highlights: • We aimed to detect signs of very early damage in peripheral cells of T1DM adolescents. • T1DM patients had high levels of oxidized cells and reduced expression of iNOS and NO. • Highly mutagenic lesions were markedly increased in the diabetic group, mainly in females. • The observed damage might increase the risk of cancer in the patients later in life. - Abstract: Oxidative stress caused by an excess of free radicals is implicated in the pathogenesis and development of type 1 diabetes mellitus (T1DM) and, in turn, it can lead to genome damage, especially in the form of DNA double-strand break (DSB). The DNA DSB is a potentially carcinogenic lesion for human cells. Thus, we aimed to evaluate whether the level of oxidative stress was increased in peripheral blood lymphocytes of a group of affected adolescents. In 35 T1DM adolescents and 19 healthy controls we assessed: (1) spontaneous and H{sub 2}O{sub 2}-induced oxidation of cell membrane using a fluorescence lipid probe; (2) spontaneous and LPS-induced expression of iNOS protein and indirect NO determination via cytofluorimetric analysis of O{sub 2}{sup −}; (3) immunofluorescent detection of the basal level of histone H2AX phosphorylation (γ-H2AX foci), a well-validated marker of DNA DSB. In T1DM, the frequencies of oxidized cells, both spontaneous and H{sub 2}O{sub 2}-induced (47.13 ± 0.02) were significantly higher than in controls (35.90 ± 0.03). Patients showed, in general, both a reduced iNOS expression and production of NO. Furthermore, the level of spontaneous nuclear damage, quantified as γ-H2AX foci, was markedly increased in T1DM adolescents (6.15 ± 1.08% of γ-H2AX{sup +} cells; 8.72 ± 2.14 γ-H2AXF/n; 9.26 ± 2.37 γ-H2AXF/np), especially in females. In the present study, we confirmed the role that oxidative stress plays in the disease damaging lipids of cell membrane and, most importantly, causing genomic damage in circulating white blood cells of affected adolescents

High levels of γ-H2AX foci and cell membrane oxidation in adolescents with type 1 diabetes

International Nuclear Information System (INIS)

Giovannini, Caterina; Piaggi, Simona; Federico, Giovanni; Scarpato, Roberto

2014-01-01

Highlights: • We aimed to detect signs of very early damage in peripheral cells of T1DM adolescents. • T1DM patients had high levels of oxidized cells and reduced expression of iNOS and NO. • Highly mutagenic lesions were markedly increased in the diabetic group, mainly in females. • The observed damage might increase the risk of cancer in the patients later in life. - Abstract: Oxidative stress caused by an excess of free radicals is implicated in the pathogenesis and development of type 1 diabetes mellitus (T1DM) and, in turn, it can lead to genome damage, especially in the form of DNA double-strand break (DSB). The DNA DSB is a potentially carcinogenic lesion for human cells. Thus, we aimed to evaluate whether the level of oxidative stress was increased in peripheral blood lymphocytes of a group of affected adolescents. In 35 T1DM adolescents and 19 healthy controls we assessed: (1) spontaneous and H 2 O 2 -induced oxidation of cell membrane using a fluorescence lipid probe; (2) spontaneous and LPS-induced expression of iNOS protein and indirect NO determination via cytofluorimetric analysis of O 2 − ; (3) immunofluorescent detection of the basal level of histone H2AX phosphorylation (γ-H2AX foci), a well-validated marker of DNA DSB. In T1DM, the frequencies of oxidized cells, both spontaneous and H 2 O 2 -induced (47.13 ± 0.02) were significantly higher than in controls (35.90 ± 0.03). Patients showed, in general, both a reduced iNOS expression and production of NO. Furthermore, the level of spontaneous nuclear damage, quantified as γ-H2AX foci, was markedly increased in T1DM adolescents (6.15 ± 1.08% of γ-H2AX + cells; 8.72 ± 2.14 γ-H2AXF/n; 9.26 ± 2.37 γ-H2AXF/np), especially in females. In the present study, we confirmed the role that oxidative stress plays in the disease damaging lipids of cell membrane and, most importantly, causing genomic damage in circulating white blood cells of affected adolescents. This also indicates that

Advanced fuel cell development in the United States

International Nuclear Information System (INIS)

Ackerman, J.P.

1984-01-01

Both molten carbonate and solid oxide fuel cells are being developed in the United States to complement and/or supplant phosphoric acid cells for commercial and utility use. This paper described the two technologies and the programs for their development

Mosaic Expression of Thyroid Hormone Regulatory Genes Defines Cell Type-Specific Dependency in the Developing Chicken Cerebellum.

Science.gov (United States)

Delbaere, Joke; Van Herck, Stijn L J; Bourgeois, Nele M A; Vancamp, Pieter; Yang, Shuo; Wingate, Richard J T; Darras, Veerle M

2016-12-01

The cerebellum is a morphologically unique brain structure that requires thyroid hormones (THs) for the correct coordination of key cellular events driving its development. Unravelling the interplay between the multiple factors that can regulate intracellular TH levels is a key step to understanding their role in the regulation of these cellular processes. We therefore investigated the regional/cell-specific expression pattern of TH transporters and deiodinases in the cerebellum using the chicken embryo as a model. In situ hybridisation revealed expression of the TH transporters monocarboxylate transporter 8 (MCT8) and 10 (MCT10), L-type amino acid transporter 1 (LAT1) and organic anion transporting polypeptide 1C1 (OATP1C1) as well as the inactivating type 3 deiodinase (D3) in the fourth ventricle choroid plexus, suggesting a possible contribution of the resulting proteins to TH exchange and subsequent inactivation of excess hormone at the blood-cerebrospinal fluid barrier. Exclusive expression of LAT1 and the activating type 2 deiodinase (D2) mRNA was found at the level of the blood-brain barrier, suggesting a concerted function for LAT1 and D2 in the direct access of active T 3 to the developing cerebellum via the capillary endothelial cells. The presence of MCT8 mRNA in Purkinje cells and cerebellar nuclei during the first 2 weeks of embryonic development points to a potential role of this transporter in the uptake of T 3 in central neurons. At later stages, together with MCT10, detection of MCT8 signal in close association with the Purkinje cell dendritic tree suggests a role of both transporters in TH signalling during Purkinje cell synaptogenesis. MCT10 was also expressed in late-born cells in the rhombic lip lineage with a clear hybridisation signal in the outer external granular layer, indicating a potential role for MCT10 in the proliferation of granule cell precursors. By contrast, expression of D3 in the first-born rhombic lip-derived population may

Development of induction cells at CAEP

International Nuclear Information System (INIS)

Wang Huacen; Zhang Kaizhi; Cheng Nian'an; Zhang Wenwei; Lai Qinggui; Wen Long; Zhang Linwen; Deng Jianjun; Ding Bonan

2002-01-01

The effects to develop induction cells for induction linac and radiography at CAEP are introduced and reviewed in this paper. During the past two decades, several kinds of cells have been designed and tested, and some of them have been used for construction of induction linac, such as Dragon-1 and 12 MeV, and a Synthetic Test Stand (STS) for comprehensive linac technology study. The structure, test results and performance in the induction linac of these cells are given

Development of Sensors and Sensing Technology for Hydrogen Fuel Cell Vehicle Applications

Energy Technology Data Exchange (ETDEWEB)

Brosha, E L; Sekhar, P K; Mukundan, R; Williamson, T; Garzon, F H; Woo, L Y; Glass, R R

2010-01-06

One related area of hydrogen fuel cell vehicle (FCV) development that cannot be overlooked is the anticipated requirement for new sensors for both the monitoring and control of the fuel cell's systems and for those devices that will be required for safety. Present day automobiles have dozens of sensors on-board including those for IC engine management/control, sensors for state-of-health monitoring/control of emissions systems, sensors for control of active safety systems, sensors for triggering passive safety systems, and sensors for more mundane tasks such as fluids level monitoring to name the more obvious. The number of sensors continues to grow every few years as a result of safety mandates but also in response to consumer demands for new conveniences and safety features. Some of these devices (e.g. yaw sensors for dynamic stability control systems or tire presure warning RF-based devices) may be used on fuel cell vehicles without any modification. However the use of hydrogen as a fuel will dictate the development of completely new technologies for such requirements as the detection of hydrogen leaks, sensors and systems to continuously monitor hydrogen fuel purity and protect the fuel cell stack from poisoning, and for the important, yet often taken for granted, tasks such as determining the state of charge of the hydrogen fuel storage and delivery system. Two such sensors that rely on different transduction mechanisms will be highlighted in this presentation. The first is an electrochemical device for monitoring hydrogen levels in air. The other technology covered in this work, is an acoustic-based approach to determine the state of charge of a hydride storage system.

Progress in the Development of Lightweight Nickel Electrode for Nickel-Hydrogen Cell

Science.gov (United States)

Britton, Doris L.

1999-01-01

Development of a high specific energy battery is one of the objectives of the lightweight nickel-hydrogen (Ni-H2) program at the NASA Glenn Research Center. The approach has been to improve the nickel electrode by continuing combined in-house and contract efforts to develop a lighter weight electrode for the nickel-hydrogen cell. Small fiber diameter nickel plaques are used as conductive supports for the nickel hydroxide active material. These plaques are commercial products and have an advantage of increased surface area available for the deposition of active material. Initial tests include activation and capacity measurements at five different discharge levels, C/2, 1.0 C, 1.37 C, 2.0 C, and 2.74 C. The electrodes are life cycle tested using a half-cell configuration at 40 and 80% depths-of-discharge (DOD) in a low-Earth-orbit regime. The electrodes that pass the initial tests are life cycle-tested in a boiler plate nickel-hydrogen cell before flight weight design are built and tested.

Development of New Technologies for Stem Cell Research

Directory of Open Access Journals (Sweden)

Xibo Ma

2012-01-01

Full Text Available Since the 1960s, the stem cells have been extensively studied including embryonic stem cells, neural stem cells, bone marrow hematopoietic stem cells, and mesenchymal stem cells. In the recent years, several stem cells have been initially used in the treatment of diseases, such as in bone marrow transplant. At the same time, isolation and culture experimental technologies for stem cell research have been widely developed in recent years. In addition, molecular imaging technologies including optical molecular imaging, positron emission tomography, single-photon emission computed tomography, and computed tomography have been developed rapidly in recent the 10 years and have also been used in the research on disease mechanism and evaluation of treatment of disease related with stem cells. This paper will focus on recent typical isolation, culture, and observation techniques of stem cells followed by a concise introduction. Finally, the current challenges and the future applications of the new technologies in stem cells are given according to the understanding of the authors, and the paper is then concluded.

Compare analysis for the nanotoxicity effects of different amounts of endocytic iron oxide nanoparticles at single cell level.

Science.gov (United States)

Huang, Chen-Yu; Ger, Tzong-Rong; Wei, Zung-Hang; Lai, Mei-Feng

2014-01-01

Developing methods that evaluate the cellular uptake of magnetic nanoparticles (MNPs) and nanotoxicity effects at single-cellular level are needed. In this study, magnetophoresis combining fluorescence based cytotoxicity assay was proposed to assess the viability and the single-cellular MNPs uptake simultaneously. Malignant cells (SKHep-1, HepG2, HeLa) were incubated with 10 nm anionic iron oxide nanoparticles. Prussian blue stain was performed to visualize the distribution of magnetic nanoparticles. MTT and fluorescence based assay analyzed the cytotoxicity effects of the bulk cell population and single cell, respectively. DAPI/PI stained was applied to evaluate death mechanism. The number of intracellular MNPs was found to be strongly correlated with the cell death. Significant differences between cellular MNP uptake in living and dead cells were observed. The method could be useful for future study of the nanotoxicity induced by MNPs.

EBV promotes human CD8 NKT cell development.

Directory of Open Access Journals (Sweden)

Yuling He

2010-05-01

Full Text Available The reports on the origin of human CD8(+ Valpha24(+ T-cell receptor (TCR natural killer T (NKT cells are controversial. The underlying mechanism that controls human CD4 versus CD8 NKT cell development is not well-characterized. In the present study, we have studied total 177 eligible patients and subjects including 128 healthy latent Epstein-Barr-virus(EBV-infected subjects, 17 newly-onset acute infectious mononucleosis patients, 16 newly-diagnosed EBV-associated Hodgkin lymphoma patients, and 16 EBV-negative normal control subjects. We have established human-thymus/liver-SCID chimera, reaggregated thymic organ culture, and fetal thymic organ culture. We here show that the average frequency of total and CD8(+ NKT cells in PBMCs from 128 healthy latent EBV-infected subjects is significantly higher than in 17 acute EBV infectious mononucleosis patients, 16 EBV-associated Hodgkin lymphoma patients, and 16 EBV-negative normal control subjects. However, the frequency of total and CD8(+ NKT cells is remarkably increased in the acute EBV infectious mononucleosis patients at year 1 post-onset. EBV-challenge promotes CD8(+ NKT cell development in the thymus of human-thymus/liver-SCID chimeras. The frequency of total (3% of thymic cells and CD8(+ NKT cells ( approximately 25% of NKT cells is significantly increased in EBV-challenged chimeras, compared to those in the unchallenged chimeras (<0.01% of thymic cells, CD8(+ NKT cells undetectable, respectively. The EBV-induced increase in thymic NKT cells is also reflected in the periphery, where there is an increase in total and CD8(+ NKT cells in liver and peripheral blood in EBV-challenged chimeras. EBV-induced thymic CD8(+ NKT cells display an activated memory phenotype (CD69(+CD45RO(hiCD161(+CD62L(lo. After EBV-challenge, a proportion of NKT precursors diverges from DP thymocytes, develops and differentiates into mature CD8(+ NKT cells in thymus in EBV-challenged human-thymus/liver-SCID chimeras or

Changes in the expression of Th17 cell-associated cytokines in the development of rheumatic heart disease.

Science.gov (United States)

Wen, Yun; Zeng, Zhiyu; Gui, Chun; Li, Lang; Li, Wenting

2015-01-01

Autoimmunity plays a critical role in the development of rheumatic heart disease (RHD). Recent studies have linked Th17 cells to the autoimmune mechanism associated with RHD. This study aimed to investigate changes in Th17 cell-related cytokine expression in acute and chronic RHD. We established a Lewis rat model of experimental RHD, which was induced by inactivated Group A streptococci and complete Freund's adjuvant. After 7- and 24-week intervention treatments, we measured serum levels of interleukin-17 (IL-17) and IL-6, key cytokines associated with Th17 cells, using a Luminex liquichip method, and levels of IL-17 and IL-6 in heart tissues using immunohistochemical assays. Moreover, expression levels of IL-17, IL-21, IL-6, and IL-23 in mitral valve tissues of human RHD patients were also measured using immunohistochemistry. Compared with the normal control group, serum IL-17 and IL-6 concentrations were significantly increased, and the expression levels of IL-17 and IL-6 in the mitral valve were also significantly increased in 7- or 24-week RHD rats (P<.017). Compared with the control group, expression of IL-17, IL-21, IL-6, and IL-23 in mitral valve tissues was significantly increased in RHD patients (P<.05). Our study suggested that the increased expression of Th17 cell-associated cytokines might play an important role in the pathogenesis and development of RHD. Copyright © 2015 Elsevier Inc. All rights reserved.

Influence and timing of arrival of murine neural crest on pancreatic beta cell development and maturation.

Science.gov (United States)

Plank, Jennifer L; Mundell, Nathan A; Frist, Audrey Y; LeGrone, Alison W; Kim, Thomas; Musser, Melissa A; Walter, Teagan J; Labosky, Patricia A

2011-01-15

the neural crest is a critical regulator of beta cell development on two levels: by negatively regulating beta cell proliferation and by promoting beta cell maturation. Copyright © 2010 Elsevier Inc. All rights reserved.

HIV-1 transgenic rats develop T cell abnormalities

International Nuclear Information System (INIS)

Reid, William; Abdelwahab, Sayed; Sadowska, Mariola; Huso, David; Neal, Ashley; Ahearn, Aaron; Bryant, Joseph; Gallo, Robert C.; Lewis, George K.; Reitz, Marvin

2004-01-01

HIV-1 infection leads to impaired antigen-specific T cell proliferation, increased susceptibility of T cells to apoptosis, progressive impairment of T-helper 1 (Th1) responses, and altered maturation of HIV-1-specific memory cells. We have identified similar impairments in HIV-1 transgenic (Tg) rats. Tg rats developed an absolute reduction in CD4 + and CD8 + T cells able to produce IFN-γ following activation and an increased susceptibility of T cells to activation-induced apoptosis. CD4 + and CD8 + effector/memory (CD45RC - CD62L - ) pools were significantly smaller in Tg rats compared to non-Tg controls, although the converse was true for the naieve (CD45RC + CD62L + ) T cell pool. Our interpretation is that the HIV transgene causes defects in the development of T cell effector function and generation of specific effector/memory T cell subsets, and that activation-induced apoptosis may be an essential factor in this process

The fuel cell; development and possibilities

Energy Technology Data Exchange (ETDEWEB)

Van Rijnsoever, J.W.M.

Activities on fuel cells and fuel cell development in the USA and Japan are surveyed. Possibilities for large scale application are mentioned. Attention is given to efficiency and environmental aspects. There are no problems about hazardous emissions. Besides electric power some heat is generated, which is not always a disadvantage. In many cases both are useful products. (A.V.)

Unraveling Natural Killer T-Cells Development

Directory of Open Access Journals (Sweden)

Sabrina Bianca Bennstein

2018-01-01

Full Text Available Natural killer T-cells are a subset of innate-like T-cells with the ability to bridge innate and adaptive immunity. There is great interest in harnessing these cells to improve tumor therapy; however, greater understanding of invariant NKT (iNKT cell biology is needed. The first step is to learn more about NKT development within the thymus. Recent studies suggest lineage separation of murine iNKT cells into iNKT1, iNKT2, and iNKT17 cells instead of shared developmental stages. This review will focus on these new studies and will discuss the evidence for lineage separation in contrast to shared developmental stages. The author will also highlight the classifications of murine iNKT cells according to identified transcription factors and cytokine production, and will discuss transcriptional and posttranscriptional regulations, and the role of mammalian target of rapamycin. Finally, the importance of these findings for human cancer therapy will be briefly discussed.

Development of endosperm transfer cells in barley.

Science.gov (United States)

Thiel, Johannes

2014-01-01

Endosperm transfer cells (ETCs) are positioned at the intersection of maternal and filial tissues in seeds of cereals and represent a bottleneck for apoplasmic transport of assimilates into the endosperm. Endosperm cellularization starts at the maternal-filial boundary and generates the highly specialized ETCs. During differentiation barley ETCs develop characteristic flange-like wall ingrowths to facilitate effective nutrient transfer. A comprehensive morphological analysis depicted distinct developmental time points in establishment of transfer cell (TC) morphology and revealed intracellular changes possibly associated with cell wall metabolism. Embedded inside the grain, ETCs are barely accessible by manual preparation. To get tissue-specific information about ETC specification and differentiation, laser microdissection (LM)-based methods were used for transcript and metabolite profiling. Transcriptome analysis of ETCs at different developmental stages by microarrays indicated activated gene expression programs related to control of cell proliferation and cell shape, cell wall and carbohydrate metabolism reflecting the morphological changes during early ETC development. Transporter genes reveal distinct expression patterns suggesting a switch from active to passive modes of nutrient uptake with the onset of grain filling. Tissue-specific RNA-seq of the differentiating ETC region from the syncytial stage until functionality in nutrient transfer identified a high number of novel transcripts putatively involved in ETC differentiation. An essential role for two-component signaling (TCS) pathways in ETC development of barley emerged from this analysis. Correlative data provide evidence for abscisic acid and ethylene influences on ETC differentiation and hint at a crosstalk between hormone signal transduction and TCS phosphorelays. Collectively, the data expose a comprehensive view on ETC development, associated pathways and identified candidate genes for ETC

Nuclear receptor TLX regulates cell cycle progression in neural stem cells of the developing brain.

Science.gov (United States)

Li, Wenwu; Sun, Guoqiang; Yang, Su; Qu, Qiuhao; Nakashima, Kinichi; Shi, Yanhong

2008-01-01

TLX is an orphan nuclear receptor that is expressed exclusively in vertebrate forebrains. Although TLX is known to be expressed in embryonic brains, the mechanism by which it influences neural development remains largely unknown. We show here that TLX is expressed specifically in periventricular neural stem cells in embryonic brains. Significant thinning of neocortex was observed in embryonic d 14.5 TLX-null brains with reduced nestin labeling and decreased cell proliferation in the germinal zone. Cell cycle analysis revealed both prolonged cell cycles and increased cell cycle exit in TLX-null embryonic brains. Increased expression of a cyclin-dependent kinase inhibitor p21 and decreased expression of cyclin D1 provide a molecular basis for the deficiency of cell cycle progression in embryonic brains of TLX-null mice. Furthermore, transient knockdown of TLX by in utero electroporation led to precocious cell cycle exit and differentiation of neural stem cells followed by outward migration. Together these results indicate that TLX plays an important role in neural development by regulating cell cycle progression and exit of neural stem cells in the developing brain.

Development of analytical cell support for vitrification at the West Valley Demonstration Project. Topical report

International Nuclear Information System (INIS)

Barber, F.H.; Borek, T.T.; Christopher, J.Z.

1997-12-01

Analytical and Process Chemistry (A ampersand PC) support is essential to the high-level waste vitrification campaign at the West Valley Demonstration Project (WVDP). A ampersand PC characterizes the waste, providing information necessary to formulate the recipe for the target radioactive glass product. High-level waste (HLW) samples are prepared and analyzed in the analytical cells (ACs) and Sample Storage Cell (SSC) on the third floor of the main plant. The high levels of radioactivity in the samples require handling them in the shielded cells with remote manipulators. The analytical hot cells and third floor laboratories were refurbished to ensure optimal uninterrupted operation during the vitrification campaign. New and modified instrumentation, tools, sample preparation and analysis techniques, and equipment and training were required for A ampersand PC to support vitrification. Analytical Cell Mockup Units (ACMUs) were designed to facilitate method development, scientist and technician training, and planning for analytical process flow. The ACMUs were fabricated and installed to simulate the analytical cell environment and dimensions. New techniques, equipment, and tools could be evaluated m in the ACMUs without the consequences of generating or handling radioactive waste. Tools were fabricated, handling and disposal of wastes was addressed, and spatial arrangements for equipment were refined. As a result of the work at the ACMUs the remote preparation and analysis methods and the equipment and tools were ready for installation into the ACs and SSC m in July 1995. Before use m in the hot cells, all remote methods had been validated and four to eight technicians were trained on each. Fine tuning of the procedures has been ongoing at the ACs based on input from A ampersand PC technicians. Working at the ACs presents greater challenges than had development at the ACMUs. The ACMU work and further refinements m in the ACs have resulted m in a reduction m in

Different roles of prepubertal and postpubertal germ cells and Sertoli cells in the regulation of serum inhibin B levels

DEFF Research Database (Denmark)

Andersson, A M; Müller, J; Skakkebaek, N E

1998-01-01

testis, intense immunostaining for the betaB-subunit was evident in germ cells from the pachytene spermatocyte to early spermatid stages and to a lesser degree in Leydig cells, but not in Sertoli cells or other stages of germ cells. Thus, surprisingly, in adult men the two subunits constituting inhibin B......-subunit. The correlation in adult men between serum inhibin B levels and spermatogenesis may be due to the fact that inhibin B in adult men is possibly a joint product of Sertoli cells and germ cells, including the stages from pachytene spermatocytes to early spermatids....

Accident sequence precursor analysis level 2/3 model development

International Nuclear Information System (INIS)

Lui, C.H.; Galyean, W.J.; Brownson, D.A.

1997-01-01

The US Nuclear Regulatory Commission's Accident Sequence Precursor (ASP) program currently uses simple Level 1 models to assess the conditional core damage probability for operational events occurring in commercial nuclear power plants (NPP). Since not all accident sequences leading to core damage will result in the same radiological consequences, it is necessary to develop simple Level 2/3 models that can be used to analyze the response of the NPP containment structure in the context of a core damage accident, estimate the magnitude of the resulting radioactive releases to the environment, and calculate the consequences associated with these releases. The simple Level 2/3 model development work was initiated in 1995, and several prototype models have been completed. Once developed, these simple Level 2/3 models are linked to the simple Level 1 models to provide risk perspectives for operational events. This paper describes the methods implemented for the development of these simple Level 2/3 ASP models, and the linkage process to the existing Level 1 models

Recent developments in the modeling of molten carbonate fuel cells

International Nuclear Information System (INIS)

Wilemski, G.

1984-01-01

Modeling of porous electrodes and overall performance of molten carbonate fuel cells is reviewed. Aspects needing improvement are discussed. Some preliminary results on internal methane reforming cells are presented. Successful modeling of molten carbonate fuel cells has been carried out at two levels. The first concerns the prediction of overall cell performance and performance decay, i.e., the calculation of current-voltage curves and their decay rates for various cell operating conditions. The second involves the determination of individual porous electrode performance, i.e., how the electrode overpotential is affected by pore structure, gas composition, degree of electrolyte fill, etc. Both levels are treated mechanistically, as opposed to empirically, using fundamental mathematical descriptions of the relevant physical and chemical phenomena, in order to provide quantitative predictive capability

A development of solid oxide fuel cell technology

Energy Technology Data Exchange (ETDEWEB)

Lim, Hee Chun; Lee, Chang Woo [Korea Electric Power Corp. (KEPCO), Taejon (Korea, Republic of). Research Center; Kim, Kwy Youl; Yoon, Moon Soo; Kim, Ho Ki; Kim, Young Sik; Mun, Sung In; Eom, Sung Wuk [Korea Electrotechnology Research Inst., Changwon (Korea, Republic of)

1996-12-31

Solid oxide fuel cell which was consisted of ceramics has high power density and is very simple in shape. The project named A development of SOFC(Solid Oxide Fuel Cell) technology is to develop the unit cell fabrication processing and to evaluate the unit cell of solid oxide full cell. In this project, a manufacturing process of cathode by citrate method and polymeric precursor methods were established. By using tape casting method, high density thin electrolyte was manufactured and has high performance. Unit cell composed with La{sub 17}Sr{sub 13}Mn{sub 3} as cathode, 8YSZ electrolyte and 50% NiYSZ anode had a performance of O.85 W/cm{sup 2} and recorded 510 hours operation time. On the basis of these results. 100 cm{sup 2} class unit cell will be fabricated and tests in next program (author). 59 refs., 120 figs.

A development of solid oxide fuel cell technology

Energy Technology Data Exchange (ETDEWEB)

Lim, Hee Chun; Lee, Chang Woo [Korea Electric Power Corp. (KEPCO), Taejon (Korea, Republic of). Research Center; Kim, Kwy Youl; Yoon, Moon Soo; Kim, Ho Ki; Kim, Young Sik; Mun, Sung In; Eom, Sung Wuk [Korea Electrotechnology Research Inst., Changwon (Korea, Republic of)

1995-12-31

Solid oxide fuel cell which was consisted of ceramics has high power density and is very simple in shape. The project named A development of SOFC(Solid Oxide Fuel Cell) technology is to develop the unit cell fabrication processing and to evaluate the unit cell of solid oxide full cell. In this project, a manufacturing process of cathode by citrate method and polymeric precursor methods were established. By using tape casting method, high density thin electrolyte was manufactured and has high performance. Unit cell composed with La{sub 17}Sr{sub 13}Mn{sub 3} as cathode, 8YSZ electrolyte and 50% NiYSZ anode had a performance of O.85 W/cm{sup 2} and recorded 510 hours operation time. On the basis of these results. 100 cm{sup 2} class unit cell will be fabricated and tests in next program (author). 59 refs., 120 figs.

Excessive amounts of mu heavy chain block B-cell development.

Science.gov (United States)

Zhu, Lingqiao; Chang, Cheong-Hee; Dunnick, Wesley

2011-09-01

Antigen-independent B-cell development occurs in several stages that depend on the expression of Ig heavy and light chain. We identified a line of mice that lacked mature B cells in the spleen. This mouse line carried approximately 11 copies of a transgene of the murine heavy chain constant region locus, and B-lineage cells expressed excessive amounts of the intracellular μ heavy chain. B-cell development failed in the bone marrow at the pro/pre B-cell transition, and examination of other lines with various copy numbers of the same transgene suggested that deficiencies in B-cell development increased with increased transgene copy number. Expression of a transgenic (Tg) light chain along with the Tg μ heavy chain led to minimal rescue of B-cell development in the bone marrow and B cells in the spleen. There are several potential mechanisms for the death of pro/pre B cells as a consequence of excess heavy chain expression.

[Th17 and Treg cell levels in patients with sarcoidosis and their relation to disease activation].

Science.gov (United States)

Weng, Yue-song; Wang, Hua-ying; Lv, Ding-feng; Fu, Zhong-ming; Yu, Wan-jun

2015-03-01

To investigate the Th17 cell and Treg cell levels in patients with sarcoidosis, and their relation to disease activation and glucocorticoids treatment. Twenty-three sarcoidosis patients admitted in Yinzhou People's Hospital from January 2009 to December 2013 and 25 healthy subjects (controls) were included in this study. The blood samples and bronchoalveolar lavage fluid (BALF) samples were collected in all patients before and after glucocorticoids treatment. The serum angiotensin converting enzyme (SACE) levels were detected. The percentages of Th17 cells and Treg cells in peripheral blood and BALF were determined by flow cytometry, the concentrations of cytokines in serum and supernatants of BALF were measured by enzyme-linked immunosorbent assay (ELISA). The levels of ROR-γt and Foxp3 mRNA transcripts in peripheral blood mononuclear cells (PBMC) were determined by real-time quantitative PCR. The potential correlation between the percentages of Th17 or Treg cells and SACE levels was evaluated. Compared with healthy controls, significantly higher frequencies of Th17 cells (4.34%±0.89% vs 1.60% ± 0.42%), lower frequencies of Treg cells (1.28% ± 0.37% vs 3.39% ± 0.50%) in peripheral blood were observed. Higher level of ROR-γt mRNA (21.31 ± 3.55 vs 3.63 ± 1.00) and lower level of Foxp3 mRNA (1.60 ± 0.24 vs 3.12 ± 0.76) in peripheral blood were detected in sarcoidosis patients in active stage (before glucocorticoids treatment) (all PSACE (r= 0.781). The imbalance of Th17 cells and Treg cells in peripheral blood and airway may be involved in the pathogenesis of sarcoidosis, which was associated with the activity of disease, and the treatment of glucocorticoids may achieve a therapeutic effect by correcting the immune imbalance.

MG132, a proteasome inhibitor, induces human pulmonary fibroblast cell death via increasing ROS levels and GSH depletion.

Science.gov (United States)

Park, Woo Hyun; Kim, Suhn Hee

2012-04-01

MG132 as a proteasome inhibitor can induce apoptotic cell death in lung cancer cells. However, little is known about the toxicological cellular effects of MG132 on normal primary lung cells. Here, we investigated the effects of N-acetyl cysteine (NAC) and vitamin C (well known antioxidants) or L-buthionine sulfoximine (BSO; an inhibitor of GSH synthesis) on MG132-treated human pulmonary fibroblast (HPF) cells in relation to cell death, reactive oxygen species (ROS) and glutathione (GSH). MG132 induced growth inhibition and death in HPF cells, accompanied by the loss of mitochondrial membrane potential (MMP; ∆ψm). MG132 increased ROS levels and GSH-depleted cell numbers in HPF cells. Both antioxidants, NAC and vitamin C, prevented growth inhibition, death and MMP (∆ψm) loss in MG132-treated HPF cells and also attenuated ROS levels in these cells. BSO showed a strong increase in ROS levels in MG132-treated HPF cells and slightly enhanced the growth inhibition, cell death, MMP (∆ψm) loss and GSH depletion. In addition, NAC decreased anonymous ubiquitinated protein levels in MG132-treated HPF cells. Furthermore, superoxide dismutase (SOD) 2, catalase (CTX) and GSH peroxidase (GPX) siRNAs enhanced HPF cell death by MG132, which was not correlated with ROS and GSH level changes. In conclusion, MG132 induced the growth inhibition and death of HPF cells, which were accompanied by increasing ROS levels and GSH depletion. Both NAC and vitamin C attenuated HPF cell death by MG132, whereas BSO slightly enhanced the death.

Human Lung Mast Cell Products Regulate Airway Smooth Muscle CXCL10 Levels.

Science.gov (United States)

Alkhouri, H; Cha, V; Tong, K; Moir, L M; Armour, C L; Hughes, J M

2014-01-01

In asthma, the airway smooth muscle (ASM) produces CXCL10 which may attract CXCR3(+) mast/T cells to it. Our aim was to investigate the effects of mast cell products on ASM cell CXCL10 production. ASM cells from people with and without asthma were stimulated with IL-1 β , TNF- α , and/or IFN γ and treated with histamine (1-100  μ M) ± chlorpheniramine (H1R antagonist; 1  μ M) or ranitidine (H2R antagonist; 50  μ M) or tryptase (1 nM) ± leupeptin (serine protease inhibitor; 50  μ M), heat-inactivated tryptase, or vehicle for 4 h or 24 h. Human lung mast cells (MC) were isolated and activated with IgE/anti-IgE and supernatants were collected after 2 h or 24 h. The supernatants were added to ASM cells for 48 h and ASM cell CXCL10 production detected using ELISA (protein) and real-time PCR (mRNA). Histamine reduced IL-1 β /TNF- α -induced CXCL10 protein, but not mRNA, levels independent of H1 and H2 receptor activation, whereas tryptase and MC 2 h supernatants reduced all cytokine-induced CXCL10. Tryptase also reduced CXCL10 levels in a cell-free system. Leupeptin inhibited the effects of tryptase and MC 2 h supernatants. MC 24 h supernatants contained TNF- α and amplified IFN γ -induced ASM cell CXCL10 production. This is the first evidence that MC can regulate ASM cell CXCL10 production and its degradation. Thus MC may regulate airway myositis in asthma.

DEVELOPMENT OF LOW-COST MANUFACTURING PROCESSES FOR PLANAR, MULTILAYER SOLID OXIDE FUEL CELL ELEMENTS

Energy Technology Data Exchange (ETDEWEB)

Scott Swartz; Matthew Seabaugh; William Dawson; Harlan Anderson; Tim Armstrong; Michael Cobb; Kirby Meacham; James Stephan; Russell Bennett; Bob Remick; Chuck Sishtla; Scott Barnett; John Lannutti

2004-06-12

This report summarizes the results of a four-year project, entitled, ''Low-Cost Manufacturing Of Multilayer Ceramic Fuel Cells'', jointly funded by the U.S. Department of Energy, the State of Ohio, and by project participants. The project was led by NexTech Materials, Ltd., with subcontracting support provided by University of Missouri-Rolla, Michael A. Cobb & Co., Advanced Materials Technologies, Inc., Edison Materials Technology Center, Gas Technology Institute, Northwestern University, and The Ohio State University. Oak Ridge National Laboratory, though not formally a subcontractor on the program, supported the effort with separate DOE funding. The objective of the program was to develop advanced manufacturing technologies for making solid oxide fuel cell components that are more economical and reliable for a variety of applications. The program was carried out in three phases. In the Phase I effort, several manufacturing approaches were considered and subjected to detailed assessments of manufacturability and development risk. Estimated manufacturing costs for 5-kW stacks were in the range of $139/kW to $179/kW. The risk assessment identified a number of technical issues that would need to be considered during development. Phase II development work focused on development of planar solid oxide fuel cell elements, using a number of ceramic manufacturing methods, including tape casting, colloidal-spray deposition, screen printing, spin-coating, and sintering. Several processes were successfully established for fabrication of anode-supported, thin-film electrolyte cells, with performance levels at or near the state-of-the-art. The work in Phase III involved scale-up of cell manufacturing methods, development of non-destructive evaluation methods, and comprehensive electrical and electrochemical testing of solid oxide fuel cell materials and components.

Advanced PEFC development for fuel cell powered vehicles

Science.gov (United States)

Kawatsu, Shigeyuki

Vehicles equipped with fuel cells have been developed with much progress. Outcomes of such development efforts include a Toyota fuel cell electric vehicle (FCEV) using hydrogen as the fuel which was developed and introduced in 1996, followed by another Toyota FCEV using methanol as the fuel, developed and introduced in 1997. In those Toyota FCEVs, a fuel cell system is installed under the floor of each RAV4L, to sports utility vehicle. It has been found that the CO concentration in the reformed gas of methanol reformer can be reduced to 100 ppm in wide ranges of catalyst temperature and gas flow rate, by using the ruthenium (Ru) catalyst as the CO selective oxidizer, instead of the platinum (Pt) catalyst known from some time ago. It has been also found that a fuel cell performance equivalent to that with pure hydrogen can be ensured even in the reformed gas with the carbon monoxide (CO) concentration of 100 ppm, by using the Pt-Ru (platinum ruthenium alloy) electrocatalyst as the anode electrocatalyst of a polymer electrolyte fuel cell (PEFC), instead of the Pt electrocatalyst known from some time ago.

Reduced expression levels of PTEN are associated with decreased sensitivity of HCC827 cells to icotinib.

Science.gov (United States)

Zhai, Yang; Zhang, Yanjun; Nan, Kejun; Liang, Xuan

2017-05-01

The clinical resistance of non-small cell lung cancer (NSCLC) to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) has been linked to EGFR T790M resistance mutations or MET amplifications. Additional mechanisms underlying EGFR-TKI drug resistance remain unclear. The present study demonstrated that icotinib significantly inhibited the proliferation and increased the apoptosis rate of HCC827 cells; the cellular mRNA and protein expression levels of phosphatase and tensin homolog (PTEN) were also significantly downregulated. To investigate the effect of PTEN expression levels on the sensitivity of HCC827 cells to icotinib, PTEN expression was silenced using a PTEN-specific small interfering RNA. The current study identified that the downregulation of PTEN expression levels may promote cellular proliferation in addition to decreasing the apoptosis of HCC827 cells, and may reduce the sensitivity of HCC827 cells to icotinib. These results suggested that reduced PTEN expression levels were associated with the decreased sensitivity of HCC827 cells to icotinib. Furthermore, PTEN expression levels may be a useful marker for predicting icotinib resistance and elucidating the resistance mechanisms underlying EGFR-mutated NSCLC.

Eimeria tenella: in vitro development in irradiated bovine kidney cells

Energy Technology Data Exchange (ETDEWEB)

Crane, M.St.J.; Schmatz, D.M.; Stevens, S.; Habbersett, M.C.; Murray, P.K. (Merck Sharp and Dohme Research Labs., Rahway, NJ (USA))

1984-06-01

The initial infection and first-generation development of Eimeria tenella was quantified using a cloned MDBK (Madin-Darby Bovine Kidney) cell line, irradiated with gamma radiation prior to infection, as the host cell. Irradiated cell cultures were found to be more susceptible to infection and had a greater capacity to support parasite development than non-irradiated cultures. It was suggested that the larger proportion of cells in the G/sub 2/ phase of the cell cycle, the larger individual cell size and the inhibition of cell division in the irradiated cultures were all factors contributing to the increased susceptibility to infection and capacity to support parasite growth and development. The application of this technique (host cell irradiation) to the cultivation of other intracellular, protozoan parasites is discussed.

Eimeria tenella: in vitro development in irradiated bovine kidney cells

International Nuclear Information System (INIS)

Crane, M. St.J.; Schmatz, D.M.; Stevens, S.; Habbersett, M.C.; Murray, P.K.

1984-01-01

The initial infection and first-generation development of Eimeria tenella was quantified using a cloned MDBK (Madin-Darby Bovine Kidney) cell line, irradiated with gamma radiation prior to infection, as the host cell. Irradiated cell cultures were found to be more susceptible to infection and had a greater capacity to support parasite development than non-irradiated cultures. It was suggested that the larger proportion of cells in the G 2 phase of the cell cycle, the larger individual cell size and the inhibition of cell division in the irradiated cultures were all factors contributing to the increased susceptibility to infection and capacity to support parasite growth and development. The application of this technique (host cell irradiation) to the cultivation of other intracellular, protozoan parasites is discussed. (author)

Altered development of NKT cells, γδ T cells, CD8 T cells and NK cells in a PLZF deficient patient.

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Maggie Eidson

Full Text Available In mice, the transcription factor, PLZF, controls the development of effector functions in invariant NKT cells and a subset of NKT cell-like, γδ T cells. Here, we show that in human lymphocytes, in addition to invariant NKT cells, PLZF was also expressed in a large percentage of CD8+ and CD4+ T cells. Furthermore, PLZF was also found to be expressed in all γδ T cells and in all NK cells. Importantly, we show that in a donor lacking functional PLZF, all of these various lymphocyte populations were altered. Therefore, in contrast to mice, PLZF appears to control the development and/or function of a wide variety of human lymphocytes that represent more than 10% of the total PBMCs. Interestingly, the PLZF-expressing CD8+ T cell population was found to be expanded in the peripheral blood of patients with metastatic melanoma but was greatly diminished in patients with autoimmune disease.

Synergism between arsenite and proteasome inhibitor MG132 over cell death in myeloid leukaemic cells U937 and the induction of low levels of intracellular superoxide anion

International Nuclear Information System (INIS)

Lombardo, Tomás; Cavaliere, Victoria; Costantino, Susana N.; Kornblihtt, Laura; Alvarez, Elida M.; Blanco, Guillermo A.

2012-01-01

Increased oxygen species production has often been cited as a mechanism determining synergism on cell death and growth inhibition effects of arsenic-combined drugs. However the net effect of drug combination may not be easily anticipated solely from available knowledge of drug-induced death mechanisms. We evaluated the combined effect of sodium arsenite with the proteasome inhibitor MG132, and the anti-leukaemic agent CAPE, on growth-inhibition and cell death effect in acute myeloid leukaemic cells U937 and Burkitt's lymphoma-derived Raji cells, by the Chou–Talalay method. In addition we explored the association of cytotoxic effect of drugs with changes in intracellular superoxide anion (O 2 − ) levels. Our results showed that combined arsenite + MG132 produced low levels of O 2 − at 6 h and 24 h after exposure and were synergic on cell death induction in U937 cells over the whole dose range, although the combination was antagonistic on growth inhibition effect. Exposure to a constant non-cytotoxic dose of 80 μM hydrogen peroxide together with arsenite + MG132 changed synergism on cell death to antagonism at all effect levels while increasing O 2 − levels. Arsenite + hydrogen peroxide also resulted in antagonism with increased O 2 − levels in U937 cells. In Raji cells, arsenite + MG132 also produced low levels of O 2 − at 6 h and 24 h but resulted in antagonism on cell death and growth inhibition. By contrast, the combination arsenite + CAPE showed high levels of O 2 − production at 6 h and 24 h post exposure but resulted in antagonism over cell death and growth inhibition effects in U937 and Raji cells. We conclude that synergism between arsenite and MG132 in U937 cells is negatively associated to O 2 − levels at early time points after exposure. -- Highlights: ► Arsenic combined cytotoxic and anti-proliferative effects by Chou–Talalay method. ► Cytotoxic effect associated with superoxide levels as assessed by flow cytometry. ► Synergism

Intraportal injection of insulin-producing cells generated from human bone marrow mesenchymal stem cells decreases blood glucose level in diabetic rats.

Science.gov (United States)

Tsai, Pei-Jiun; Wang, Hwai-Shi; Lin, Chi-Hung; Weng, Zen-Chung; Chen, Tien-Hua; Shyu, Jia-Fwu

2014-01-01

We studied the process of trans-differentiation of human bone marrow mesenchymal stem cells (hBM-MSCs) into insulin-producing cells. Streptozotocin (STZ)-induced diabetic rat model was used to study the effect of portal vein transplantation of these insulin-producing cells on blood sugar levels. The BM-MSCs were differentiated into insulin-producing cells under defined conditions. Real-time PCR, immunocytochemistry and glucose challenge were used to evaluate in vitro differentiation. Flow cytometry showed that hBM-MSCs were strongly positive for CD44, CD105 and CD73 and negative for hematopoietic markers CD34, CD38 and CD45. Differentiated cells expressed C-peptide as well as β-cells specific genes and hormones. Glucose stimulation increased C-peptide secretion in these cells. The insulin-producing, differentiated cells were transplanted into the portal vein of STZ-induced diabetic rats using a Port-A catheter. The insulin-producing cells were localized in the liver of the recipient rat and expressed human C-peptide. Blood glucose levels were reduced in diabetic rats transplanted with insulin-producing cells. We concluded that hBM-MSCs could be trans-differentiated into insulin-producing cells in vitro. Portal vein transplantation of insulin-producing cells alleviated hyperglycemia in diabetic rats.

Angiotensin II potentiates prostaglandin stimulation of cyclic AMP levels in intact bovine adrenal medulla cells but not adenylate cyclase in permeabilized cells.

Science.gov (United States)

Boarder, M R; Plevin, R; Marriott, D B

1988-10-25

The level of cyclic AMP in primary cultures of bovine adrenal medulla cells is elevated by prostaglandin E1. Angiotensin II is commonly reported to act on receptors linked to phosphoinositide metabolism or to inhibition of adenylate cyclase. We have investigated the effect of angiotensin II on prostaglandin E1-stimulated cyclic AMP levels in these primary cultures. Rather than reducing cyclic AMP levels, we have found that angiotensin II powerfully potentiates prostaglandin E1-stimulated cyclic AMP accumulation in intact cells, both in the presence and absence of phosphodiesterase inhibitors. The 50% maximal response was similar to that for stimulation of phosphoinositide breakdown by angiotensin II in these cultures. The potentiation of stimulated cyclic AMP levels was seen, although to a smaller maximum, with the protein kinase C (Ca2+/phospholipid-dependent enzyme) activating phorbol ester tetradecanoyl phorbolacetate and with the synthetic diacylglycerol 1-oleoyl-2-acetylglycerol; pretreatment (24 h) with active phorbol ester, which would be expected to diminish protein kinase C levels, attenuated the angiotensin II potentiation of cyclic AMP. Using digitonin-permeabilized cells we showed that adenylate cyclase activity was stimulated by prostaglandin E1 with the same dose-response relationship as was cyclic AMP accumulation in intact cells, but the permeabilized cells showed no response to angiotensin II. The results are discussed with respect to the hypothesis that the angiotensin II influence on cyclic AMP levels is mediated, in part, by diacylglycerol stimulation of protein kinase C.

Correlation between residual level of DNA double-strand breaks and the radiosensitivity of cancer cells

International Nuclear Information System (INIS)

Sun Jianxiang; Sun Weijian; Sui Jianli; Zhou Pingkun

2008-01-01

Objective: To understand the variation of the DNA double-strand break rejoining capacity among different cultured cancer cell lines and the primary cancer cells from brain cancer patients, and to explore the predictor of radiotherapy responses of cancers. Methods: DNA double-strand breaks (DSBs) were induced by 60 Co γ-irradiation. Pulsed-field gel electrophoresis was used to analyze the initial production and rejoining of DNA DSBs. Radiosensitivity was determined by in vitro assay of clonogenic-forming capacity. Results: A wide variation of radiosensitivity, e.g. the survival parameter of Do varied from 0.65 to 2.15 Gy, was displayed among the eight cell lines derived from different type of cancers. Although differential level of initial DNA DSBs induced by 20 Gy γ-rays was observed among various cell lines, it was not correlated with the radiosensitivity. The deficiency of DNA DSB rejoining in radiosensitive cell lines was shown either in the early rapid-rejoining phase (SX-10 cells) or in the late slow-rejoining phase (A2780 cells). A significant relationship was observed between the residual level of DNA DSBs measured at 2 h post-20 Gy irradiation and the cellular radiosensitivity (D 0 or SF 2 ). The kinetic curves of rejoining DNA DSBs in the primary human brain tumor cells indicated a variation on DSB rejoining capacity among different individual tumor. The residual level of DNA DSBs after 2 h of rejoining post 20 Gy irradiation in primary human brain tumor cells is compatible to the results obtained in vitro culture cancer cell lines. Conclusions: The residual level of DNA DSBs is correlated with radioresistance of cancer cells, and the residual DNA damage is a useful parameter in predicting the response of tumor tissue to radiotherapy. (authors)

Development of maintenance equipment for nuclear material fabrication equipment in a highly active hot cell

International Nuclear Information System (INIS)

Park, J. J.; Yang, M. S.; Kim, K. H. and others

2000-09-01

This report presents the development of a maintenance system for a highly contaminated nuclear material handling equipment at a hot-cell. This maintenance system has mainly three subsystems - a gamma-radiation measurement module for detecting a gamma-radiation level and identifying its distribution in-situ, a dry-type decontamination device for cleaning up contaminated particles, and a maintenance chamber for isolating contaminated equipment. The mechanical design considerations, controller, capabilities and remote operation and manipulation of the maintenance system are described. Such subsystems developed were installed and tested in the IMEF (Irradiated Material Examination Facility) M6 hot-cell after mock-up tests and performed their specific tasks successfully

Development of maintenance equipment for nuclear material fabrication equipment in a highly active hot cell

Energy Technology Data Exchange (ETDEWEB)

Park, J. J.; Yang, M. S.; Kim, K. H. and others

2000-09-01

This report presents the development of a maintenance system for a highly contaminated nuclear material handling equipment at a hot-cell. This maintenance system has mainly three subsystems - a gamma-radiation measurement module for detecting a gamma-radiation level and identifying its distribution in-situ, a dry-type decontamination device for cleaning up contaminated particles, and a maintenance chamber for isolating contaminated equipment. The mechanical design considerations, controller, capabilities and remote operation and manipulation of the maintenance system are described. Such subsystems developed were installed and tested in the IMEF (Irradiated Material Examination Facility) M6 hot-cell after mock-up tests and performed their specific tasks successfully.

Spatially monitoring oxygen level in 3D microfabricated cell culture systems using optical oxygen sensing beads.

Science.gov (United States)

Wang, Lin; Acosta, Miguel A; Leach, Jennie B; Carrier, Rebecca L

2013-04-21

Capability of measuring and monitoring local oxygen concentration at the single cell level (tens of microns scale) is often desirable but difficult to achieve in cell culture. In this study, biocompatible oxygen sensing beads were prepared and tested for their potential for real-time monitoring and mapping of local oxygen concentration in 3D micro-patterned cell culture systems. Each oxygen sensing bead is composed of a silica core loaded with both an oxygen sensitive Ru(Ph2phen3)Cl2 dye and oxygen insensitive Nile blue reference dye, and a poly-dimethylsiloxane (PDMS) shell rendering biocompatibility. Human intestinal epithelial Caco-2 cells were cultivated on a series of PDMS and type I collagen based substrates patterned with micro-well arrays for 3 or 7 days, and then brought into contact with oxygen sensing beads. Using an image analysis algorithm to convert florescence intensity of beads to partial oxygen pressure in the culture system, tens of microns-size oxygen sensing beads enabled the spatial measurement of local oxygen concentration in the microfabricated system. Results generally indicated lower oxygen level inside wells than on top of wells, and local oxygen level dependence on structural features of cell culture surfaces. Interestingly, chemical composition of cell culture substrates also appeared to affect oxygen level, with type-I collagen based cell culture systems having lower oxygen concentration compared to PDMS based cell culture systems. In general, results suggest that oxygen sensing beads can be utilized to achieve real-time and local monitoring of micro-environment oxygen level in 3D microfabricated cell culture systems.

Development of Inorganic Solar Cells by Nanotechnology

Institute of Scientific and Technical Information of China (English)

Yafei Zhang; Huijuan Geng; Zhihua Zhou; Jiang Wu; Zhiming Wang; Yaozhong Zhang; Zhongli Li; Liying Zhang; Zhi Yang; Huey Liang Hwang

2012-01-01

Inorganic solar cells, as durable photovoltaic devices for harvesting electric energy from sun light,have received tremendous attention due to the fear of exhausting the earth’s energy resources and damaging the living environment due to greenhouse gases. Some recent developments in nanotechnology have opened up new avenues for more relevant inorganic solar cells produced by new photovoltaic conversion concepts and effective solar energy harvesting nanostructures. In this review, the multiple exciton generation effect solar cells, hot carrier solar cells, one dimensional material constructed asymmetrical schottky barrier arrays, noble nanoparticle induced plasmonic enhancement, and light trapping nanostructured semiconductor solar cells are highlighted.

Fluorescent reporter signals, EGFP and DsRed, encoded in HIV-1 facilitate the detection of productively infected cells and cell-associated viral replication levels

Directory of Open Access Journals (Sweden)

Kazutaka eTerahara

2012-01-01

Full Text Available Flow cytometric analysis is a reliable and convenient method for investigating molecules at the single cell level. Previously, recombinant human immunodeficiency virus type 1 (HIV-1 strains were constructed that express a fluorescent reporter, either enhanced green fluorescent protein or DsRed, which allow the monitoring of HIV-1-infected cells by flow cytometry. The present study further investigated the potential of these recombinant viruses in terms of whether the HIV-1 fluorescent reporters would be helpful in evaluating viral replication based on fluorescence intensity. When primary CD4+ T cells were infected with recombinant viruses, the fluorescent reporter intensity measured by flow cytometry was associated with the level of CD4 downmodulation and Gag p24 expression in infected cells. Interestingly, some HIV-1-infected cells, in which CD4 was only moderately downmodulated, were reporter-positive but Gag p24-negative. Furthermore, when the activation status of primary CD4+ T cells was modulated by T cell receptor-mediated stimulation, we confirmed the preferential viral production upon strong stimulation and showed that the intensity of the fluorescent reporter within a proportion of HIV-1-infected cells was correlated with the viral replication level. These findings indicate that a fluorescent reporter encoded within HIV-1 is useful for the sensitive detection of productively-infected cells at different stages of infection and for evaluating cell-associated viral replication at the single cell level.

High serotonin levels during brain development alter the structural input-output connectivity of neural networks in the rat somatosensory layer IV

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Stéphanie eMiceli

2013-06-01

Full Text Available Homeostatic regulation of serotonin (5-HT concentration is critical for normal topographical organization and development of thalamocortical (TC afferent circuits. Down-regulation of the serotonin transporter (SERT and the consequent impaired reuptake of 5-HT at the synapse, results in a reduced terminal branching of developing TC afferents within the primary somatosensory cortex (S1. Despite the presence of multiple genetic models, the effect of high extracellular 5-HT levels on the structure and function of developing intracortical neural networks is far from being understood. Here, using juvenile SERT knockout (SERT-/- rats we investigated, in vitro, the effect of increased 5-HT levels on the structural organization of (i the thalamocortical projections of the ventroposteromedial thalamic nucleus towards S1, (ii the general barrel-field pattern and (iii the electrophysiological and morphological properties of the excitatory cell population in layer IV of S1 (spiny stellate and pyramidal cells. Our results confirmed previous findings that high levels of 5-HT during development lead to a reduction of the topographical precision of TCA projections towards the barrel cortex. Also, the barrel pattern was altered but not abolished in SERT-/- rats. In layer IV, both excitatory spiny stellate and pyramidal cells showed a significantly reduced intracolumnar organization of their axonal projections. In addition, the layer IV spiny stellate cells gave rise to a prominent projection towards the infragranular layer Vb. Our findings point to a structural and functional reorganization, of TCAs, as well as early stage intracortical microcircuitry, following the disruption of 5-HT reuptake during critical developmental periods. The increased projection pattern of the layer IV neurons suggests that the intracortical network changes are not limited to the main entry layer IV but may also affect the subsequent stages of the canonical circuits of the barrel

Kluyveromyces marxianus and Saccharomyces boulardii Induce Distinct Levels of Dendritic Cell Cytokine Secretion and Significantly Different T Cell Responses In Vitro.

Directory of Open Access Journals (Sweden)

Ida M Smith

Full Text Available Interactions between members of the intestinal microbiota and the mucosal immune system can significantly impact human health, and in this context, fungi and food-related yeasts are known to influence intestinal inflammation through direct interactions with specialized immune cells in vivo. The aim of the present study was to characterize the immune modulating properties of the food-related yeast Kluyveromyces marxianus in terms of adaptive immune responses indicating inflammation versus tolerance and to explore the mechanisms behind the observed responses. Benchmarking against a Saccharomyces boulardii strain with probiotic effects documented in clinical trials, we evaluated the ability of K. marxianus to modulate human dendritic cell (DC function in vitro. Further, we assessed yeast induced DC modulation of naive T cells toward effector responses dominated by secretion of IFNγ and IL-17 versus induction of a Treg response characterized by robust IL-10 secretion. In addition, we blocked relevant DC surface receptors and investigated the stimulating properties of β-glucan containing yeast cell wall extracts. K. marxianus and S. boulardii induced distinct levels of DC cytokine secretion, primarily driven by Dectin-1 recognition of β-glucan components in their cell walls. Upon co-incubation of yeast exposed DCs and naive T cells, S. boulardii induced a potent IFNγ response indicating TH1 mobilization. In contrast, K. marxianus induced a response dominated by Foxp3+ Treg cells, a characteristic that may benefit human health in conditions characterized by excessive inflammation and positions K. marxianus as a strong candidate for further development as a novel yeast probiotic.

Kluyveromyces marxianus and Saccharomyces boulardii Induce Distinct Levels of Dendritic Cell Cytokine Secretion and Significantly Different T Cell Responses In Vitro.

Science.gov (United States)

Smith, Ida M; Baker, Adam; Christensen, Jeffrey E; Boekhout, Teun; Frøkiær, Hanne; Arneborg, Nils; Jespersen, Lene

2016-01-01

Interactions between members of the intestinal microbiota and the mucosal immune system can significantly impact human health, and in this context, fungi and food-related yeasts are known to influence intestinal inflammation through direct interactions with specialized immune cells in vivo. The aim of the present study was to characterize the immune modulating properties of the food-related yeast Kluyveromyces marxianus in terms of adaptive immune responses indicating inflammation versus tolerance and to explore the mechanisms behind the observed responses. Benchmarking against a Saccharomyces boulardii strain with probiotic effects documented in clinical trials, we evaluated the ability of K. marxianus to modulate human dendritic cell (DC) function in vitro. Further, we assessed yeast induced DC modulation of naive T cells toward effector responses dominated by secretion of IFNγ and IL-17 versus induction of a Treg response characterized by robust IL-10 secretion. In addition, we blocked relevant DC surface receptors and investigated the stimulating properties of β-glucan containing yeast cell wall extracts. K. marxianus and S. boulardii induced distinct levels of DC cytokine secretion, primarily driven by Dectin-1 recognition of β-glucan components in their cell walls. Upon co-incubation of yeast exposed DCs and naive T cells, S. boulardii induced a potent IFNγ response indicating TH1 mobilization. In contrast, K. marxianus induced a response dominated by Foxp3+ Treg cells, a characteristic that may benefit human health in conditions characterized by excessive inflammation and positions K. marxianus as a strong candidate for further development as a novel yeast probiotic.

Cell Death in C. elegans Development.

Science.gov (United States)

Malin, Jennifer Zuckerman; Shaham, Shai

2015-01-01

Cell death is a common and important feature of animal development, and cell death defects underlie many human disease states. The nematode Caenorhabditis elegans has proven fertile ground for uncovering molecular and cellular processes controlling programmed cell death. A core pathway consisting of the conserved proteins EGL-1/BH3-only, CED-9/BCL2, CED-4/APAF1, and CED-3/caspase promotes most cell death in the nematode, and a conserved set of proteins ensures the engulfment and degradation of dying cells. Multiple regulatory pathways control cell death onset in C. elegans, and many reveal similarities with tumor formation pathways in mammals, supporting the idea that cell death plays key roles in malignant progression. Nonetheless, a number of observations suggest that our understanding of developmental cell death in C. elegans is incomplete. The interaction between dying and engulfing cells seems to be more complex than originally appreciated, and it appears that key aspects of cell death initiation are not fully understood. It has also become apparent that the conserved apoptotic pathway is dispensable for the demise of the C. elegans linker cell, leading to the discovery of a previously unexplored gene program promoting cell death. Here, we review studies that formed the foundation of cell death research in C. elegans and describe new observations that expand, and in some cases remodel, this edifice. We raise the possibility that, in some cells, more than one death program may be needed to ensure cell death fidelity. © 2015 Elsevier Inc. All rights reserved.

lck-Driven Cre Expression Alters T Cell Development in the Thymus and the Frequencies and Functions of Peripheral T Cell Subsets.

Science.gov (United States)

Carow, Berit; Gao, Yu; Coquet, Jonathan; Reilly, Marie; Rottenberg, Martin E

2016-09-15

Conditional gene targeting using the bacteriophage-derived Cre recombinase is widely applied for functional gene studies in mice. Mice transgenic for Cre under the control of the lck gene promoter are used to study the role of loxP-targeted genes in T cell development and function. In this article, we show a striking 65% reduction in cellularity, preferential development of γδ versus αβ T cells, and increased expression of IL-7R in the thymus of mice expressing Cre under the proximal lck promoter (lck-cre(+) mice). The transition from CD4/CD8 double-negative to double-positive cells was blocked, and lck-cre(+) double-positive cells were more prone to apoptosis and showed higher levels of Cre expression. Importantly, numbers of naive T cells were reduced in spleens and lymph nodes of lck-cre(+) mice. In contrast, frequencies of γδ T cells, CD44(+)CD62L(-) effector T cells, and Foxp3(+) regulatory T cells were elevated, as was the frequency of IFN-γ-secreting CD4(+) and CD8(+) T cells. A literature survey of 332 articles that used lck-cre(+) mice for deletion of floxed genes indicated that results are statistically influenced by the control used (lck-cre(+) or lck-cre(-)), more frequently resembling the lck-cre(+) phenotype described in this article if lck-cre(-) controls were used. Altogether, care should be taken when interpreting published results and to properly control targeted gene deletions using the lck-cre(+) strain. Copyright © 2016 by The American Association of Immunologists, Inc.

Regulation of Arabidopsis Early Anther Development by Putative Cell-Cell Signaling Molecules and Transcriptional Regulators

Institute of Scientific and Technical Information of China (English)

Yu-Jin Sun; Carey LH Hord; Chang-Bin Chen; Hong Ma

2007-01-01

Anther development in flowering plants involves the formation of several cell types, including the tapetal and pollen mother cells. The use of genetic and molecular tools has led to the identification and characterization of genes that are critical for normal cell division and differentiation in Arabidopsis early anther development. We review here several recent studies on these genes, including the demonstration that the putative receptor protein kinases BAM1 and BAM2 together play essential roles in the control of early cell division and differentiation. In addition, we discuss the hypothesis that BAM1/2 may form a positive-negative feedback regulatory loop with a previously identified key regulator, SPOROCYTELESS (also called NOZZLE),to control the balance between sporogenous and somatic cell types in the anther. Furthermore, we summarize the isolation and functional analysis of the DYSFUNCTIONAL TAPETUM1 (DYT1) gene in promoting proper tapetal cell differentiation. Our finding that DYT1 encodes a putative transcription factor of the bHLH family, as well as relevant expression analyses, strongly supports a model that DYT1 serves as a critical link between upstream factors and downstream target genes that are critical for normal tapetum development and function. These studies, together with other recently published works, indicate that cell-cell communication and transcriptional control are key processes essential for cell fate specification in anther development.

Chronophin coordinates cell leading edge dynamics by controlling active cofilin levels

Science.gov (United States)

Delorme-Walker, Violaine; Seo, Ji-Yeon; Gohla, Antje; Fowler, Bruce; Bohl, Ben; DerMardirossian, Céline

2015-01-01

Cofilin, a critical player of actin dynamics, is spatially and temporally regulated to control the direction and force of membrane extension required for cell locomotion. In carcinoma cells, although the signaling pathways regulating cofilin activity to control cell direction have been established, the molecular machinery required to generate the force of the protrusion remains unclear. We show that the cofilin phosphatase chronophin (CIN) spatiotemporally regulates cofilin activity at the cell edge to generate persistent membrane extension. We show that CIN translocates to the leading edge in a PI3-kinase–, Rac1-, and cofilin-dependent manner after EGF stimulation to activate cofilin, promotes actin free barbed end formation, accelerates actin turnover, and enhances membrane protrusion. In addition, we establish that CIN is crucial for the balance of protrusion/retraction events during cell migration. Thus, CIN coordinates the leading edge dynamics by controlling active cofilin levels to promote MTLn3 cell protrusion. PMID:26324884

Development of portable fuel cells

Energy Technology Data Exchange (ETDEWEB)

Nakatou, K.; Sumi, S.; Nishizawa, N. [Sanyo Electric Co., Ltd., Osaka (Japan)

1996-12-31

Sanyo Electric has been concentrating on developing a marketable portable fuel cell using phosphoric acid fuel cells (PAFC). Due to the fact that this power source uses PAFC that operate at low temperature around 100{degrees} C, they are easier to handle compared to conventional fuel cells that operate at around 200{degrees} C , they can also be expected to provide extended reliable operation because corrosion of the electrode material and deterioration of the electrode catalyst are almost completely nonexistent. This power source is meant to be used independently and stored at room temperature. When it is started up, it generates electricity itself using its internal load to raise the temperature. As a result, the phosphoric acid (the electolyte) absorbs the reaction water when the temperature starts to be raised (around room temperature). At the same time the concentration and volume of the phosphoric acid changes, which may adversely affect the life time of the cell. We have studied means for starting, operating PAFC stack using methods that can simply evaluate changes in the concentration of the electrolyte in the stack with the aim of improving and extending cell life and report on them in this paper.

EVI and MDS/EVI are required for adult intestinal stem cell formation during postembryonic vertebrate development.

Science.gov (United States)

Okada, Morihiro; Shi, Yun-Bo

2018-01-01

The gene ectopic viral integration site 1 (EVI) and its variant myelodysplastic syndrome 1 (MDS)/EVI encode zinc-finger proteins that have been recognized as important oncogenes in various types of cancer. In contrast to the established role of EVI and MDS/EVI in cancer development, their potential function during vertebrate postembryonic development, especially in organ-specific adult stem cells, is unclear. Amphibian metamorphosis is strikingly similar to postembryonic development around birth in mammals, with both processes taking place when plasma thyroid hormone (T3) levels are high. Using the T3-dependent metamorphosis in Xenopus tropicalis as a model, we show here that high levels of EVI and MDS/EVI are expressed in the intestine at the climax of metamorphosis and are induced by T3. By using the transcription activator-like effector nuclease gene editing technology, we have knocked out both EVI and MDS/EVI and have shown that EVI and MDS/EVI are not essential for embryogenesis and premetamorphosis in X. tropicalis On the other hand, knocking out EVI and MDS/EVI causes severe retardation in the growth and development of the tadpoles during metamorphosis and leads to tadpole lethality at the climax of metamorphosis. Furthermore, the homozygous-knockout animals have reduced adult intestinal epithelial stem cell proliferation at the end of metamorphosis (for the few that survive through metamorphosis) or during T3-induced metamorphosis. These findings reveal a novel role of EVI and/or MDS/EVI in regulating the formation and/or proliferation of adult intestinal adult stem cells during postembryonic development in vertebrates.-Okada, M., Shi, Y.-B. EVI and MDS/EVI are required for adult intestinal stem cell formation during postembryonic vertebrate development. © FASEB.

Effects of environmental Bisphenol A exposures on germ cell development and Leydig cell function in the human fetal testis.

Directory of Open Access Journals (Sweden)

Soria Eladak

Full Text Available Using an organotypic culture system termed human Fetal Testis Assay (hFeTA we previously showed that 0.01 μM BPA decreases basal, but not LH-stimulated, testosterone secreted by the first trimester human fetal testis. The present study was conducted to determine the potential for a long-term antiandrogenic effect of BPA using a xenograft model, and also to study the effect of BPA on germ cell development using both the hFETA and xenograft models.Using the hFeTA system, first trimester testes were cultured for 3 days with 0.01 to 10 μM BPA. For xenografts, adult castrate male nude mice were injected with hCG and grafted with first trimester testes. Host mice received 10 μM BPA (~ 500 μg/kg/day in their drinking water for 5 weeks. Plasma levels of total and unconjugated BPA were 0.10 μM and 0.038 μM respectively. Mice grafted with second trimester testes received 0.5 and 50 μg/kg/day BPA by oral gavage for 5 weeks.With first trimester human testes, using the hFeTA model, 10 μM BPA increased germ cell apoptosis. In xenografts, germ cell density was also reduced by BPA exposure. Importantly, BPA exposure significantly decreased the percentage of germ cells expressing the pluripotency marker AP-2γ, whilst the percentage of those expressing the pre-spermatogonial marker MAGE-A4 significantly increased. BPA exposure did not affect hCG-stimulated androgen production in first and second trimester xenografts as evaluated by both plasma testosterone level and seminal vesicle weight in host mice.Exposure to BPA at environmentally relevant concentrations impairs germ cell development in first trimester human fetal testis, whilst gonadotrophin-stimulated testosterone production was unaffected in both first and second trimester testis. Studies using first trimester human fetal testis demonstrate the complementarity of the FeTA and xenograft models for determining the respective short-term and long term effects of environmental exposures.

[{sup 131}I]FIAU labeling of genetically transduced, tumor-reactive lymphocytes: cell-level dosimetry and dose-dependent toxicity

Energy Technology Data Exchange (ETDEWEB)

Zanzonico, Pat [Memorial Sloan-Kettering Cancer Center, Department of Medical Physics, New York, NY (United States); Koehne, Guenther; Doubrovina, Ekaterina; O' Reilly, Richard J. [Memorial Sloan-Kettering Cancer Center, Allogeneic Transplantation Service, New York, NY (United States); Memorial Sloan-Kettering Cancer Center, Immunology Program, New York, NY (United States); Gallardo, Humilidad F. [Memorial Sloan-Kettering Cancer Center, Gene Transfer and Somatic Cell Engineering Facility, New York, NY (United States); Doubrovin, Mikhail; Blasberg, Ronald G. [Memorial Sloan-Kettering Cancer Center, Department of Radiology, New York, NY (United States); Memorial Sloan-Kettering Cancer Center, Department of Neurology, New York, NY (United States); Finn, Ronald [Memorial Sloan-Kettering Cancer Center, Radiochemistry and Cyclotron Core Facility, New York, NY (United States); Riviere, Isabelle; Sadelain, Michel [Memorial Sloan-Kettering Cancer Center, Immunology Program, New York, NY (United States); Memorial Sloan-Kettering Cancer Center, Gene Transfer and Somatic Cell Engineering Facility, New York, NY (United States); Larson, Steven M. [Memorial Sloan-Kettering Cancer Center, Department of Radiology, New York, NY (United States)

2006-09-15

Donor T cells have been shown to be reactive against and effective in adoptive immunotherapy of Epstein-Barr virus (EBV) lymphomas which develop in some leukemia patients post marrow transplantation. These T cells may be genetically modified by incorporation of a replication-incompetent viral vector (NIT) encoding both an inactive mutant nerve growth factor receptor (LNGFR), as an immunoselectable surface marker, and a herpes simplex virus thymidine kinase (HSV-TK), rendering the cells sensitive to ganciclovir. The current studies are based on the selective HSV-TK-catalyzed trapping (phosphorylation) of the thymidine analog [{sup 131}I]-2'-fluoro-2'-deoxy-1-{beta}-D-arabinofuransyl-5-iodo-uracil (FIAU) as a means of stably labeling such T cells for in vivo trafficking (including tumor targeting) studies. Because of the radiosensitivity of lymphocytes and the potentially high absorbed dose to the nucleus from intracellular {sup 131}I (even at tracer levels), the nucleus absorbed dose (D{sub n}) and dose-dependent immune functionality were evaluated for NIT {sup +} T cells labeled ex vivo in [{sup 131}I ]FIAU-containing medium. Based on in vitro kinetic studies of [{sup 131}I ]FIAU uptake by NIT {sup +} T cells, D{sub n} was calculated using an adaptation of the MIRD formalism and the recently published MIRD cellular S factors. Immune cytotoxicity of [{sup 131}I ]FIAU-labeled cells was assayed against {sup 51}Cr-labeled target cells [B-lymphoblastoid cells (BLCLs) ] in a standard 4-h release assay. At median nuclear absorbed doses up to 830 cGy, a {sup 51}Cr-release assay against BLCLs showed no loss of immune cytotoxicity, thus demonstrating the functional integrity of genetically transduced, tumor-reactive T cells labeled at this dose level for in vivo cell trafficking and tumor targeting studies. (orig.)

The effect of additives on red cell 2,3 diphosphoglycerate levels in CPDA preservatives.

Science.gov (United States)

Vora, S; West, C; Beutler, E

1989-01-01

Forty-two chemical substances, chosen because they might influence red cell metabolism, were screened for effect on red cell adenosine triphosphate and 2,3 diphosphoglycerate (2,3 DPG) levels during storage in CPD or CPDA-1 at 4 degrees C. Of these substances, six appeared on initial screening to increase 2,3 DPG levels during storage; on repeated examination, four compounds, i.e., oxalate, glyoxalate, ethyl oxaloacetate, and L-phenylalanyl-L-alanine, consistently increased 2,3 DPG levels during storage. It was shown that glyoxalate was converted rapidly to oxalate in blood, presumably through the lactate dehydrogenase reaction. Ethyl oxaloacetate is known to hydrolyze, giving rise to oxalate. Thus, the effect of both glyoxalate and ethyl oxaloacetate can be explained by the formation of oxalate, a compound already known to increase 2,3 DPG levels. The effect of L-phenylalanyl-L-alanine remains to be explained, but it may be hydrolyzed to L-alanine and L-phenylalanine, both of which are thought to have the capacity to increase red cell 2,3 DPG levels by inhibiting pyruvate kinase activity.

Regulation of myogenesis and skeletal muscle regeneration: effects of oxygen levels on satellite cell activity.

Science.gov (United States)

Chaillou, Thomas; Lanner, Johanna T

2016-12-01

Reduced oxygen (O 2 ) levels (hypoxia) are present during embryogenesis and exposure to altitude and in pathologic conditions. During embryogenesis, myogenic progenitor cells reside in a hypoxic microenvironment, which may regulate their activity. Satellite cells are myogenic progenitor cells localized in a local environment, suggesting that the O 2 level could affect their activity during muscle regeneration. In this review, we present the idea that O 2 levels regulate myogenesis and muscle regeneration, we elucidate the molecular mechanisms underlying myogenesis and muscle regeneration in hypoxia and depict therapeutic strategies using changes in O 2 levels to promote muscle regeneration. Severe hypoxia (≤1% O 2 ) appears detrimental for myogenic differentiation in vitro, whereas a 3-6% O 2 level could promote myogenesis. Hypoxia impairs the regenerative capacity of injured muscles. Although it remains to be explored, hypoxia may contribute to the muscle damage observed in patients with pathologies associated with hypoxia (chronic obstructive pulmonary disease, and peripheral arterial disease). Hypoxia affects satellite cell activity and myogenesis through mechanisms dependent and independent of hypoxia-inducible factor-1α. Finally, hyperbaric oxygen therapy and transplantation of hypoxia-conditioned myoblasts are beneficial procedures to enhance muscle regeneration in animals. These therapies may be clinically relevant to treatment of patients with severe muscle damage.-Chaillou, T. Lanner, J. T. Regulation of myogenesis and skeletal muscle regeneration: effects of oxygen levels on satellite cell activity. © FASEB.

Determination of NAD⁺ and NADH level in a Single Cell Under H₂O₂ Stress by Capillary Electrophoresis