Targeted cellular ablation based on the morphology of malignant cells

NASA Astrophysics Data System (ADS)

Ivey, Jill W.; Latouche, Eduardo L.; Sano, Michael B.; Rossmeisl, John H.; Davalos, Rafael V.; Verbridge, Scott S.

2015-11-01

Treatment of glioblastoma multiforme (GBM) is especially challenging due to a shortage of methods to preferentially target diffuse infiltrative cells, and therapy-resistant glioma stem cell populations. Here we report a physical treatment method based on electrical disruption of cells, whose action depends strongly on cellular morphology. Interestingly, numerical modeling suggests that while outer lipid bilayer disruption induced by long pulses (~100 μs) is enhanced for larger cells, short pulses (~1 μs) preferentially result in high fields within the cell interior, which scale in magnitude with nucleus size. Because enlarged nuclei represent a reliable indicator of malignancy, this suggested a means of preferentially targeting malignant cells. While we demonstrate killing of both normal and malignant cells using pulsed electric fields (PEFs) to treat spontaneous canine GBM, we proposed that properly tuned PEFs might provide targeted ablation based on nuclear size. Using 3D hydrogel models of normal and malignant brain tissues, which permit high-resolution interrogation during treatment testing, we confirmed that PEFs could be tuned to preferentially kill cancerous cells. Finally, we estimated the nuclear envelope electric potential disruption needed for cell death from PEFs. Our results may be useful in safely targeting the therapy-resistant cell niches that cause recurrence of GBM tumors.

Targeted cellular ablation based on the morphology of malignant cells

PubMed Central

Ivey, Jill W.; Latouche, Eduardo L.; Sano, Michael B.; Rossmeisl, John H.; Davalos, Rafael V.; Verbridge, Scott S.

2015-01-01

Treatment of glioblastoma multiforme (GBM) is especially challenging due to a shortage of methods to preferentially target diffuse infiltrative cells, and therapy-resistant glioma stem cell populations. Here we report a physical treatment method based on electrical disruption of cells, whose action depends strongly on cellular morphology. Interestingly, numerical modeling suggests that while outer lipid bilayer disruption induced by long pulses (~100 μs) is enhanced for larger cells, short pulses (~1 μs) preferentially result in high fields within the cell interior, which scale in magnitude with nucleus size. Because enlarged nuclei represent a reliable indicator of malignancy, this suggested a means of preferentially targeting malignant cells. While we demonstrate killing of both normal and malignant cells using pulsed electric fields (PEFs) to treat spontaneous canine GBM, we proposed that properly tuned PEFs might provide targeted ablation based on nuclear size. Using 3D hydrogel models of normal and malignant brain tissues, which permit high-resolution interrogation during treatment testing, we confirmed that PEFs could be tuned to preferentially kill cancerous cells. Finally, we estimated the nuclear envelope electric potential disruption needed for cell death from PEFs. Our results may be useful in safely targeting the therapy-resistant cell niches that cause recurrence of GBM tumors. PMID:26596248

A difunctional squarylium indocyanine dye distinguishes dead cells through diverse staining of the cell nuclei/membranes.

PubMed

Li, Jie; Guo, Kunru; Shen, Jie; Yang, Wantai; Yin, Meizhen

2014-04-09

Functionalized fluorescent dyes have attracted great interest for the specific staining of subcellular organelles in multicellular organisms. A novel nanometer-sized water-soluble multi-functional squarylium indocyanine dye (D1) that contains four primary amines is synthesized. The dye exhibits good photostability, non-toxicity and biocompatibility. Isothermal titration calorimetry demonstrates that an affinity between D1 and DNA is higher than that between D1 and analogue of phospholipids. Analysis of circular dichroism spectra indicates that D1 targets to the DNA minor groove and aggregates to a helix. Because of the distinct affinity between the dye and subcellular organelles, the dye exhibits difunctional abilities to label the cell nuclei in fixed cells/tissue and the cell membranes in live cells/tissue. By combination of the two staining capabilities, the dye is further explored as a specific marker to distinguish apoptotic cells in live cells/tissue. The research opens a new way to design novel multifunctional dyes for life science applications. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Target fragments in collisions of 1.8 GeV/nucleon Fe-56 nuclei with photoemulsion nuclei, and the cascade-evaporation model

NASA Technical Reports Server (NTRS)

Dudkin, V. E.; Kovalev, E. E.; Nefedov, N. A.; Antonchik, V. A.; Bogdanov, S. D.; Ostroumov, V. I.; Benton, E. V.; Crawford, H. J.

1995-01-01

Nuclear photographic emulsion is used to study the dependence of the characteristics of target-nucleus fragments on the masses and impact parameters of interacting nuclei. The data obtained are compared in all details with the calculation results made in terms of the Dubna version of the cascade-evaporation model (DCM).

A generic nuclei detection method for histopathological breast images

NASA Astrophysics Data System (ADS)

Kost, Henning; Homeyer, André; Bult, Peter; Balkenhol, Maschenka C. A.; van der Laak, Jeroen A. W. M.; Hahn, Horst K.

2016-03-01

The detection of cell nuclei plays a key role in various histopathological image analysis problems. Considering the high variability of its applications, we propose a novel generic and trainable detection approach. Adaption to specific nuclei detection tasks is done by providing training samples. A trainable deconvolution and classification algorithm is used to generate a probability map indicating the presence of a nucleus. The map is processed by an extended watershed segmentation step to identify the nuclei positions. We have tested our method on data sets with different stains and target nuclear types. We obtained F1-measures between 0.83 and 0.93.

Optimized Methods for the Isolation of Arabidopsis Female Central Cells and Their Nuclei

PubMed Central

Park, Kyunghyuk; Frost, Jennifer M.; Adair, Adam James; Kim, Dong Min; Yun, Hyein; Brooks, Janie S.; Fischer, Robert L.; Choi, Yeonhee

2016-01-01

The Arabidopsis female gametophyte contains seven cells with eight haploid nuclei buried within layers of sporophytic tissue. Following double fertilization, the egg and central cells of the gametophyte develop into the embryo and endosperm of the seed, respectively. The epigenetic status of the central cell has long presented an enigma due both to its inaccessibility, and the fascinating epigenome of the endosperm, thought to have been inherited from the central cell following activity of the DEMETER demethylase enzyme, prior to fertilization. Here, we present for the first time, a method to isolate pure populations of Arabidopsis central cell nuclei. Utilizing a protocol designed to isolate leaf mesophyll protoplasts, we systematically optimized each step in order to efficiently separate central cells from the female gametophyte. We use initial manual pistil dissection followed by the derivation of central cell protoplasts, during which process the central cell emerges from the micropylar pole of the embryo sac. Then, we use a modified version of the Isolation of Nuclei TAgged in specific Cell Types (INTACT) protocol to purify central cell nuclei, resulting in a purity of 75–90% and a yield sufficient to undertake downstream molecular analyses. We find that the process is highly dependent on the health of the original plant tissue used, and the efficiency of protoplasting solution infiltration into the gametophyte. By isolating pure central cell populations, we have enabled elucidation of the physiology of this rare cell type, which in the future will provide novel insights into Arabidopsis reproduction. PMID:27788573

Automated segmentation and isolation of touching cell nuclei in cytopathology smear images of pleural effusion using distance transform watershed method

NASA Astrophysics Data System (ADS)

Win, Khin Yadanar; Choomchuay, Somsak; Hamamoto, Kazuhiko

2017-06-01

The automated segmentation of cell nuclei is an essential stage in the quantitative image analysis of cell nuclei extracted from smear cytology images of pleural fluid. Cell nuclei can indicate cancer as the characteristics of cell nuclei are associated with cells proliferation and malignancy in term of size, shape and the stained color. Nevertheless, automatic nuclei segmentation has remained challenging due to the artifacts caused by slide preparation, nuclei heterogeneity such as the poor contrast, inconsistent stained color, the cells variation, and cells overlapping. In this paper, we proposed a watershed-based method that is capable to segment the nuclei of the variety of cells from cytology pleural fluid smear images. Firstly, the original image is preprocessed by converting into the grayscale image and enhancing by adjusting and equalizing the intensity using histogram equalization. Next, the cell nuclei are segmented using OTSU thresholding as the binary image. The undesirable artifacts are eliminated using morphological operations. Finally, the distance transform based watershed method is applied to isolate the touching and overlapping cell nuclei. The proposed method is tested with 25 Papanicolaou (Pap) stained pleural fluid images. The accuracy of our proposed method is 92%. The method is relatively simple, and the results are very promising.

Automated recognition of cell phenotypes in histology images based on membrane- and nuclei-targeting biomarkers

PubMed Central

Karaçalı, Bilge; Vamvakidou, Alexandra P; Tözeren, Aydın

2007-01-01

Background Three-dimensional in vitro culture of cancer cells are used to predict the effects of prospective anti-cancer drugs in vivo. In this study, we present an automated image analysis protocol for detailed morphological protein marker profiling of tumoroid cross section images. Methods Histologic cross sections of breast tumoroids developed in co-culture suspensions of breast cancer cell lines, stained for E-cadherin and progesterone receptor, were digitized and pixels in these images were classified into five categories using k-means clustering. Automated segmentation was used to identify image regions composed of cells expressing a given biomarker. Synthesized images were created to check the accuracy of the image processing system. Results Accuracy of automated segmentation was over 95% in identifying regions of interest in synthesized images. Image analysis of adjacent histology slides stained, respectively, for Ecad and PR, accurately predicted regions of different cell phenotypes. Image analysis of tumoroid cross sections from different tumoroids obtained under the same co-culture conditions indicated the variation of cellular composition from one tumoroid to another. Variations in the compositions of cross sections obtained from the same tumoroid were established by parallel analysis of Ecad and PR-stained cross section images. Conclusion Proposed image analysis methods offer standardized high throughput profiling of molecular anatomy of tumoroids based on both membrane and nuclei markers that is suitable to rapid large scale investigations of anti-cancer compounds for drug development. PMID:17822559

Inheritance of gene density–related higher order chromatin arrangements in normal and tumor cell nuclei

PubMed Central

Cremer, Marion; Küpper, Katrin; Wagler, Babett; Wizelman, Leah; Hase, Johann v.; Weiland, Yanina; Kreja, Ludwika; Diebold, Joachim; Speicher, Michael R.; Cremer, Thomas

2003-01-01

A gene density–related difference in the radial arrangement of chromosome territories (CTs) was previously described for human lymphocyte nuclei with gene-poor CT #18 located toward the nuclear periphery and gene-dense CT #19 in the nuclear interior (Croft, J.A., J.M. Bridger, S. Boyle, P. Perry, P. Teague, and W.A. Bickmore. 1999. J. Cell Biol. 145:1119–1131). Here, we analyzed the radial distribution of chromosome 18 and 19 chromatin in six normal cell types and in eight tumor cell lines, some of them with imbalances and rearrangements of the two chromosomes. Our findings demonstrate that a significant difference in the radial distribution of #18 and #19 chromatin is a common feature of higher order chromatin architecture in both normal and malignant cell types. However, in seven of eight tumor cell lines, the difference was less pronounced compared with normal cell nuclei due to a higher fraction of nuclei showing an inverted CT position, i.e., a CT #18 located more internally than a CT #19. This observation emphasizes a partial loss of radial chromatin order in tumor cell nuclei. PMID:12952935

Plant parasitic nematode effectors target host defense and nuclear functions to establish feeding cells.

PubMed

Quentin, Michaëel; Abad, Pierre; Favery, Bruno

2013-01-01

Plant parasitic nematodes are microscopic worms, the most damaging species of which have adopted a sedentary lifestyle within their hosts. These obligate endoparasites have a biotrophic relationship with plants, in which they induce the differentiation of root cells into hypertrophied, multinucleate feeding cells (FCs). Effectors synthesized in the esophageal glands of the nematode are injected into the plant cells via the syringe-like stylet and play a key role in manipulating the host machinery. The establishment of specialized FCs requires these effectors to modulate many aspects of plant cell morphogenesis and physiology, including defense responses. This cell reprogramming requires changes to host nuclear processes. Some proteins encoded by parasitism genes target host nuclei. Several of these proteins were immunolocalized within FC nuclei or shown to interact with host nuclear proteins. Comparative genomics and functional analyses are gradually revealing the roles of nematode effectors. We describe here these effectors and their hypothesized roles in the unique feeding behavior of these pests.

Release of cell-free ice nuclei from Halomonas elongata expressing the ice nucleation gene inaZ of Pseudomonas syringae.

PubMed

Tegos, G; Vargas, C; Perysinakis, A; Koukkou, A I; Christogianni, A; Nieto, J J; Ventosa, A; Drainas, C

2000-11-01

Release of ice nuclei in the growth medium of recombinant Halomonas elongata cells expressing the inaZ gene of Pseudomonas syringae was studied in an attempt to produce cell-free active ice nuclei for biotechnological applications. Cell-free ice nuclei were not retained by cellulose acetate filters of 0.2 microm pore size. Highest activity of cell-free ice nuclei was obtained when cells were grown in low salinity (0.5-5% NaCl, w/v). Freezing temperature threshold, estimated to be below -7 degrees C indicating class C nuclei, was not affected by medium salinity. Their density, as estimated by Percoll density centrifugation, was 1.018 +/- 0.002 gml(-1) and they were found to be free of lipids. Ice nuclei are released in the growth medium of recombinant H. elongata cells probably because of inefficient anchoring of the ice-nucleation protein aggregates in the outer membrane. The ice+ recombinant H. elongata cells could be useful for future use as a source of active cell-free ice nucleation protein.

Refractometry of melanocyte cell nuclei using optical scatter images recorded by digital Fourier microscopy.

PubMed

Seet, Katrina Y T; Nieminen, Timo A; Zvyagin, Andrei V

2009-01-01

The cell nucleus is the dominant optical scatterer in the cell. Neoplastic cells are characterized by cell nucleus polymorphism and polychromism-i.e., the nuclei exhibits an increase in the distribution of both size and refractive index. The relative size parameter, and its distribution, is proportional to the product of the nucleus size and its relative refractive index and is a useful discriminant between normal and abnormal (cancerous) cells. We demonstrate a recently introduced holographic technique, digital Fourier microscopy (DFM), to provide a sensitive measure of this relative size parameter. Fourier holograms were recorded and optical scatter of individual scatterers were extracted and modeled with Mie theory to determine the relative size parameter. The relative size parameter of individual melanocyte cell nuclei were found to be 16.5+/-0.2, which gives a cell nucleus refractive index of 1.38+/-0.01 and is in good agreement with previously reported data. The relative size parameters of individual malignant melanocyte cell nuclei are expected to be greater than 16.5.

Glioma grading using cell nuclei morphologic features in digital pathology images

NASA Astrophysics Data System (ADS)

Reza, Syed M. S.; Iftekharuddin, Khan M.

2016-03-01

This work proposes a computationally efficient cell nuclei morphologic feature analysis technique to characterize the brain gliomas in tissue slide images. In this work, our contributions are two-fold: 1) obtain an optimized cell nuclei segmentation method based on the pros and cons of the existing techniques in literature, 2) extract representative features by k-mean clustering of nuclei morphologic features to include area, perimeter, eccentricity, and major axis length. This clustering based representative feature extraction avoids shortcomings of extensive tile [1] [2] and nuclear score [3] based methods for brain glioma grading in pathology images. Multilayer perceptron (MLP) is used to classify extracted features into two tumor types: glioblastoma multiforme (GBM) and low grade glioma (LGG). Quantitative scores such as precision, recall, and accuracy are obtained using 66 clinical patients' images from The Cancer Genome Atlas (TCGA) [4] dataset. On an average ~94% accuracy from 10 fold crossvalidation confirms the efficacy of the proposed method.

Physical insight into light scattering by photoreceptor cell nuclei.

PubMed

Kreysing, Moritz; Boyde, Lars; Guck, Jochen; Chalut, Kevin J

2010-08-01

A recent study showed that the rod photoreceptor cell nuclei in the retina of nocturnal and diurnal mammals differ considerably in architecture: the location of euchromatin and heterochromatin in the nucleus is interchanged. This inversion has significant implications for the refractive index distribution and the light scattering properties of the nucleus. Here, we extend previous two-dimensional analysis to three dimensions (3D) by using both a numerical finite-difference time-domain and an analytic Mie theory approach. We find that the specific arrangement of the chromatin phases in the nuclear core-shell models employed have little impact on the far-field scattering cross section. However, scattering in the near field, which is the relevant regime inside the retina, shows a significant difference between the two architectures. The "inverted" photoreceptor cell nuclei of nocturnal mammals act as collection lenses, with the lensing effect being much more pronounced in 3D than in two dimensions. This lensing helps to deliver light efficiently to the light-sensing outer segments of the rod photoreceptor cells and thereby improve night vision.

Search for α-Cluster Structure in Exotic Nuclei with the Prototype Active-Target Time-Projection Chamber

NASA Astrophysics Data System (ADS)

Fritsch, A.; Ayyad, Y.; Bazin, D.; Beceiro-Novo, S.; Bradt, J.; Carpenter, L.; Cortesi, M.; Mittig, W.; Suzuki, D.; Ahn, T.; Kolata, J. J.; Becchetti, F. D.; Howard, A. M.

2016-03-01

Some exotic nuclei appear to exhibit α-cluster structure. While various theoretical models currently describe such clustering, more experimental data are needed to constrain model predictions. The Prototype Active-Target Time-Projection Chamber (PAT-TPC) has low-energy thresholds for charged-particle decay and a high luminosity due to its thick gaseous active target volume, making it well-suited to search for low-energy α-cluster reactions. Radioactive-ion beams produced by the TwinSol facility at the University of Notre Dame were delivered to the PAT-TPC to study nuclei including 14C and 14O via α-resonant scattering. Differential cross sections and excitation functions were measured. Preliminary results from our recent experiments will be presented. This work is supported by the U.S. National Science Foundation.

Cell nuclei and cytoplasm joint segmentation using the sliding band filter.

PubMed

Quelhas, Pedro; Marcuzzo, Monica; Mendonça, Ana Maria; Campilho, Aurélio

2010-08-01

Microscopy cell image analysis is a fundamental tool for biological research. In particular, multivariate fluorescence microscopy is used to observe different aspects of cells in cultures. It is still common practice to perform analysis tasks by visual inspection of individual cells which is time consuming, exhausting and prone to induce subjective bias. This makes automatic cell image analysis essential for large scale, objective studies of cell cultures. Traditionally the task of automatic cell analysis is approached through the use of image segmentation methods for extraction of cells' locations and shapes. Image segmentation, although fundamental, is neither an easy task in computer vision nor is it robust to image quality changes. This makes image segmentation for cell detection semi-automated requiring frequent tuning of parameters. We introduce a new approach for cell detection and shape estimation in multivariate images based on the sliding band filter (SBF). This filter's design makes it adequate to detect overall convex shapes and as such it performs well for cell detection. Furthermore, the parameters involved are intuitive as they are directly related to the expected cell size. Using the SBF filter we detect cells' nucleus and cytoplasm location and shapes. Based on the assumption that each cell has the same approximate shape center in both nuclei and cytoplasm fluorescence channels, we guide cytoplasm shape estimation by the nuclear detections improving performance and reducing errors. Then we validate cell detection by gathering evidence from nuclei and cytoplasm channels. Additionally, we include overlap correction and shape regularization steps which further improve the estimated cell shapes. The approach is evaluated using two datasets with different types of data: a 20 images benchmark set of simulated cell culture images, containing 1000 simulated cells; a 16 images Drosophila melanogaster Kc167 dataset containing 1255 cells, stained for DNA and

Development of activity pencil beam algorithm using measured distribution data of positron emitter nuclei generated by proton irradiation of targets containing (12)C, (16)O, and (40)Ca nuclei in preparation of clinical application.

PubMed

Miyatake, Aya; Nishio, Teiji; Ogino, Takashi

2011-10-01

The purpose of this study is to develop a new calculation algorithm that is satisfactory in terms of the requirements for both accuracy and calculation time for a simulation of imaging of the proton-irradiated volume in a patient body in clinical proton therapy. The activity pencil beam algorithm (APB algorithm), which is a new technique to apply the pencil beam algorithm generally used for proton dose calculations in proton therapy to the calculation of activity distributions, was developed as a calculation algorithm of the activity distributions formed by positron emitter nuclei generated from target nuclear fragment reactions. In the APB algorithm, activity distributions are calculated using an activity pencil beam kernel. In addition, the activity pencil beam kernel is constructed using measured activity distributions in the depth direction and calculations in the lateral direction. (12)C, (16)O, and (40)Ca nuclei were determined as the major target nuclei that constitute a human body that are of relevance for calculation of activity distributions. In this study, "virtual positron emitter nuclei" was defined as the integral yield of various positron emitter nuclei generated from each target nucleus by target nuclear fragment reactions with irradiated proton beam. Compounds, namely, polyethylene, water (including some gelatin) and calcium oxide, which contain plenty of the target nuclei, were irradiated using a proton beam. In addition, depth activity distributions of virtual positron emitter nuclei generated in each compound from target nuclear fragment reactions were measured using a beam ON-LINE PET system mounted a rotating gantry port (BOLPs-RGp). The measured activity distributions depend on depth or, in other words, energy. The irradiated proton beam energies were 138, 179, and 223 MeV, and measurement time was about 5 h until the measured activity reached the background level. Furthermore, the activity pencil beam data were made using the activity pencil

Hit rates and radiation doses to nuclei of bone lining cells from alpha-particle-emitting radionuclides

NASA Technical Reports Server (NTRS)

Polig, E.; Jee, W. S.; Kruglikov, I. L.

1992-01-01

Factors relating the local concentration of a bone-seeking alpha-particle emitter to the mean hit rate have been determined for nuclei of bone lining cells using a Monte Carlo procedure. Cell nuclei were approximated by oblate spheroids with dimensions and location taken from a previous histomorphometric study. The Monte Carlo simulation is applicable for planar and diffuse labels at plane or cylindrical bone surfaces. Additionally, the mean nuclear dose per hit, the dose mean per hit, the mean track segment length and its second moment, the percentage of stoppers, and the frequency distribution of the dose have been determined. Some basic features of the hit statistics for bone lining cells have been outlined, and the consequences of existing standards of radiation protection with regard to the hit frequency to cell nuclei are discussed.

FACTORS INFLUENCING THE ABILITY OF ISOLATED CELL NUCLEI TO FORM GELS IN DILUTE ALKALI

PubMed Central

Dounce, Alexander L.; Monty, Kenneth J.

1955-01-01

1. Known methods for isolating cell nuclei are divided into two classes, depending on whether or not the nuclei are capable of forming gels in dilute alkali or strong saline solutions. Methods which produce nuclei that can form gels apparently prevent the action of an intramitochondrial enzyme capable of destroying the gel-forming capacity of the nuclei. Methods in the other class are believed to permit this enzyme to act on the nuclei during the isolation procedure, causing detachment of DNA from some nuclear constituent (probably protein). 2. It is shown that heating in alkaline solution and x-irradiation can destroy nuclear gels. Heating in acid or neutral solutions can destroy the capacity of isolated nuclei to form gels. 3. Chemical and biological evidence is summarized in favor of the hypothesis that DNA is normally bound firmly to some nuclear component by non-ionic linkages. PMID:14381437

Spiral Ganglion Neuron Projection Development to the Hindbrain in Mice Lacking Peripheral and/or Central Target Differentiation

PubMed Central

Elliott, Karen L.; Kersigo, Jennifer; Pan, Ning; Jahan, Israt; Fritzsch, Bernd

2017-01-01

We investigate the importance of the degree of peripheral or central target differentiation for mouse auditory afferent navigation to the organ of Corti and auditory nuclei in three different mouse models: first, a mouse in which the differentiation of hair cells, but not central auditory nuclei neurons is compromised (Atoh1-cre; Atoh1f/f); second, a mouse in which hair cell defects are combined with a delayed defect in central auditory nuclei neurons (Pax2-cre; Atoh1f/f), and third, a mouse in which both hair cells and central auditory nuclei are absent (Atoh1−/−). Our results show that neither differentiated peripheral nor the central target cells of inner ear afferents are needed (hair cells, cochlear nucleus neurons) for segregation of vestibular and cochlear afferents within the hindbrain and some degree of base to apex segregation of cochlear afferents. These data suggest that inner ear spiral ganglion neuron processes may predominantly rely on temporally and spatially distinct molecular cues in the region of the targets rather than interaction with differentiated target cells for a crude topological organization. These developmental data imply that auditory neuron navigation properties may have evolved before auditory nuclei. PMID:28450830

Self-Affinity and Lacunarity of Chromatin Texture in Benign and Malignant Breast Epithelial Cell Nuclei

NASA Astrophysics Data System (ADS)

Einstein, Andrew J.; Wu, Hai-Shan; Gil, Joan

1998-01-01

Methods are presented for characterizing the self-affinity and lacunarity of arbitrarily shaped images. Chromatin appearance in breast epithelial cell nuclei is shown to be statistically self-affine. Spectral and Minkowski dimensions are lesser in nuclei of malignant cases than in nuclei of benign cases, and lacunarity further quantifies morphologic differences such as chromatin clumping and nucleoli. Fractal texture features are used as the basis for an accurate cytologic diagnosis of breast cancer.

Vascular endothelial cells minimize the total force on their nuclei.

PubMed Central

Hazel, A L; Pedley, T J

2000-01-01

The vascular endothelium is a cellular monolayer that lines the arterial walls. It plays a vital role in the initiation and development of atherosclerosis, an occlusive arterial disease responsible for 50% of deaths in the Western world. The focal nature of the disease suggests that hemodynamic forces are an important factor in its pathogenesis. This has led to the investigation of the effects of mechanical forces on the endothelial cells themselves. It has been found that endothelial cells do respond to stresses induced by the flowing blood; in particular, they elongate and align with an imposed flow direction. In this paper, we calculate the distribution of force exerted on a three-dimensional hump, representing the raised cell nucleus, by a uniform shear flow. It is found that, for a nonaxisymmetric ellipsoidal hump, the least total force is experienced when the hump is aligned with the flow. Furthermore, for a hump of fixed volume, there is a specific aspect ratio combination that results in the least total force upon the hump, (0.38:2.2:1.0; height:length:width). This is approximately the same as the average aspect ratio taken up by the cell nuclei in vivo (0.27:2.23:1.0). It is possible, therefore, that the cells respond to the flow in such a way as to minimize the total force on their nuclei. PMID:10620272

Estradiol-promoted accumulation of receptor in nuclei of porcine endometrium cells. Immunogold electron microscopy of resting and estradiol-stimulated cells.

PubMed

Sierralta, W D; Jakob, F; Thole, H; Engel, P; Jungblut, P W

1992-01-01

Endometrium was collected by curettage from castrated pigs, either untreated or exposed to estradiol in vivo by intrauterine injection, and processed for electron microscopy. The resin LR Gold was used for embedding, and sections were floated on droplets of 10 nm diameter gold particles, coated with the immunoglobulin-G1 (IgG1) fraction or its Fab2 fragment of a monospecific polyclonal antiserum raised in goats against the C-terminal half of the estradiol receptor. On average, only one gold particle per microns 2 became attached in the cytoplasmic area of untreated cells, whereas four were found over the nuclear area. These figures rose to 2-3/microns 2 and 15-26/microns 2, respectively, within 10 min after exposure to estradiol. The labeling intensities of nuclei in cell clusters and of coprocessed nuclei released from cells ruptured during curettage were identical in all situations. Nuclear pores were frequently tagged after estradiol treatment. The proportions of tagging densities in nuclei of untreated and estradiol-exposed cells corresponded to those of receptor contents measured in extracts of isolated nuclei by ligand binding. This correlation was not seen for the cytoplasmic compartment of untreated cells, the scarce tagging of which is interpreted by hidden antigenic determinants. Our morphological analyses support the conclusions drawn from biochemical data (Sierralta et al., 1992) of an estradiol-promoted translocation of receptor from the cytoplasm into the nucleus.

Robust and automated three-dimensional segmentation of densely packed cell nuclei in different biological specimens with Lines-of-Sight decomposition.

PubMed

Mathew, B; Schmitz, A; Muñoz-Descalzo, S; Ansari, N; Pampaloni, F; Stelzer, E H K; Fischer, S C

2015-06-08

Due to the large amount of data produced by advanced microscopy, automated image analysis is crucial in modern biology. Most applications require reliable cell nuclei segmentation. However, in many biological specimens cell nuclei are densely packed and appear to touch one another in the images. Therefore, a major difficulty of three-dimensional cell nuclei segmentation is the decomposition of cell nuclei that apparently touch each other. Current methods are highly adapted to a certain biological specimen or a specific microscope. They do not ensure similarly accurate segmentation performance, i.e. their robustness for different datasets is not guaranteed. Hence, these methods require elaborate adjustments to each dataset. We present an advanced three-dimensional cell nuclei segmentation algorithm that is accurate and robust. Our approach combines local adaptive pre-processing with decomposition based on Lines-of-Sight (LoS) to separate apparently touching cell nuclei into approximately convex parts. We demonstrate the superior performance of our algorithm using data from different specimens recorded with different microscopes. The three-dimensional images were recorded with confocal and light sheet-based fluorescence microscopes. The specimens are an early mouse embryo and two different cellular spheroids. We compared the segmentation accuracy of our algorithm with ground truth data for the test images and results from state-of-the-art methods. The analysis shows that our method is accurate throughout all test datasets (mean F-measure: 91%) whereas the other methods each failed for at least one dataset (F-measure≤69%). Furthermore, nuclei volume measurements are improved for LoS decomposition. The state-of-the-art methods required laborious adjustments of parameter values to achieve these results. Our LoS algorithm did not require parameter value adjustments. The accurate performance was achieved with one fixed set of parameter values. We developed a novel and

Identification of cardiomyocyte nuclei and assessment of ploidy for the analysis of cell turnover.

PubMed

Bergmann, Olaf; Zdunek, Sofia; Alkass, Kanar; Druid, Henrik; Bernard, Samuel; Frisén, Jonas

2011-01-15

Assays to quantify myocardial renewal rely on the accurate identification of cardiomyocyte nuclei. We previously ¹⁴C birth dated human cardiomyocytes based on the nuclear localization of cTroponins T and I. A recent report by Kajstura et al. suggested that cTroponin I is only localized to the nucleus in a senescent subpopulation of cardiomyocytes, implying that ¹⁴C birth dating of cTroponin T and I positive cell populations underestimates cardiomyocyte renewal in humans. We show here that the isolation of cell nuclei from the heart by flow cytometry with antibodies against cardiac Troponins T and I, as well as pericentriolar material 1 (PCM-1), allows for isolation of close to all cardiomyocyte nuclei, based on ploidy and marker expression. We also present a reassessment of cardiomyocyte ploidy, which has important implications for the analysis of cell turnover, and iododeoxyuridine (IdU) incorporation data. These data provide the foundation for reliable analysis of cardiomyocyte turnover in humans. Copyright © 2010 Elsevier Inc. All rights reserved.

Activation of raphe nuclei triggers rapid and distinct effects on parallel olfactory bulb output channels

PubMed Central

Kapoor, Vikrant; Provost, Allison; Agarwal, Prateek; Murthy, Venkatesh N.

2015-01-01

The serotonergic raphe nuclei are involved in regulating brain states over time-scales of minutes and hours. We examined more rapid effects of serotonergic activation on two classes of principal neurons in the mouse olfactory bulb, mitral and tufted cells, which send olfactory information to distinct targets. Brief stimulation of the raphe nuclei led to excitation of tufted cells at rest and potentiation of their odor responses. While mitral cells at rest were also excited by raphe activation, their odor responses were bidirectionally modulated, leading to improved pattern separation of odors. In vitro whole-cell recordings revealed that specific optogenetic activation of raphe axons affected bulbar neurons through dual release of serotonin and glutamate. Therefore, the raphe nuclei, in addition to their role in neuromodulation of brain states, are also involved in fast, sub-second top-down modulation, similar to cortical feedback. This modulation can selectively and differentially sensitize or decorrelate distinct output channels. PMID:26752161

Does flat epithelial atypia have rounder nuclei than columnar cell change/hyperplasia? A morphometric approach to columnar cell lesions of the breast.

PubMed

Yamashita, Yoshiko; Ichihara, Shu; Moritani, Suzuko; Yoon, Han-Seung; Yamaguchi, Masahiro

2016-06-01

Columnar cell lesions of the breast encompass columnar cell change/hyperplasia (CCC/CCH) and flat epithelial atypia (FEA). These have attracted researchers because emerging data suggest that FEA may represent the earliest histologically detectable non-obligate precursor of breast cancer. However, it is occasionally difficult to distinguish FEA from CCC/CCH because of similar histology. Although the nuclei of FEA are frequently described as relatively round compared with those of CCC/CCH, there are few morphometric studies to support this statement. The aim of this study was to provide objective data as to the nuclear shape in columnar cell lesions. As a shape descriptor, we adopted ellipticity that is defined by the formula 2b/2a, where a is the length of the long axis of the ellipse and b is the length of the short axis. Contrary to circularity, ellipticity reflects the overall configuration of an ellipse irrespective of surface irregularity. Our image analysis included generating whole slide images, extracting glandular cell nuclei, measuring nuclear ellipticity, and superimposing graded colors based on execution of results on the captured images. A total of 7917 nuclei extracted from 22 FEA images and 5010 nuclei extracted from 13 CCC/CCH images were analyzed. There was a significant difference in nuclear roundness between FEA and CCC/CCH with mean ellipticity values of 0.723 and 0.679, respectively (p < 0.001, Welch's t test). Furthermore, FEA with malignancy had significantly rounder nuclei than FEA without malignancy (p < 0.001). Our preliminary results suggest that nuclear ellipticity is a key parameter in reproducibly classifying columnar cell lesions of the breast.

Preparation of isolated nuclei from K 562 haemopoietic cell line for high resolution scanning electron microscopy.

PubMed

Reipert, S; Reipert, B M; Allen, T D

1994-09-01

The aim of the work is to visualise nuclear pore complexes (NPCs) in mammalian cells by high resolution scanning electron microscopy. A detergent-free isolation protocol was employed to obtain clean nuclei from the haemopoietic cell line K 562. Nuclear isolation was performed by mechanical homogenisation under hypotonic conditions followed by purification of the nuclear fraction. The isolated nuclei were attached to silicon chips, fixed, critical point dried, and sputter coated with a thin film (3-4 nm) of tantalum. Analysis of the nuclear surface by scanning electron microscopy (SEM) revealed a strong sensitivity of the outer nuclear membrane (ONM) to disruption during the isolation procedure. A significant reduction of the characteristic pattern of damage to the ONM was achieved by means of an isopicnic centrifugation on an isoosmolar balanced Percoll gradient. Analysis of the population of isolated nuclei by flow cytometry showed no signs of cell cycle specific losses of nuclei during isolation. The SEM investigations of the morphology of the nuclear envelope (NE) and of substructural details of NPCs and polyribosomes were performed using an in-lens field emission scanning electron microscope.

Segmentation of whole cells and cell nuclei from 3-D optical microscope images using dynamic programming.

PubMed

McCullough, D P; Gudla, P R; Harris, B S; Collins, J A; Meaburn, K J; Nakaya, M A; Yamaguchi, T P; Misteli, T; Lockett, S J

2008-05-01

Communications between cells in large part drive tissue development and function, as well as disease-related processes such as tumorigenesis. Understanding the mechanistic bases of these processes necessitates quantifying specific molecules in adjacent cells or cell nuclei of intact tissue. However, a major restriction on such analyses is the lack of an efficient method that correctly segments each object (cell or nucleus) from 3-D images of an intact tissue specimen. We report a highly reliable and accurate semi-automatic algorithmic method for segmenting fluorescence-labeled cells or nuclei from 3-D tissue images. Segmentation begins with semi-automatic, 2-D object delineation in a user-selected plane, using dynamic programming (DP) to locate the border with an accumulated intensity per unit length greater that any other possible border around the same object. Then the two surfaces of the object in planes above and below the selected plane are found using an algorithm that combines DP and combinatorial searching. Following segmentation, any perceived errors can be interactively corrected. Segmentation accuracy is not significantly affected by intermittent labeling of object surfaces, diffuse surfaces, or spurious signals away from surfaces. The unique strength of the segmentation method was demonstrated on a variety of biological tissue samples where all cells, including irregularly shaped cells, were accurately segmented based on visual inspection.

Organization of projections from the raphe nuclei to the vestibular nuclei in rats

NASA Technical Reports Server (NTRS)

Halberstadt, A. L.; Balaban, C. D.

2003-01-01

Previous anatomic and electrophysiological evidence suggests that serotonin modulates processing in the vestibular nuclei. This study examined the organization of projections from serotonergic raphe nuclei to the vestibular nuclei in rats. The distribution of serotonergic axons in the vestibular nuclei was visualized immunohistochemically in rat brain slices using antisera directed against the serotonin transporter. The density of serotonin transporter-immunopositive fibers is greatest in the superior vestibular nucleus and the medial vestibular nucleus, especially along the border of the fourth ventricle; it declines in more lateral and caudal regions of the vestibular nuclear complex. After unilateral iontophoretic injections of Fluoro-Gold into the vestibular nuclei, retrogradely labeled neurons were found in the dorsal raphe nucleus (including the dorsomedial, ventromedial and lateral subdivisions) and nucleus raphe obscurus, and to a minor extent in nucleus raphe pallidus and nucleus raphe magnus. The combination of retrograde tracing with serotonin immunohistofluorescence in additional experiments revealed that the vestibular nuclei receive both serotonergic and non-serotonergic projections from raphe nuclei. Tracer injections in densely innervated regions (especially the medial and superior vestibular nuclei) were associated with the largest numbers of Fluoro-Gold-labeled cells. Differences were observed in the termination patterns of projections from the individual raphe nuclei. Thus, the dorsal raphe nucleus sends projections that terminate predominantly in the rostral and medial aspects of the vestibular nuclear complex, while nucleus raphe obscurus projects relatively uniformly throughout the vestibular nuclei. Based on the topographical organization of raphe input to the vestibular nuclei, it appears that dense projections from raphe nuclei are colocalized with terminal fields of flocculo-nodular lobe and uvula Purkinje cells. It is hypothesized that

Parental genomes mix in mule and human cell nuclei.

PubMed

Hepperger, Claudia; Mayer, Andreas; Merz, Julia; Vanderwall, Dirk K; Dietzel, Steffen

2009-06-01

Whether chromosome sets inherited from father and mother occupy separate spaces in the cell nucleus is a question first asked over 110 years ago. Recently, the nuclear organization of the genome has come increasingly into focus as an important level of epigenetic regulation. In this context, it is indispensable to know whether or not parental genomes are spatially separated. Genome separation had been demonstrated for plant hybrids and for the early mammalian embryo. Conclusive studies for somatic mammalian cell nuclei are lacking because homologous chromosomes from the two parents cannot be distinguished within a species. We circumvented this problem by investigating the three-dimensional distribution of chromosomes in mule lymphocytes and fibroblasts. Genomic DNA of horse and donkey was used as probes in fluorescence in situ hybridization under conditions where only tandem repetitive sequences were detected. We thus could determine the distribution of maternal and paternal chromosome sets in structurally preserved interphase nuclei for the first time. In addition, we investigated the distribution of several pairs of chromosomes in human bilobed granulocytes. Qualitative and quantitative image evaluation did not reveal any evidence for the separation of parental genomes. On the contrary, we observed mixing of maternal and paternal chromosome sets.

Fluorescent Magnesium Nanocomplex in Protein Scaffold for Cell Nuclei Imaging Application

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pandya, Alok; Tripathi, Apritam; Purohit, Rahul

2015-10-27

Here in, we report a facile strategy for the synthesis of water-soluble ultra-fine blue emitting fluorescent Magnesium nanoparticles-protein complex (MgNC). This MgNC is demonstrated to exhibit excellent photo stability and biocompatibility. It was also observed that MgNC stain cell nuclei with high specifcity.

Investigation to synthesis more isotopes of superheavy nuclei Z = 118

NASA Astrophysics Data System (ADS)

Manjunatha, H. C.; Sridhar, K. N.

2018-07-01

We have studied the α-decay properties of superheavy nuclei Z = 118 in the range 275 ≤ A ≤ 325. Most of the predicted, unknown nuclei in the range 291 ≤ A ≤ 301 were found to have α-decay chains. Of these the nuclei 293-301118 were found to have long half-lives and hence could be sufficient to detect them if synthesized in a laboratory. Fusion barries for different projectile-target combinations to synthesis superheavy nuclei Z = 118 are studied and are also represented in simple relations. We have also studied the evaporation residue cross section, compound nucleus formation probability (PCN) and survival probability (PSurv) of different projectile-target combinations to synthesis superheavy element Z = 118. The selected most probable projectile-target combinations are Ca+Cf, Ti+Cm, Sc+Bk, V+Am, Cr+Pu, Fe+U, Mn+Np, Ni+Th and Kr+Pb. We have formulated simple relations for maximum evaporation residue cross sections and its corresponding energies. This helps to identify the projectile-target combinations quickly. Hence, we have identified the most probable projectile-target combinations to synthesis these superheavy nuclei. We hope that our predictions may be a guide for the future experiments in the synthesis of more isotopes of superheavy nuclei Z = 118.

Automatic Detection of Mitosis and Nuclei From Cytogenetic Images by CellProfiler Software for Mitotic Index Estimation.

PubMed

González, Jorge Ernesto; Radl, Analía; Romero, Ivonne; Barquinero, Joan Francesc; García, Omar; Di Giorgio, Marina

2016-12-01

Mitotic Index (MI) estimation expressed as percentage of mitosis plays an important role as quality control endpoint. To this end, MI is applied to check the lot of media and reagents to be used throughout the assay and also to check cellular viability after blood sample shipping, indicating satisfactory/unsatisfactory conditions for the progression of cell culture. The objective of this paper was to apply the CellProfiler open-source software for automatic detection of mitotic and nuclei figures from digitized images of cultured human lymphocytes for MI assessment, and to compare its performance to that performed through semi-automatic and visual detection. Lymphocytes were irradiated and cultured for mitosis detection. Sets of images from cultures were analyzed visually and findings were compared with those using CellProfiler software. The CellProfiler pipeline includes the detection of nuclei and mitosis with 80% sensitivity and more than 99% specificity. We conclude that CellProfiler is a reliable tool for counting mitosis and nuclei from cytogenetic images, saves considerable time compared to manual operation and reduces the variability derived from the scoring criteria of different scorers. The CellProfiler automated pipeline achieves good agreement with visual counting workflow, i.e. it allows fully automated mitotic and nuclei scoring in cytogenetic images yielding reliable information with minimal user intervention. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Identification of a nuclear-localized nuclease from wheat cells undergoing programmed cell death that is able to trigger DNA fragmentation and apoptotic morphology on nuclei from human cells

PubMed Central

Domínguez, Fernando; Cejudo, Francisco J.

2006-01-01

PCD (programmed cell death) in plants presents important morphological and biochemical differences compared with apoptosis in animal cells. This raises the question of whether PCD arose independently or from a common ancestor in plants and animals. In the present study we describe a cell-free system, using wheat grain nucellar cells undergoing PCD, to analyse nucleus dismantling, the final stage of PCD. We have identified a Ca2+/Mg2+ nuclease and a serine protease localized to the nucleus of dying nucellar cells. Nuclear extracts from nucellar cells undergoing PCD triggered DNA fragmentation and other apoptotic morphology in nuclei from different plant tissues. Inhibition of the serine protease did not affect DNA laddering. Furthermore, we show that the nuclear extracts from plant cells triggered DNA fragmentation and apoptotic morphology in nuclei from human cells. The inhibition of the nucleolytic activity with Zn2+ or EDTA blocked the morphological changes of the nucleus. Moreover, nuclear extracts from apoptotic human cells triggered DNA fragmentation and apoptotic morphology in nuclei from plant cells. These results show that degradation of the nucleus is morphologically and biochemically similar in plant and animal cells. The implication of this finding on the origin of PCD in plants and animals is discussed. PMID:16613587

Blue two-photon fluorescence metal cluster probe precisely marking cell nuclei of two cell lines.

PubMed

Wang, Yaling; Cui, Yanyan; Liu, Ru; Wei, Yueteng; Jiang, Xinglu; Zhu, Huarui; Gao, Liang; Zhao, Yuliang; Chai, Zhifang; Gao, Xueyun

2013-11-25

A bifunctional peptide was designed to in situ reduce Cu ions and anchor a Cu cluster. The peptide-Cu cluster probe, mainly composed of Cu14, emitted blue two-photon fluorescence under femtosecond laser excitation. Most important, the probe can specifically mark the nuclei of HeLa and A549 cells, respectively.

On the occurrence of nuclei in mature sieve elements.

PubMed

Event, R F; Davis, J D; Tucker, C M; Alfieri, F J

1970-12-01

The secondary phloem of 3 species of the Taxodiaceae and 13 species of woody dicotyledons was examined for the occurrence of nuclei in mature sieve elements. Nuclei were found in all mature sieve cells of Metasequoia glyptostroboides, Sequoia sempervirens and Taxodium distichum, and in some mature sieve-tube members in 12 of the 13 species of woody dicotyledons. Except for nuclei of sieve cells undergoing cessation of function, the nuclei in mature sieve cells of M. glyptostroboides, S. sempervirens and T. distichum were normal in appearance. The occurrence and morphology of nuclei in mature sieve-tube members of the woody dicotyledons were quite variable. Only 3 species, Robinia pseudoacacia, Ulmus americana and Vitis riparia, contained some mature sieve elements with apparently normal nuclei.

Isolation of Nuclei and Nucleoli.

PubMed

Pendle, Alison F; Shaw, Peter J

2017-01-01

Here we describe methods for producing nuclei from Arabidopsis suspension cultures or root tips of Arabidopsis, wheat, or pea. These methods could be adapted for other species and cell types. The resulting nuclei can be further purified for use in biochemical or proteomic studies, or can be used for microscopy. We also describe how the nuclei can be used to obtain a preparation of nucleoli.

Interaction of 160-GeV muon with emulsion nuclei

NASA Astrophysics Data System (ADS)

Othman, S. M.; Ghoneim, M. T.; Hussein, M. T.; El-Samman, H.; Hussein, A.

In this work we present some results of the interaction of high-energy muons with emulsion nuclei. The interaction results in emission of a number of fragments as a consequence of electromagnetic dissociation of the excited target nuclei. This excitation is attributed to absorption of photons by the target nuclei due to the intense electric field of the very fast incident muon particles. The interactions take place at impact parameters that allow ultra-peripheral collisions to take place, leading to giant resonances and hence multifragmentation of emulsion targets. Charge identification, range, energy spectra, angular distribution and topological cross-section of the produced fragments are measured and evaluated.

Targeting Photoinduced DNA Destruction by Ru(II) Tetraazaphenanthrene in Live Cells by Signal Peptide.

PubMed

Burke, Christopher S; Byrne, Aisling; Keyes, Tia E

2018-06-06

Exploiting NF-κB transcription factor peptide conjugation, a Ru(II)-bis-tap complex (tap = 1,4,5,8-tetraazaphenanthrene) was targeted specifically to the nuclei of live HeLa and CHO cells for the first time. DNA binding of the complex  within the nucleus of live cells was evident from gradual extinction of the metal complex luminescence after it had crossed the nuclear envelope, attributed to guanine quenching of the ruthenium emission via photoinduced electron transfer. Resonance Raman imaging confirmed that the complex remained in the nucleus after emission is extinguished. In the dark and under imaging conditions the cells remain viable, but efficient cellular destruction was induced with precise spatiotemporal control by applying higher irradiation intensities to selected cells. Solution studies indicate that the peptide conjugated complex associates strongly with calf thymus DNA ex-cellulo and gel electrophoresis confirmed that the peptide conjugate is capable of singlet oxygen independent photodamage to plasmid DNA. This indicates that the observed efficient cellular destruction likely operates via direct DNA oxidation by photoinduced electron transfer between guanine and the precision targeted Ru(II)-tap probe. The discrete targeting of polyazaaromatic complexes to the cell nucleus and confirmation that they are photocytotoxic after nuclear delivery is an important step toward their application in cellular phototherapy.

Computational efficient segmentation of cell nuclei in 2D and 3D fluorescent micrographs

NASA Astrophysics Data System (ADS)

De Vylder, Jonas; Philips, Wilfried

2011-02-01

This paper proposes a new segmentation technique developed for the segmentation of cell nuclei in both 2D and 3D fluorescent micrographs. The proposed method can deal with both blurred edges as with touching nuclei. Using a dual scan line algorithm its both memory as computational efficient, making it interesting for the analysis of images coming from high throughput systems or the analysis of 3D microscopic images. Experiments show good results, i.e. recall of over 0.98.

Review of metastable states in heavy nuclei

DOE PAGES

Dracoulis, G. D.; Walker, P. M.; Kondev, F. G.

2016-05-31

Here, the structure of nuclear isomeric states is reviewed in the context of their role in contemporary nuclear physics research. Emphasis is given to high-spin isomers in heavy nuclei, with A ≳ 150. The possibility to exploit isomers to study some of the most exotic nuclei is a recurring theme. In spherical nuclei, the role of octupole collectivity is discussed in detail, while in deformed nuclei the limitations of the K quantum number are addressed. Isomer targets and isomer beams are considered, along with applications related to energy storage, astrophysics, medicine, and experimental advances.

Impact of physical confinement on nuclei geometry and cell division dynamics in 3D spheroids.

PubMed

Desmaison, Annaïck; Guillaume, Ludivine; Triclin, Sarah; Weiss, Pierre; Ducommun, Bernard; Lobjois, Valérie

2018-06-08

Multicellular tumour spheroids are used as a culture model to reproduce the 3D architecture, proliferation gradient and cell interactions of a tumour micro-domain. However, their 3D characterization at the cell scale remains challenging due to size and cell density issues. In this study, we developed a methodology based on 3D light sheet fluorescence microscopy (LSFM) image analysis and convex hull calculation that allows characterizing the 3D shape and orientation of cell nuclei relative to the spheroid surface. By using this technique and optically cleared spheroids, we found that in freely growing spheroids, nuclei display an elongated shape and are preferentially oriented parallel to the spheroid surface. This geometry is lost when spheroids are grown in conditions of physical confinement. Live 3D LSFM analysis of cell division revealed that confined growth also altered the preferential cell division axis orientation parallel to the spheroid surface and induced prometaphase delay. These results provide key information and parameters that help understanding the impact of physical confinement on cell proliferation within tumour micro-domains.

[A morphometric analysis of the nuclei and nucleoli in tumor cells in lymphogranulomatosis, diffuse large B-cell lymphoma and anaplastic large cell lymphoma].

PubMed

Gorgidze, L A; Vorob'ev, I A

2009-01-01

To make a comparative morphometric analysis of the nuclei and nucleoli of tumor cells in lymphogranulomatosis (LGM), diffuse large B-cell lymphoma (DLBCL) and anaplastic large cell lymphoma (ALCL) for differential diagnosis of these lymphomas. Biopsy material (lymph node biopsies) was frozen in hexane, fixed and stained, then microscopic pictures were made. Mean area of tumor cell nuclei in LGM was 97.25 +/- 68.77 mcm2, in DLBCL and ALCL--55.89 +/- 20.13 mcm2 and 70.31 +/- 34.64 mcm2, respectively. The area differences were significant (p < 0.001). Hodgkin's and Berezovsky-Rid-Sternberg cell bucleoli area was the largest (11.44 +/- 7.83 mcm2). The nucleoli of the former are larger than those of the latter. Mean area of the nucleoli in DLBCL was 3.05 +/- 1.58, in ALCL--5.53 +/- 4.94 mcm2. The differences are significant (p < 0.001). Nucleoli in Hodgkin 's cells are significantly larger than those in the tumor cells in ALCL and DLBCL and the nucleoli with the area more than 12 mcm2 can be used in differential diagnosis between LGM and DLBCL but not between LGM and ALCL.

Cell nuclei attributed relational graphs for efficient representation and classification of gastric cancer in digital histopathology

NASA Astrophysics Data System (ADS)

Sharma, Harshita; Zerbe, Norman; Heim, Daniel; Wienert, Stephan; Lohmann, Sebastian; Hellwich, Olaf; Hufnagl, Peter

2016-03-01

This paper describes a novel graph-based method for efficient representation and subsequent classification in histological whole slide images of gastric cancer. Her2/neu immunohistochemically stained and haematoxylin and eosin stained histological sections of gastric carcinoma are digitized. Immunohistochemical staining is used in practice by pathologists to determine extent of malignancy, however, it is laborious to visually discriminate the corresponding malignancy levels in the more commonly used haematoxylin and eosin stain, and this study attempts to solve this problem using a computer-based method. Cell nuclei are first isolated at high magnification using an automatic cell nuclei segmentation strategy, followed by construction of cell nuclei attributed relational graphs of the tissue regions. These graphs represent tissue architecture comprehensively, as they contain information about cell nuclei morphology as vertex attributes, along with knowledge of neighborhood in the form of edge linking and edge attributes. Global graph characteristics are derived and ensemble learning is used to discriminate between three types of malignancy levels, namely, non-tumor, Her2/neu positive tumor and Her2/neu negative tumor. Performance is compared with state of the art methods including four texture feature groups (Haralick, Gabor, Local Binary Patterns and Varma Zisserman features), color and intensity features, and Voronoi diagram and Delaunay triangulation. Texture, color and intensity information is also combined with graph-based knowledge, followed by correlation analysis. Quantitative assessment is performed using two cross validation strategies. On investigating the experimental results, it can be concluded that the proposed method provides a promising way for computer-based analysis of histopathological images of gastric cancer.

Computer vision approach to morphometric feature analysis of basal cell nuclei for evaluating malignant potentiality of oral submucous fibrosis.

PubMed

Muthu Rama Krishnan, M; Pal, Mousumi; Paul, Ranjan Rashmi; Chakraborty, Chandan; Chatterjee, Jyotirmoy; Ray, Ajoy K

2012-06-01

This research work presents a quantitative approach for analysis of histomorphometric features of the basal cell nuclei in respect to their size, shape and intensity of staining, from surface epithelium of Oral Submucous Fibrosis showing dysplasia (OSFD) to that of the Normal Oral Mucosa (NOM). For all biological activity, the basal cells of the surface epithelium form the proliferative compartment and therefore their morphometric changes will spell the intricate biological behavior pertaining to normal cellular functions as well as in premalignant and malignant status. In view of this, the changes in shape, size and intensity of staining of the nuclei in the basal cell layer of the NOM and OSFD have been studied. Geometric, Zernike moments and Fourier descriptor (FD) based as well as intensity based features are extracted for histomorphometric pattern analysis of the nuclei. All these features are statistically analyzed along with 3D visualization in order to discriminate the groups. Results showed increase in the dimensions (area and perimeter), shape parameters and decreasing mean nuclei intensity of the nuclei in OSFD in respect to NOM. Further, the selected features are fed to the Bayesian classifier to discriminate normal and OSFD. The morphometric and intensity features provide a good sensitivity of 100%, specificity of 98.53% and positive predicative accuracy of 97.35%. This comparative quantitative characterization of basal cell nuclei will be of immense help for oral onco-pathologists, researchers and clinicians to assess the biological behavior of OSFD, specially relating to their premalignant and malignant potentiality. As a future direction more extensive study involving more number of disease subjects is observed.

Nuclear spectroscopy of r-process nuclei around N = 126 using KISS

NASA Astrophysics Data System (ADS)

Hirayama, Y.; Watanabe, Y. X.; Miyatake, H.; Schury, P.; Wada, M.; Oyaizu, M.; Kakiguchi, Y.; Mukai, M.; Kimura, S.; Ahmed, M.; Jeong, S. C.; Moon, J. Y.; Park, J. H.

2017-09-01

The beta-decay properties and atomic mass of nuclei with neutron magic number of N = 126 are considered critical for understanding the production of heavy elements such as gold and platinum at astrophysical sites. We will produce and measure the half-lives and masses of the nuclei with Z = 74-77 around N = 126 by using the multinucleon transfer (MNT) reaction of ^{136} Xe/ ^{238} U beams and ^{198} Pt target system. For this purpose, we have constructed the KEK Isotope Separation System (KISS) at RIKEN RIBF facility. KISS consists of an argon gas cell based laser ion source (atomic number selection) and an isotope separation on-line (ISOL) (mass number selection), to produce pure low-energy beams of neutron-rich isotopes around N = 126 . We performed the on-line tests to study the basic properties of the KISS and, successfully extracted laser-ionized nuclei around N = 126.

Enlarged squamous cell nuclei in cervical cytologic specimens from perimenopausal women ("PM Cells") : a cause of ASC overdiagnosis.

PubMed

Cibas, Edmund S; Browne, Tara-Jane; Bassichis, Michelle H Mantel; Lee, Kenneth R

2005-07-01

We studied the appropriateness of interpreting squamous cells with enlarged, smooth, bland nuclei in perimenopausal women ("PM cells") as atypical squamous cells (ASCs). Papanicolaou smears (Paps) from 100 women (40-55 years old) with a cytologic interpretation of ASC of undetermined significance (ASCUS) and human papillomavirus (HPV) testing or a biopsy within 6 months were reviewed by 2 observers without knowledge of the biopsy diagnosis or HPV results. Cases in which both reviewers agreed that the Paps were diagnosed more properly as "negative for intraepithelial lesion or malignancy" were compared with cases of "true ASCUS," using histologic squamous intraepithelial lesion and/or a positive high-risk HPV test as a positive outcome (abnormal follow-up). Of 100 cases, 28 were reclassified as benign by both observers. In 15 of these, the original ASCUS interpretation was based on cells with bland nuclear enlargement (2-3 times the area of intermediate cell nuclei), smooth nuclear membranes, and fine chromatin. Abnormal follow-up was identified in 1 (7%) of 15 benign cases but in 30 (42%) of 72 true ASCUS cases (P = .023). PM cells are a significant cause of ASC overdiagnosis in women 40 to 55 years old. Cervical Paps with cells no more atypical than these can be interpreted safely as negative for intraepithelial lesion or malignancy.

Efficient Subcellular Targeting to the Cell Nucleus of Quantum Dots Densely Decorated with a Nuclear Localization Sequence Peptide.

PubMed

Maity, Amit Ranjan; Stepensky, David

2016-01-27

Organelle-targeted drug delivery can enhance the efficiency of the intracellularly acting drugs and reduce their toxicity. We generated core-shell type CdSe-ZnS quantum dots (QDs) densely decorated with NLS peptidic targeting residues using a 3-stage decoration approach and investigated their endocytosis and nuclear targeting efficiencies. The diameter of the generated QDs increased following the individual decoration stages (16.3, 18.9, and 21.9 nm), the ζ-potential became less negative (-33.2, -17.5, and -11.9 mV), and characteristic changes appeared in the FTIR spectra following decoration with the linker and NLS peptides. Quantitative analysis of the last decoration stage revealed that 37.9% and 33.2% of the alkyne-modified NLS groups that were added to the reaction mix became covalently attached or adsorbed to the QDs surface, respectively. These numbers correspond to 63.6 and 55.7 peptides conjugated or adsorbed to a single QD (the surface density of 42 and 37 conjugated and adsorbed peptides per 1000 nm(2) of the QDs surface), which is higher than in the majority of previous studies that reported decoration efficiencies of formulations intended for nuclear-targeted drug delivery. QDs decorated with NLS peptides undergo more efficient endocytosis, as compared to other investigated QDs formulations, and accumulated to a higher extent in the cell nucleus or in close vicinity to it (11.9%, 14.6%, and 56.1% of the QDs endocytosed by an average cell for the QD-COOH, QD-azide, and QD-NLS formulations, respectively). We conclude that dense decoration of QDs with NLS residues increased their endocytosis and led to their nuclear targeting (preferential accumulation in the cells nuclei or in close vicinity to them). The experimental system and research tools that were used in this study allow quantitative investigation of the mechanisms that govern the QDs nuclear targeting and their dependence on the formulation properties. These findings will contribute to the

Hydroxyl-HIF2-alpha is potential therapeutic target for renal cell carcinomas

PubMed Central

Isono, Takahiro; Chano, Tokuhiro; Yoshida, Tetsuya; Kageyama, Susumu; Kawauchi, Akihiro; Suzaki, Masafumi; Yuasa, Takeshi

2016-01-01

Dormant cancer cells are deprivation-resistant, and cause a number of problems for therapeutic approaches for cancers. Renal cell carcinomas (RCCs) include deprivation-resistant cells that are resistant to various treatments. In this study, the specific characteristics of deprivation-resistant cells were transcriptionally identified by next generation sequencing. The hypoxia-inducible factors (HIF) transcription factor network was significantly enhanced in deprivation-resistant RCCs compared to the sensitive RCCs. Deprivation-resistant RCCs, that had lost Von Hippel-Lindau tumor suppressor expression, expressed hydroxyl-HIF2-alpha in the nucleus, but not sensitive-RCCs. Hydroxyl-HIF-alpha was also expressed in nuclei of RCC tissue samples. Knockdown for HIF2-alpha, but not HIF1-alpha, induced cell death related to a reduction in HIF-related gene expression in deprivation-resistant RCC cells. Chetomin, a nuclear HIF-inhibitor, induced marked level of cytotoxicity in deprivation-resistant cells, similar to the knockdown of HIF2-alpha. Therefore, hydroxyl-HIF2-alpha might be a potential therapeutic target for RCCs. PMID:27822416

Cell cycle S phase markers are expressed in cerebral neuron nuclei of cats infected by the Feline Panleukopenia Virus.

PubMed

Poncelet, Luc; Garigliany, Mutien; Ando, Kunie; Franssen, Mathieu; Desmecht, Daniel; Brion, Jean-Pierre

2016-12-16

The cell cycle-associated neuronal death hypothesis, which has been proposed as a common mechanism for most neurodegenerative diseases, is notably supported by evidencing cell cycle effectors in neurons. However, in naturally occurring nervous system diseases, these markers are not expressed in neuron nuclei but in cytoplasmic compartments. In other respects, the Feline Panleukopenia Virus (FPV) is able to complete its cycle in mature brain neurons in the feline species. As a parvovirus, the FPV is strictly dependent on its host cell reaching the cell cycle S phase to start its multiplication. In this retrospective study on the whole brain of 12 cats with naturally-occurring, FPV-associated cerebellar atrophy, VP2 capsid protein expression was detected by immunostaining not only in some brain neuronal nuclei but also in neuronal cytoplasm in 2 cats, suggesting that viral mRNA translation was still occurring. In these cats, double immunostainings demonstrated the expression of cell cycle S phase markers cyclin A, cdk2 and PCNA in neuronal nuclei. Parvoviruses are able to maintain their host cells in S phase by triggering the DNA damage response. S139 phospho H2A1, a key player in the cell cycle arrest, was detected in some neuronal nuclei, supporting that infected neurons were also blocked into the S phase. PCR studies did not support a co-infection with an adeno or herpes virus. ERK1/2 nuclear accumulation was observed in some neurons suggesting that the ERK signaling pathway might be involved as a mechanism driving these neurons far into the cell cycle.

Synchronization of Mammalian Cells and Nuclei by Centrifugal Elutriation.

PubMed

Banfalvi, Gaspar

2017-01-01

Synchronized populations of large numbers of cells can be obtained by centrifugal elutriation on the basis of sedimentation properties of small round particles, with minimal perturbation of cellular functions. The physical characteristics of cell size and sedimentation velocity are operative in the technique of centrifugal elutriation also known as counterstreaming centrifugation. The elutriator is an advanced device for increasing the sedimentation rate to yield enhanced resolution of cell separation. A random population of cells is introduced into the elutriation chamber of an elutriator rotor running in a specially designed centrifuge. By increasing step-by-step the flow rate of the elutriation fluid, successive populations of relatively homogeneous cell size can be removed from the elutriation chamber and used as synchronized subpopulations. For cell synchronization by centrifugal elutriation, early log S phase cell populations are most suitable where most of the cells are in G1 and S phase (>80 %). Apoptotic cells can be found in the early elutriation fractions belonging to the sub-Go window. Protocols for the synchronization of nuclei of murine pre-B cells and high-resolution centrifugal elutriation of CHO cells are given. The verification of purity and cell cycle positions of cells in elutriated fractions includes the measurement of DNA synthesis by [ 3 H]-thymidine incorporation and DNA content by propidium iodide flow cytometry.

Magnesium and Calcium in Isolated Cell Nuclei

PubMed Central

Naora, H.; Naora, H.; Mirsky, A. E.; Allfrey, V. G.

1961-01-01

The calcium and magnesium contents of thymus nuclei have been determined and the nuclear sites of attachment of these two elements have been studied. The nuclei used for these purposes were isolated in non-aqueous media and in sucrose solutions. Non-aqueous nuclei contain 0.024 per cent calcium and 0.115 per cent magnesium. Calcium and magnesium are held at different sites. The greater part of the magnesium is bound to DNA, probably to its phosphate groups. Evidence is presented that the magnesium atoms combined with the phosphate groups of DNA are also attached to mononucleotides. There is reason to believe that those DNA-phosphate groups to which magnesium is bound, less than 1/10th of the total, are metabolically active, while those to which histones are attached seem to be inactive. PMID:13727745

Crystalline inclusions in the cytoplasm and nuclei of cells of acute myeloid leukaemia.

PubMed

Pearson, E C

1989-01-01

In a survey by electron microscopy of peripheral blood and/or bone marrow from 230 adult patients with acute myeloid leukaemia, five were observed to contain crystalline inclusions in the cytoplasm of the leukaemic cells and a sixth contained crystals in the nuclei. In four cases, two of FAB type M2 and two of M4, the cytoplasmic crystals were hexagonal in section and 1-2 micron long. Two examples showed internal periodicities in the range 3.3-4.0 nm when the electronmicrographs were analysed by optical diffractometry. A single case of M1 contained smaller trapezoidal crystals with a 4.9nm periodicity. The sixth patient, with unusual cytological abnormalities and a rare t(3; 6) chromosomal translocation, contained six-sided crystals in the nuclei of some relatively undifferentiated cells. To the best of our knowledge such intranuclear crystals have not previously been reported in leukaemia. The relevance of the crystals to the leukaemic process is discussed.

Three-Dimensional, Live-Cell Imaging of Chromatin Dynamics in Plant Nuclei Using Chromatin Tagging Systems.

PubMed

Hirakawa, Takeshi; Matsunaga, Sachihiro

2016-01-01

In plants, chromatin dynamics spatiotemporally change in response to various environmental stimuli. However, little is known about chromatin dynamics in the nuclei of plants. Here, we introduce a three-dimensional, live-cell imaging method that can monitor chromatin dynamics in nuclei via a chromatin tagging system that can visualize specific genomic loci in living plant cells. The chromatin tagging system is based on a bacterial operator/repressor system in which the repressor is fused to fluorescent proteins. A recent refinement of promoters for the system solved the problem of gene silencing and abnormal pairing frequencies between operators. Using this system, we can detect the spatiotemporal dynamics of two homologous loci as two fluorescent signals within a nucleus and monitor the distance between homologous loci. These live-cell imaging methods will provide new insights into genome organization, development processes, and subnuclear responses to environmental stimuli in plants.

Production of neutron-rich nuclei approaching r-process by gamma-induced fission of 238U at ELI-NP

NASA Astrophysics Data System (ADS)

Mei, Bo; Balabanski, Dimiter; Constantin, Paul; Anh Le, Tuan; Viet Cuong, Phan

2018-05-01

The investigation of neutron-rich exotic nuclei is crucial not only for nuclear physics but also for nuclear astrophysics. Experimentally, only few neutron-rich nuclei near the stability have been studied, however, most neutron-rich nuclei have not been measured due to their small production cross sections as well as short half-lives. At ELI-NP, gamma beams with high intensities will open new opportunities to investigate very neutron-rich fragments produced by photofission of 238U targets in a gas cell. Based on some simulations, a novel gas cell has been designed to produce, stop and extract 238U photofission fragments. The extraction time and efficiency of photofission fragments have been optimized by using SIMION simulations. According to these simulations, a high extraction efficiency and a short extraction time can be achieved for 238U photofission fragments in the gas cell, which will allow one to measure very neutron-rich fragments with short half-lives by using the IGISOL facility proposed at ELI-NP.

76 FR 63702 - In the Matter of the Designation of Conspiracy of Fire Nuclei, aka Conspiracy of the Nuclei of...

Federal Register 2010, 2011, 2012, 2013, 2014

2011-10-13

... DEPARTMENT OF STATE [Public Notice: 7643] In the Matter of the Designation of Conspiracy of Fire Nuclei, aka Conspiracy of the Nuclei of Fire, aka Conspiracy of Cells of Fire, aka Synomosia of Pyrinon Tis Fotias, aka Thessaloniki-Athens Fire Nuclei Conspiracy, as a Specially Designated Global Terrorist...

[Effect of a glutamate and glutamine excess on the nucleic acid content of the spleen cell nuclei in rats].

PubMed

Vorontsova, E N; Okunev, V N

1976-01-01

In tests conducted with albino rats subject to investigation was the effect of sodium glutamate, or glutamine, daily introduced into the stomach in doses of 300 and 150 mg/kg, on the nucleic acids content in the splenic cell nuclei. All the animals taken in the experiment demonstrated a clearcut quantity of nucleonic RNA. By using a maximum dose of sodium glutamate and minimal one of glutamine a rise in the amount of DNA occurs in the nuclei of the splenic cells.

Development of a stained cell nuclei counting system

NASA Astrophysics Data System (ADS)

Timilsina, Niranjan; Moffatt, Christopher; Okada, Kazunori

2011-03-01

This paper presents a novel cell counting system which exploits the Fast Radial Symmetry Transformation (FRST) algorithm [1]. The driving force behind our system is a research on neurogenesis in the intact nervous system of Manduca Sexta or the Tobacco Hornworm, which was being studied to assess the impact of age, food and environment on neurogenesis. The varying thickness of the intact nervous system in this species often yields images with inhomogeneous background and inconsistencies such as varying illumination, variable contrast, and irregular cell size. For automated counting, such inhomogeneity and inconsistencies must be addressed, which no existing work has done successfully. Thus, our goal is to devise a new cell counting algorithm for the images with non-uniform background. Our solution adapts FRST: a computer vision algorithm which is designed to detect points of interest on circular regions such as human eyes. This algorithm enhances the occurrences of the stained-cell nuclei in 2D digital images and negates the problems caused by their inhomogeneity. Besides FRST, our algorithm employs standard image processing methods, such as mathematical morphology and connected component analysis. We have evaluated the developed cell counting system with fourteen digital images of Tobacco Hornworm's nervous system collected for this study with ground-truth cell counts by biology experts. Experimental results show that our system has a minimum error of 1.41% and mean error of 16.68% which is at least forty-four percent better than the algorithm without FRST.

Analysis of growth of tetraploid nuclei in roots of Vicia faba.

PubMed

Bansal, J; Davidson, D

1978-03-01

Growth of nuclei of a marked population of cells was determined from G1 to prophase in roots of Vicia faba. The cells were marked by inducing them to become tetraploid by treatment with 0.002% colchicine for 1 hr. Variation in nuclear volume is large; it is established in early G1 and maintained through interphase and into prophase. One consequence of this variation is that there is considerable overlap between volumes of nuclei of different ages in the cell cycle; nuclear volume, we suggest, cannot be used as an accurate indicator of the age of the cell in its growth cycle. Nuclei exhibit considerable variation in their growth rate through the cell cycle. Of the marked population of cells, about 65% had completed a cell cycle 14--15 hr after they were formed. These tetraploid nuclei have a cell cycle duration similar to that of fast cycling diploid cells of the same roots. Since they do complete a cell cycle, at least 65% of the nuclei studied must come from rapidly proliferating cells, showing that variability in nuclear volumes must be present in growing cells and cannot be attributed solely to the presence, in our samples, of non-cycling cells.

Searching for non-transposable targets of planarian nuclear PIWI in pluripotent stem cells and differentiated cells.

PubMed

Kashima, Makoto; Agata, Kiyokazu; Shibata, Norito

2018-06-01

Nuclear PIWIs together with their guide RNAs (piRNAs) epigenetically silence various genes including transposons in many organisms. In planarians, the nuclear piwi family gene, DjpiwiB is specifically transcribed in adult pluripotent stem cells (adult PSC, neoblast), but not in differentiated cells. However, the protein accumulates in the nuclei of both neoblasts and their descendant differentiated cells. Interestingly, PIWI(DjPiwiB)-piRNA complexes are indispensable for the repression of transposable genes at the onset of differentiation from neoblasts. Here, we conducted a comparative transcriptome analysis between control and DjpiwiB(RNAi) animals to identify non-transposable target genes of the DjPiwiB-piRNA complexes. Using bioinformatic analyses and RNAi we demonstrate that DjPiwiB-piRNA complexes are required for the proper expression of Djmcm2 and Djhistone h4 in neoblasts and that DjPiwiB-piRNA complexes regulate the transient expression of Djcalu during neoblast differentiation. Thus, DjPiwiB-piRNA complexes regulate the correct expression patterns during neoblast self-renewal and differentiation. © 2018 Japanese Society of Developmental Biologists.

Targeting dendritic cells--why bother?

PubMed

Kreutz, Martin; Tacken, Paul J; Figdor, Carl G

2013-04-11

Vaccination is among the most efficient forms of immunotherapy. Although sometimes inducing lifelong protective B-cell responses, T-cell-mediated immunity remains challenging. Targeting antigen to dendritic cells (DCs) is an extensively explored concept aimed at improving cellular immunity. The identification of various DC subsets with distinct functional characteristics now allows for the fine-tuning of targeting strategies. Although some of these DC subsets are regarded as superior for (cross-) priming of naive T cells, controversies still remain about which subset represents the best target for immunotherapy. Because targeting the antigen alone may not be sufficient to obtain effective T-cell responses, delivery systems have been developed to target multiple vaccine components to DCs. In this Perspective, we discuss the pros and cons of targeting DCs: if targeting is beneficial at all and which vaccine vehicles and immunization routes represent promising strategies to reach and activate DCs.

An "ASYMPTOTIC FRACTAL" Approach to the Morphology of Malignant Cell Nuclei

NASA Astrophysics Data System (ADS)

Landini, Gabriel; Rippin, John W.

To investigate quantitatively nuclear membrane irregularity, 672 nuclei from 10 cases of oral cancer (squamous cell carcinoma) and normal cells from oral mucosa were studied in transmission electron micrographs. The nuclei were photographed at ×1400 magnification and transferred to computer memory (1 pixel = 35 nm). The perimeter of the profiles was analysed using the "yardstick method" of fractal dimension estimation, and the log-log plot of ruler size vs. boundary length demonstrated that there exists a significant effect of resolution on length measurement. However, this effect seems to disappear at higher resolutions. As this observation is compatible with the concept of asymptotic fractal, we estimated the parameters c, L and Bm from the asymptotic fractal formula Br = Bm {1 + (r / L)c}-1 , where Br is the boundary length measured with a ruler of size r, Bm is the maximum boundary for r → 0, L is a constant, and c = asymptotic fractal dimension minus topological dimension (D - Dt) for r → ∞. Analyses of variance showed c to be significantly higher in the normal than malignant cases (P < 0.001), but log(L) and Bm to be significantly higher in the malignant cases (P < 0.001). A multivariate linear discrimination analysis on c, log(L) and Bm re-classified 76.6% of the cells correctly (84.8% of the normal and 67.5% of the tumor). Furthermore, this shows that asymptotic fractal analysis applied to nuclear profiles has great potential for shape quantification in diagnosis of oral cancer.

Flavanol binding of nuclei from tree species.

PubMed

Feucht, W; Treutter, D; Polster, J

2004-01-01

Light microscopy was used to examine the nuclei of five tree species with respect to the presence of flavanols. Flavanols develop a blue colouration in the presence of a special p-dimethylaminocinnamaldehyde (DMACA) reagent that enables those nuclei loaded with flavanols to be recognized. Staining of the nuclei was most pronounced in both Tsuga canadensis and Taxus baccata, variable in Metasequoia glyptostroboides, faint in Coffea arabica and minimal in Prunus avium. HPLC analysis showed that the five species contained substantial amounts of different flavanols such as catechin, epicatechin and proanthocyanidins. Quantitatively, total flavanols were quite different among the species. The nuclei themselves, as studied in Tsuga seed wings, were found to contain mainly catechin, much lower amounts of epicatechin and traces of proanthocyanidins. Blue-coloured nuclei located centrally in small cells were often found to maximally occupy up to 90% of a cell's radius, and the surrounding small rim of cytoplasm was visibly free of flavanols. A survey of 34 gymnosperm and angiosperm species indicated that the first group has much higher nuclear binding capacities for flavanols than the second group.

Cell cycle-regulated proteolysis of mitotic target proteins.

PubMed

Bastians, H; Topper, L M; Gorbsky, G L; Ruderman, J V

1999-11-01

The ubiquitin-dependent proteolysis of mitotic cyclin B, which is catalyzed by the anaphase-promoting complex/cyclosome (APC/C) and ubiquitin-conjugating enzyme H10 (UbcH10), begins around the time of the metaphase-anaphase transition and continues through G1 phase of the next cell cycle. We have used cell-free systems from mammalian somatic cells collected at different cell cycle stages (G0, G1, S, G2, and M) to investigate the regulated degradation of four targets of the mitotic destruction machinery: cyclins A and B, geminin H (an inhibitor of S phase identified in Xenopus), and Cut2p (an inhibitor of anaphase onset identified in fission yeast). All four are degraded by G1 extracts but not by extracts of S phase cells. Maintenance of destruction during G1 requires the activity of a PP2A-like phosphatase. Destruction of each target is dependent on the presence of an N-terminal destruction box motif, is accelerated by additional wild-type UbcH10 and is blocked by dominant negative UbcH10. Destruction of each is terminated by a dominant activity that appears in nuclei near the start of S phase. Previous work indicates that the APC/C-dependent destruction of anaphase inhibitors is activated after chromosome alignment at the metaphase plate. In support of this, we show that addition of dominant negative UbcH10 to G1 extracts blocks destruction of the yeast anaphase inhibitor Cut2p in vitro, and injection of dominant negative UbcH10 blocks anaphase onset in vivo. Finally, we report that injection of dominant negative Ubc3/Cdc34, whose role in G1-S control is well established and has been implicated in kinetochore function during mitosis in yeast, dramatically interferes with congression of chromosomes to the metaphase plate. These results demonstrate that the regulated ubiquitination and destruction of critical mitotic proteins is highly conserved from yeast to humans.

Superheavy nuclei from 48Ca-induced reactions

NASA Astrophysics Data System (ADS)

Oganessian, Yu. Ts.; Utyonkov, V. K.

2015-12-01

The discovery and investigation of the new region of superheavy nuclei at the DGFRS separator based on fusion reactions of 48Ca with 238U-249Cf target nuclei are reviewed. The production cross sections and summaries of the decay properties, including the results of the posterior experiments performed at the SHIP, BGS, and TASCA separators, as well as at the chemistry setups, are discussed and compared with the theoretical calculations and the systematic trends in the α-decay and spontaneous fission properties. The properties of the new nuclei, isotopes of elements 112-118, and their decay products demonstrate significant increases in the stability of the heaviest nuclei with increasing neutron number and closer approach to magic number N = 184.

Fragmentation of relativistic nuclei in peripheral interactions in nuclear track emulsion

DOE Office of Scientific and Technical Information (OSTI.GOV)

Artemenkov, D. A., E-mail: artemenkov@lhe.jinr.ru; Bradnova, V.; Chernyavsky, M. M.

2008-09-15

The technique of nuclear track emulsions is used to explore the fragmentation of light relativistic nuclei down to the most peripheral interactions: nuclear 'white' stars. A complete pattern of the relativistic dissociation of a 8B nucleus with target fragment accompaniment is presented. Relativistic dissociation {sup 9}Be {yields} 2{alpha} is explored using significant statistics, and a relative contribution of {sup 8}Be decays from 0+ and 2+ states is established. Target fragment accompaniments are shown for relativistic fragmentation {sup 14}N {yields} 3He +H and {sup 22}Ne {yields} 5He. The leading role of the electromagnetic dissociation on heavy nuclei with respect to breakupsmore » on target protons is demonstrated in all these cases. It is possible to conclude that the peripheral dissociation of relativistic nuclei in nuclear track emulsion is a unique tool to study many-body systems composed of the lightest nuclei and nucleons in the energy scale relevant for nuclear astrophysics.« less

Automatic extraction of nuclei centroids of mouse embryonic cells from fluorescence microscopy images.

PubMed

Bashar, Md Khayrul; Komatsu, Koji; Fujimori, Toshihiko; Kobayashi, Tetsuya J

2012-01-01

Accurate identification of cell nuclei and their tracking using three dimensional (3D) microscopic images is a demanding task in many biological studies. Manual identification of nuclei centroids from images is an error-prone task, sometimes impossible to accomplish due to low contrast and the presence of noise. Nonetheless, only a few methods are available for 3D bioimaging applications, which sharply contrast with 2D analysis, where many methods already exist. In addition, most methods essentially adopt segmentation for which a reliable solution is still unknown, especially for 3D bio-images having juxtaposed cells. In this work, we propose a new method that can directly extract nuclei centroids from fluorescence microscopy images. This method involves three steps: (i) Pre-processing, (ii) Local enhancement, and (iii) Centroid extraction. The first step includes two variations: first variation (Variant-1) uses the whole 3D pre-processed image, whereas the second one (Variant-2) modifies the preprocessed image to the candidate regions or the candidate hybrid image for further processing. At the second step, a multiscale cube filtering is employed in order to locally enhance the pre-processed image. Centroid extraction in the third step consists of three stages. In Stage-1, we compute a local characteristic ratio at every voxel and extract local maxima regions as candidate centroids using a ratio threshold. Stage-2 processing removes spurious centroids from Stage-1 results by analyzing shapes of intensity profiles from the enhanced image. An iterative procedure based on the nearest neighborhood principle is then proposed to combine if there are fragmented nuclei. Both qualitative and quantitative analyses on a set of 100 images of 3D mouse embryo are performed. Investigations reveal a promising achievement of the technique presented in terms of average sensitivity and precision (i.e., 88.04% and 91.30% for Variant-1; 86.19% and 95.00% for Variant-2), when compared

Natural Killer Cell Immunotherapy Targeting Cancer Stem Cells

PubMed Central

Luna, Jesus I; Grossenbacher, Steven K.; Murphy, William J; Canter, Robert J

2017-01-01

Introduction Standard cytoreductive cancer therapy, such as chemotherapy and radiotherapy, are frequently resisted by a small portion of cancer cells with “stem-cell” like properties including quiescence and repopulation. Immunotherapy represents a breakthrough modality for improving oncologic outcomes in cancer patients. Since the success of immunotherapy is not contingent on target cell proliferation, it may also be uniquely suited to address the problem of resistance and repopulation exerted by cancer stem cells (CSCs). Areas covered Natural killer (NK) cells have long been known for their ability to reject allogeneic hematopoietic stem cells, and there are increasing data demonstrating that NK cells can selectively identify and lyse CSCs. In this report, we review the current knowledge of CSCs and NK cells and highlight recent studies that support the concept that NK cells are capable of targeting CSC in solid tumors, especially in the context of combination therapy simultaneously targeting non-CSCs and CSCs. Expert Opinion Unlike cytotoxic cancer treatments, NK cells are able to target and eliminate quiescent/non-proliferating cells such as CSCs, and these enigmatic cells are an important source of relapse and metastasis. NK targeting of CSCs represents a novel and potentially high impact method to capitalize on the intrinsic therapeutic potential of NK cells. PMID:27960589

Search for α -Cluster Structure in Exotic Nuclei with the Prototype Active-Target Time-Projection Chamber

NASA Astrophysics Data System (ADS)

Fritsch, A.; Ayyad, Y.; Bazin, D.; Beceiro-Novo, S.; Bradt, J.; Carpenter, L.; Cortesi, M.; Mittig, W.; Suzuki, D.; Ahn, T.; Kolata, J. J.; Howard, A. M.; Becchetti, F. D.; Wolff, M.

Some exotic nuclei appear to exhibit α -cluster structure, which may impact nucleosynthesis reaction rates. While various theoretical models currently describe such clustering, more experimental data are needed to constrain model predictions. The Prototype Active-Target Time-Projection Chamber (PAT-TPC) has low-energy thresholds for charged-particle decay and a high detection efficiency due to its thick gaseous active target volume, making it well-suited to search for low-energy α -cluster reactions. Radioactive-ion beams produced by the TwinSol facility at the University of Notre Dame were delivered to the PAT-TPC to study 14C via α -resonant scattering. Differential cross sections and excitation functions were measured and show evidence of three-body exit channels. Additional data were measured with an updated Micromegas detector more sensitive to three-body decay. Preliminary results are presented.

Plant nuclei can contain extensive grooves and invaginations

NASA Technical Reports Server (NTRS)

Collings, D. A.; Carter, C. N.; Rink, J. C.; Scott, A. C.; Wyatt, S. E.; Allen, N. S.; Brown, C. S. (Principal Investigator)

2000-01-01

Plant cells can exhibit highly complex nuclear organization. Through dye-labeling experiments in untransformed onion epidermal and tobacco culture cells and through the expression of green fluorescent protein targeted to either the nucleus or the lumen of the endoplasmic reticulum/nuclear envelope in these cells, we have visualized deep grooves and invaginations into the large nuclei of these cells. In onion, these structures, which are similar to invaginations seen in some animal cells, form tubular or planelike infoldings of the nuclear envelope. Both grooves and invaginations are stable structures, and both have cytoplasmic cores containing actin bundles that can support cytoplasmic streaming. In dividing tobacco cells, invaginations seem to form during cell division, possibly from strands of the endoplasmic reticulum trapped in the reforming nucleus. The substantial increase in nuclear surface area resulting from these grooves and invaginations, their apparent preference for association with nucleoli, and the presence in them of actin bundles that support vesicle motility suggest that the structures might function both in mRNA export from the nucleus and in protein import from the cytoplasm to the nucleus.

Over-expression of GFP-FEZ1 causes generation of multi-lobulated nuclei mediated by microtubules in HEK293 cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lanza, Daniel C.F.; Trindade, Daniel M.; Instituto de Biologia, Universidade Estadual de Campinas, Campinas, SP

2008-06-10

FEZ1 (Fasciculation and elongation protein zeta 1) is an ortholog of the Caenorhabditis elegans protein UNC-76, involved in neuronal development and axon outgrowth, in that worm. Mammalian FEZ1 has already been reported to cooperate with PKC-zeta in the differentiation and polarization of PC12 neuronal cells. Furthermore, FEZ1 is associated with kinesin 1 and JIP1 to form a cargo-complex responsible for microtubule based transport of mitochondria along axons. FEZ1 can also be classified as a hub protein, since it was reported to interact with over 40 different proteins in yeast two-hybrid screens, including at least nine nuclear proteins. Here, we transientlymore » over-expressed GFP-FEZ1full in human HEK293 and HeLa cells in order to study the sub-cellular localization of GFP-FEZ1. We observed that over 40% of transiently transfected cells at 3 days post-transfection develop multi-lobulated nuclei, which are also called flower-like nuclei. We further demonstrated that GFP-FEZ1 localizes either to the cytoplasm or the nuclear fraction, and that the appearance of the flower-like nuclei depends on intact microtubule function. Finally, we show that FEZ1 co-localizes with both, {alpha}- and especially with {gamma}-tubulin, which localizes as a centrosome like structure at the center of the multiple lobules. In summary, our data suggest that FEZ1 has an important centrosomal function and supply new mechanistic insights to the formation of flower-like nuclei, which are a phenotypical hallmark of human leukemia cells.« less

A unique combination of anatomy and physiology in cells of the rat paralaminar thalamic nuclei adjacent to the medial geniculate body

PubMed Central

Smith, Philip H.; Bartlett, Edward L.; Kowalkowski, Anna

2010-01-01

The medial geniculate body (MGB) has three major subdivisions - ventral (MGV), dorsal (MGD) and medial (MGM). MGM is linked with paralaminar nuclei that are situated medial and ventral to MGV/MGD. Paralaminar nuclei have unique inputs and outputs when compared with MGV and MGD and have been linked to circuitry underlying some important functional roles. We recorded intracellularly from cells in the paralaminar nuclei in vitro. We found that they possess an unusual combination of anatomical and physiological features when compared to those reported for “standard” thalamic neurons seen in the MGV/MGD and elsewhere in the thalamus. Compared to MGV/MGD neurons, anatomically, 1) paralaminar cell dendrites can be long, branch sparingly and encompass a much larger area. 2) their dendrites may be smooth but can have well defined spines and 3) their axons can have collaterals that branch locally within the same or nearby paralaminar nuclei. When compared to MGV/MGD neurons physiologically 1) their spikes are larger in amplitude and can be shorter in duration and 2) can have dual afterhyperpolarizations with fast and slow components and 3) they can have a reduction or complete absence of the low threshold, voltage-sensitive calcium conductance that reduces or eliminates the voltage-dependent burst response. We also recorded from cells in the parafascicular nucleus, a nucleus of the posterior intralaminar nuclear group, because they have unusual anatomical features that are similar to some of our paralaminar cells. Like the labeled paralaminar cells, parafascicular cells had physiological features distinguishing them from typical thalamic neurons. PMID:16566009

Intralaminar nuclei of the thalamus in Lewy body diseases.

PubMed

Brooks, Daniel; Halliday, Glenda M

2009-02-16

Although the intralaminar thalamus is a target of alpha-synuclein pathology in Parkinson's disease, the degree of neuronal loss in Lewy body diseases has not been assessed. We have used unbiased stereological techniques to quantify neuronal loss in intralaminar thalamic nuclei concentrating alpha-synuclein pathology (the anterodorsal, cucullar, parataenial, paraventricular, central medial, central lateral and centre-median/parafascicular complex) in different clinical forms of Lewy body disease (Parkinson's disease with and without dementia, and dementia with Lewy bodies, N=21) compared with controls (N=5). Associations were performed in the Lewy body cases between intralaminar cell loss and the main diagnostic clinical (parkinsonism, dementia, fluctuation in consciousness, and visual hallucinations) and pathological (Braak stage of Parkinson's disease) features of these diseases, as well as between cell loss and the scaled severity of the alpha-synuclein deposition within the intralaminar thalamus. As expected, significant alpha-synuclein accumulation occurred in the intralaminar thalamus in the cases with Lewy body disease. Pathology concentrated anteriorly and in the central lateral and paraventricular nuclei was related to the Braak stage of Parkinson's disease, ageing, and the presence of dementia. Across all types of Lewy body cases there was substantial atrophy and neuronal loss in the central lateral, cucullar and parataenial nuclei, and neuronal loss without atrophy in the centre-median/parafascicular complex. Cases with visual hallucinations showed a greater degree of atrophy of the cucullar nucleus, possibly due to amygdala denervation. The significant degeneration demonstrated in the intralaminar thalamus is likely to contribute to the movement and cognitive dysfunction observed in Lewy body disorders.

Calculation of (n,α) reaction cross sections by using some Skyrme force parameters for Potassium (41K) target nuclei

NASA Astrophysics Data System (ADS)

Tel, Eyyup; Sahan, Muhittin; Alkanli, Hasancan; Sahan, Halide; Yigit, Mustafa

2017-09-01

In this study, the (n,α) nuclear reaction cross section was calculated for 41K target nuclei for neutron and proton density parameters using SKa, SKb, SLy5, and SLy6 Skyrme force. Theoretical cross section for the (n,α) nuclear reaction was obtained using a formula constituted by Tel et al. (2008). Results are compared with experimental data from EXFOR. The calculated results from formula was found in a close agreement with experimental data.

Cooperative tumour cell membrane targeted phototherapy

NASA Astrophysics Data System (ADS)

Kim, Heegon; Lee, Junsung; Oh, Chanhee; Park, Ji-Ho

2017-06-01

The targeted delivery of therapeutics using antibodies or nanomaterials has improved the precision and safety of cancer therapy. However, the paucity and heterogeneity of identified molecular targets within tumours have resulted in poor and uneven distribution of targeted agents, thus compromising treatment outcomes. Here, we construct a cooperative targeting system in which synthetic and biological nanocomponents participate together in the tumour cell membrane-selective localization of synthetic receptor-lipid conjugates (SR-lipids) to amplify the subsequent targeting of therapeutics. The SR-lipids are first delivered selectively to tumour cell membranes in the perivascular region using fusogenic liposomes. By hitchhiking with extracellular vesicles secreted by the cells, the SR-lipids are transferred to neighbouring cells and further spread throughout the tumour tissues where the molecular targets are limited. We show that this tumour cell membrane-targeted delivery of SR-lipids leads to uniform distribution and enhanced phototherapeutic efficacy of the targeted photosensitizer.

Release of specific proteins from nuclei of HL-60 and MOLT-4 cells by antitumor drugs having affinity to nucleic acids.

PubMed

Lassota, P; Melamed, M R; Darzynkiewicz, Z

The binding sites for mitoxantrone (MIT), Ametantrone (AMT), doxorubicin (DOX), actinomycin D (AMD) and ethidium bromide (EB) in nuclei from exponentially growing and differentiating human promyelocytic HL-60 and lymphocytic leukemic MOLT-4 cells were studied by gel electrophoresis of proteins selectively released during titration of these nuclei with the drugs. Each drug at different drug: DNA binding ratios resulted in a characteristic pattern of protein elution and/or retention. For example, in nuclei from exponentially growing HL-60 cells, MIT affected 44 nuclear proteins that were different from those affected by EB; of these 29 were progressively released at increasing MIT:DNA ratios, 11 were transiently released (i.e. only at a low MIT:DNA ratio) and 4 entrapped. Patterns of proteins displaced from nuclei of exponentially growing HL-60 cells differed from those of cells undergoing myeloid differentiation as well as from those of exponentially growing MOLT-4 cells. The first effects were seen at a binding density of approximately one drug molecule per 10-50 base pairs of DNA. The observed selective displacement of proteins may reflect drug-altered affinity of the binding sites for those proteins, for example due to a change of nucleic acid or protein conformation upon binding the ligand. The data show that the binding site(s) for each of the ligands studied is different and the differences correlate with variability in chemical structure between the ligands. The nature of the drug-affected proteins may provide clues regarding antitumor or cytotoxic mechanisms of drug action.

A cell-targeted, size-photocontrollable, nuclear-uptake nanodrug delivery system for drug-resistant cancer therapy.

PubMed

Qiu, Liping; Chen, Tao; Öçsoy, Ismail; Yasun, Emir; Wu, Cuichen; Zhu, Guizhi; You, Mingxu; Han, Da; Jiang, Jianhui; Yu, Ruqin; Tan, Weihong

2015-01-14

The development of multidrug resistance (MDR) has become an increasingly serious problem in cancer therapy. The cell-membrane overexpression of P-glycoprotein (P-gp), which can actively efflux various anticancer drugs from the cell, is a major mechanism of MDR. Nuclear-uptake nanodrug delivery systems, which enable intranuclear release of anticancer drugs, are expected to address this challenge by bypassing P-gp. However, before entering the nucleus, the nanocarrier must pass through the cell membrane, necessitating coordination between intracellular and intranuclear delivery. To accommodate this requirement, we have used DNA self-assembly to develop a nuclear-uptake nanodrug system carried by a cell-targeted near-infrared (NIR)-responsive nanotruck for drug-resistant cancer therapy. Via DNA hybridization, small drug-loaded gold nanoparticles (termed nanodrugs) can self-assemble onto the side face of a silver-gold nanorod (NR, termed nanotruck) whose end faces were modified with a cell type-specific internalizing aptamer. By using this size-photocontrollable nanodrug delivery system, anticancer drugs can be efficiently accumulated in the nuclei to effectively kill the cancer cells.

Structure of interphase chromosomes in the nuclei of Drosophila cells.

PubMed

Banfalvi, Gaspar

2006-10-01

Fluorescent images of interphase chromatin structures and chromosome structures isolated from reversibly permeable Drosophila cells were analyzed. Decondensed chromatin in early S phase (2.0-2.5 C-value) consisted of a veil-like fibrillary network. Fibrillar chromatin formed rodlets later in the early S phase (2.5-2.75 C). Drosophila chromosomes contain several smaller subunits called rodlets. Fibrillar chromatin turned to chromatin ribbon and the early mid-S-phase globular chromosomes (2.75-3.0 C), then to opened fibrous globular forms later in the mid-S-phase (3.0-3.25 C), to late-S-phase supercoiled ribbons (3.25-3.5 C), end-S-phase elongated prechromosomes (3.5-3.75 C), bent and linear chromosomes (3.75-4.0 C). Early-S phase chromatin fibrils in the nuclei of Drosophila cells are thinner than the veil-like structures in mammalian cells. The connectivity of chromosomes shows linear arrangement (3, 1, 2, 4), with larger chromosomes (1 and 2) inside and smaller chromosomes (3, 4) at the two ends in the chromosomal chain.

Systematic Morphometry of Catecholamine Nuclei in the Brainstem.

PubMed

Bucci, Domenico; Busceti, Carla L; Calierno, Maria T; Di Pietro, Paola; Madonna, Michele; Biagioni, Francesca; Ryskalin, Larisa; Limanaqi, Fiona; Nicoletti, Ferdinando; Fornai, Francesco

2017-01-01

Catecholamine nuclei within the brainstem reticular formation (RF) play a pivotal role in a variety of brain functions. However, a systematic characterization of these nuclei in the very same experimental conditions is missing so far. Tyrosine hydroxylase (TH) immune-positive cells of the brainstem correspond to dopamine (DA)-, norepinephrine (NE)-, and epinephrine (E)-containing cells. Here, we report a systematic count of TH-positive neurons in the RF of the mouse brainstem by using stereological morphometry. All these nuclei were analyzed for anatomical localization, rostro-caudal extension, volume, neuron number, neuron density, and mean neuronal area for each nucleus. The present data apart from inherent informative value wish to represent a reference for neuronal mapping in those studies investigating the functional anatomy of the brainstem RF. These include: the sleep-wake cycle, movement control, muscle tone modulation, mood control, novelty orienting stimuli, attention, archaic responses to internal and external stressful stimuli, anxiety, breathing, blood pressure, and innumerable activities modulated by the archaic iso-dendritic hard core of the brainstem RF. Most TH-immune-positive cells fill the lateral part of the RF, which indeed possesses a high catecholamine content. A few nuclei are medial, although conventional nosography considers all these nuclei as part of the lateral column of the RF. Despite the key role of these nuclei in psychiatric and neurological disorders, only a few of them aspired a great attention in biomedical investigation, while most of them remain largely obscure although intense research is currently in progress. A simultaneous description of all these nuclei is not simply key to comprehend the variety of brainstem catecholamine reticular neurons, but probably represents an intrinsically key base for understanding brain physiology and physiopathology.

Systematic Morphometry of Catecholamine Nuclei in the Brainstem

PubMed Central

Bucci, Domenico; Busceti, Carla L.; Calierno, Maria T.; Di Pietro, Paola; Madonna, Michele; Biagioni, Francesca; Ryskalin, Larisa; Limanaqi, Fiona; Nicoletti, Ferdinando; Fornai, Francesco

2017-01-01

Catecholamine nuclei within the brainstem reticular formation (RF) play a pivotal role in a variety of brain functions. However, a systematic characterization of these nuclei in the very same experimental conditions is missing so far. Tyrosine hydroxylase (TH) immune-positive cells of the brainstem correspond to dopamine (DA)-, norepinephrine (NE)-, and epinephrine (E)-containing cells. Here, we report a systematic count of TH-positive neurons in the RF of the mouse brainstem by using stereological morphometry. All these nuclei were analyzed for anatomical localization, rostro-caudal extension, volume, neuron number, neuron density, and mean neuronal area for each nucleus. The present data apart from inherent informative value wish to represent a reference for neuronal mapping in those studies investigating the functional anatomy of the brainstem RF. These include: the sleep-wake cycle, movement control, muscle tone modulation, mood control, novelty orienting stimuli, attention, archaic responses to internal and external stressful stimuli, anxiety, breathing, blood pressure, and innumerable activities modulated by the archaic iso-dendritic hard core of the brainstem RF. Most TH-immune-positive cells fill the lateral part of the RF, which indeed possesses a high catecholamine content. A few nuclei are medial, although conventional nosography considers all these nuclei as part of the lateral column of the RF. Despite the key role of these nuclei in psychiatric and neurological disorders, only a few of them aspired a great attention in biomedical investigation, while most of them remain largely obscure although intense research is currently in progress. A simultaneous description of all these nuclei is not simply key to comprehend the variety of brainstem catecholamine reticular neurons, but probably represents an intrinsically key base for understanding brain physiology and physiopathology. PMID:29163071

Cell-Type-Specific Modulation of Sensory Responses in Olfactory Bulb Circuits by Serotonergic Projections from the Raphe Nuclei

PubMed Central

Brunert, Daniela; Tsuno, Yusuke; Rothermel, Markus; Shipley, Michael T.

2016-01-01

Serotonergic neurons in the brainstem raphe nuclei densely innervate the olfactory bulb (OB), where they can modulate the initial representation and processing of olfactory information. Serotonergic modulation of sensory responses among defined OB cell types is poorly characterized in vivo. Here, we used cell-type-specific expression of optical reporters to visualize how raphe stimulation alters sensory responses in two classes of GABAergic neurons of the mouse OB glomerular layer, periglomerular (PG) and short axon (SA) cells, as well as mitral/tufted (MT) cells carrying OB output to piriform cortex. In PG and SA cells, brief (1–4 s) raphe stimulation elicited a large increase in the magnitude of responses linked to inhalation of ambient air, as well as modest increases in the magnitude of odorant-evoked responses. Near-identical effects were observed when the optical reporter of glutamatergic transmission iGluSnFR was expressed in PG and SA cells, suggesting enhanced excitatory input to these neurons. In contrast, in MT cells imaged from the dorsal OB, raphe stimulation elicited a strong increase in resting GCaMP fluorescence with only a slight enhancement of inhalation-linked responses to odorant. Finally, optogenetically stimulating raphe serotonergic afferents in the OB had heterogeneous effects on presumptive MT cells recorded extracellularly, with an overall modest increase in resting and odorant-evoked responses during serotonergic afferent stimulation. These results suggest that serotonergic afferents from raphe dynamically modulate olfactory processing through distinct effects on multiple OB targets, and may alter the degree to which OB output is shaped by inhibition during behavior. SIGNIFICANCE STATEMENT Modulation of the circuits that process sensory information can profoundly impact how information about the external world is represented and perceived. This study investigates how the serotonergic system modulates the initial processing of olfactory

Targeted Knock-Down of miR21 Primary Transcripts Using snoMEN Vectors Induces Apoptosis in Human Cancer Cell Lines.

PubMed

Ono, Motoharu; Yamada, Kayo; Avolio, Fabio; Afzal, Vackar; Bensaddek, Dalila; Lamond, Angus I

2015-01-01

We have previously reported an antisense technology, 'snoMEN vectors', for targeted knock-down of protein coding mRNAs using human snoRNAs manipulated to contain short regions of sequence complementarity with the mRNA target. Here we characterise the use of snoMEN vectors to target the knock-down of micro RNA primary transcripts. We document the specific knock-down of miR21 in HeLa cells using plasmid vectors expressing miR21-targeted snoMEN RNAs and show this induces apoptosis. Knock-down is dependent on the presence of complementary sequences in the snoMEN vector and the induction of apoptosis can be suppressed by over-expression of miR21. Furthermore, we have also developed lentiviral vectors for delivery of snoMEN RNAs and show this increases the efficiency of vector transduction in many human cell lines that are difficult to transfect with plasmid vectors. Transduction of lentiviral vectors expressing snoMEN targeted to pri-miR21 induces apoptosis in human lung adenocarcinoma cells, which express high levels of miR21, but not in human primary cells. We show that snoMEN-mediated suppression of miRNA expression is prevented by siRNA knock-down of Ago2, but not by knock-down of Ago1 or Upf1. snoMEN RNAs colocalise with Ago2 in cell nuclei and nucleoli and can be co-immunoprecipitated from nuclear extracts by antibodies specific for Ago2.

Condensin I and II behaviour in interphase nuclei and cells undergoing premature chromosome condensation.

PubMed

Zhang, Tao; Paulson, James R; Bakhrebah, Muhammed; Kim, Ji Hun; Nowell, Cameron; Kalitsis, Paul; Hudson, Damien F

2016-05-01

Condensin is an integral component of the mitotic chromosome condensation machinery, which ensures orderly segregation of chromosomes during cell division. In metazoans, condensin exists as two complexes, condensin I and II. It is not yet clear what roles these complexes may play outside mitosis, and so we have examined their behaviour both in normal interphase and in premature chromosome condensation (PCC). We find that a small fraction of condensin I is retained in interphase nuclei, and our data suggests that this interphase nuclear condensin I is active in both gene regulation and chromosome condensation. Furthermore, live cell imaging demonstrates condensin II dramatically increases on G1 nuclei following completion of mitosis. Our PCC studies show condensins I and II and topoisomerase II localise to the chromosome axis in G1-PCC and G2/M-PCC, while KIF4 binding is altered. Individually, condensins I and II are dispensable for PCC. However, when both are knocked out, G1-PCC chromatids are less well structured. Our results define new roles for the condensins during interphase and provide new information about the mechanism of PCC.

Neuronal nuclei isolation from human postmortem brain tissue.

PubMed

Matevossian, Anouch; Akbarian, Schahram

2008-10-01

Neurons in the human brain become postmitotic largely during prenatal development, and thus maintain their nuclei throughout the full lifespan. However, little is known about changes in neuronal chromatin and nuclear organization during the course of development and aging, or in chronic neuropsychiatric disease. However, to date most chromatin and DNA based assays (other than FISH) lack single cell resolution. To this end, the considerable cellular heterogeneity of brain tissue poses a significant limitation, because typically various subpopulations of neurons are intermingled with different types of glia and other non-neuronal cells. One possible solution would be to grow cell-type specific cultures, but most CNS cells, including neurons, are ex vivo sustainable, at best, for only a few weeks and thus would provide an incomplete model for epigenetic mechanisms potentially operating across the full lifespan. Here, we provide a protocol to extract and purify nuclei from frozen (never fixed) human postmortem brain. The method involves extraction of nuclei in hypotonic lysis buffer, followed by ultracentrifugation and immunotagging with anti-NeuN antibody. Labeled neuronal nuclei are then collected separately using fluorescence-activated sorting. This method should be applicable to any brain region in a wide range of species and suitable for chromatin immunoprecipitation studies with site- and modification-specific anti-histone antibodies, and for DNA methylation and other assays.

Age-related retention of fiber cell nuclei and nuclear fragments in the lens cortices of multiple species

PubMed Central

Pendergrass, William; Zitnik, Galynn; Urfer, Silvan R.

2011-01-01

Purpose To determine the differences between species in the retention of lens fiber cell nuclei and nuclear fragments in the aging lens cortex and the relationship of nuclear retention to lens opacity. For this purpose old human, monkey, dog, and rat lenses were compared to those of three strains of mouse. We also investigated possible mechanisms leading to nuclear retention. Methods Fixed specimens of the species referred to above were obtained from immediate on site sacrifice of mice and rats, or from recently fixed lenses of other species, dogs, monkeys, and humans, obtained from collaborators. The retention of undegraded nuclei and nuclear fragments was graded 1–4 from histologic observation. All species lenses were examined microscopically in fixed sections stained with hematoxylin and eosin (H&E) or 4',6-diamidino-2-phenylindole (DAPI). Slit lamp observations were made only on the mice and rats before sacrifice and lens fixation. Values of 0 to 4 (clear lens to cataract) were given to degree of opacity. MRNA content in young versus old C57BL/6 mouse lenses was determined by quantitative PCR (qPCR) for DNase II-like acid DNase (DLAD) and other proteins. DLAD protein was determined by immunofluorescence of fixed eye sections. Results In old C57BL/6 and DBA mice and, to a lesser degree, in old CBA mice and old Brown Norway (BN) rats lenses were seen to contain a greatly expanded pool of unresolved whole nuclei or fragments of nuclei in differentiating lens fiber cells. This generally correlated with increased slit lamp opacities in these mice. Most old dog lenses also had an increase in retained cortical nuclei, as did a few old humans. However, a second rat strain, BNF1, in which opacity was quite high had no increase in retained nuclei with age nor did any of the old monkeys, indicating that retained nuclei could not be a cause of opacity in these animals. The nuclei and nuclear fragments were located at all levels in the outer cortex extending inward from

Targeting B Cells and Plasma Cells in Autoimmune Diseases

PubMed Central

Hofmann, Katharina; Clauder, Ann-Katrin; Manz, Rudolf Armin

2018-01-01

Success with B cell depletion using rituximab has proven the concept that B lineage cells represent a valid target for the treatment of autoimmune diseases, and has promoted the development of other B cell targeting agents. Present data confirm that B cell depletion is beneficial in various autoimmune disorders and also show that it can worsen the disease course in some patients. These findings suggest that B lineage cells not only produce pathogenic autoantibodies, but also significantly contribute to the regulation of inflammation. In this review, we will discuss the multiple pro- and anti-inflammatory roles of B lineage cells play in autoimmune diseases, in the context of recent findings using B lineage targeting therapies. PMID:29740441

Distribution of zebrin-immunoreactive Purkinje cell terminals in the cerebellar and vestibular nuclei of birds.

PubMed

Wylie, Douglas R; Pakan, Janelle M P; Huynh, Hang; Graham, David J; Iwaniuk, Andrew N

2012-05-01

Zebrin II (aldolase C) is expressed in a subset of Purkinje cells in the mammalian and avian cerebella such that there is a characteristic parasagittal organization of zebrin-immunopositive stripes alternating with zebrin-immunonegative stripes. Zebrin is expressed not only in the soma and dendrites of Purkinje cells but also in their axonal terminals. Here we describe the distribution of zebrin immunoreactivity in both the vestibular and the cerebellar nuclei of pigeons (Columba livia) and hummingbirds (Calypte anna, Selasphorus rufus). In the medial cerebellar nucleus, zebrin-positive labeling was particularly heavy in the “shell,” whereas the “core” was zebrin negative. In the lateral cerebellar nucleus, labeling was not as heavy, but a positive shell and negative core were also observed. In the vestibular nuclear complex, zebrin-positive terminal labeling was heavy in the dorsolateral vestibular nucleus and the lateral margin of the superior vestibular nucleus. The central and medial regions of the superior nucleus were generally zebrin negative. Labeling was moderate to heavy in the medial vestibular nucleus, particulary the rostral half of the parvocellular subnucleus. A moderate amount of zebrin-positive labeling was present in the descending vestibular nucleus: this was heaviest laterally, and the central region was generally zebrin negative. Zebrin-positive terminals were also observed in the the cerebellovestibular process, prepositus hypoglossi, and lateral tangential nucleus. We discuss our findings in light of similar studies in rats and with respect to the corticonuclear projections to the cerebellar nuclei and the functional connections of the vestibulocerebellum with the vestibular nuclei. Copyright © 2011 Wiley Periodicals, Inc.

Neutron-rich nuclei produced at zero degrees in damped collisions induced by a beam of 18O on a 238U target

NASA Astrophysics Data System (ADS)

Stefan, I.; Fornal, B.; Leoni, S.; Azaiez, F.; Portail, C.; Thomas, J. C.; Karpov, A. V.; Ackermann, D.; Bednarczyk, P.; Blumenfeld, Y.; Calinescu, S.; Chbihi, A.; Ciemala, M.; Cieplicka-Oryńczak, N.; Crespi, F. C. L.; Franchoo, S.; Hammache, F.; Iskra, Ł. W.; Jacquot, B.; Janssens, R. V. F.; Kamalou, O.; Lauritsen, T.; Lewitowicz, M.; Olivier, L.; Lukyanov, S. M.; Maccormick, M.; Maj, A.; Marini, P.; Matea, I.; Naumenko, M. A.; de Oliveira Santos, F.; Petrone, C.; Penionzhkevich, Yu. E.; Rotaru, F.; Savajols, H.; Sorlin, O.; Stanoiu, M.; Szpak, B.; Tarasov, O. B.; Verney, D.

2018-04-01

Cross sections and corresponding momentum distributions have been measured for the first time at zero degrees for the exotic nuclei obtained from a beam of 18O at 8.5 MeV/A impinging on a 1 mg/cm2238U target. Sizable cross sections were found for the production of exotic species arising from the neutron transfer and proton removal from the projectile. Comparisons of experimental results with calculations based on deep-inelastic reaction models, taking into account the particle evaporation process, indicate that zero degree is a scattering angle at which the differential reaction cross section for production of exotic nuclei is at its maximum. This result is important in view of the new generation of zero degrees spectrometers under construction, such as the S3 separator at GANIL, for example.

Plant Nuclei Can Contain Extensive Grooves and InvaginationsW⃞W⃞

PubMed Central

Collings, David A.; Carter, Crystal N.; Rink, Jochen C.; Scott, Amie C.; Wyatt, Sarah E.; Allen, Nina Strömgren

2000-01-01

Plant cells can exhibit highly complex nuclear organization. Through dye-labeling experiments in untransformed onion epidermal and tobacco culture cells and through the expression of green fluorescent protein targeted to either the nucleus or the lumen of the endoplasmic reticulum/nuclear envelope in these cells, we have visualized deep grooves and invaginations into the large nuclei of these cells. In onion, these structures, which are similar to invaginations seen in some animal cells, form tubular or planelike infoldings of the nuclear envelope. Both grooves and invaginations are stable structures, and both have cytoplasmic cores containing actin bundles that can support cytoplasmic streaming. In dividing tobacco cells, invaginations seem to form during cell division, possibly from strands of the endoplasmic reticulum trapped in the reforming nucleus. The substantial increase in nuclear surface area resulting from these grooves and invaginations, their apparent preference for association with nucleoli, and the presence in them of actin bundles that support vesicle motility suggest that the structures might function both in mRNA export from the nucleus and in protein import from the cytoplasm to the nucleus. PMID:11148288

The complex pericentriolar material 1 protein allows differentiation between myonuclei and nuclei of satellite cells of the skeletal muscle.

PubMed

Brunn, Anna

2018-05-27

The original article by Winje et al., entitled "Specific labelling of myonuclei by an antibody against pericentriolar material 1 (PCM1) on skeletal muscle tissue sections" 1 , sheds new light on the issue of heterogeneity of skeletal muscle and, thus, the problem to reliably distinguish between myonuclei versus nuclei of satellite cells of the skeletal muscle which are intimately associated. At the light microscopical level this differentiation is particularly difficult since only nuclei inside the muscle fiber are defined as true myonuclei. This is a major problem in analyses that use tissue homogenates, while in situ immunohistochemical studies using appropriate antibodies usually allow differentiation of cell populations. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

DNA methylation profiles of donor nuclei cells and tissues of cloned bovine fetuses.

PubMed

Kremenskoy, Maksym; Kremenska, Yuliya; Suzuki, Masako; Imai, Kei; Takahashi, Seiya; Hashizume, Kazuyoshi; Yagi, Shintaro; Shiota, Kunio

2006-04-01

Methylation of DNA in CpG islands plays an important role during fetal development and differentiation because CpG islands are preferentially located in upstream regions of mammalian genomic DNA, including the transcription start site of housekeeping genes and are also associated with tissue-specific genes. Somatic nuclear transfer (NT) technology has been used to generate live clones in numerous mammalian species, but only a low percentage of nuclear transferred animals develop to term. Abnormal epigenetic changes in the CpG islands of donor nuclei after nuclear transfer could contribute to a high rate of abortion during early gestation and increase perinatal death. These changes have yet to be explored. Thus, we investigated the genome-wide DNA methylation profiles of CpG islands in nuclei donor cells and NT animals. Using Restriction Landmark Genomic Scanning (RLGS), we showed, for the first time, the epigenetic profile formation of tissues from NT bovine fetuses produced from cumulus cells. From approximately 2600 unmethylated NotI sites visualized on the RLGS profile, at least 35 NotI sites showed different methylation statuses. Moreover, we proved that fetal and placental tissues from artificially inseminated and cloned cattle have tissue-specific differences in the genome-wide methylation profiles of the CpG islands. We also found that possible abnormalities occurred in the fetal brain and placental tissues of cloned animals.

In situ surface-enhanced Raman scattering spectroscopy exploring molecular changes of drug-treated cancer cell nucleus.

PubMed

Liang, Lijia; Huang, Dianshuai; Wang, Hailong; Li, Haibo; Xu, Shuping; Chang, Yixin; Li, Hui; Yang, Ying-Wei; Liang, Chongyang; Xu, Weiqing

2015-02-17

Investigating the molecular changes of cancer cell nucleus with drugs treatment is crucial for the design of new anticancer drugs, the development of novel diagnostic strategies, and the advancement of cancer therapy efficiency. In order to better understand the action effects of drugs, accurate location and in situ acquisition of the molecular information of the cell nuclei are necessary. In this work, we report a microspectroscopic technique called dark-field and fluorescence coimaging assisted surface-enhanced Raman scattering (SERS) spectroscopy, combined with nuclear targeting nanoprobes, to in situ study Soma Gastric Cancer (SGC-7901) cell nuclei treated with two model drugs, e.g., DNA binder (Hoechst33342) and anticancer drug (doxorubicin, Dox) via spectral analysis at the molecular level. Nuclear targeting nanoprobes with an assembly structure of thiol-modified polyethylene glycol polymers (PEG) and nuclear localizing signal peptides (NLS) around gold nanorods (AuNRs) were prepared to achieve the amplified SERS signals of biomolecules in the cell nuclei. With the assistance of dark field/fluorescence imaging with simultaneous location, in situ SERS spectra in one cell nucleus were measured and analyzed to disclose the effects of Hoechst33342 and Dox on main biomolecules in the cell nuclei. The experimental results show that this method possesses great potential to investigate the targets of new anticancer drugs and the real-time monitoring of the dynamic changes of cells caused by exogenous molecules.

Pharmacologic suppression of target cell recognition by engineered T cells expressing chimeric T-cell receptors.

PubMed

Alvarez-Vallina, L; Yañez, R; Blanco, B; Gil, M; Russell, S J

2000-04-01

Adoptive therapy with autologous T cells expressing chimeric T-cell receptors (chTCRs) is of potential interest for the treatment of malignancy. To limit possible T-cell-mediated damage to normal tissues that weakly express the targeted tumor antigen (Ag), we have tested a strategy for the suppression of target cell recognition by engineered T cells. Jurkat T cells were transduced with an anti-hapten chTCR tinder the control of a tetracycline-suppressible promoter and were shown to respond to Ag-positive (hapten-coated) but not to Ag-negative target cells. The engineered T cells were then reacted with hapten-coated target cells at different effector to target cell ratios before and after exposure to tetracycline. When the engineered T cells were treated with tetracycline, expression of the chTCR was greatly decreased and recognition of the hapten-coated target cells was completely suppressed. Tetracycline-mediated suppression of target cell recognition by engineered T cells may be a useful strategy to limit the toxicity of the approach to cancer gene therapy.

α-Amanitin-Resistant Viral RNA Synthesis in Nuclei Isolated from Nuclear Polyhedrosis Virus-Infected Heliothis zea Larvae and Spodoptera frugiperda Cells

PubMed Central

Grula, Marjori A.; Buller, Patricia L.; Weaver, Robert F.

1981-01-01

[3H]RNA was synthesized in nuclei isolated at various times postinfection from the fat bodies of Heliothis zea larvae infected with H. zea nuclear polyhedrosis virus and from cultured Spodoptera frugiperda cells infected with Autographa californica nuclear polyhedrosis virus. To detect virus-specific RNA synthesis, the [3H]RNA was hybridized to denatured viral DNA immobilized on nitrocellulose filters. Nuclear polyhedrosis virus-specific RNA synthesis in the infected nuclei isolated from H. zea larval fat bodies and S. frugiperda cells was only inhibited 20 to 25% by concentrations of α-amanitin sufficient to inhibit the host RNA polymerase II. In addition, a productive nuclear polyhedrosis virus infection was obtained in S. frugiperda cells grown in the presence of an α-amanitin concentration that inhibited 90% of the cellular RNA polymerase II activity. The cellular RNA polymerase II enzyme remained sensitive to α-amanitin during infection, and there was no evidence that a virus-coded, α-amanitin-resistant enzyme was synthesized after the onset of infection. The data suggest that the bulk of nuclear polyhedrosis virus-specific RNA synthesis in isolated nuclei is transcribed by an enzyme other than the host RNA polymerase II. PMID:16789208

Spectroscopy of neutron rich nuclei using cold neutron induced fission of actinide targets at the ILL: The EXILL campaign

NASA Astrophysics Data System (ADS)

Blanc, A.; de France, G.; Drouet, F.; Jentschel, M.; Köster, U.; Mancuso, C.; Mutti, P.; Régis, J. M.; Simpson, G.; Soldner, T.; Ur, C. A.; Urban, W.; Vancraeyenest, A.

2013-12-01

One way to explore exotic nuclei is to study their structure by performing γ-ray spectroscopy. At the ILL, we exploit a high neutron flux reactor to induce the cold fission of actinide targets. In this process, fission products that cannot be accessed using standard spontaneous fission sources are produced with a yield allowing their detailed study using high resolution γ-ray spectroscopy. This is what was pursued at the ILL with the EXILL (for EXOGAM at the ILL) campaign. In the present work, the EXILL setup and performance will be presented.

Human immune cell targeting of protein nanoparticles - caveospheres

NASA Astrophysics Data System (ADS)

Glass, Joshua J.; Yuen, Daniel; Rae, James; Johnston, Angus P. R.; Parton, Robert G.; Kent, Stephen J.; de Rose, Robert

2016-04-01

Nanotechnology has the power to transform vaccine and drug delivery through protection of payloads from both metabolism and off-target effects, while facilitating specific delivery of cargo to immune cells. However, evaluation of immune cell nanoparticle targeting is conventionally restricted to monocultured cell line models. We generated human caveolin-1 nanoparticles, termed caveospheres, which were efficiently functionalized with monoclonal antibodies. Using this platform, we investigated CD4+ T cell and CD20+ B cell targeting within physiological mixtures of primary human blood immune cells using flow cytometry, imaging flow cytometry and confocal microscopy. Antibody-functionalization enhanced caveosphere binding to targeted immune cells (6.6 to 43.9-fold) within mixed populations and in the presence of protein-containing fluids. Moreover, targeting caveospheres to CCR5 enabled caveosphere internalization by non-phagocytic CD4+ T cells--an important therapeutic target for HIV treatment. This efficient and flexible system of immune cell-targeted caveosphere nanoparticles holds promise for the development of advanced immunotherapeutics and vaccines.

Population of Nuclei Via 7Li-Induced Binary Reactions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Clark, Rodney M.; Phair, Larry W.; Descovich, M.

2005-08-08

The authors have investigated the population of nuclei formed in binary reactions involving {sup 7}Li beams on targets of {sup 160}Gd and {sup 184}W. The {sup 7}Li + {sup 184}W data were taken in the first experiment using the LIBERACE Ge-array in combination with the STARS Si {Delta}E-E telescope system at the 88-Inch Cyclotron of the Lawrence Berkeley National Laboratory. By using the Wilczynski binary transfer model, in combination with a standard evaporation model, they are able to reproduce the experimental results. This is a useful method for predicting the population of neutron-rich heavy nuclei formed in binary reactions involvingmore » beams of weakly bound nuclei formed in binary reactions involving beams of weakly bound nuclei and will be of use in future spectroscopic studies.« less

Localization of 14-3-3 proteins in the nuclei of arabidopsis and maize.

PubMed

Bihn, E A; Paul, A L; Wang, S W; Erdos, G W; Ferl, R J

1997-12-01

It has been demonstrated that 14-3-3 proteins are present in the nuclei of Arabidopsis thaliana and Zea mays cells using laser scanning confocal microscopy and immunocytochemistry with monoclonal antibodies against plant 14-3-3 proteins. Confirmation of nuclear localization provides insight into the range of functions normally attributed to 14-3-3 proteins, especially since the association of 14-3-3s with transcription factors is (thus far) a phenomenon unique to plants, and since 14-3-3 proteins do not possess a recognizable nuclear targeting sequence.

Outer nuclear membrane fusion of adjacent nuclei in varicella-zoster virus-induced syncytia.

PubMed

Wang, Wei; Yang, Lianwei; Huang, Xiumin; Fu, Wenkun; Pan, Dequan; Cai, Linli; Ye, Jianghui; Liu, Jian; Xia, Ningshao; Cheng, Tong; Zhu, Hua

2017-12-01

Syncytia formation has been considered important for cell-to-cell spread and pathogenesis of many viruses. As a syncytium forms, individual nuclei often congregate together, allowing close contact of nuclear membranes and possibly fusion to occur. However, there is currently no reported evidence of nuclear membrane fusion between adjacent nuclei in wild-type virus-induced syncytia. Varicella-zoster virus (VZV) is one typical syncytia-inducing virus that causes chickenpox and shingles in humans. Here, we report, for the first time, an interesting observation of apparent fusion of the outer nuclear membranes from juxtaposed nuclei that comprise VZV syncytia both in ARPE-19 human epithelial cells in vitro and in human skin xenografts in the SCID-hu mouse model in vivo. This work reveals a novel aspect of VZV-related cytopathic effect in the context of multinucleated syncytia. Additionally, the information provided by this study could be helpful for future studies on interactions of viruses with host cell nuclei. Copyright © 2017 Elsevier Inc. All rights reserved.

Impact of the accuracy of automatic segmentation of cell nuclei clusters on classification of thyroid follicular lesions.

PubMed

Jung, Chanho; Kim, Changick

2014-08-01

Automatic segmentation of cell nuclei clusters is a key building block in systems for quantitative analysis of microscopy cell images. For that reason, it has received a great attention over the last decade, and diverse automatic approaches to segment clustered nuclei with varying levels of performance under different test conditions have been proposed in literature. To the best of our knowledge, however, so far there is no comparative study on the methods. This study is a first attempt to fill this research gap. More precisely, the purpose of this study is to present an objective performance comparison of existing state-of-the-art segmentation methods. Particularly, the impact of their accuracy on classification of thyroid follicular lesions is also investigated "quantitatively" under the same experimental condition, to evaluate the applicability of the methods. Thirteen different segmentation approaches are compared in terms of not only errors in nuclei segmentation and delineation, but also their impact on the performance of system to classify thyroid follicular lesions using different metrics (e.g., diagnostic accuracy, sensitivity, specificity, etc.). Extensive experiments have been conducted on a total of 204 digitized thyroid biopsy specimens. Our study demonstrates that significant diagnostic errors can be avoided using more advanced segmentation approaches. We believe that this comprehensive comparative study serves as a reference point and guide for developers and practitioners in choosing an appropriate automatic segmentation technique adopted for building automated systems for specifically classifying follicular thyroid lesions. © 2014 International Society for Advancement of Cytometry.

Membrane nanotubes facilitate long-distance interactions between natural killer cells and target cells

PubMed Central

Chauveau, Anne; Aucher, Anne; Eissmann, Philipp; Vivier, Eric; Davis, Daniel M.

2010-01-01

Membrane nanotubes are membranous tethers that physically link cell bodies over long distances. Here, we present evidence that nanotubes allow human natural killer (NK) cells to interact functionally with target cells over long distances. Nanotubes were formed when NK cells contacted target cells and moved apart. The frequency of nanotube formation was dependent on the number of receptor/ligand interactions and increased on NK cell activation. Most importantly, NK cell nanotubes contained a submicron scale junction where proteins accumulated, including DAP10, the signaling adaptor that associates with the activating receptor NKG2D, and MHC class I chain-related protein A (MICA), a cognate ligand for NKG2D, as occurs at close intercellular synapses between NK cells and target cells. Quantitative live-cell fluorescence imaging suggested that MICA accumulated at small nanotube synapses in sufficient numbers to trigger cell activation. In addition, tyrosine-phosphorylated proteins and Vav-1 accumulated at such junctions. Functionally, nanotubes could aid the lysis of distant target cells either directly or by moving target cells along the nanotube path into close contact for lysis via a conventional immune synapse. Target cells moving along the nanotube path were commonly polarized such that their uropods faced the direction of movement. This is the opposite polarization than for normal cell migration, implying that nanotubes can specifically drive target cell movement. Finally, target cells that remained connected to an NK cell by a nanotube were frequently lysed, whereas removing the nanotube using a micromanipulator reduced lysis of these target cells. PMID:20212116

Cell-specific targeting by heterobivalent ligands.

PubMed

Josan, Jatinder S; Handl, Heather L; Sankaranarayanan, Rajesh; Xu, Liping; Lynch, Ronald M; Vagner, Josef; Mash, Eugene A; Hruby, Victor J; Gillies, Robert J

2011-07-20

Current cancer therapies exploit either differential metabolism or targeting to specific individual gene products that are overexpressed in aberrant cells. The work described herein proposes an alternative approach--to specifically target combinations of cell-surface receptors using heteromultivalent ligands ("receptor combination approach"). As a proof-of-concept that functionally unrelated receptors can be noncovalently cross-linked with high avidity and specificity, a series of heterobivalent ligands (htBVLs) were constructed from analogues of the melanocortin peptide ligand ([Nle(4), dPhe(7)]-α-MSH) and the cholecystokinin peptide ligand (CCK-8). Binding of these ligands to cells expressing the human Melanocortin-4 receptor and the Cholecystokinin-2 receptor was analyzed. The MSH(7) and CCK(6) were tethered with linkers of varying rigidity and length, constructed from natural and/or synthetic building blocks. Modeling data suggest that a linker length of 20-50 Å is needed to simultaneously bind these two different G-protein coupled receptors (GPCRs). These ligands exhibited up to 24-fold enhancement in binding affinity to cells that expressed both (bivalent binding), compared to cells with only one (monovalent binding) of the cognate receptors. The htBVLs had up to 50-fold higher affinity than that of a monomeric CCK ligand, i.e., Ac-CCK(6)-NH(2). Cell-surface targeting of these two cell types with labeled heteromultivalent ligand demonstrated high avidity and specificity, thereby validating the receptor combination approach. This ability to noncovalently cross-link heterologous receptors and target individual cells using a receptor combination approach opens up new possibilities for specific cell targeting in vivo for therapy or imaging.

Cell-Specific Targeting by Heterobivalent Ligands

PubMed Central

Josan, Jatinder S.; Handl, Heather L.; Sankaranarayanan, Rajesh; Xu, Liping; Lynch, Ronald M.; Vagner, Josef; Mash, Eugene A.; Hruby, Victor J.; Gillies, Robert J.

2012-01-01

Current cancer therapies exploit either differential metabolism or targeting to specific individual gene products that are overexpressed in aberrant cells. The work described herein proposes an alternative approach—to specifically target combinations of cell-surface receptors using heteromultivalent ligands (“receptor combination approach”). As a proof-of-concept that functionally unrelated receptors can be noncovalently cross-linked with high avidity and specificity, a series of heterobivalent ligands (htBVLs) were constructed from analogues of the melanocortin peptide ligand ([Nle4, DPhe7]-α-MSH) and the cholecystokinin peptide ligand (CCK-8). Binding of these ligands to cells expressing the human Melanocortin-4 receptor and the Cholecystokinin-2 receptor was analyzed. The MSH(7) and CCK(6) were tethered with linkers of varying rigidity and length, constructed from natural and/or synthetic building blocks. Modeling data suggest that a linker length of 20–50 Å is needed to simultaneously bind these two different G-protein coupled receptors (GPCRs). These ligands exhibited up to 24-fold enhancement in binding affinity to cells that expressed both (bivalent binding), compared to cells with only one (monovalent binding) of the cognate receptors. The htBVLs had up to 50-fold higher affinity than that of a monomeric CCK ligand, i.e., Ac-CCK(6)-NH2. Cell-surface targeting of these two cell types with labeled heteromultivalent ligand demonstrated high avidity and specificity, thereby validating the receptor combination approach. This ability to noncovalently cross-link heterologous receptors and target individual cells using a receptor combination approach opens up new possibilities for specific cell targeting in vivo for therapy or imaging. PMID:21639139

Accurate Segmentation of Cervical Cytoplasm and Nuclei Based on Multiscale Convolutional Network and Graph Partitioning.

PubMed

Song, Youyi; Zhang, Ling; Chen, Siping; Ni, Dong; Lei, Baiying; Wang, Tianfu

2015-10-01

In this paper, a multiscale convolutional network (MSCN) and graph-partitioning-based method is proposed for accurate segmentation of cervical cytoplasm and nuclei. Specifically, deep learning via the MSCN is explored to extract scale invariant features, and then, segment regions centered at each pixel. The coarse segmentation is refined by an automated graph partitioning method based on the pretrained feature. The texture, shape, and contextual information of the target objects are learned to localize the appearance of distinctive boundary, which is also explored to generate markers to split the touching nuclei. For further refinement of the segmentation, a coarse-to-fine nucleus segmentation framework is developed. The computational complexity of the segmentation is reduced by using superpixel instead of raw pixels. Extensive experimental results demonstrate that the proposed cervical nucleus cell segmentation delivers promising results and outperforms existing methods.

Binding and internalization of NGR-peptide-targeted liposomal doxorubicin (TVT-DOX) in CD13-expressing cells and its antitumor effects.

PubMed

Garde, Seema V; Forté, André J; Ge, Michael; Lepekhin, Eugene A; Panchal, Chandra J; Rabbani, Shafaat A; Wu, Jinzi J

2007-11-01

In an effort to develop new agents and molecular targets for the treatment of cancer, aspargine-glycine-arginine (NGR)-targeted liposomal doxorubicin (TVT-DOX) is being studied. The NGR peptide on the surface of liposomal doxorubicin (DOX) targets an aminopeptidase N (CD13) isoform, specific to the tumor neovasculature, making it a promising strategy. To further understand the molecular mechanisms of action, we investigated cell binding, kinetics of internalization as well as cytotoxicity of TVT-DOX in vitro. We demonstrate the specific binding of TVT-DOX to CD13-expressing endothelial [human umbilical vein endothelial cells (HUVEC) and Kaposi sarcoma-derived endothelial cells (SLK)] and tumor (fibrosarcoma, HT-1080) cells in vitro. Following binding, the drug was shown to internalize through the endosomal pathway, eventually leading to the localization of doxorubicin in cell nuclei. TVT-DOX showed selective toxicity toward CD13-expressing HUVEC, sparing the CD13-negative colon-cancer cells, HT-29. Additionally, the nontargeted counterpart of TVT-DOX, Caelyx, was less cytotoxic to the CD13-positive HUVECs demonstrating the advantages of NGR targeting in vitro. The antitumor activity of TVT-DOX was tested in nude mice bearing human prostate-cancer xenografts (PC3). A significant growth inhibition (up to 60%) of PC3 tumors in vivo was observed. Reduction of tumor vasculature following treatment with TVT-DOX was also apparent. We further compared the efficacies of TVT-DOX and free doxorubicin in the DOX-resistant colon-cancer model, HCT-116, and observed the more pronounced antitumor effects of the TVT-DOX formulation over free DOX. The potential utility of TVT-DOX in a variety of vascularized solid tumors is promising.

I-motif DNA structures are formed in the nuclei of human cells

NASA Astrophysics Data System (ADS)

Zeraati, Mahdi; Langley, David B.; Schofield, Peter; Moye, Aaron L.; Rouet, Romain; Hughes, William E.; Bryan, Tracy M.; Dinger, Marcel E.; Christ, Daniel

2018-06-01

Human genome function is underpinned by the primary storage of genetic information in canonical B-form DNA, with a second layer of DNA structure providing regulatory control. I-motif structures are thought to form in cytosine-rich regions of the genome and to have regulatory functions; however, in vivo evidence for the existence of such structures has so far remained elusive. Here we report the generation and characterization of an antibody fragment (iMab) that recognizes i-motif structures with high selectivity and affinity, enabling the detection of i-motifs in the nuclei of human cells. We demonstrate that the in vivo formation of such structures is cell-cycle and pH dependent. Furthermore, we provide evidence that i-motif structures are formed in regulatory regions of the human genome, including promoters and telomeric regions. Our results support the notion that i-motif structures provide key regulatory roles in the genome.

Targeting Cell Polarity Machinery to Exhaust Breast Cancer Stem Cells

DTIC Science & Technology

2016-10-01

AWARD NUMBER: W81XWH-15-1-0644 TITLE: Targeting Cell Polarity Machinery to Exhaust Breast Cancer Stem Cells PRINCIPAL INVESTIGATOR: Chun-Ju...U.S. Army Medical Research and Materiel Command Fort Detrick, Maryland 21702-5012 DISTRIBUTION STATEMENT: Approved for Public Release...Targeting Cell Polarity Machinery to Exhaust Breast Cancer Stem Cells 5a. CONTRACT NUMBER 5b. GRANT NUMBER W81XWH-15-1-0644 5c. PROGRAM ELEMENT

A hypothesis of target cell formation in sickle cell disease.

PubMed

Wong, P

2016-08-01

A fraction of erythrocytes appear as target cells in stained blood smears in sickle cell disease, due to a inheritance of the hemoglobin variant Hb S, polymerizing upon deoxygenation. These cells appear in a three dimension as thin cups. A process of their formation in this disease is proposed based on a band 3-based mechanism of the erythrocyte shape control, able to explain the erythrocyte echinocytosis by glucose depletion. It indicates that their formation is due to a stomatocytogenic slow outward transport of the dibasic form of endogenous Pi with an H(+) by band 3, promoted by the decrease of the Donnan ratio, which decreases cell pH and volume, attributed by a decrease of cell KCl concentration by the higher efflux of K(+)Cl(-) cotransport and Ca(2+) activation of the Gardos channel. Its implications are briefly discussed with respect to target cells per se, target cell formation in other hemoglobinopathies, acquired and inherited disorders of the lipid metabolism and dehydrated hereditary stomatocytosis as well as a stomatocyte presence in a double heterozygote of Hb S and Hb C and of an involvement of the process of target cell formation in acanthocytosis in acquired and inherited disorders. Copyright © 2016 Elsevier Ltd. All rights reserved.

Generation of high-energy neutron beam by fragmentation of relativistic heavy nuclei

NASA Astrophysics Data System (ADS)

Yurevich, Vladimir

2016-09-01

The phenomenon of multiple production of neutrons in reactions with heavy nuclei induced by high-energy protons and light nuclei is analyzed using a Moving Source Model. The Lorentz transformation of the obtained neutron distributions is used to study the neutron characteristics in the inverse kinematics where relativistic heavy nuclei bombard a light-mass target. The neutron beam generated at 0∘has a Gaussian shape with a maximum at the energy of the projectile nucleons and an energy resolution σE/E < 4% above 6 GeV.

The Equilibrium and Pre-equilibrium Triton Emission Spectra of Some Target Nuclei for ( n, xt) Reactions up to 45 MeV Energy

NASA Astrophysics Data System (ADS)

Tel, E.; Kaplan, A.; Aydın, A.; Özkorucuklu, S.; Büyükuslu, H.; Yıldırım, G.

2010-08-01

Although there have been significant research and development studies on the inertial and magnetic fusion reactor technology, there is still a long way to go to penetrate commercial fusion reactors to the energy market. Tritium self-sufficiency must be maintained for a commercial power plant. For self-sustaining (D-T) fusion driver tritium breeding ratio should be greater than 1.05. So, working out the systematics of ( n,t) reaction cross sections and triton emission differential data are important for the given reaction taking place on various nuclei at different energies. In this study, ( n,xt) reactions for some target nuclei as 16O, 27Al, 59Co and 209Bi have been investigated up to 45 MeV incident neutron energy. In the calculations of the triton emission spectra, the pre-equilibrium and equilibrium effects have been used. The calculated results have been compared with the experimental data taken from the literature.

Evolving phage vectors for cell targeted gene delivery.

PubMed

Larocca, David; Burg, Michael A; Jensen-Pergakes, Kristen; Ravey, Edward Prenn; Gonzalez, Ana Maria; Baird, Andrew

2002-03-01

We adapted filamentous phage vectors for targeted gene delivery to mammalian cells by inserting a mammalian reporter gene expression cassette (GFP) into the vector backbone and fusing the pIII coat protein to a cell targeting ligand (i.e. FGF2, EGF). Like transfection with animal viral vectors, targeted phage gene delivery is concentration, time, and ligand dependent. Importantly, targeted phage particles are specific for the appropriate target cell surface receptor. Phage have distinct advantages over existing gene therapy vectors because they are simple, economical to produce at high titer, have no intrinsic tropism for mammalian cells, and are relatively simple to genetically modify and evolve. Initially transduction by targeted phage particles was low resulting in foreign gene expression in 1-2% of transfected cells. We increased transduction efficiency by modifying both the transfection protocol and vector design. For example, we stabilized the display of the targeting ligand to create multivalent phagemid-based vectors with transduction efficiencies of up to 45% in certain cell lines when combined with genotoxic treatment. Taken together, these studies establish that the efficiency of phage-mediated gene transfer can be significantly improved through genetic modification. We are currently evolving phage vectors with enhanced cell targeting, increased stability, reduced immunogenicity and other properties suitable for gene therapy.

[Morphometric analysis of lymphocyte nuclei in chronic lymphocytic leukemia].

PubMed

Ostapenko, V A; Kruchinskiĭ, N G; Smirnova, L A; Cherednik, A B; Nesterov, V N; Tepliakov, A I

1994-01-01

This work is dedicated to the study of use of quantitative analysis of cell nucleus structure for the analysis of peripheral blood lymphocytes in patients with chronic lymphocytic leukaemia. The structure of lymphocytic nuclei of healthy donors was evaluated by means of staining by toluidine blue purified cell suspensions smears. The preparations were analysed on the television measuring system "omnicon" with measurements of the following parameters: square of the nucleus, euchromatin, heterochromatin, and the ratio of heterochromatin and euchromatin squares. Actuarial analysis and nuclei classification of the previously mentioned parameters showed, that in peripheral blood of patients with chronic lymphocytic leukemia a large amount of atypical lymphocytes is present with reduced nucleus sizes. Atypical cells retain the ratio of structural components of chromatine, characteristic to normal cells, which show their low proliferative activity.

The change is length and width of the Sertoli cell nuclei in cytologic smears of testes with depopulation of the seminiferous epithelium.

PubMed

Banek, L; Posinovec, J

1980-09-15

The appearance of the Sertoli cells in cytological smears of tests with depopulation of the seminiferous epithelium is described. The mean values of the lengths and widths of the Sertoli cell nuclei in smears differed significantly between the depopulation and the control group (p < 0.01).

Modeling multi-nucleon transfer in symmetric collisions of massive nuclei

DOE Office of Scientific and Technical Information (OSTI.GOV)

Welsh, T.; Loveland, W.; Yanez, R.

We propose symmetric collisions of massive nuclei, such as 238U + 248Cm, as ways to make new n-rich heavy nuclei via multi-nucleon transfer (MNT) reactions. We have measured the yields of several projectile-like and target-like fragments from the reaction of 1360 MeV 204Hg + 198Pt. We also find that current models for this symmetric collision (GRAZING, DNS, ImQMD) significantly underestimate the yields of these transfer products, even for small transfers.

Modeling multi-nucleon transfer in symmetric collisions of massive nuclei

DOE PAGES

Welsh, T.; Loveland, W.; Yanez, R.; ...

2017-05-18

We propose symmetric collisions of massive nuclei, such as 238U + 248Cm, as ways to make new n-rich heavy nuclei via multi-nucleon transfer (MNT) reactions. We have measured the yields of several projectile-like and target-like fragments from the reaction of 1360 MeV 204Hg + 198Pt. We also find that current models for this symmetric collision (GRAZING, DNS, ImQMD) significantly underestimate the yields of these transfer products, even for small transfers.

[The frequency of sex chromatine occurring in cell nuclei of internal organs determined by the smear method (author's transl)].

PubMed

Michailow, R

1975-09-05

The frequency of sex chromatine occurring in cell nuclei of twelve organs from 25 male and female corpses was determined using the smear method. It was found to be about 60% in the case of female, and about 6% in the case of male corpses.

Improved and Robust Detection of Cell Nuclei from Four Dimensional Fluorescence Images

PubMed Central

Bashar, Md. Khayrul; Yamagata, Kazuo; Kobayashi, Tetsuya J.

2014-01-01

Segmentation-free direct methods are quite efficient for automated nuclei extraction from high dimensional images. A few such methods do exist but most of them do not ensure algorithmic robustness to parameter and noise variations. In this research, we propose a method based on multiscale adaptive filtering for efficient and robust detection of nuclei centroids from four dimensional (4D) fluorescence images. A temporal feedback mechanism is employed between the enhancement and the initial detection steps of a typical direct method. We estimate the minimum and maximum nuclei diameters from the previous frame and feed back them as filter lengths for multiscale enhancement of the current frame. A radial intensity-gradient function is optimized at positions of initial centroids to estimate all nuclei diameters. This procedure continues for processing subsequent images in the sequence. Above mechanism thus ensures proper enhancement by automated estimation of major parameters. This brings robustness and safeguards the system against additive noises and effects from wrong parameters. Later, the method and its single-scale variant are simplified for further reduction of parameters. The proposed method is then extended for nuclei volume segmentation. The same optimization technique is applied to final centroid positions of the enhanced image and the estimated diameters are projected onto the binary candidate regions to segment nuclei volumes.Our method is finally integrated with a simple sequential tracking approach to establish nuclear trajectories in the 4D space. Experimental evaluations with five image-sequences (each having 271 3D sequential images) corresponding to five different mouse embryos show promising performances of our methods in terms of nuclear detection, segmentation, and tracking. A detail analysis with a sub-sequence of 101 3D images from an embryo reveals that the proposed method can improve the nuclei detection accuracy by 9 over the previous methods

Statistical Modeling of Single Target Cell Encapsulation

PubMed Central

Moon, SangJun; Ceyhan, Elvan; Gurkan, Umut Atakan; Demirci, Utkan

2011-01-01

High throughput drop-on-demand systems for separation and encapsulation of individual target cells from heterogeneous mixtures of multiple cell types is an emerging method in biotechnology that has broad applications in tissue engineering and regenerative medicine, genomics, and cryobiology. However, cell encapsulation in droplets is a random process that is hard to control. Statistical models can provide an understanding of the underlying processes and estimation of the relevant parameters, and enable reliable and repeatable control over the encapsulation of cells in droplets during the isolation process with high confidence level. We have modeled and experimentally verified a microdroplet-based cell encapsulation process for various combinations of cell loading and target cell concentrations. Here, we explain theoretically and validate experimentally a model to isolate and pattern single target cells from heterogeneous mixtures without using complex peripheral systems. PMID:21814548

Nuclei of plants as a sink for flavanols.

PubMed

Feucht, W; Polster, J

2001-01-01

Onion cepa (L.) and Tsuga canadensis (L.) Carr. were investigated histochemically on the association of flavanols to nuclei. The young roots of Onion cepa are totally devoid of flavanol structures. Therefore, the excised roots tips were directly incubated into different solutions of flavanols. After 3 h of incubation a flavanol binding on the nuclei was recognizable, as seen by a yellowish-brown tanning reaction. Still to ensure the presence of flavanols on the nuclei, subsequent staining with the p-dimethylaminocinnamaldehyde reagent (DMACA) resulted in an intense blue colouration. Tsuga canadensis has significant amounts of vacuolar flavanol deposits in all parts of the tree as indicated by the DMACA reagent. It is obvious that also the nuclei were associated strongly with flavanols which can be demonstrated particularly elegant in the cells of the seed wings by histochemical methods. However, the mode of flavanol release from the original deposits is not yet clear.

Oxidant-induced DNA damage of target cells.

PubMed Central

Schraufstätter, I; Hyslop, P A; Jackson, J H; Cochrane, C G

1988-01-01

In this study we examined the leukocytic oxidant species that induce oxidant damage of DNA in whole cells. H2O2 added extracellularly in micromolar concentrations (10-100 microM) induced DNA strand breaks in various target cells. The sensitivity of a specific target cell was inversely correlated to its catalase content and the rate of removal of H2O2 by the target cell. Oxidant species produced by xanthine oxidase/purine or phorbol myristate acetate-stimulated monocytes induced DNA breakage of target cells in proportion to the amount of H2O2 generated. These DNA strand breaks were prevented by extracellular catalase, but not by superoxide dismutase. Cytotoxic doses of HOCl, added to target cells, did not induce DNA strand breakage, and myeloperoxidase added extracellularly in the presence of an H2O2-generating system, prevented the formation of DNA strand breaks in proportion to its H2O2 degrading capacity. The studies also indicated that H2O2 formed hydroxyl radical (.OH) intracellularly, which appeared to be the most likely free radical responsible for DNA damage: .OH was detected in cells exposed to H2O2; the DNA base, deoxyguanosine, was hydroxylated in cells exposed to H2O2; and intracellular iron was essential for induction of DNA strand breaks. PMID:2843565

Population of Nuclei Via 7Li-Induced Binary Reactions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Clark, R M; Phair, L W; Descovich, M

2005-08-09

The authors have investigated the population of nuclei formed in binary reactions involving {sup 7}Li beams on targets of {sup 160}Gd and {sup 184}W. The {sup 7}Li + {sup 184}W data were taken in the first experiment using the LIBERACE Ge-array in combination with the STARS Si {Delta}E-E telescope system at the 88-Inch Cyclotron of the Lawrence Berkeley National Laboratory. By using the Wilczynski binary transfer model, in combination with a standard evaporation model, they are able to reproduce the experimental results. This is a useful method for predicting the population of neutron-rich heavy nuclei formed in binary reactions involvingmore » beams of weakly bound nuclei and will be of use in future spectroscopic studies.« less

DNA synthesis in HeLa cells and isolated nuclei after treatment with an inhibitor of spermidine synthesis, methyl glyoxal bis(guanylhydrazone).

PubMed

Krokan, H; Eriksen, A

1977-02-01

Addition of methyl glyoxal bis(guanylhydrazone) to HeLa S3 suspension cultures resulted in increased putrescine levels and decreased spermidine and spermine levels preceding a drop in incorporation of [3H]thymidine, [3H]uridine and [14C]leucine into macromolecules. When putrescine, spermidine, spermine or cadaverine was added simultaneously with methyl glyoxal bis(guanylhydrazone), the drug had no detectable effect on the synthesis of macromolecules. In nuclei isolated from cells treated with methyl glyoxal bis(guanylhydrazone) the reduction in the rate of DNA synthesis was equal to the reduction of [3H]thymidine incorporation in the corresponding whole cells. The capability of the nuclei to synthesize DNA could not be restored by adding spermidine or spermine to the system in vitro. The rate of DNA chain elongation was only reduced slightly by methyl glyoxal bis(guanylhydrazone) indicating that decreased levels of spermidine and spermine lead to a decrease in the number of replication units active in DNA synthesis within each cell.

[Some morphometric parameters of nucleoli and nuclei in invasive ductal breast carcinomas in women].

PubMed

Karpinska-Kaczmarczyk, Katarzyna

2009-01-01

The purpose of this study was to correlate seven morphometric parameters of nucleoli and nuclei of invasive ductal cancer cells with some clinico-pathological factors such as age, tumor size, axillary lymph node status, MIB-1 proliferation index, and estrogen receptor expression in tumor cells. Methyl green-pyronin Y (MG-PY) was used for simultaneous staining of nuclei and nucleoli in histological sections of 150 invasive ductal breast carcinomas. Next, morphometric parameters of nucleoli and nuclei of tumor cells were measured with computerized image analysis. Nuclear area and number of nucleoli in breast tumor cells were greater in younger axillary node-negative patients. The number of nucleoli and nucleolar shape polymorphism were reduced in tumors measuring 20 mm or less or with lower histological grade. Nuclear area, nucleolar number, and nucleolar polymorphism in carcinomas with low proliferation index and estrogen receptor expression were smaller than in carcinomas with high proliferation index and no estrogen receptor expression. Nucleolar area in primary tumors without axillary node involvement was greater than in tumors with more than three axillary nodes positive. MG-PY selectively and simultaneously stains nucleoli and nuclei of tumor cells enabling standardized and reproducible examination of these structures with computerized image analysis. Univariate statistical analysis disclosed that some morphometric parameters of nucleoli and nuclei of tumor cells correlated with several established clinico-pathological prognostic factors. Therefore, the prognostic significance of these parameters should be studied in a larger group of patients with invasive ductal breast carcinomas.

Designing oral vaccines targeting intestinal dendritic cells.

PubMed

Devriendt, Bert; De Geest, Bruno G; Cox, Eric

2011-04-01

Most pathogens colonize and invade the host at mucosal surfaces, such as the lung and the intestine. To combat intestinal pathogens the induction of local adaptive immune responses is required, which is mainly achieved through oral vaccination. However, most vaccines are ineffective when given orally owing to the hostile environment in the gastrointestinal tract. The encapsulation of antigens in biodegradable microparticulate delivery systems enhances their immunogenicity; however, the uptake of these delivery systems by intestinal immune cells is rather poor. Surface decoration of the particulates with targeting ligands could increase the uptake and mediate the selective targeting of the vaccine to intestinal antigen-presenting cells, including dendritic cells. In this review, current knowledge on dendritic cell subsets is discussed, along with progress in the development of selective antigen targeting to these cells, in addition to focusing on data obtained in mice and, where possible, the pig, as a non-rodent animal model for humans. Moreover, the potential use and benefits of Fcγ receptor-mediated targeting of antigen delivery systems are highlighted. In conclusion, dendritic cell targeting ligands grafted on antigen carrier systems should preferably bind to a conserved endocytotic receptor, facilitating the design of a multispecies vaccine platform, which could elicit robust protective immune responses against enteric pathogens.

Shared target antigens on cancer cells and tissue stem cells: go or no-go for CAR T cells?

PubMed

Hombach, Andreas A; Abken, Hinrich

2017-02-01

Adoptive therapy with chimeric antigen receptor (CAR) T cells redirected towards CD19 produces remissions of B cell malignancies, however, it also eradicates healthy B cells sharing the target antigen. Such 'on-target off-tumor' toxicity raises serious safety concerns when the target antigen is also expressed by tissue stem cells, with the risk of lasting tissue destruction. Areas covered: We discuss CAR T cell targeting of activation antigens versus lineage associated antigens on the basis of recent experimental and animal data and the literature in the field. Expert commentary: Targeting an activation associated antigen which is transiently expressed by stem cells seems to be safe, like CAR T cells targeting CD30 spare CD30 + hematopoietic stem and progenitor cells while eliminating CD30 + lymphoma cells, whereas targeting lineage associated antigens which increase in expression during cell maturation, like folate receptor-β and CD123, is of risk to destruct tissue stem cells.

Isolation of the constitutive heterochromatin from mouse liver nuclei.

PubMed

Zatsepina, Olga V; Zharskaya, Oxana O; Prusov, Andrei N

2008-01-01

A method for isolation of constitutive heterochromatin (chromocenters) from nuclei of mouse liver cells is described. This method is based on the higher resistance of chromocenters to low ionic strength treatment as compared with that of nucleoli and euchromatin. The method allows separation of chromocenters that are essentially free of nucleoli and other nuclear contaminants. In contrast to nuclei and nucleoli, isolated chromocenters are characterized by a simpler protein composition and contain a smaller number of proteins (especially of high molecular weight proteins). They possess telomeric DNA and telomerase activity that suggests a tight association of chromocenters with the telomerase complex in mouse hepatocyte nuclei.

PNA-COMBO-FISH: From combinatorial probe design in silico to vitality compatible, specific labelling of gene targets in cell nuclei.

PubMed

Müller, Patrick; Rößler, Jens; Schwarz-Finsterle, Jutta; Schmitt, Eberhard; Hausmann, Michael

2016-07-01

Recently, advantages concerning targeting specificity of PCR constructed oligonucleotide FISH probes in contrast to established FISH probes, e.g. BAC clones, have been demonstrated. These techniques, however, are still using labelling protocols with DNA denaturing steps applying harsh heat treatment with or without further denaturing chemical agents. COMBO-FISH (COMBinatorial Oligonucleotide FISH) allows the design of specific oligonucleotide probe combinations in silico. Thus, being independent from primer libraries or PCR laboratory conditions, the probe sequences extracted by computer sequence data base search can also be synthesized as single stranded PNA-probes (Peptide Nucleic Acid probes) or TINA-DNA (Twisted Intercalating Nucleic Acids). Gene targets can be specifically labelled with at least about 20 probes obtaining visibly background free specimens. By using appropriately designed triplex forming oligonucleotides, the denaturing procedures can completely be omitted. These results reveal a significant step towards oligonucleotide-FISH maintaining the 3d-nanostructure and even the viability of the cell target. The method is demonstrated with the detection of Her2/neu and GRB7 genes, which are indicators in breast cancer diagnosis and therapy. Copyright © 2016. Published by Elsevier Inc.

Studies of Heavy-Ion Reactions and Transuranic Nuclei

DOE Office of Scientific and Technical Information (OSTI.GOV)

Schroeder, W. Udo

2016-07-28

Studies of heavy-ion reactions and transuranic nuclei performed by the University of Rochester Nuclear Science Research Group have been successful in furthering experimental systematics and theoretical understanding of the behavior of nuclear systems excited to their limits of stability. The theoretical results explain specifically the “boiling” and “vaporization” of atomic nuclei, but are more generally applicable to isolated, quantal many-particle systems which, under thermal or mechanical stresses, all disintegrate by evaporation, via surface cluster emission, or via fission-like processes. Accompanying experimental investigations by the group have demonstrated several new types of dynamical instability of nuclei: In central, “head-on” collisions, targetmore » nuclei exhibit limited ability to stop energetic projectile nuclei and to dissipate the imparted linear momentum. Substantial matter overlap (“neck”) between projectile and target nuclei, which is observed at elevated collision energies, can be stretched considerably and break at several places simultaneously. These results provide new testing grounds for microscopic theory of the cohesion of nuclear matter. This property has remained elusive, even though the elementary nucleon-nucleon forces are well known since some time. Technical R&D has resulted in a detailed characterization of a novel plastic material, which can now be used in the design of sensitive diagnostic systems for various types of radio-activity. Innovative application of powerful laser systems has produced intense, controllable sources of exotic particle radioactivity for nuclear investigations. Several students have received their Ph.D. degree in experimental nuclear science for their work on basic nuclear research or R&D projects.« less

Targeting tumor cell motility to prevent metastasis

PubMed Central

Palmer, Trenis D.; Ashby, William J.; Lewis, John D.; Zijlstra, Andries

2011-01-01

Mortality and morbidity in patients with solid tumors invariably results from the disruption of normal biological function caused by disseminating tumor cells. Tumor cell migration is under intense investigation as the underlying cause of cancer metastasis. The need for tumor cell motility in the progression of metastasis has been established experimentally and is supported empirically by basic and clinical research implicating a large collection of migration-related genes. However, there are few clinical interventions designed to specifically target the motility of tumor cells and adjuvant therapy to specifically prevent cancer cell dissemination is severely limited. In an attempt to define motility targets suitable for treating metastasis, we have parsed the molecular determinants of tumor cell motility into five underlying principles including cell autonomous ability, soluble communication, cell-cell adhesion, cell-matrix adhesion, and integrating these determinants of migration on molecular scaffolds. The current challenge is to implement meaningful and sustainable inhibition of metastasis by developing clinically viable disruption of molecular targets that control these fundamental capabilities. PMID:21664937

Probing Neutron-Skin Thickness of Unstable Nuclei with Total Reaction Cross Sections

NASA Astrophysics Data System (ADS)

Horiuchi, Wataru; Suzuki, Yasuyuki; Inakura, Tsunenori

We present our recent analysis of the total reaction cross sections, σR, of unstable nuclei and discuss their sensitivity to the neutron-skin thickness. The σR is calculated with the Glauber model using projectile densities obtained with the Skyrme-Hartree-Fock method on the three-dimensional coordinate space. We cover 91 nuclei of O, Ne, Mg, Si, S, Ca, and Ni isotopes. Defining a reaction radius, aR = √{σ R/π } , to characterize the nuclear size and target (proton or 12C) dependence, we see the 12C target probes the matter radius while the proton target is sensitive to the skin-thickness. We find an empirical formula for expressing aR with the point matter radius and the skin thickness, which can be used to determine the skin thickness.

Synthesis of Superheavy Nuclei in 48CA-INDUCED Reactions

NASA Astrophysics Data System (ADS)

Oganessian, Yu. Ts.; Utyonkov, V. K.; Lobanov, Yu. V.; Abdullin, F. Sh.; Polyakov, A. N.; Sagaidak, R. N.; Shirokovsky, I. V.; Tsyganov, Yu. S.; Voinov, A. A.; Gulbekian, G. G.; Bogomolov, S. L.; Gikal, B. N.; Mezentsev, A. N.; Iliev, S.; Subbotin, V. G.; Sukhov, A. M.; Subotic, K.; Zagrebaev, V. I.; Vostokin, G. K.; Itkis, M. G.; Moody, K. J.; Patin, J. B.; Shaughnessy, D. A.; Stoyer, M. A.; Stoyer, N. J.; Wilk, P. A.; Kenneally, J. M.; Landrum, J. H.; Wild, J. F.; Lougheed, R. W.

2008-11-01

Thirty-four new nuclides with Z = 104-116, 118 and N = 161-177 have been synthesized in the complete-fusion reactions of 238U, 237Np, 242,244Pu, 243Am, 245,248Cm, and 249Cf targets with 48Ca beams. The masses of evaporation residues were identified through measurements of the excitation functions of the xn-evaporation channels and from cross bombardments. The decay properties of the new nuclei agree with those of previously known heavy nuclei and with predictions from different theoretical models. A discussion of self-consistent interpretations of all observed decay chains originating from the parent isotopes 282,283112, 282113, 286-289114, 287,288115, 290-293116, and 294118 is presented. Decay energies and lifetimes of the neutron-rich superheavy nuclei as well as their production cross sections indicate a considerable increase in the stability of nuclei with an increasing number of neutrons, which agrees with the predictions of theoretical models concerning the decisive dependence of the structure and radioactive properties of superheavy elements on their proximity to the nuclear shells with N = 184 and Z = 114.

Human Cytomegalovirus Nuclear Egress Proteins Ectopically Expressed in the Heterologous Environment of Plant Cells are Strictly Targeted to the Nuclear Envelope.

PubMed

Lamm, Christian E; Link, Katrin; Wagner, Sabrina; Milbradt, Jens; Marschall, Manfred; Sonnewald, Uwe

2016-03-10

In all eukaryotic cells, the nucleus forms a prominent cellular compartment containing the cell's nuclear genome. Although structurally similar, animal and plant nuclei differ substantially in details of their architecture. One example is the nuclear lamina, a layer of tightly interconnected filament proteins (lamins) underlying the nuclear envelope of metazoans. So far no orthologous lamin genes could be detected in plant genomes and putative lamin-like proteins are only poorly described in plants. To probe for potentially conserved features of metazoan and plant nuclear envelopes, we ectopically expressed the core nuclear egress proteins of human cytomegalovirus pUL50 and pUL53 in plant cells. pUL50 localizes to the inner envelope of metazoan nuclei and recruits the nuclear localized pUL53 to it, forming heterodimers. Upon expression in plant cells, a very similar localization pattern of both proteins could be determined. Notably, pUL50 is specifically targeted to the plant nuclear envelope in a rim-like fashion, a location to which coexpressed pUL53 becomes strictly corecruited from its initial nucleoplasmic distribution. Using pUL50 as bait in a yeast two-hybrid screening, the cytoplasmic re-initiation supporting protein RISP could be identified. Interaction of pUL50 and RISP could be confirmed by coexpression and coimmunoprecipitation in mammalian cells and by confocal laser scanning microscopy in plant cells, demonstrating partial pUL50-RISP colocalization in areas of the nuclear rim and other intracellular compartments. Thus, our study provides strong evidence for conserved structural features of plant and metazoan nuclear envelops and identifies RISP as a potential pUL50-interacting plant protein.

Cytotoxic T cells use mechanical force to potentiate target cell killing

PubMed Central

Basu, Roshni; Whitlock, Benjamin M.; Husson, Julien; Le Floc’h, Audrey; Jin, Weiyang; Oyler-Yaniv, Alon; Dotiwala, Farokh; Giannone, Gregory; Hivroz, Claire; Biais, Nicolas; Lieberman, Judy; Kam, Lance C.; Huse, Morgan

2016-01-01

SUMMARY The immunological synapse formed between a cytotoxic T lymphocyte (CTL) and an infected or transformed target cell is a physically active structure capable of exerting mechanical force. Here, we investigated whether synaptic forces promote the destruction of target cells. CTLs kill by secreting toxic proteases and the pore forming protein perforin into the synapse. Biophysical experiments revealed a striking correlation between the magnitude of force exertion across the synapse and the speed of perforin pore formation on the target cell, implying that force potentiates cytotoxicity by enhancing perforin activity. Consistent with this interpretation, we found that increasing target cell tension augmented pore formation by perforin and killing by CTLs. Our data also indicate that CTLs coordinate perforin release and force exertion in space and time. These results reveal an unappreciated physical dimension to lymphocyte function and demonstrate that cells use mechanical forces to control the activity of outgoing chemical signals. PMID:26924577

Cytotoxic T Cells Use Mechanical Force to Potentiate Target Cell Killing.

PubMed

Basu, Roshni; Whitlock, Benjamin M; Husson, Julien; Le Floc'h, Audrey; Jin, Weiyang; Oyler-Yaniv, Alon; Dotiwala, Farokh; Giannone, Gregory; Hivroz, Claire; Biais, Nicolas; Lieberman, Judy; Kam, Lance C; Huse, Morgan

2016-03-24

The immunological synapse formed between a cytotoxic T lymphocyte (CTL) and an infected or transformed target cell is a physically active structure capable of exerting mechanical force. Here, we investigated whether synaptic forces promote the destruction of target cells. CTLs kill by secreting toxic proteases and the pore forming protein perforin into the synapse. Biophysical experiments revealed a striking correlation between the magnitude of force exertion across the synapse and the speed of perforin pore formation on the target cell, implying that force potentiates cytotoxicity by enhancing perforin activity. Consistent with this interpretation, we found that increasing target cell tension augmented pore formation by perforin and killing by CTLs. Our data also indicate that CTLs coordinate perforin release and force exertion in space and time. These results reveal an unappreciated physical dimension to lymphocyte function and demonstrate that cells use mechanical forces to control the activity of outgoing chemical signals. Copyright © 2016 Elsevier Inc. All rights reserved.

Extracting nuclear sizes of medium to heavy nuclei from total reaction cross sections

NASA Astrophysics Data System (ADS)

Horiuchi, W.; Hatakeyama, S.; Ebata, S.; Suzuki, Y.

2016-04-01

Background: Proton and neutron radii are fundamental quantities of atomic nuclei. To study the sizes of short-lived unstable nuclei, there is a need for an alternative to electron scattering. Purpose: The recent paper by Horiuchi et al. [Phys. Rev. C 89, 011601(R) (2014)], 10.1103/PhysRevC.89.011601 proposed a possible way of extracting the matter and neutron-skin thickness of light- to medium-mass nuclei using total reaction cross section, σR. The analysis is extended to medium to heavy nuclei up to lead isotopes with due attention to Coulomb breakup contributions as well as density distributions improved by paring correlation. Methods: We formulate a quantitative calculation of σR based on the Glauber model including the Coulomb breakup. To substantiate the treatment of the Coulomb breakup, we also evaluate the Coulomb breakup cross section due to the electric dipole field in a canonical-basis-time-dependent-Hartree-Fock-Bogoliubov theory in the three-dimensional coordinate space. Results: We analyze σR's of 103 nuclei with Z =20 , 28, 40, 50, 70, and 82 incident on light targets, H,21, 4He, and 12C. Three kinds of Skyrme interactions are tested to generate those wave functions. To discuss possible uncertainty due to the Coulomb breakup, we examine its dependence on the target, the incident energy, and the Skyrme interaction. The proton is a most promising target for extracting the nuclear sizes as the Coulomb excitation can safely be neglected. We find that the so-called reaction radius, aR=√{σR/π } , for the proton target is very well approximated by a linear function of two variables, the matter radius and the skin thickness, in which three constants depend only on the incident energy. We quantify the accuracy of σR measurements needed to extract the nuclear sizes. Conclusions: The proton is the best target because, once the incident energy is set, its aR is very accurately determined by only the matter radius and neutron-skin thickness. If σR's at

Cell-to-Cell Transmission Can Overcome Multiple Donor and Target Cell Barriers Imposed on Cell-Free HIV

PubMed Central

Ilinskaya, Anna; Dorjbal, Batsukh; Truong, Rosaline; Derse, David; Uchil, Pradeep D.; Heidecker, Gisela; Mothes, Walther

2013-01-01

Virus transmission can occur either by a cell-free mode through the extracellular space or by cell-to-cell transmission involving direct cell-to-cell contact. The factors that determine whether a virus spreads by either pathway are poorly understood. Here, we assessed the relative contribution of cell-free and cell-to-cell transmission to the spreading of the human immunodeficiency virus (HIV). We demonstrate that HIV can spread by a cell-free pathway if all the steps of the viral replication cycle are efficiently supported in highly permissive cells. However, when the cell-free path was systematically hindered at various steps, HIV transmission became contact-dependent. Cell-to-cell transmission overcame barriers introduced in the donor cell at the level of gene expression and surface retention by the restriction factor tetherin. Moreover, neutralizing antibodies that efficiently inhibit cell-free HIV were less effective against cell-to-cell transmitted virus. HIV cell-to-cell transmission also efficiently infected target T cells that were relatively poorly susceptible to cell-free HIV. Importantly, we demonstrate that the donor and target cell types influence critically the extent by which cell-to-cell transmission can overcome each barrier. Mechanistically, cell-to-cell transmission promoted HIV spread to more cells and infected target cells with a higher proviral content than observed for cell-free virus. Our data demonstrate that the frequently observed contact-dependent spread of HIV is the result of specific features in donor and target cell types, thus offering an explanation for conflicting reports on the extent of cell-to-cell transmission of HIV. PMID:23308151

Single-Cell Droplet Microfluidic Screening for Antibodies Specifically Binding to Target Cells.

PubMed

Shembekar, Nachiket; Hu, Hongxing; Eustace, David; Merten, Christoph A

2018-02-20

Monoclonal antibodies are a main player in modern drug discovery. Many antibody screening formats exist, each with specific advantages and limitations. Nonetheless, it remains challenging to screen antibodies for the binding of cell-surface receptors (the most important class of all drug targets) or for the binding to target cells rather than purified proteins. Here, we present a high-throughput droplet microfluidics approach employing dual-color normalized fluorescence readout to detect antibody binding. This enables us to obtain quantitative data on target cell recognition, using as little as 33 fg of IgG per assay. Starting with an excess of hybridoma cells releasing unspecific antibodies, individual clones secreting specific binders (of target cells co-encapsulated into droplets) could be enriched 220-fold after sorting 80,000 clones in a single experiment. This opens the way for therapeutic antibody discovery, especially since the single-cell approach is in principle also applicable to primary human plasma cells. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Heavy neutron rich nuclei: production and investigation

NASA Astrophysics Data System (ADS)

Zemlyanoy, S.; Avvakumov, K.; Kazarinov, N.; Fedosseev, V.; Bark, R.; Blazczak, Z.; Janas, Z.

2018-05-01

For production and investigation of heavy neutron rich nuclei devoted the new setup, which is under construction at Flerov Laboratory for Nuclear Reactions (FLNR) - JINR, Dubna now. This setup is planned to exploit available beams from the U-400M cyclotron in low energy multi-nucleon transfer reactions to study exotic neutron-rich nuclei located in the “north-east” region of nuclear map. Products from 4.5 to 9 MeV/nucleon heavy-ion collisions, such as 136Xe on 208Pb, are to be captured in a gas cell and selectively laser-ionized in a sextupole (quadrupole) ion guide extraction system.

The major nucleoside triphosphatase in pea (Pisum sativum L.) nuclei and in rat liver nuclei share common epitopes also present in nuclear lamins

NASA Technical Reports Server (NTRS)

Tong, C. G.; Dauwalder, M.; Clawson, G. A.; Hatem, C. L.; Roux, S. J.

1993-01-01

The major nucleoside triphosphatase (NTPase) activities in mammalian and pea (Pisum sativum L.) nuclei are associated with enzymes that are very similar both biochemically and immunochemically. The major NTPase from rat liver nuclei appears to be a 46-kD enzyme that represents the N-terminal portion of lamins A and C, two lamina proteins that apparently arise from the same gene by alternate splicing. Monoclonal antibody (MAb) G2, raised to human lamin C, both immunoprecipitates the major (47 kD) NTPase in pea nuclei and recognizes it in western blot analyses. A polyclonal antibody preparation raised to the 47-kD pea NTPase (pc480) reacts with the same lamin bands that are recognized by MAb G2 in mammalian nuclei. The pc480 antibodies also bind to the same lamin-like bands in pea nuclear envelope-matrix preparations that are recognized by G2 and three other MAbs known to bind to mammalian lamins. In immunofluorescence assays, pc480 and anti-lamin antibodies stain both cytoplasmic and nuclear antigens in plant cells, with slightly enhanced staining along the periphery of the nuclei. These results indicate that the pea and rat liver NTPases are structurally similar and that, in pea nuclei as in rat liver nuclei, the major NTPase is probably derived from a lamin precursor by proteolysis.

Modeling multi-nucleon transfer in symmetric collisions of massive nuclei

DOE Office of Scientific and Technical Information (OSTI.GOV)

Welsh, T.; Loveland, W.; Yanez, R.

Symmetric collisions of massive nuclei, such as U-238 + Cm-248, have been proposed as ways to make new n-rich heavy nuclei via multi-nucleon transfer (MNT) reactions. We have measured the yields of several projectile-like and target-like fragments from the reaction of 1360 MeV Hg-204 + Pt-198. We find that current models for this symmetric collision (GRAZING, DNS, ImQMD) significantly underestimate the yields of these transfer products, even for small transfers. (C) 2017 The Author(s). Published by Elsevier B.V.

Integrity of chromatin and replicating DNA in nuclei released from fission yeast by semi-automated grinding in liquid nitrogen

PubMed Central

2011-01-01

Background Studies of nuclear function in many organisms, especially those with tough cell walls, are limited by lack of availability of simple, economical methods for large-scale preparation of clean, undamaged nuclei. Findings Here we present a useful method for nuclear isolation from the important model organism, the fission yeast, Schizosaccharomyces pombe. To preserve in vivo molecular configurations, we flash-froze the yeast cells in liquid nitrogen. Then we broke their tough cell walls, without damaging their nuclei, by grinding in a precision-controlled motorized mortar-and-pestle apparatus. The cryo-ground cells were resuspended and thawed in a buffer designed to preserve nuclear morphology, and the nuclei were enriched by differential centrifugation. The washed nuclei were free from contaminating nucleases and have proven well-suited as starting material for genome-wide chromatin analysis and for preparation of fragile DNA replication intermediates. Conclusions We have developed a simple, reproducible, economical procedure for large-scale preparation of endogenous-nuclease-free, morphologically intact nuclei from fission yeast. With appropriate modifications, this procedure may well prove useful for isolation of nuclei from other organisms with, or without, tough cell walls. PMID:22088094

Integrity of chromatin and replicating DNA in nuclei released from fission yeast by semi-automated grinding in liquid nitrogen.

PubMed

Givens, Robert M; Mesner, Larry D; Hamlin, Joyce L; Buck, Michael J; Huberman, Joel A

2011-11-16

Studies of nuclear function in many organisms, especially those with tough cell walls, are limited by lack of availability of simple, economical methods for large-scale preparation of clean, undamaged nuclei. Here we present a useful method for nuclear isolation from the important model organism, the fission yeast, Schizosaccharomyces pombe. To preserve in vivo molecular configurations, we flash-froze the yeast cells in liquid nitrogen. Then we broke their tough cell walls, without damaging their nuclei, by grinding in a precision-controlled motorized mortar-and-pestle apparatus. The cryo-ground cells were resuspended and thawed in a buffer designed to preserve nuclear morphology, and the nuclei were enriched by differential centrifugation. The washed nuclei were free from contaminating nucleases and have proven well-suited as starting material for genome-wide chromatin analysis and for preparation of fragile DNA replication intermediates. We have developed a simple, reproducible, economical procedure for large-scale preparation of endogenous-nuclease-free, morphologically intact nuclei from fission yeast. With appropriate modifications, this procedure may well prove useful for isolation of nuclei from other organisms with, or without, tough cell walls.

Research with Radioactive Targets

NASA Astrophysics Data System (ADS)

Ahle, Larry

2004-10-01

Obtaining precise information about neutron capture cross-sections for s-process branch points is a key goal of nuclear astrophysics. Since these nuclei are unstable and neutron targets do not exist, performing these measurements require a facility that can produce the nuclei of interest at a sufficient rate to allow formation of a meaningful target (at least 1015 atoms). The Rare Isotope Accelerator (RIA) promises such rates, often enabling collection of greater than 1016 atoms after only of few days of production running. By properly designing both the ISOL and fragmentation lines, these collections will often be possible to obtained parasitically to other radioactive ion beam production. But given a target, performing the neutron capture cross-section measurement also presents its own challenges. In many cases, activation measurements are feasible, providing one obtains a target of sufficient purity. But for many branch point nuclei, the capture product is stable or long enough lived that no radiation signature is available for detection. Measurements for these nuclei will require a BaF2 array like DANCE at Los Alamos National Laboratory, which uses gamma calorimetry to detect neutron capture events. Plans and issues associated with isotope harvesting will be discussed, as well as challenges associated with performing theses measurements. Current plans for doing DANCE type measurements at RIA will also be discussed. This work was performed under the auspices of the U.S. Department of Energy by the University of California, Lawrence Livermore National Laboratory under contract No. W-7405-Eng-48.

Phloem-Conducting Cells in Haustoria of the Root-Parasitic Plant Phelipanche aegyptiaca Retain Nuclei and Are Not Mature Sieve Elements.

PubMed

Ekawa, Minako; Aoki, Koh

2017-12-05

Phelipanche aegyptiaca parasitizes a wide range of plants, including important crops, and causes serious damage to their production. P. aegyptiaca develops a specialized intrusive organ called a haustorium that establishes connections to the host's xylem and phloem. In parallel with the development of xylem vessels, the differentiation of phloem-conducting cells has been demonstrated by the translocation of symplasmic tracers from the host to the parasite. However, it is unclear yet whether haustorial phloem-conducting cells are sieve elements. In this study, we identified phloem-conducting cells in haustoria by the host-to-parasite translocation of green fluorescent protein (GFP) from AtSUC2pro::GFP tomato sieve tubes. Haustorial GFP-conducting cells contained nuclei but not callose-rich sieve plates, indicating that phloem-conducting cells in haustoria differ from conventional sieve elements. To ascertain why the nuclei were not degenerated, expression of the P. aegyptiaca homologs NAC-domain containing transcription factor ( NAC45 ), NAC45/86-dependent exonuclease-domain protein 1 ( NEN1 ), and NEN4 was examined. However, these genes were more highly expressed in the haustorium than in tubercle protrusion, implying that nuclear degradation in haustoria may not be exclusively controlled by the NAC45 / 86 - NEN regulatory pathway. Our results also suggest that the formation of plasmodesmata with large size exclusion limits is independent of nuclear degradation and callose deposition.

Targeted Mutagenesis in Rice Using TALENs and the CRISPR/Cas9 System.

PubMed

Endo, Masaki; Nishizawa-Yokoi, Ayako; Toki, Seiichi

2016-01-01

Sequence-specific nucleases (SSNs), such as zinc-finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and the clustered regularly interspersed short palindromic repeats (CRISPR)/CRISPR-associated protein 9 nuclease (Cas9) system, are powerful tools for understanding gene function and for developing novel traits in plants. In plant species for which transformation and regeneration systems using protoplasts are not yet established, direct delivery to nuclei of SSNs either in the form of RNA or protein is difficult. Thus, Agrobacterium-mediated transformation of SSN expression constructs in cultured cells is a practical means of delivering targeted mutagenesis in some plant species including rice. Because targeted mutagenesis occurs stochastically in transgenic cells and SSN-mediated targeted mutagenesis often leads to no selectable phenotype, identification of highly mutated cell lines is a critical step in obtaining regenerated plants with desired mutations.

Controversies in targeted therapy of adult T cell leukemia/lymphoma: ON target or OFF target effects?

PubMed

Nasr, Rihab; El Hajj, Hiba; Kfoury, Youmna; de Thé, Hugues; Hermine, Olivier; Bazarbachi, Ali

2011-06-01

Adult T cell leukemia/lymphoma (ATL) represents an ideal model for targeted therapy because of intrinsic chemo-resistance of ATL cells and the presence of two well identified targets: the HTLV-I retrovirus and the viral oncoprotein Tax. The combination of zidovudine (AZT) and interferon-alpha (IFN) has a dramatic impact on survival of ATL patients. Although the mechanism of action remains unclear, arguments in favor or against a direct antiviral effect will be discussed. Yet, most patients relapse and alternative therapies are mandatory. IFN and arsenic trioxide induce Tax proteolysis, synergize to induce apoptosis in ATL cells and cure Tax-driven ATL in mice through specific targeting of leukemia initiating cell activity. These results provide a biological basis for the clinical success of arsenic/IFN/AZT therapy in ATL patients and suggest that both extinction of viral replication (AZT) and Tax degradation (arsenic/IFN) are needed to cure ATL.

Nuclei pulposi formation from the embryonic notochord occurs normally in GDF-5-deficient mice.

PubMed

Maier, Jennifer A; Harfe, Brian D

2011-11-15

The transition of the mouse embryonic notochord into nuclei pulposi was determined ("fate mapped") in vivo in growth and differentiating factor-5 (GDF-5)-null mice using the Shhcre and R26R alleles. To determine whether abnormal nuclei pulposi formation from the embryonic notochord was responsible for defects present in adult nuclei pulposi of Gdf-5-null mice. The development, maintenance, and degeneration of the intervertebral disc are not understood. Previously, we demonstrated that all cells in the adult nucleus pulposus of normal mice are derived from the embryonic notochord. Gdf-5-null mice have been reported to contain intervertebral discs in which the nucleus pulposus is abnormal. It is currently unclear if disc defects in Gdf-5-null mice arise during the formation of nuclei pulposi from the notochord during embryogenesis or result from progressive postnatal degeneration of nuclei pulposi. Gdf-5 messenger RNA expression was examined in the discs of wild-type embryos by RNA in situ hybridization to determine when and where this gene was expressed. To examine nucleus pulposus formation in Gdf-5-null mice, intervertebral discs in which embryonic notochord cells were marked were analyzed in newborn and 24-week-old mice. Our Gdf-5 messenger RNA in situ experiments determined that this gene is localized to the annulus fibrosus and not the nucleus pulposus in mouse embryos. Notochord fate-mapping experiments revealed that notochord cells in Gdf-5-null mice correctly form nuclei pulposi. Our data suggest that the defects reported in the nucleus pulposus of adult Gdf-5-null mice do not result from abnormal patterning of the embryonic notochord. The use of mouse alleles to mark cells that produce all cell types that reside in the adult nucleus pulposus will allow for a detailed examination of disc formation in other mouse mutants that have been reported to contain disc defects.

Antibody-targeted interleukin 2 stimulates T-cell killing of autologous tumor cells.

PubMed Central

Gillies, S D; Reilly, E B; Lo, K M; Reisfeld, R A

1992-01-01

A genetically engineered fusion protein consisting of a chimeric anti-ganglioside GD2 antibody (ch14.18) and interleukin 2 (IL2) was tested for its ability to enhance the killing of autologous GD2-expressing melanoma target cells by a tumor-infiltrating lymphocyte line (660 TIL). The fusion of IL2 to the carboxyl terminus of the immunoglobulin heavy chain did not reduce IL2 activity as measured in a standard proliferation assay using either mouse or human T-cell lines. Antigen-binding activity was greater than that of the native chimeric antibody. The ability of resting 660 TIL cells to kill their autologous GD2-positive target cells was enhanced if the target cells were first coated with the fusion protein. This stimulation of killing was greater than that of uncoated cells in the presence of equivalent or higher concentrations of free IL2. Such antibody-cytokine fusion proteins may prove useful in targeting the biological effect of IL2 and other cytokines to tumor cells and in this way stimulate their immune destruction. Images PMID:1741398

Boron neutron capture therapy: Moving toward targeted cancer therapy.

PubMed

Mirzaei, Hamid Reza; Sahebkar, Amirhossein; Salehi, Rasoul; Nahand, Javid Sadri; Karimi, Ehsan; Jaafari, Mahmoud Reza; Mirzaei, Hamed

2016-01-01

Boron neutron capture therapy (BNCT) occurs when a stable isotope, boton-10, is irradiated with low-energy thermal neutrons to yield stripped down helium-4 nuclei and lithium-7 nuclei. It is a binary therapy in the treatment of cancer in which a cytotoxic event is triggered when an atom placed in a cancer cell. Here, we provide an overview on the application of BNCT in cancer therapy as well as current preclinical and clinical evidence on the efficacy of BNCT in the treatment of melanoma, brain tumors, head and neck cancer, and thyroid cancer. Several studies have shown that BNCT is effective in patients who had been treated with a full dose of conventional radiotherapy, because of its selectivity. In addition, BNCT is dependent on the normal/tumor tissue ratio of boron distribution. Increasing evidence has shown that BNCT can be combined with different drug delivery systems to enhance the delivery of boron to cancer cells. The flexibility of BNCT to be used in combination with different tumor-targeting approaches has made this strategy a promising option for cancer therapy. This review aims to provide a state-of-the-art overview of the recent advances in the use of BNCT for targeted therapy of cancer.

Cell-targeted platinum nanoparticles and nanoparticle clusters.

PubMed

Papst, Stefanie; Brimble, Margaret A; Evans, Clive W; Verdon, Daniel J; Feisst, Vaughan; Dunbar, P Rod; Tilley, Richard D; Williams, David E

2015-06-21

Herein, we report the facile preparation of cell-targeted platinum nanoparticles (PtNPs), through the design of peptides that, as a single molecule added in small concentration during the synthesis, control the size of PtNP clusters during their growth, stabilise the PtNPs in aqueous suspension and enable the functionalisation of the PtNPs with a versatile range of cell-targeting ligands. Water-soluble PtNPs targeted respectively at blood group antigens and at integrin receptors are demonstrated.

Controversies in Targeted Therapy of Adult T Cell Leukemia/Lymphoma: ON Target or OFF Target Effects?

PubMed Central

Nasr, Rihab; Hajj, Hiba El; Kfoury, Youmna; de Thé, Hugues; Hermine, Olivier; Bazarbachi, Ali

2011-01-01

Adult T cell leukemia/lymphoma (ATL) represents an ideal model for targeted therapy because of intrinsic chemo-resistance of ATL cells and the presence of two well identified targets: the HTLV-I retrovirus and the viral oncoprotein Tax. The combination of zidovudine (AZT) and interferon-alpha (IFN) has a dramatic impact on survival of ATL patients. Although the mechanism of action remains unclear, arguments in favor or against a direct antiviral effect will be discussed. Yet, most patients relapse and alternative therapies are mandatory. IFN and arsenic trioxide induce Tax proteolysis, synergize to induce apoptosis in ATL cells and cure Tax-driven ATL in mice through specific targeting of leukemia initiating cell activity. These results provide a biological basis for the clinical success of arsenic/IFN/AZT therapy in ATL patients and suggest that both extinction of viral replication (AZT) and Tax degradation (arsenic/IFN) are needed to cure ATL. PMID:21994752

Central Topography of Cranial Motor Nuclei Controlled by Differential Cadherin Expression

PubMed Central

Astick, Marc; Tubby, Kristina; Mubarak, Waleed M.; Guthrie, Sarah; Price, Stephen R.

2014-01-01

Summary Neuronal nuclei are prominent, evolutionarily conserved features of vertebrate central nervous system (CNS) organization [1]. Nuclei are clusters of soma of functionally related neurons and are located in highly stereotyped positions. Establishment of this CNS topography is critical to neural circuit assembly. However, little is known of either the cellular or molecular mechanisms that drive nucleus formation during development, a process termed nucleogenesis [2–5]. Brainstem motor neurons, which contribute axons to distinct cranial nerves and whose functions are essential to vertebrate survival, are organized exclusively as nuclei. Cranial motor nuclei are composed of two main classes, termed branchiomotor/visceromotor and somatomotor [6]. Each of these classes innervates evolutionarily distinct structures, for example, the branchial arches and eyes, respectively. Additionally, each class is generated by distinct progenitor cell populations and is defined by differential transcription factor expression [7, 8]; for example, Hb9 distinguishes somatomotor from branchiomotor neurons. We characterized the time course of cranial motornucleogenesis, finding that despite differences in cellular origin, segregation of branchiomotor and somatomotor nuclei occurs actively, passing through a phase of each being intermingled. We also found that differential expression of cadherin cell adhesion family members uniquely defines each motor nucleus. We show that cadherin expression is critical to nucleogenesis as its perturbation degrades nucleus topography predictably. PMID:25308074

Development of a resonant laser ionization gas cell for high-energy, short-lived nuclei

NASA Astrophysics Data System (ADS)

Sonoda, T.; Wada, M.; Tomita, H.; Sakamoto, C.; Takatsuka, T.; Furukawa, T.; Iimura, H.; Ito, Y.; Kubo, T.; Matsuo, Y.; Mita, H.; Naimi, S.; Nakamura, S.; Noto, T.; Schury, P.; Shinozuka, T.; Wakui, T.; Miyatake, H.; Jeong, S.; Ishiyama, H.; Watanabe, Y. X.; Hirayama, Y.; Okada, K.; Takamine, A.

2013-01-01

A new laser ion source configuration based on resonant photoionization in a gas cell has been developed at RIBF RIKEN. This system is intended for the future PArasitic RI-beam production by Laser Ion-Source (PALIS) project which will be installed at RIKEN's fragment separator, BigRIPS. A novel implementation of differential pumping, in combination with a sextupole ion beam guide (SPIG), has been developed. A few small scroll pumps create a pressure difference from 1000 hPa-10-3 Pa within a geometry drastically miniaturized compared to conventional systems. This system can utilize a large exit hole for fast evacuation times, minimizing the decay loss for short-lived nuclei during extraction from a buffer gas cell, while sufficient gas cell pressure is maintained for stopping high energy RI-beams. In spite of the motion in a dense pressure gradient, the photo-ionized ions inside the gas cell are ejected with an assisting force gas jet and successfully transported to a high-vacuum region via SPIG followed by a quadrupole mass separator. Observed behaviors agree with the results of gas flow and Monte Carlo simulations.

Development and Testing of a 212Pb/212Bi Peptide for Targeting Metastatic Melanoma

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fisher, Darrell R.

2012-10-25

The purpose of this project is to develop a new radiolabeled peptide for imaging and treating metastatic melanoma. The immunoconjugate consists of a receptor-specific peptide that targets melanoma cells. The beta-emitter lead-212 (half-life = 10.4 hours) is linked by coordination chemistry to the peptide. After injection, the peptide targets melanoma receptors on the surfaces of melanoma cells. Lead-212 decays to the alpha-emitter bismuth-212 (half-life = 60 minutes). Alpha-particles that hit melanoma cell nuclei are likely to kill the melanoma cell. For cancer cell imaging, the lead-212 is replaced by lead-203 (half-life = 52 hours). Lead-203 emits 279 keV photons (80.1%more » abundance) that can be imaged and measured for biodistribution analysis, cancer imaging, and quantitative dosimetry.« less

Cell-permeable nanobodies for targeted immunolabelling and antigen manipulation in living cells

NASA Astrophysics Data System (ADS)

Herce, Henry D.; Schumacher, Dominik; Schneider, Anselm F. L.; Ludwig, Anne K.; Mann, Florian A.; Fillies, Marion; Kasper, Marc-André; Reinke, Stefan; Krause, Eberhard; Leonhardt, Heinrich; Cardoso, M. Cristina; Hackenberger, Christian P. R.

2017-08-01

Functional antibody delivery in living cells would enable the labelling and manipulation of intracellular antigens, which constitutes a long-thought goal in cell biology and medicine. Here we present a modular strategy to create functional cell-permeable nanobodies capable of targeted labelling and manipulation of intracellular antigens in living cells. The cell-permeable nanobodies are formed by the site-specific attachment of intracellularly stable (or cleavable) cyclic arginine-rich cell-penetrating peptides to camelid-derived single-chain VHH antibody fragments. We used this strategy for the non-endocytic delivery of two recombinant nanobodies into living cells, which enabled the relocalization of the polymerase clamp PCNA (proliferating cell nuclear antigen) and tumour suppressor p53 to the nucleolus, and thereby allowed the detection of protein-protein interactions that involve these two proteins in living cells. Furthermore, cell-permeable nanobodies permitted the co-transport of therapeutically relevant proteins, such as Mecp2, into the cells. This technology constitutes a major step in the labelling, delivery and targeted manipulation of intracellular antigens. Ultimately, this approach opens the door towards immunostaining in living cells and the expansion of immunotherapies to intracellular antigen targets.

Robust nuclei segmentation in cyto-histopathological images using statistical level set approach with topology preserving constraint

NASA Astrophysics Data System (ADS)

Taheri, Shaghayegh; Fevens, Thomas; Bui, Tien D.

2017-02-01

Computerized assessments for diagnosis or malignancy grading of cyto-histopathological specimens have drawn increased attention in the field of digital pathology. Automatic segmentation of cell nuclei is a fundamental step in such automated systems. Despite considerable research, nuclei segmentation is still a challenging task due noise, nonuniform illumination, and most importantly, in 2D projection images, overlapping and touching nuclei. In most published approaches, nuclei refinement is a post-processing step after segmentation, which usually refers to the task of detaching the aggregated nuclei or merging the over-segmented nuclei. In this work, we present a novel segmentation technique which effectively addresses the problem of individually segmenting touching or overlapping cell nuclei during the segmentation process. The proposed framework is a region-based segmentation method, which consists of three major modules: i) the image is passed through a color deconvolution step to extract the desired stains; ii) then the generalized fast radial symmetry transform is applied to the image followed by non-maxima suppression to specify the initial seed points for nuclei, and their corresponding GFRS ellipses which are interpreted as the initial nuclei borders for segmentation; iii) finally, these nuclei border initial curves are evolved through the use of a statistical level-set approach along with topology preserving criteria for segmentation and separation of nuclei at the same time. The proposed method is evaluated using Hematoxylin and Eosin, and fluorescent stained images, performing qualitative and quantitative analysis, showing that the method outperforms thresholding and watershed segmentation approaches.

Production and investigation of heavy neutron rich nuclei

NASA Astrophysics Data System (ADS)

Zemlyanoy, Sergey; Avvakumov, Konstantin; Kozulin, Eduard; Fedosseev, Valentin; Bark, Robert; Janas, Zenon

2017-11-01

A project devoted to the production and study of neutron rich heavy nuclei (GALS - project) is being realized at Flerov Laboratory for Nuclear Reactions (FLNR) - JINR. GALS is planned to exploit available beams from the U-400M cyclotron in low energy multi-nucleon transfer reactions to study exotic neutron rich nuclei located in the "north-east" region of nuclear map. Products from 4.5 to 9 MeV/nucleon heavy-ion collisions, such as 136Xe on 208Pb, are to be captured in a gas cell and selectively laser-ionized in a sextupole (quadrupole) ion guide extraction system.

Automatic segmentation and supervised learning-based selection of nuclei in cancer tissue images.

PubMed

Nandy, Kaustav; Gudla, Prabhakar R; Amundsen, Ryan; Meaburn, Karen J; Misteli, Tom; Lockett, Stephen J

2012-09-01

Analysis of preferential localization of certain genes within the cell nuclei is emerging as a new technique for the diagnosis of breast cancer. Quantitation requires accurate segmentation of 100-200 cell nuclei in each tissue section to draw a statistically significant result. Thus, for large-scale analysis, manual processing is too time consuming and subjective. Fortuitously, acquired images generally contain many more nuclei than are needed for analysis. Therefore, we developed an integrated workflow that selects, following automatic segmentation, a subpopulation of accurately delineated nuclei for positioning of fluorescence in situ hybridization-labeled genes of interest. Segmentation was performed by a multistage watershed-based algorithm and screening by an artificial neural network-based pattern recognition engine. The performance of the workflow was quantified in terms of the fraction of automatically selected nuclei that were visually confirmed as well segmented and by the boundary accuracy of the well-segmented nuclei relative to a 2D dynamic programming-based reference segmentation method. Application of the method was demonstrated for discriminating normal and cancerous breast tissue sections based on the differential positioning of the HES5 gene. Automatic results agreed with manual analysis in 11 out of 14 cancers, all four normal cases, and all five noncancerous breast disease cases, thus showing the accuracy and robustness of the proposed approach. Published 2012 Wiley Periodicals, Inc.

Bovine oocytes with the potential to reprogram somatic cell nuclei have a unique 23-kDa protein, phosphorylated transcriptionally controlled tumor protein (TCTP).

PubMed

Tani, Tetsuya; Shimada, Hiroaki; Kato, Yoko; Tsunoda, Yukio

2007-01-01

Despite the long-held assumption that reprogramming factors are present in mammalian oocytes at the second metaphase stage, the molecular nature of these factors is not known. Here, we demonstrated that oocytes with the potential to reprogram somatic cell nuclei have a unique 23-kDa protein, phosphorylated transcriptionally controlled tumor protein (TCTP). Injection of TCTP double-stranded RNA into germinal vesicle oocytes decreased the potential of nuclear-transferred (NT) oocytes, but not in vitro fertilized oocytes, to develop into blastocysts. Phosphorylated TCTP is considered to facilitate the first step of somatic cell reprogramming. After transfer of blastocysts that developed from NT oocytes fused with cumulus cells in which phosphorylated TCTP peptide was previously incorporated, the recipient pregnancy rate (47%) increased and the abortion rate (13%) decreased. Moreover, all seven cloned calves survived for at least 1 month after parturition, and had no morphologic abnormalities. The present study demonstrated that pretreatment of donor cells with phosphorylated TCTP peptide has a beneficial effect on the potential of bovine somatic cell nuclei to develop into normal cloned calves. Before widespread application of TCTP for bovine cloning, however, a large-scale embryo transfer study using different donor cell lines of various origins is necessary.

Induction of viral interference by IPNV-carrier cells on target cells: A cell co-culture study.

PubMed

Parreño, Ricardo; Torres, Susana; Almagro, Lucía; Belló-Pérez, Melissa; Estepa, Amparo; Perez, Luis

2016-11-01

IPNV is a salmonid birnavirus that possesses the ability to establish asymptomatic persistent infections in a number of valuable fish species. The presence of IPNV may interfere with subsequent infection by other viruses. In the present study we show that an IPNV-carrier cell line (EPC IPNV ) can induce an antiviral state in fresh EPC by co-cultivating both cell types in three different ways: a "droplet" culture system, a plastic chamber setup, and a transmembrane (Transwell ® ) system. All three cell co-culture methods were proven useful to study donor/target cell interaction. Naïve EPC cells grown in contact with EPC IPNV cells develop resistance to VHSV superinfection. The transmembrane system seems best suited to examine gene expression in donor and target cells separately. Our findings point to the conclusion that one or more soluble factors produced by the IPNV carrier culture induce the innate immune response within the target cells. This antiviral response is associated to the up-regulation of interferon (ifn) and mx gene expression in target EPC cells. To our knowledge this is the first article describing co-culture systems to study the interplay between virus-carrier cells and naive cells in fish. Copyright © 2016 The Author(s). Published by Elsevier Ltd.. All rights reserved.

Synthetic mRNA is a more reliable tool for the delivery of DNA-targeting proteins into the cell nucleus than fusion with a protein transduction domain.

PubMed

Leontovyc, Ivan; Habart, David; Loukotova, Sarka; Kosinova, Lucie; Kriz, Jan; Saudek, Frantisek; Koblas, Tomas

2017-01-01

Cell reprogramming requires efficient delivery of reprogramming transcription factors into the cell nucleus. Here, we compared the robustness and workload of two protein delivery methods that avoid the risk of genomic integration. The first method is based on fusion of the protein of interest to a protein transduction domain (PTD) for delivery across the membranes of target cells. The second method relies on de novo synthesis of the protein of interest inside the target cells utilizing synthetic mRNA (syn-mRNA) as a template. We established a Cre/lox reporter system in three different cell types derived from human (PANC-1, HEK293) and rat (BRIN-BD11) tissues and used Cre recombinase to model a protein of interest. The system allowed constitutive expression of red fluorescence protein (RFP), while green fluorescence protein (GFP) was expressed only after the genomic action of Cre recombinase. The efficiency of protein delivery into cell nuclei was quantified as the frequency of GFP+ cells in the total cell number. The PTD method showed good efficiency only in BRIN-BD11 cells (68%), whereas it failed in PANC-1 and HEK293 cells. By contrast, the syn-mRNA method was highly effective in all three cell types (29-71%). We conclude that using synthetic mRNA is a more robust and less labor-intensive approach than using the PTD-fusion alternative.

Nuclear localization signal targeting to macronucleus and micronucleus in binucleated ciliate Tetrahymena thermophila.

PubMed

Iwamoto, Masaaki; Mori, Chie; Osakada, Hiroko; Koujin, Takako; Hiraoka, Yasushi; Haraguchi, Tokuko

2018-06-08

Ciliated protozoa possess two morphologically and functionally distinct nuclei: a macronucleus (MAC) and a micronucleus (MIC). The MAC is transcriptionally active and functions in all cellular events. The MIC is transcriptionally inactive during cell growth, but functions in meiotic events to produce progeny nuclei. Thus, these two nuclei must be distinguished by the nuclear proteins required for their distinct functions during cellular events such as cell proliferation and meiosis. To understand the mechanism of the nuclear transport specific to either MAC or MIC, we identified specific nuclear localization signals (NLSs) in two MAC- and MIC-specific nuclear proteins, macronuclear histone H1 and micronuclear linker histone-like protein (Mlh1), respectively. By expressing GFP-fused fragments of these proteins in Tetrahymena thermophila cells, two distinct regions in macronuclear histone H1 protein were assigned as independent MAC-specific NLSs and two distinct regions in Mlh1 protein were assigned as independent MIC-specific NLSs. These NLSs contain several essential lysine residues responsible for the MAC- and MIC-specific nuclear transport, but neither contains any consensus sequence with known monopartite or bipartite NLSs in other model organisms. Our findings contribute to understanding how specific nuclear targeting is achieved to perform distinct nuclear functions in binucleated ciliates. © 2018 The Authors. Genes to Cells published by Molecular Biology Society of Japan and John Wiley & Sons Australia, Ltd.

Selection of Phage Display Peptides Targeting Human Pluripotent Stem Cell-Derived Progenitor Cell Lines.

PubMed

Bignone, Paola A; Krupa, Rachel A; West, Michael D; Larocca, David

2016-01-01

The ability of human pluripotent stem cells (hPS) to both self-renew and differentiate into virtually any cell type makes them a promising source of cells for cell-based regenerative therapies. However, stem cell identity, purity, and scalability remain formidable challenges that need to be overcome for translation of pluripotent stem cell research into clinical applications. Directed differentiation from hPS cells is inefficient and residual contamination with pluripotent cells that have the potential to form tumors remains problematic. The derivation of scalable (self-renewing) embryonic progenitor stem cell lines offers a solution because they are well defined and clonally pure. Clonally pure progenitor stem cell lines also provide a means for identifying cell surface targeting reagents that are useful for identification, tracking, and repeated derivation of the corresponding progenitor stem cell types from additional hPS cell sources. Such stem cell targeting reagents can then be applied to the manufacture of genetically diverse banks of human embryonic progenitor cell lines for drug screening, disease modeling, and cell therapy. Here we present methods to identify human embryonic progenitor stem cell targeting peptides by selection of phage display libraries on clonal embryonic progenitor cell lines and demonstrate their use for targeting quantum dots (Qdots) for stem cell labeling.

A method for estimating the accuracy of measurements of optical characteristics of the nuclei of blood cells in the diagnosis of acute leukemia

NASA Astrophysics Data System (ADS)

Polyakov, E. V.; Nikitaev, V. G.

2017-01-01

The work is devoted to investigation of the random component of the measurement error of the nuclei structure characteristics, which are used in the method of structural elements to measure the differences of blood cells of different types. This method is realized in information-measuring system of the analysis of micropreparations of blood cells in the diagnosis of acute leukemia and its variants.

Nuclei pulposi formation from the embryonic notochord occurs normally in GDF5-deficient mice

PubMed Central

Maier, Jennifer A.; Harfe, Brian D.

2011-01-01

Study Design The transition of the mouse embryonic notochord into nuclei pulposi was determined (“fate mapped”) in vivo in GDF-5 null mice using the Shhcre and R26R alleles. Objective To determine if abnormal nuclei pulposi formation from the embryonic notochord was responsible for defects present in adult nuclei pulposi of Gdf-5 null mice. Summary of Background Data The development, maintenance, and degeneration of the intervertebral disc are not understood. Previously, we demonstrated that all cells in the adult nucleus pulposus of normal mice are derived from the embryonic notochord. Gdf-5 null mice have been reported to contain intervertebral discs in which the nucleus pulposus is abnormal. It is currently unclear if disc defects in Gdf-5 null mice arise during the formation of nuclei pulposi from the notochord during embryogenesis or resulted from progressive postnatal degeneration of nuclei pulposi. Methods Gdf-5 mRNA expression was examined in the discs of wild-type embryos by RNA in situ hybridization to determine when and where this gene was expressed. To examine nucleus pulposus formation in Gdf-5 null mice, intervertebral discs in which embryonic notochord cells were marked were analyzed in newborn and 24 week old mice. Results Our Gdf-5 mRNA in situ experiments determined that this gene is localized to the annulus fibrosus and not the nucleus pulposus in mouse embryos. Notochord fate mapping experiments revealed that notochord cells in Gdf-5 null mice correctly form nuclei pulposi. Conclusion Our data suggest that the defects reported in the nucleus pulposus of adult Gdf-5 null mice do not result from abnormal patterning of the embryonic notochord. The use of mouse alleles to mark cells that produce all cell types that reside in the adult nucleus pulposus will allow for a detailed examination of disc formation in other mouse mutants that have been reported to contain disc defects. PMID:21278629

Coherent dissociation of relativistic {sup 9}C nuclei in nuclear track emulsion

DOE Office of Scientific and Technical Information (OSTI.GOV)

Krivenkov, D. O.; Artemenkov, D. A.; Bradnova, V.

2010-04-30

For the first time nuclear track emulsion is exposed to relativistic {sup 9}C nuclei. A systematic pattern of the distributions of charge combinations of fragments in the peripheral interactions of {sup 9}C nuclei in a nuclear track emulsion has been obtained. The main conclusion is that the contribution of the channel {sup 9}C->{sup 8}B+p and {sup 9}C->{sup 7}Be+2p is most important in events that do not involve the production of target-nucleus fragments or mesons (coherent dissociation). It can be concluded that in the peripheral {sup 9}C dissociation the picture hitherto obtained for {sup 8}B and {sup 7}Be with the additionmore » of one or two protons, respectively, is reproduced. Three coherent dissociation events {sup 9}C->3{sup 3}He accompanied by neither target fragments of the nucleus target nor charged mesons are identified.« less

Targeted gene disruption of Hsp70-2 results in failed meiosis, germ cell apoptosis, and male infertility.

PubMed Central

Dix, D J; Allen, J W; Collins, B W; Mori, C; Nakamura, N; Poorman-Allen, P; Goulding, E H; Eddy, E M

1996-01-01

In addition to the five 70-kDa heat shock proteins (HSP70) common to germ cells and somatic tissues of mammals, spermatogenic cells synthesize HSP70-2 during meiosis. To determine if this unique stress protein has a critical role in meiosis, we used gene-targeting techniques to disrupt Hsp70-2 in mice. Male mice homozygous for the mutant allele (Hsp70-2 -/-) did not synthesize HSP70-2, lacked postmeiotic spermatids and mature sperm, and were infertile. However, neither meiosis nor fertility was affected in female Hsp70-2 -/- mice. We previously found that HSP70-2 is associated with synaptonemal complexes in the nucleus of meiotic spermatocytes from mice and hamsters. While synaptonemal complexes assembled in Hsp70-2 -/- spermatocytes, structural abnormalities became apparent in these cells by late prophase, and development rarely progressed to the meiotic divisions. Furthermore, analysis of nuclei and genomic DNA indicated that the failure of meiosis in Hsp70-2 -/- mice was coincident with a dramatic increase in spermatocyte apoptosis. These results suggest that HSP70-2 participates in synaptonemal complex function during meiosis in male germ cells and is linked to mechanisms that inhibit apoptosis. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:8622925

The Quest for Targets Executing MYC-Dependent Cell Transformation.

PubMed

Hartl, Markus

2016-01-01

MYC represents a transcription factor with oncogenic potential converting multiple cellular signals into a broad transcriptional response, thereby controlling the expression of numerous protein-coding and non-coding RNAs important for cell proliferation, metabolism, differentiation, and apoptosis. Constitutive activation of MYC leads to neoplastic cell transformation, and deregulated MYC alleles are frequently observed in many human cancer cell types. Multiple approaches have been performed to isolate genes differentially expressed in cells containing aberrantly activated MYC proteins leading to the identification of thousands of putative targets. Functional analyses of genes differentially expressed in MYC-transformed cells had revealed that so far more than 40 upregulated or downregulated MYC targets are actively involved in cell transformation or tumorigenesis. However, further systematic and selective approaches are required for determination of the known or yet unidentified targets responsible for processing the oncogenic MYC program. The search for critical targets in MYC-dependent tumor cells is exacerbated by the fact that during tumor development, cancer cells progressively evolve in a multistep process, thereby acquiring their characteristic features in an additive manner. Functional expression cloning, combinatorial gene expression, and appropriate in vivo tests could represent adequate tools for dissecting the complex scenario of MYC-specified cell transformation. In this context, the central goal is to identify a minimal set of targets that suffices to phenocopy oncogenic MYC. Recently developed genomic editing tools could be employed to confirm the requirement of crucial transformation-associated targets. Knowledge about essential MYC-regulated genes is beneficial to expedite the development of specific inhibitors to interfere with growth and viability of human tumor cells in which MYC is aberrantly activated. Approaches based on the principle of

The Quest for Targets Executing MYC-Dependent Cell Transformation

PubMed Central

Hartl, Markus

2016-01-01

MYC represents a transcription factor with oncogenic potential converting multiple cellular signals into a broad transcriptional response, thereby controlling the expression of numerous protein-coding and non-coding RNAs important for cell proliferation, metabolism, differentiation, and apoptosis. Constitutive activation of MYC leads to neoplastic cell transformation, and deregulated MYC alleles are frequently observed in many human cancer cell types. Multiple approaches have been performed to isolate genes differentially expressed in cells containing aberrantly activated MYC proteins leading to the identification of thousands of putative targets. Functional analyses of genes differentially expressed in MYC-transformed cells had revealed that so far more than 40 upregulated or downregulated MYC targets are actively involved in cell transformation or tumorigenesis. However, further systematic and selective approaches are required for determination of the known or yet unidentified targets responsible for processing the oncogenic MYC program. The search for critical targets in MYC-dependent tumor cells is exacerbated by the fact that during tumor development, cancer cells progressively evolve in a multistep process, thereby acquiring their characteristic features in an additive manner. Functional expression cloning, combinatorial gene expression, and appropriate in vivo tests could represent adequate tools for dissecting the complex scenario of MYC-specified cell transformation. In this context, the central goal is to identify a minimal set of targets that suffices to phenocopy oncogenic MYC. Recently developed genomic editing tools could be employed to confirm the requirement of crucial transformation-associated targets. Knowledge about essential MYC-regulated genes is beneficial to expedite the development of specific inhibitors to interfere with growth and viability of human tumor cells in which MYC is aberrantly activated. Approaches based on the principle of

Targeting human breast cancer cells by an oncolytic adenovirus using microRNA-targeting strategy.

PubMed

Shayestehpour, Mohammad; Moghim, Sharareh; Salimi, Vahid; Jalilvand, Somayeh; Yavarian, Jila; Romani, Bizhan; Mokhtari-Azad, Talat

2017-08-15

MicroRNA-targeting strategy is a promising approach that enables oncolytic viruses to replicate in tumor cells but not in normal cells. In this study, we targeted adenoviral replication toward breast cancer cells by inserting ten complementary binding sites for miR-145-5p downstream of E1A gene. In addition, we evaluated the effect of increasing miR-145 binding sites on inhibition of virus replication. Ad5-control and adenoviruses carrying five or ten copies of miR145-5p target sites (Ad5-5miR145T, Ad5-10miR145T) were generated and inoculated into MDA-MB-453, BT-20, MCF-7 breast cancer cell lines and human mammary epithelial cells (HMEpC). Titer of Ad5-10miR145T in HMEpC was significantly lower than Ad5-control titer. Difference between the titer of these two viruses at 12, 24, 36, and 48h after infection was 1.25, 2.96, 3.06, and 3.77 log TCID 50 . No significant difference was observed between the titer of both adenoviruses in MDA-MB-453, BT-20 and MCF-7 cells. The infectious titer of adenovirus containing 10 miR-145 binding sites in HMEpC cells at 24, 36, and 48h post-infection was 1.7, 2.08, and 4-fold, respectively, lower than the titer of adenovirus carrying 5 miR-145 targets. Our results suggest that miR-145-targeting strategy provides selectivity for adenovirus replication in breast cancer cells. Increasing the number of miRNA binding sites within the adenoviral genome confers more selectivity for viral replication in cancer cells. Copyright © 2017. Published by Elsevier B.V.

Cell cycle-tailored targeting of metastatic melanoma: Challenges and opportunities.

PubMed

Haass, Nikolas K; Gabrielli, Brian

2017-07-01

The advent of targeted therapies of metastatic melanoma, such as MAPK pathway inhibitors and immune checkpoint antagonists, has turned dermato-oncology from the "bad guy" to the "poster child" in oncology. Current targeted therapies are effective, although here is a clear need to develop combination therapies to delay the onset of resistance. Many antimelanoma drugs impact on the cell cycle but are also dependent on certain cell cycle phases resulting in cell cycle phase-specific drug insensitivity. Here, we raise the question: Have combination trials been abandoned prematurely as ineffective possibly only because drug scheduling was not optimized? Firstly, if both drugs of a combination hit targets in the same melanoma cell, cell cycle-mediated drug insensitivity should be taken into account when planning combination therapies, timing of dosing schedules and choice of drug therapies in solid tumors. Secondly, if the combination is designed to target different tumor cell subpopulations of a heterogeneous tumor, one drug effective in a particular subpopulation should not negatively impact on the other drug targeting another subpopulation. In addition to the role of cell cycle stage and progression on standard chemotherapeutics and targeted drugs, we discuss the utilization of cell cycle checkpoint control defects to enhance chemotherapeutic responses or as targets themselves. We propose that cell cycle-tailored targeting of metastatic melanoma could further improve therapy outcomes and that our real-time cell cycle imaging 3D melanoma spheroid model could be utilized as a tool to measure and design drug scheduling approaches. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Sonoporation of endothelial cells by vibrating targeted microbubbles.

PubMed

Kooiman, Klazina; Foppen-Harteveld, Miranda; van der Steen, Antonius F W; de Jong, Nico

2011-08-25

Molecular imaging using ultrasound makes use of targeted microbubbles. In this study we investigated whether these microbubbles could also be used to induce sonoporation in endothelial cells. Lipid-coated microbubbles were targeted to CD31 and insonified at 1 MHz at low peak negative acoustic pressures at six sequences of 10 cycle sine-wave bursts. Vibration of the targeted microbubbles was recorded with the Brandaris-128 high-speed camera (~13 million frames per second). In total, 31 cells were studied that all had one microbubble (1.2-4.2 micron in diameter) attached per cell. After insonification at 80 kPa, 30% of the cells (n=6) had taken up propidium iodide, while this was 20% (n=1) at 120 kPa and 83% (n=5) at 200 kPa. Irrespective of the peak negative acoustic pressure, uptake of propidium iodide was observed when the relative vibration amplitude of targeted microbubbles was greater than 0.5. No relationship was found between the position of the microbubble on the cell and induction of sonoporation. This study shows that targeted microbubbles can also be used to induce sonoporation, thus making it possible to combine molecular imaging and drug delivery. Copyright © 2011 Elsevier B.V. All rights reserved.

Targeted silver nanoparticles for ratiometric cell phenotyping

NASA Astrophysics Data System (ADS)

Willmore, Anne-Mari A.; Simón-Gracia, Lorena; Toome, Kadri; Paiste, Päärn; Kotamraju, Venkata Ramana; Mölder, Tarmo; Sugahara, Kazuki N.; Ruoslahti, Erkki; Braun, Gary B.; Teesalu, Tambet

2016-04-01

Affinity targeting is used to deliver nanoparticles to cells and tissues. For efficient targeting, it is critical to consider the expression and accessibility of the relevant receptors in the target cells. Here, we describe isotopically barcoded silver nanoparticles (AgNPs) as a tool for auditing affinity ligand receptors in cells. Tumor penetrating peptide RPARPAR (receptor: NRP-1) and tumor homing peptide GKRK (receptor: p32) were used as affinity ligands on the AgNPs. The binding and uptake of the peptide-functionalized AgNPs by cultured PPC-1 prostate cancer and M21 melanoma cells was dependent on the cell surface expression of the cognate peptide receptors. Barcoded peptide-functionalized AgNPs were synthesized from silver and palladium isotopes. The cells were incubated with a cocktail of the barcoded nanoparticles [RPARPAR (R), GKRK (K), and control], and cellular binding and internalization of each type of nanoparticle was assessed by inductively coupled plasma mass spectrometry. The results of isotopic analysis were in agreement with data obtained using optical methods. Using ratiometric measurements, we were able to classify the PPC-1 cell line as mainly NRP-1-positive, with 75 +/- 5% R-AgNP uptake, and the M21 cell line as only p32-positive, with 89 +/- 9% K-AgNP uptake. The isotopically barcoded multiplexed AgNPs are useful as an in vitro ratiometric phenotyping tool and have potential uses in functional evaluation of the expression of accessible homing peptide receptors in vivo.Affinity targeting is used to deliver nanoparticles to cells and tissues. For efficient targeting, it is critical to consider the expression and accessibility of the relevant receptors in the target cells. Here, we describe isotopically barcoded silver nanoparticles (AgNPs) as a tool for auditing affinity ligand receptors in cells. Tumor penetrating peptide RPARPAR (receptor: NRP-1) and tumor homing peptide GKRK (receptor: p32) were used as affinity ligands on the AgNPs. The

A novel double-targeted nondrug delivery system for targeting cancer stem cells

PubMed Central

Qiao, Shupei; Zhao, Yufang; Geng, Shuai; Li, Yong; Hou, Xiaolu; Liu, Yi; Lin, Feng-Huei; Yao, Lifen; Tian, Weiming

2016-01-01

Instead of killing cancer stem cells (CSCs), the conventional chemotherapy used for cancer treatment promotes the enrichment of CSCs, which are responsible for tumor growth, metastasis, and recurrence. However, most therapeutic agents are only able to kill a small proportion of CSCs by targeting one or two cell surface markers or dysregulated CSC pathways, which are usually shared with normal stem cells (NSCs). In this study, we developed a novel nondrug delivery system for the dual targeting of CSCs by conjugating hyaluronic acid (HA) and grafting the doublecortin-like kinase 1 (DCLK1) monoclonal antibody to the surface of poly(ethylene glycol) (PEG)–poly(d,l-lactide-co-glycolide) (PLGA) nanoparticles (NPs), which can specifically target CD44 receptors and the DCLK1 surface marker – the latter was shown to possess the capacity to distinguish between CSCSs and NSCs. The size and morphology of these NPs were characterized by dynamic light scattering (DLS), transmission electron microscopy (TEM), and scanning electron microscopy (SEM). This was followed by studies of NP encapsulation efficiency and in vitro drug release properties. Then, the cytotoxicity of the NPs was tested via Cell Counting Kit-8 assay. Finally, the 4T1 CSCs were obtained from the alginate-based platform, which we developed as an in vitro tumor model. Tumor-bearing nude mice were used as in vivo models to systematically detect the ability of NPs to target CSCs. Our results showed that the DCLK1–HA–PEG–PLGA NPs exhibited a targeting effect toward CSCs both in vitro and in vivo. These findings have important implications for the rational design of drug delivery systems that target CSCs with high efficacy. PMID:27994463

Bigger Brains or Bigger Nuclei? Regulating the Size of Auditory Structures in Birds

PubMed Central

Kubke, M. Fabiana; Massoglia, Dino P.; Carr, Catherine E.

2012-01-01

Increases in the size of the neuronal structures that mediate specific behaviors are believed to be related to enhanced computational performance. It is not clear, however, what developmental and evolutionary mechanisms mediate these changes, nor whether an increase in the size of a given neuronal population is a general mechanism to achieve enhanced computational ability. We addressed the issue of size by analyzing the variation in the relative number of cells of auditory structures in auditory specialists and generalists. We show that bird species with different auditory specializations exhibit variation in the relative size of their hindbrain auditory nuclei. In the barn owl, an auditory specialist, the hind-brain auditory nuclei involved in the computation of sound location show hyperplasia. This hyperplasia was also found in songbirds, but not in non-auditory specialists. The hyperplasia of auditory nuclei was also not seen in birds with large body weight suggesting that the total number of cells is selected for in auditory specialists. In barn owls, differences observed in the relative size of the auditory nuclei might be attributed to modifications in neurogenesis and cell death. Thus, hyperplasia of circuits used for auditory computation accompanies auditory specialization in different orders of birds. PMID:14726625

Formation of Neutron-Enriched Heavy and Superheavy Nuclei in Fusion Reactions

NASA Astrophysics Data System (ADS)

Karpov, A. V.; Rachkov, V. A.; Saiko, V. V.

2018-05-01

The formation of new isotopes of heavy and superheavy elements in the fusion of neutron-enriched projectiles with actinide targets is discussed. Cross sections for the formation of evaporation residues in fusion reactions is predicted for several combinations of colliding nuclei.

[Automated morphometric evaluation of the chromatin structure of liver cell nuclei after vagotomy].

PubMed

Butusova, N N; Zhukotskiĭ, A V; Sherbo, I V; Gribkov, E N; Dubovaia, T K

1989-05-01

The morphometric analysis of the interphase chromatine structure of the hepatic cells nuclei was carried out on the automated TV installation for the quantitative analysis of images "IBAS-2" (by the OPTON firm, the FRG) according to 50 optical and geometric parameters during various periods (1.2 and 4 weeks) after the vagotomy operation. It is determined that upper-molecular organisation of chromatine undergoes the biggest changes one week after operation, and changes of granular component are more informative than changes of the nongranular component (with the difference 15-20%). It was also revealed that chromatine components differ in tinctorial properties, which are evidently dependent on physicochemical characteristics of the chromatine under various functional conditions of the cell. As a result of the correlation analysis the group of morphometric indices of chromatine structure was revealed, which are highly correlated with level of transcription activity of chromatine during various terms after denervation. The correlation quotient of these parameters is 0.85-0.97. The summing up: vagus denervation of the liver causes changes in the morphofunctional organisation of the chromatine.

Advanced cell therapies: targeting, tracking and actuation of cells with magnetic particles.

PubMed

Connell, John J; Patrick, P Stephen; Yu, Yichao; Lythgoe, Mark F; Kalber, Tammy L

2015-01-01

Regenerative medicine would greatly benefit from a new platform technology that enabled measurable, controllable and targeting of stem cells to a site of disease or injury in the body. Superparamagnetic iron-oxide nanoparticles offer attractive possibilities in biomedicine and can be incorporated into cells, affording a safe and reliable means of tagging. This review describes three current and emerging methods to enhance regenerative medicine using magnetic particles to guide therapeutic cells to a target organ; track the cells using MRI and assess their spatial localization with high precision and influence the behavior of the cell using magnetic actuation. This approach is complementary to the systemic injection of cell therapies, thus expanding the horizon of stem cell therapeutics.

Microscopic few-body and Gaussian-shaped density distributions for the analysis of the 6He exotic nucleus with different target nuclei

NASA Astrophysics Data System (ADS)

Aygun, M.; Kucuk, Y.; Boztosun, I.; Ibraheem, Awad A.

2010-12-01

The elastic scattering angular distributions of 6He projectile on different medium and heavy mass target nuclei including 12C, 27Al, 58Ni, 64Zn, 65Cu, 197Au, 208Pb and 209Bi have been examined by using the few-body and Gaussian-shaped density distributions at various energies. The microscopic real parts of the complex nuclear optical potential have been obtained by using the double-folding model for each of the density distributions and the phenomenological imaginary potentials have been taken as the Woods-Saxon type. Comparative results of the few-body and Gaussian-shaped density distributions together with the experimental data are presented within the framework of the optical model.

A flexible and robust approach for segmenting cell nuclei from 2D microscopy images using supervised learning and template matching

PubMed Central

Chen, Cheng; Wang, Wei; Ozolek, John A.; Rohde, Gustavo K.

2013-01-01

We describe a new supervised learning-based template matching approach for segmenting cell nuclei from microscopy images. The method uses examples selected by a user for building a statistical model which captures the texture and shape variations of the nuclear structures from a given dataset to be segmented. Segmentation of subsequent, unlabeled, images is then performed by finding the model instance that best matches (in the normalized cross correlation sense) local neighborhood in the input image. We demonstrate the application of our method to segmenting nuclei from a variety of imaging modalities, and quantitatively compare our results to several other methods. Quantitative results using both simulated and real image data show that, while certain methods may work well for certain imaging modalities, our software is able to obtain high accuracy across several imaging modalities studied. Results also demonstrate that, relative to several existing methods, the template-based method we propose presents increased robustness in the sense of better handling variations in illumination, variations in texture from different imaging modalities, providing more smooth and accurate segmentation borders, as well as handling better cluttered nuclei. PMID:23568787

Self-targeted salinomycin-loaded DSPE-PEG-methotrexate nanomicelles for targeting both head and neck squamous cell carcinoma cancer cells and cancer stem cells.

PubMed

Zhu, Minhui; Chen, Shicai; Hua, Libo; Zhang, Caiyun; Chen, Mengjie; Chen, Donghui; Dong, Yinmei; Zhang, Yingying; Li, Meng; Song, Xianmin; Chen, Huaiwen; Zheng, Hongliang

2017-02-01

To target both head and neck squamous cell carcinoma (HNSCC) cells and cancer stem cells (CSCs) by salinomycin-loaded DSPE-PEG-MTX (synthesized using DSPE-PEG2000-NH2 and methotrexate) nanomicelles (M-SAL-MTX). The characterization, antitumor activity and mechanism of M-SAL-MTX were evaluated. M-SAL-MTX showed enhanced inhibitory effect toward both HNSCC CSCs and non-CSCs compared with a single treatment of methotrexate and salinomycin. In nude mice-bearing HNSCC xenografts, M-SAL-MTX suppressed tumor growth more effectively than other controls including combination of methotrexate and salinomycin. Therefore, M-SAL-MTX may provide a strategy for treating HNSCC by targeting both HNSCC CSCs and HNSCC cells.

Detection of high-grade atypia nuclei in breast cancer imaging

NASA Astrophysics Data System (ADS)

Noël, Henri; Roux, Ludovic; Lu, Shijian; Boudier, Thomas

2015-03-01

Along with mitotic count, nuclear pleomorphism or nuclear atypia is an important criterion for the grading of breast cancer in histopathology. Though some works have been done in mitosis detection (ICPR 2012,1 MICCAI 2013,2 and ICPR 2014), not much work has been dedicated to automated nuclear atypia grading, especially the most difficult task of detection of grade 3 nuclei. We propose the use of Convolutional Neural Networks for the automated detection of cell nuclei, using images from the three grades of breast cancer for training. The images were obtained from ICPR contests. Additional manual annotation was performed to classify pixels into five classes: stroma, nuclei, lymphocytes, mitosis and fat. At total of 3,000 thumbnail images of 101 × 101 pixels were used for training. By dividing this training set in an 80/20 ratio we could obtain good training results (around 90%). We tested our CNN on images of the three grades which were not in the training set. High grades nuclei were correctly classified. We then thresholded the classification map and performed basic analysis to keep only rounded objects. Our results show that mostly all atypical nuclei were correctly detected.

An innovative pre-targeting strategy for tumor cell specific imaging and therapy

NASA Astrophysics Data System (ADS)

Qin, Si-Yong; Peng, Meng-Yun; Rong, Lei; Jia, Hui-Zhen; Chen, Si; Cheng, Si-Xue; Feng, Jun; Zhang, Xian-Zheng

2015-08-01

A programmed pre-targeting system for tumor cell imaging and targeting therapy was established based on the ``biotin-avidin'' interaction. In this programmed functional system, transferrin-biotin can be actively captured by tumor cells with the overexpression of transferrin receptors, thus achieving the pre-targeting modality. Depending upon avidin-biotin recognition, the attachment of multivalent FITC-avidin to biotinylated tumor cells not only offered the rapid fluorescence labelling, but also endowed the pre-targeted cells with targeting sites for the specifically designed biotinylated peptide nano-drug. Owing to the successful pre-targeting, tumorous HepG2 and HeLa cells were effectively distinguished from the normal 3T3 cells via fluorescence imaging. In addition, the self-assembled peptide nano-drug resulted in enhanced cell apoptosis in the observed HepG2 cells. The tumor cell specific pre-targeting strategy is applicable for a variety of different imaging and therapeutic agents for tumor treatments.A programmed pre-targeting system for tumor cell imaging and targeting therapy was established based on the ``biotin-avidin'' interaction. In this programmed functional system, transferrin-biotin can be actively captured by tumor cells with the overexpression of transferrin receptors, thus achieving the pre-targeting modality. Depending upon avidin-biotin recognition, the attachment of multivalent FITC-avidin to biotinylated tumor cells not only offered the rapid fluorescence labelling, but also endowed the pre-targeted cells with targeting sites for the specifically designed biotinylated peptide nano-drug. Owing to the successful pre-targeting, tumorous HepG2 and HeLa cells were effectively distinguished from the normal 3T3 cells via fluorescence imaging. In addition, the self-assembled peptide nano-drug resulted in enhanced cell apoptosis in the observed HepG2 cells. The tumor cell specific pre-targeting strategy is applicable for a variety of different imaging

Stromal cells in breast cancer as a potential therapeutic target

PubMed Central

Dykes, Samantha S.; Hughes, Veronica S.; Wiggins, Jennifer M.; Fasanya, Henrietta O.; Tanaka, Mai; Siemann, Dietmar

2018-01-01

Breast cancer in the United States is the second most commonly diagnosed cancer in women. About 1 in 8 women will develop invasive breast cancer over the course of her lifetime and breast cancer remains the second leading cause of cancer-related death. In pursuit of novel therapeutic strategies, researchers have examined the tumor microenvironment as a potential anti-cancer target. In addition to neoplastic cells, the tumor microenvironment is composed of several critical normal cell types, including fibroblasts, vascular and lymph endothelial cells, osteoclasts, adipocytes, and immune cells. These cells have important roles in healthy tissue stasis, which frequently are altered in tumors. Indeed, tumor-associated stromal cells often contribute to tumorigenesis, tumor progression, and metastasis. Consequently, these host cells may serve as a possible target in anti-tumor and anti-metastatic therapeutic strategies. Targeting the tumor associated host cells offers the benefit that such cells do not mutate and develop resistance in response to treatment, a major cause of failure in cancer therapeutics targeting neoplastic cells. This review discusses the role of host cells in the tumor microenvironment during tumorigenesis, progression, and metastasis, and provides an overview of recent developments in targeting these cell populations to enhance cancer therapy efficacy.

Isolation of Cardiomyocyte Nuclei from Post-mortem Tissue

PubMed Central

Bergmann, Olaf; Jovinge, Stefan

2012-01-01

Identification of cardiomyocyte nuclei has been challenging in tissue sections as most strategies rely only on cytoplasmic marker proteins1. Rare events in cardiac myocytes such as proliferation and apoptosis require an accurate identification of cardiac myocyte nuclei to analyze cellular renewal in homeostasis and in pathological conditions2. Here, we provide a method to isolate cardiomyocyte nuclei from post mortem tissue by density sedimentation and immunolabeling with antibodies against pericentriolar material 1 (PCM-1) and subsequent flow cytometry sorting. This strategy allows a high throughput analysis and isolation with the advantage of working equally well on fresh tissue and frozen archival material. This makes it possible to study material already collected in biobanks. This technique is applicable and tested in a wide range of species and suitable for multiple downstream applications such as carbon-14 dating3, cell-cycle analysis4, visualization of thymidine analogues (e.g. BrdU and IdU)4, transcriptome and epigenetic analysis. PMID:22805241

Protective Effects of Scutellarin on Human Cardiac Microvascular Endothelial Cells against Hypoxia-Reoxygenation Injury and Its Possible Target-Related Proteins

PubMed Central

Shi, Meina; Liu, Yingting; Feng, Lixing; Cui, Yingbo; Chen, Yajuan; Wang, Peng; Wu, Wenjuan; Chen, Chen; Liu, Xuan; Yang, Weimin

2015-01-01

Scutellarin (SCU) is one of the main components of traditional Chinese medicine plant Erigeron breviscapus (Vant.) Hand.-Mazz. In this paper, we studied the protective effects of SCU on human cardiac microvascular endothelial cells (HCMECs) against hypoxia-reoxygenation (HR) injury and its possible target-related proteins. Results of MTT assay showed that pretreatment of SCU at doses of 1, 5, and 10 μM for 2 h could significantly inhibit the decrease in cell viability of HCMECs induced by HR injury. Subcellular fractions of cells treated with vehicle control, 1 μM SCU, HR injury, or 1 μM SCU + HR injury were separated by ultracentrifugation. The protein expression profiles of cytoplasm and membrane/nuclei fractions were checked using protein two-dimensional electrophoresis (2-DE). Proteins differentially expressed between control and SCU-treated group, control and HR group, or HR and SCU + HR group were identified using mass spectrometry (MS/MS). Possible interaction network of these target-related proteins was predicted using bioinformatic analysis. The influence of SCU on the expression levels of these proteins was confirmed using Western blotting assay. The results indicated that proteins such as p27BBP protein (EIF6), heat shock 60 kDa protein 1 (HSPD1), and chaperonin containing TCP1 subunit 6A isoform (CCT6A) might play important roles in the effects of SCU. PMID:26557144

Protective Effects of Scutellarin on Human Cardiac Microvascular Endothelial Cells against Hypoxia-Reoxygenation Injury and Its Possible Target-Related Proteins.

PubMed

Shi, Meina; Liu, Yingting; Feng, Lixing; Cui, Yingbo; Chen, Yajuan; Wang, Peng; Wu, Wenjuan; Chen, Chen; Liu, Xuan; Yang, Weimin

2015-01-01

Scutellarin (SCU) is one of the main components of traditional Chinese medicine plant Erigeron breviscapus (Vant.) Hand.-Mazz. In this paper, we studied the protective effects of SCU on human cardiac microvascular endothelial cells (HCMECs) against hypoxia-reoxygenation (HR) injury and its possible target-related proteins. Results of MTT assay showed that pretreatment of SCU at doses of 1, 5, and 10 μM for 2 h could significantly inhibit the decrease in cell viability of HCMECs induced by HR injury. Subcellular fractions of cells treated with vehicle control, 1 μM SCU, HR injury, or 1 μM SCU + HR injury were separated by ultracentrifugation. The protein expression profiles of cytoplasm and membrane/nuclei fractions were checked using protein two-dimensional electrophoresis (2-DE). Proteins differentially expressed between control and SCU-treated group, control and HR group, or HR and SCU + HR group were identified using mass spectrometry (MS/MS). Possible interaction network of these target-related proteins was predicted using bioinformatic analysis. The influence of SCU on the expression levels of these proteins was confirmed using Western blotting assay. The results indicated that proteins such as p27BBP protein (EIF6), heat shock 60 kDa protein 1 (HSPD1), and chaperonin containing TCP1 subunit 6A isoform (CCT6A) might play important roles in the effects of SCU.

Mast cell proteases as pharmacological targets

PubMed Central

Caughey, George H.

2015-01-01

Mast cells are rich in proteases, which are the major proteins of intracellular granules and are released with histamine and heparin by activated cells. Most of these proteases are active in the granule as well outside of the mast cell when secreted, and can cleave targets near degranulating mast cells and in adjoining tissue compartments. Some proteases released from mast cells reach the bloodstream and may have far-reaching actions. In terms of relative amounts, the major mast cell proteases include the tryptases, chymases, cathepsin G, carboxypeptidase A3, dipeptidylpeptidase I/cathepsin C, and cathepsins L and S. Some mast cells also produce granzyme B, plasminogen activators, and matrix metalloproteinases. Tryptases and chymases are almost entirely mast cell-specific, whereas other proteases, such as cathepsins G, C, and L are expressed by a variety of inflammatory cells. Carboxypeptidase A3 expression is a property shared by basophils and mast cells. Other proteases, such as mastins, are largely basophil-specific, although human basophils are protease-deficient compared with their murine counterparts. The major classes of mast cell proteases have been targeted for development of therapeutic inhibitors. Also, a human β-tryptase has been proposed as a potential drug itself, to inactivate of snake venins. Diseases linked to mast cell proteases include allergic diseases, such as asthma, eczema, and anaphylaxis, but also include non-allergic diseases such inflammatory bowel disease, autoimmune arthritis, atherosclerosis, aortic aneurysms, hypertension, myocardial infarction, heart failure, pulmonary hypertension and scarring diseases of lungs and other organs. In some cases, studies performed in mouse models suggest protective or homeostatic roles for specific proteases (or groups of proteases) in infections by bacteria, worms and other parasites, and even in allergic inflammation. At the same time, a clearer picture has emerged of differences in the properties

The dynamic shuttling of SIRT1 between cytoplasm and nuclei in bronchial epithelial cells by single and repeated cigarette smoke exposure

PubMed Central

Yanagisawa, Satoru; Baker, Jonathan R.; Vuppusetty, Chaitanya; Koga, Takeshi; Colley, Thomas; Fenwick, Peter; Donnelly, Louise E.; Barnes, Peter J.

2018-01-01

SIRT1 (silent information regulator 2 homolog 1) is a crucial cellular survival protein especially in oxidative stress environments, and has been thought to locate within the nuclei, but also known to shuttle between cytoplasm and nuclei in some cell types. Here, we show for the first time the dynamics of SIRT1 in the presence of single or concurrent cigarette smoke extract (CSE) exposure in human bronchial epithelial cells (HBEC). In BEAS-2B HBEC or primary HBEC, SIRT1 was localized predominantly in cytoplasm, and the CSE (3%) induced nuclear translocation of SIRT1 from cytoplasm in the presence of L-buthionine sulfoximine (an irreversible inhibitor of γ-glutamylcystein synthetase), mainly through the activation of phosphatidylinositol 3-kinase (PI3K) α subunit. This SIRT1 nuclear shuttling was associated with FOXO3a nuclear translocation and the strong induction of several anti-oxidant genes including superoxide dismutase (SOD) 2 and 3; therefore seemed to be an adaptive response. When BEAS-2B cells were pretreated with repeated exposure to a lower concentration of CSE (0.3%), the CSE-induced SIRT1 shuttling and resultant SOD2/3 mRNA induction were significantly impaired. Thus, this result offers a useful cell model to mimic the impaired anti-oxidant capacity in cigarette smoking-associated lung disease such as chronic obstructive pulmonary disease. PMID:29509781

Targeting Stromal Recruitment by Prostate Cancer Cells

DTIC Science & Technology

2006-03-01

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

Magnetic stem cell targeting to the inner ear

NASA Astrophysics Data System (ADS)

Le, T. N.; Straatman, L.; Yanai, A.; Rahmanian, R.; Garnis, C.; Häfeli, U. O.; Poblete, T.; Westerberg, B. D.; Gregory-Evans, K.

2017-12-01

Severe sensorineural deafness is often accompanied by a loss of auditory neurons in addition to injury of the cochlear epithelium and hair cell loss. Cochlear implant function however depends on a healthy complement of neurons and their preservation is vital in achieving optimal results. We have developed a technique to target mesenchymal stem cells (MSCs) to a deafened rat cochlea. We then assessed the neuroprotective effect of systematically delivered MSCs on the survival and function of spiral ganglion neurons (SGNs). MSCs were labeled with superparamagnetic nanoparticles, injected via the systemic circulation, and targeted using a magnetized cochlea implant and external magnet. Neurotrophic factor concentrations, survival of SGNs, and auditory function were assessed at 1 week and 4 weeks after treatments and compared against multiple control groups. Significant numbers of magnetically targeted MSCs (>30 MSCs/section) were present in the cochlea with accompanied elevation of brain-derived neurotrophic factor and glial cell-derived neurotrophic factor levels (p < 0.001). In addition we saw improved survival of SGNs (approximately 80% survival at 4 weeks). Hearing threshold levels in magnetically targeted rats were found to be significantly better than those of control rats (p < 0.05). These results indicate that magnetic targeting of MSCs to the cochlea can be accomplished with a magnetized cochlear permalloy implant and an external magnet. The targeted stem cells release neurotrophic factors which results in improved SGN survival and hearing recovery. Combining magnetic cell-based therapy and cochlear implantation may improve cochlear implant function in treating deafness.

Surface-modified gold nanorods for specific cell targeting

NASA Astrophysics Data System (ADS)

Wang, Chan-Ung; Arai, Yoshie; Kim, Insun; Jang, Wonhee; Lee, Seonghyun; Hafner, Jason H.; Jeoung, Eunhee; Jung, Deokho; Kwon, Youngeun

2012-05-01

Gold nanoparticles (GNPs) have unique properties that make them highly attractive materials for developing functional reagents for various biomedical applications including photothermal therapy, targeted drug delivery, and molecular imaging. For in vivo applications, GNPs need to be prepared with very little or negligible cytotoxicitiy. Most GNPs are, however, prepared using growth-directing surfactants such as cetyl trimethylammonium bromide (CTAB), which are known to have considerable cytotoxicity. In this paper, we describe an approach to remove CTAB to a non-toxic concentration. We optimized the conditions for surface modification with methoxypolyethylene glycol thiol (mPEG), which replaced CTAB and formed a protective layer on the surface of gold nanorods (GNRs). The cytotoxicities of pristine and surface-modified GNRs were measured in primary human umbilical vein endothelial cells and human cell lines derived from hepatic carcinoma cells, embryonic kidney cells, and thyroid papillary carcinoma cells. Cytotoxicity assays revealed that treating cells with GNRs did not significantly affect cell viability except for thyroid papillary carcinoma cells. Thyroid cancer cells were more susceptible to residual CTAB, so CTAB had to be further removed by dialysis in order to use GNRs for thyroid cell targeting. PEGylated GNRs are further modified to present monoclonal antibodies that recognize a specific surface marker, Na-I symporter, for thyroid cells. Antibody-conjugated GNRs specifically targeted human thyroid cells in vitro.

Chimeric antigen receptor T cells targeting Fc μ receptor selectively eliminate CLL cells while sparing healthy B cells.

PubMed

Faitschuk, Elena; Hombach, Andreas A; Frenzel, Lukas P; Wendtner, Clemens-Martin; Abken, Hinrich

2016-09-29

Adoptive cell therapy of chronic lymphocytic leukemia (CLL) with chimeric antigen receptor (CAR)-modified T cells targeting CD19 induced lasting remission of this refractory disease in a number of patients. However, the treatment is associated with prolonged "on-target off-tumor" toxicities due to the targeted elimination of healthy B cells demanding more selectivity in targeting CLL cells. We identified the immunoglobulin M Fc receptor (FcμR), also known as the Fas apoptotic inhibitory molecule-3 or TOSO, as a target for a more selective treatment of CLL by CAR T cells. FcμR is highly and consistently expressed by CLL cells; only minor levels are detected on healthy B cells or other hematopoietic cells. T cells with a CAR specific for FcμR efficiently responded toward CLL cells, released a panel of proinflammatory cytokines and lytic factors, like soluble FasL and granzyme B, and eliminated the leukemic cells. In contrast to CD19 CAR T cells, anti-FcμR CAR T cells did not attack healthy B cells. T cells with anti-FcμR CAR delayed outgrowth of Mec-1-induced leukemia in a xenograft mouse model. T cells from CLL patients in various stages of the disease, modified by the anti-FcμR CAR, purged their autologous CLL cells in vitro without reducing the number of healthy B cells, which is the case with anti-CD19 CAR T cells. Compared with the currently used therapies, the data strongly imply a superior therapeutic index of anti-FcμR CAR T cells for the treatment of CLL. © 2016 by The American Society of Hematology.

Curcumin targets fibroblast–tumor cell interactions in oral squamous cell carcinoma

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dudás, József, E-mail: jozsef.dudas@i-med.ac.at; Fullár, Alexandra, E-mail: fullarsz@gmail.com; 1st Department of Pathology and Experimental Cancer Research, Semmelweis University, Üllői út 26, 1085 Budapest

Co-culture of periodontal ligament fibroblasts (PDLs) and SCC-25 oral squamous carcinoma cells (OSCC) results in conversion of PDLs into carcinoma-associated fibroblasts (CAFs) and induces epithelial-to mesenchymal transition (EMT) of OSCC tumor cells. We hypothesized that Curcumin targets this dynamic mutual interaction between CAFs and tumor cells. Normal and 2 μM Curcumin-treated co-culture were performed for 4 days, followed by analysis of tumor cell invasivity, mRNA/protein expression of EMT-markers and mediators, activity measure of matrix metalloproteinase 9 (MMP-9), and western blot analysis of signal transduction in tumor cells and fibroblasts. In Curcumin-treated co-culture, in tumor cells, the levels of nuclear factormore » κB (NFκBα) and early response kinase (ERK)—decreased, in fibroblasts, integrin αv protein synthesis decreased compared to corresponding cells in normal co-culture. The signal modulatory changes induced by Curcumin caused decreased release of EMT-mediators in CAFs and reversal of EMT in tumor cells, which was associated with decreased invasion. These data confirm the palliative potential of Curcumin in clinical application. - Graphical abstract: Co-culture of periodontal ligament fibroblasts (PDLs) and SCC-25 oral squamous carcinoma cells (OSCC) results in conversion of PDLs into carcinoma-associated fibroblasts (CAFs) and induces epithelial-to mesenchymal transition (EMT) of tumor cells. Curcumin targets this dynamic mutual interaction between CAFs and tumor cells by inhibiting the production of EMT mediators in CAFs and by modification of intracellular signaling in tumor cells. This causes less invasivity and reversal of EMT in tumor cells. Highlights: ► Curcumin targets tumor–fibroblast interaction in head and neck cancer. ► Curcumin suppresses mediators of epithelial–mesenchymal transition. ► Curcumin decreases the invasivity of tumor cells.« less

Three-Dimensional Maps of All Chromosomes in Human Male Fibroblast Nuclei and Prometaphase Rosettes

PubMed Central

Bolzer, Andreas; Kreth, Gregor; Solovei, Irina; Koehler, Daniela; Saracoglu, Kaan; Fauth, Christine; Müller, Stefan; Eils, Roland; Cremer, Christoph; Speicher, Michael R

2005-01-01

Studies of higher-order chromatin arrangements are an essential part of ongoing attempts to explore changes in epigenome structure and their functional implications during development and cell differentiation. However, the extent and cell-type-specificity of three-dimensional (3D) chromosome arrangements has remained controversial. In order to overcome technical limitations of previous studies, we have developed tools that allow the quantitative 3D positional mapping of all chromosomes simultaneously. We present unequivocal evidence for a probabilistic 3D order of prometaphase chromosomes, as well as of chromosome territories (CTs) in nuclei of quiescent (G0) and cycling (early S-phase) human diploid fibroblasts (46, XY). Radial distance measurements showed a probabilistic, highly nonrandom correlation with chromosome size: small chromosomes—independently of their gene density—were distributed significantly closer to the center of the nucleus or prometaphase rosette, while large chromosomes were located closer to the nuclear or rosette rim. This arrangement was independently confirmed in both human fibroblast and amniotic fluid cell nuclei. Notably, these cell types exhibit flat-ellipsoidal cell nuclei, in contrast to the spherical nuclei of lymphocytes and several other human cell types, for which we and others previously demonstrated gene-density-correlated radial 3D CT arrangements. Modeling of 3D CT arrangements suggests that cell-type-specific differences in radial CT arrangements are not solely due to geometrical constraints that result from nuclear shape differences. We also found gene-density-correlated arrangements of higher-order chromatin shared by all human cell types studied so far. Chromatin domains, which are gene-poor, form a layer beneath the nuclear envelope, while gene-dense chromatin is enriched in the nuclear interior. We discuss the possible functional implications of this finding. PMID:15839726

Cell-type-specific, Aptamer-functionalized Agents for Targeted Disease Therapy

PubMed Central

Zhou, Jiehua; Rossi, John J.

2014-01-01

One hundred years ago, Dr. Paul Ehrlich popularized the “magic bullet” concept for cancer therapy in which an ideal therapeutic agent would only kill the specific tumor cells it targeted. Since then, “targeted therapy” that specifically targets the molecular defects responsible for a patient's condition has become a long-standing goal for treating human disease. However, safe and efficient drug delivery during the treatment of cancer and infectious disease remains a major challenge for clinical translation and the development of new therapies. The advent of SELEX technology has inspired many groundbreaking studies that successfully adapted cell-specific aptamers for targeted delivery of active drug substances in both in vitro and in vivo models. By covalently linking or physically functionalizing the cell-specific aptamers with therapeutic agents, such as siRNA, microRNA, chemotherapeutics or toxins, or delivery vehicles, such as organic or inorganic nanocarriers, the targeted cells and tissues can be specifically recognized and the therapeutic compounds internalized, thereby improving the local concentration of the drug and its therapeutic efficacy. Currently, many cell-type-specific aptamers have been developed that can target distinct diseases or tissues in a cell-type-specific manner. In this review, we discuss recent advances in the use of cell-specific aptamers for targeted disease therapy, as well as conjugation strategies and challenges. PMID:24936916

Trispecific antibodies for CD16A-directed NK cell engagement and dual-targeting of tumor cells.

PubMed

Gantke, Thorsten; Weichel, Michael; Herbrecht, Carmen; Reusch, Uwe; Ellwanger, Kristina; Fucek, Ivica; Eser, Markus; Müller, Thomas; Griep, Remko; Molkenthin, Vera; Zhukovsky, Eugene A; Treder, Martin

2017-09-01

Bispecific antibodies that redirect the lytic activity of cytotoxic immune effector cells, such as T- and NK cells, onto tumor cells have emerged as a highly attractive and clinically validated treatment modality for hematological malignancies. Advancement of this therapeutic concept into solid tumor indications, however, is hampered by the scarcity of targetable antigens that are surface-expressed on tumor cells but demonstrate only limited expression on healthy tissues. To overcome this limitation, the concept of dual-targeting, i.e. the simultaneous targeting of two tumor-expressed surface antigens with limited co-expression on non-malignant cells, with multispecific antibodies has been proposed to increase tumor selectivity of antibody-induced effector cell cytotoxicity. Here, a novel CD16A (FcγRIIIa)-directed trispecific, tetravalent antibody format, termed aTriFlex, is described, that is capable of redirecting NK cell cytotoxicity to two surface-expressed antigens. Using a BCMA/CD200-based in vitro model system, the potential use of aTriFlex antibodies for dual-targeting and selective induction of NK cell-mediated target cell lysis was investigated. Bivalent bispecific target cell binding was found to result in significant avidity gains and up to 17-fold increased in vitro potency. These data suggest trispecific aTriFlex antibodies may support dual-targeting strategies to redirect NK cell cytotoxicity with increased selectivity to enable targeting of solid tumor antigens. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Dendritic cell targeted vaccines: Recent progresses and challenges

PubMed Central

Chen, Pengfei; Liu, Xinsheng; Sun, Yuefeng; Zhou, Peng; Wang, Yonglu; Zhang, Yongguang

2016-01-01

ABSTRACT Dendritic cells (DCs) are known to be a set of morphology, structure and function of heterogeneous professional antigen presenting cells (APCs), as well as the strongest functional antigen presenting cells, which can absorb, process and present antigens. As the key regulators of innate and adaptive immune responses, DCs are at the center of the immune system and capable of interacting with both B cells and T cells, thereby manipulating the humoral and cellular immune responses. DCs provide an essential link between the innate and adaptive immunity, and the strong immune activation function of DCs and their properties of natural adjuvants, make them a valuable target for antigen delivery. Targeting antigens to DC-specific endocytic receptors in combination with the relevant antibodies or ligands along with immunostimulatory adjuvants has been recently recognized as a promising strategy for designing an effective vaccine that elicits a strong and durable T cell response against intracellular pathogens and cancer. This opinion article provides a brief summary of the rationales, superiorities and challenges of existing DC-targeting approaches. PMID:26513200

Integral refractive index imaging of flowing cell nuclei using quantitative phase microscopy combined with fluorescence microscopy.

PubMed

Dardikman, Gili; Nygate, Yoav N; Barnea, Itay; Turko, Nir A; Singh, Gyanendra; Javidi, Barham; Shaked, Natan T

2018-03-01

We suggest a new multimodal imaging technique for quantitatively measuring the integral (thickness-average) refractive index of the nuclei of live biological cells in suspension. For this aim, we combined quantitative phase microscopy with simultaneous 2-D fluorescence microscopy. We used 2-D fluorescence microscopy to localize the nucleus inside the quantitative phase map of the cell, as well as for measuring the nucleus radii. As verified offline by both 3-D confocal fluorescence microscopy and 2-D fluorescence microscopy while rotating the cells during flow, the nucleus of cells in suspension that are not during division can be assumed to be an ellipsoid. The entire shape of a cell in suspension can be assumed to be a sphere. Then, the cell and nucleus 3-D shapes can be evaluated based on their in-plain radii available from the 2-D phase and fluorescent measurements, respectively. Finally, the nucleus integral refractive index profile is calculated. We demonstrate the new technique on cancer cells, obtaining nucleus refractive index values that are lower than those of the cytoplasm, coinciding with recent findings. We believe that the proposed technique has the potential to be used for flow cytometry, where full 3-D refractive index tomography is too slow to be implemented during flow.

Selective tumor cell targeting by the disaccharide moiety of bleomycin.

PubMed

Yu, Zhiqiang; Schmaltz, Ryan M; Bozeman, Trevor C; Paul, Rakesh; Rishel, Michael J; Tsosie, Krystal S; Hecht, Sidney M

2013-02-27

In a recent study, the well-documented tumor targeting properties of the antitumor agent bleomycin (BLM) were studied in cell culture using microbubbles that had been derivatized with multiple copies of BLM. It was shown that BLM selectively targeted MCF-7 human breast carcinoma cells but not the "normal" breast cell line MCF-10A. Furthermore, it was found that the BLM analogue deglycobleomycin, which lacks the disaccharide moiety of BLM, did not target either cell line, indicating that the BLM disaccharide moiety is necessary for tumor selectivity. Not resolved in the earlier study were the issues of whether the BLM disaccharide moiety alone is sufficient for tumor cell targeting and the possible cellular uptake of the disaccharide. In the present study, we conjugated BLM, deglycoBLM, and BLM disaccharide to the cyanine dye Cy5**. It was found that the BLM and BLM disaccharide conjugates, but not the deglycoBLM conjugate, bound selectively to MCF-7 cells and were internalized. The same was also true for the prostate cancer cell line DU-145 (but not for normal PZ-HPV-7 prostate cells) and for the pancreatic cancer cell line BxPC-3 (but not for normal SVR A221a pancreas cells). The targeting efficiency of the disaccharide was only slightly less than that of BLM in MCF-7 and DU-145 cells and comparable to that of BLM in BxPC-3 cells. These results establish that the BLM disaccharide is both necessary and sufficient for tumor cell targeting, a finding with obvious implications for the design of novel tumor imaging and therapeutic agents.

Using single nuclei for RNA-seq to capture the transcriptome of postmortem neurons

PubMed Central

Krishnaswami, Suguna Rani; Grindberg, Rashel V; Novotny, Mark; Venepally, Pratap; Lacar, Benjamin; Bhutani, Kunal; Linker, Sara B; Pham, Son; Erwin, Jennifer A; Miller, Jeremy A; Hodge, Rebecca; McCarthy, James K; Kelder, Martin; McCorrison, Jamison; Aevermann, Brian D; Fuertes, Francisco Diez; Scheuermann, Richard H; Lee, Jun; Lein, Ed S; Schork, Nicholas; McConnell, Michael J; Gage, Fred H; Lasken, Roger S

2016-01-01

A protocol is described for sequencing the transcriptome of a cell nucleus. Nuclei are isolated from specimens and sorted by FACS, cDNA libraries are constructed and RNA-seq is performed, followed by data analysis. Some steps follow published methods (Smart-seq2 for cDNA synthesis and Nextera XT barcoded library preparation) and are not described in detail here. Previous single-cell approaches for RNA-seq from tissues include cell dissociation using protease treatment at 30 °C, which is known to alter the transcriptome. We isolate nuclei at 4 °C from tissue homogenates, which cause minimal damage. Nuclear transcriptomes can be obtained from postmortem human brain tissue stored at −80 °C, making brain archives accessible for RNA-seq from individual neurons. The method also allows investigation of biological features unique to nuclei, such as enrichment of certain transcripts and precursors of some noncoding RNAs. By following this procedure, it takes about 4 d to construct cDNA libraries that are ready for sequencing. PMID:26890679

An innovative pre-targeting strategy for tumor cell specific imaging and therapy.

PubMed

Qin, Si-Yong; Peng, Meng-Yun; Rong, Lei; Jia, Hui-Zhen; Chen, Si; Cheng, Si-Xue; Feng, Jun; Zhang, Xian-Zheng

2015-09-21

A programmed pre-targeting system for tumor cell imaging and targeting therapy was established based on the "biotin-avidin" interaction. In this programmed functional system, transferrin-biotin can be actively captured by tumor cells with the overexpression of transferrin receptors, thus achieving the pre-targeting modality. Depending upon avidin-biotin recognition, the attachment of multivalent FITC-avidin to biotinylated tumor cells not only offered the rapid fluorescence labelling, but also endowed the pre-targeted cells with targeting sites for the specifically designed biotinylated peptide nano-drug. Owing to the successful pre-targeting, tumorous HepG2 and HeLa cells were effectively distinguished from the normal 3T3 cells via fluorescence imaging. In addition, the self-assembled peptide nano-drug resulted in enhanced cell apoptosis in the observed HepG2 cells. The tumor cell specific pre-targeting strategy is applicable for a variety of different imaging and therapeutic agents for tumor treatments.

Strategies to target non-T-cell HIV reservoirs.

PubMed

Sacha, Jonah B; Ndhlovu, Lishomwa C

2016-07-01

A central question for the HIV cure field is to determine new ways to target clinically relevant, latently and actively replicating HIV-infected cells beyond resting memory CD4 T cells, particularly in anatomical areas of low drug penetrability. HIV eradication strategies being positioned for targeting HIV for extinction in the CD4 T-cell compartment may also show promise in non-CD4 T-cells reservoirs. Furthermore, several exciting novel therapeutic approaches specifically focused on HIV clearance from non-CD4 T-cell populations are being developed. Although reservoir validity in these non-CD4 T cells continues to remain debated, this review will highlight recent advances and make an argument as to their clinical relevancy as we progress towards an HIV cure.

Potential targets for lung squamous cell carcinoma

Cancer.gov

Researchers have identified potential therapeutic targets in lung squamous cell carcinoma, the second most common form of lung cancer. The Cancer Genome Atlas (TCGA) Research Network study comprehensively characterized the lung squamous cell carcinoma gen

N-acetylgalactosamine-functionalized dendrimers as hepatic cancer cell-targeted carriers.

PubMed

Medina, Scott H; Tekumalla, Venkatesh; Chevliakov, Maxim V; Shewach, Donna S; Ensminger, William D; El-Sayed, Mohamed E H

2011-06-01

There is an urgent need for novel polymeric carriers that can selectively deliver a large dose of chemotherapeutic agents into hepatic cancer cells to achieve high therapeutic activity with minimal systemic side effects. PAMAM dendrimers are characterized by a unique branching architecture and a large number of chemical surface groups suitable for coupling of chemotherapeutic agents. In this article, we report the coupling of N-acetylgalactosamine (NAcGal) to generation 5 (G5) of poly(amidoamine) (PAMAM-NH₂) dendrimers via peptide and thiourea linkages to prepare NAcGal-targeted carriers used for targeted delivery of chemotherapeutic agents into hepatic cancer cells. We describe the uptake of NAcGal-targeted and non-targeted G5 dendrimers into hepatic cancer cells (HepG2) as a function of G5 concentration and incubation time. We examine the contribution of the asialoglycoprotein receptor (ASGPR) to the internalization of NAcGal-targeted dendrimers into hepatic cancer cells through a competitive inhibition assay. Our results show that uptake of NAcGal-targeted G5 dendrimers into hepatic cancer cells occurs via ASGPR-mediated endocytosis. Internalization of these targeted carriers increased with the increase in G5 concentration and incubation time following Michaelis-Menten kinetics characteristic of receptor-mediated endocytosis. These results collectively indicate that G5-NAcGal conjugates function as targeted carriers for selective delivery of chemotherapeutic agents into hepatic cancer cells. Copyright © 2011 Elsevier Ltd. All rights reserved.

Cell cycle proteins as promising targets in cancer therapy.

PubMed

Otto, Tobias; Sicinski, Piotr

2017-01-27

Cancer is characterized by uncontrolled tumour cell proliferation resulting from aberrant activity of various cell cycle proteins. Therefore, cell cycle regulators are considered attractive targets in cancer therapy. Intriguingly, animal models demonstrate that some of these proteins are not essential for proliferation of non-transformed cells and development of most tissues. By contrast, many cancers are uniquely dependent on these proteins and hence are selectively sensitive to their inhibition. After decades of research on the physiological functions of cell cycle proteins and their relevance for cancer, this knowledge recently translated into the first approved cancer therapeutic targeting of a direct regulator of the cell cycle. In this Review, we focus on proteins that directly regulate cell cycle progression (such as cyclin-dependent kinases (CDKs)), as well as checkpoint kinases, Aurora kinases and Polo-like kinases (PLKs). We discuss the role of cell cycle proteins in cancer, the rationale for targeting them in cancer treatment and results of clinical trials, as well as the future therapeutic potential of various cell cycle inhibitors.

Otolith-Canal Convergence In Vestibular Nuclei Neurons

NASA Technical Reports Server (NTRS)

Dickman, J. David; Si, Xiao-Hong

2002-01-01

The current final report covers the period from June 1, 1999 to May 31, 2002. The primary objective of the investigation was to determine how information regarding head movements and head position relative to gravity is received and processed by central vestibular nuclei neurons in the brainstem. Specialized receptors in the vestibular labyrinths of the inner ear function to detect angular and linear accelerations of the head, with receptors located in the semicircular canals transducing rotational head movements and receptors located in the otolith organs transducing changes in head position relative to gravity or linear accelerations of the head. The information from these different receptors is then transmitted to central vestibular nuclei neurons which process the input signals, then project the appropriate output information to the eye, head, and body musculature motor neurons to control compensatory reflexes. Although a number of studies have reported on the responsiveness of vestibular nuclei neurons, it has not yet been possible to determine precisely how these cells combine the information from the different angular and linear acceleration receptors into a correct neural output signal. In the present project, rotational and linear motion stimuli were separately delivered while recording responses from vestibular nuclei neurons that were characterized according to direct input from the labyrinth and eye movement sensitivity. Responses from neurons receiving convergent input from the semicircular canals and otolith organs were quantified and compared to non-convergent neurons.

Involvement of microtubules and 10-nm filaments in the movement and positioning of nuclei in syncytia

PubMed Central

1979-01-01

Previous studies (Holmes, K.V., and P.W. Choppin. J. Exp. Med. 124:501- 520; J. Cell Biol. 39:526-543) showed that infection of baby hamster kidney (BHK21-F) cells with the parainfluenza virus SV5 causes extensive cell fusion, that nuclei migrate in the syncytial cytoplasm and align in tightly-packed rows, and that microtubules are involved in nuclear movement and alignment. The role of microtubules, 10-nm filaments, and actin-containing microfilaments in this process has been investigated by immunofluorescence microscopy using specific antisera, time-lapse cinematography, and electron microscopy. During cell fusion, micro tubules and 10-nm filaments from many cells form large bundles which are localized between rows of nuclei. No organized bundles of actin fibers were detected in these areas, although actin fibers were observed in regions away from the aligned nuclei. Although colchicine disrupts microtubules and inhibits nuclear movement, cytochalasin B (CB; 20-50 microgram/ml) does not inhibit cell fusion or nuclear movement. However, CB alters the shape of the syncytium, resulting in long filamentous processes extending from a central region. When these processes from neighboring cells make contact, fusion occurs, and nuclei migrate through the channels which are formed. Electron and immunofluorescence microscopy reveal bundles of microtubules and 10-nm filaments in parallel arrays within these processes, but no bundles of microfilaments were detected. The effect of CB on the structural integrity of microfilaments at this high concentration (20 microgram/ml) was demonstrated by the disappearance of filaments interacting with heavy meromyosin. Cycloheximide (20 microgram/ml) inhibits protein synthesis but does not affect cell fusion, the formation of microtubules and 10-nm filament bundles, or nuclear migration and alignment; thus, continued protein synthesis is not required. The association of microtubules and 10-nm filaments with nuclear migration and

Physics of Unstable Nuclei

NASA Astrophysics Data System (ADS)

Khoa, Dao Tien; Egelhof, Peter; Gales, Sydney; Giai, Nguyen Van; Motobayashi, Tohru

2008-04-01

Studies at the RIKEN RI beam factory / T. Motobayashi -- Dilute nuclear states / M. Freer -- Studies of exotic systems using transfer reactions at GANIL / D. Beaumel et al. -- First results from the Magnex large-acceptance spectrometer / A. Cunsolo et al. -- The ICHOR project and spin-isospin physics with unstable beams / H. Sakai -- Structure and low-lying states of the [symbol]He exotic nucleus via direct reactions on proton / V. Lapoux et al. -- Shell gap below [symbol]Sn based on the excited states in [symbol]Cd and [symbol]In / M. Górska -- Heavy neutron-rich nuclei produced in the fragmentation of a [symbol]Pb beam / Zs. Podolyák et al. -- Breakup and incomplete fusion in reactions of weakly-bound nuclei / D.J. Hinde et al. -- Excited states of [symbol]B and [symbol]He and their cluster aspect / Y. Kanada-En'yo et al. -- Nuclear reactions with weakly-bound systems: the treatment of the continuum / C. H. Dasso, A. Vitturi -- Dynamic evolution of three-body decaying resonances / A. S. Jensen et al. -- Prerainbow oscillations in [symbol]He scattering from the Hoyle state of [symbol]C and alpha particle condensation / S. Ohkubo, Y. Hirabayashi -- Angular dispersion behavior in heavy ion elastic scattering / Q. Wang et al. -- Microscopic optical potential in relativistic approach / Z.Yu. Ma et al. -- Exotic nuclei studied in direct reactions at low momentum transfer - recent results and future perspectives at fair / P. Egelhof -- Isotopic temperatures and symmetry energy in spectator fragmentation / M. De Napoli et al. -- Multi-channel algebraic scattering theory and the structure of exotic compound nuclei / K. Amos et al. -- Results for the first feasibility study for the EXL project at the experimental storage ring at GSI / N. Kalantar-Nayestanaki et al. -- Coulomb excitation of ISOLDE neutron-rich beams along the Z = 28 chain / P. Van Duppen -- The gamma decay of the pygmy resonance far from stability and the GDR at finite temperature / G. Benzoni et al

Folate-conjugated immunoglobulin targets melanoma tumor cells for NK cell effector functions

PubMed Central

Skinner, Cassandra C.; McMichael, Elizabeth L.; Jaime-Ramirez, Alena C.; Abrams, Zachary B.; Lee, Robert J.; Carson, William E.

2016-01-01

The folate receptor (FR) is over-expressed on the vascular side of cancerous cells including those of the breast, ovaries, testes, and cervix. We hypothesized that a folate-conjugated immunoglobulin (F-IgG) would bind to the FR that is over-expressed on melanoma tumor cells to target these cells for lysis by natural killer (NK) cells. Folate receptor expression was confirmed in the Mel-39 (human melanoma) cell line by flow cytometry and immunoblot analysis, using KB (human oral epithelial) and F01 (human melanoma) as a positive and negative control, respectively. FR-positive and negative cell lines were treated with F-IgG or control immunoglobulin G (C-IgG) in the presence or absence of cytokines in order to determine NK cell ability to lyse FR-positive cell lines. NK cell activation was significantly upregulated and lysis of Mel 39 tumor cells enhanced following treatment with F-IgG, as compared to C-IgG at all effector:target (E:T) ratios (p<0.01). This trend was further enhanced by NK cell stimulation with the activating cytokine interleukin-12 (IL-12). NK cell production of cytokines such as interferon-gamma (IFN-γ), macrophage inflammatory protein 1 alpha (MIP-1α), and regulated on activation normal T-cell expressed and secreted (RANTES) were also significantly increased in response to co-stimulation with IL-12 stimulation and F-IgG-coated Mel 39 target cells, as compared to controls (p<0.01). In contrast, F-IgG did not bind to the FR-negative cell line F01 and had no significant effect on NK cell lysis or cytokine production. This research indicates the potential use of F-IgG for its ability to induce an immune response from NK cells against FR-positive melanoma tumor cells which can be further enhanced by the addition of cytokines. PMID:27035691

Curcumin: a promising agent targeting cancer stem cells.

PubMed

Zang, Shufei; Liu, Tao; Shi, Junping; Qiao, Liang

2014-01-01

Cancer stem cells are a subset of cells that are responsible for cancer initiation and relapse. They are generally resistant to the current anticancer agents. Successful anticancer therapy must consist of approaches that can target not only the differentiated cancer cells, but also cancer stem cells. Emerging evidence suggested that the dietary agent curcumin exerted its anti-cancer activities via targeting cancer stem cells of various origins such as those of colorectal cancer, pancreatic cancer, breast cancer, brain cancer, and head and neck cancer. In order to enhance the therapeutic potential of curcumin, this agent has been modified or used in combination with other agents in the experimental therapy for many cancers. In this mini-review, we discussed the effect of curcumin and its derivatives in eliminating cancer stem cells and the possible underlying mechanisms.

The mechanism of T-cell mediated cytotoxicity. VI. T-cell projections and their role in target cell killing.

PubMed Central

Sanderson, C J; Glauert, A M

1979-01-01

Electron micrographs of material fixed during the first 10 min of a T-cell cytotoxic system showed T-cell projections and T-cell burrowing into target cells. These observations were made possible by using a system with a very high rate of killing. The projections vary in shape and size, and can push deeply into the target cell, distorting organelles in their path, including the nucleus. The projections contain fine fibrillar material, to the exclusion of organelles. They push the target cell membrane in front of them to form pockets approximating to the shape of the projection. Areas of close contact occur between the projections and the target cell membrane, particularly at the leading edges. The likelihood that these projections develop as a result of contact with specific antigen, and are involved in the cytotoxic mechanism is discussed. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 Figure 10 Figure 11 Figure 12 Figure 13 Figure 14 Figure 15 Figure 16 PMID:311336

Targeting survival pathways in chronic myeloid leukaemia stem cells

PubMed Central

Sinclair, A; Latif, A L; Holyoake, T L

2013-01-01

Chronic myeloid leukaemia (CML) is a clonal myeloproliferative disorder characterized by the presence of a fusion oncogene BCR-ABL, which encodes a protein with constitutive TK activity. The implementation of tyrosine kinase inhibitors (TKIs) marked a major advance in CML therapy; however, there are problems with current treatment. For example, relapse occurs when these drugs are discontinued in the majority of patients who have achieved a complete molecular response on TKI and these agents are less effective in patients with mutations in the BCR-ABL kinase domain. Importantly, TKI can effectively target proliferating mature cells, but do not eradicate quiescent leukaemic stem cells (LSCs), therefore allowing disease persistence despite treatment. It is essential that alternative strategies are used to target the LSC population. BCR-ABL activation is responsible for the modulation of different signalling pathways, which allows the LSC fraction to evade cell death. Several pathways have been shown to be modulated by BCR-ABL, including PI3K/AKT/mTOR, JAK-STAT and autophagy signalling pathways. Targeting components of these survival pathways, alone or in combination with TKI, therefore represents an attractive potential therapeutic approach for targeting the LSC. However, many pathways are also active in normal stem cells. Therefore, potential targets must be validated to effectively eradicate CML stem cells while sparing normal counterparts. This review summarizes the main pathways modulated in CML stem cells, the recent developments and the use of novel drugs to target components in these pathways which may be used to target the LSC population. Linked Articles This article is part of a themed section on Emerging Therapeutic Aspects in Oncology. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2013.169.issue-8 PMID:23517124

Breast cancer stem cells, EMT and therapeutic targets

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kotiyal, Srishti; Bhattacharya, Susinjan, E-mail: s.bhattacharya@jiit.ac.in

Highlights: • Therapeutic targeting or inhibition of the key molecules of signaling pathways can control growth of breast cancer stem cells (BCSCs). • Development of BCSCs also involves miRNA interactions. • Therapeutic achievement can be done by targeting identified targets in the BCSC pathways. - Abstract: A small heterogeneous population of breast cancer cells acts as seeds to induce new tumor growth. These seeds or breast cancer stem cells (BCSCs) exhibit great phenotypical plasticity which allows them to undergo “epithelial to mesenchymal transition” (EMT) at the site of primary tumor and a future reverse transition. Apart from metastasis they aremore » also responsible for maintaining the tumor and conferring it with drug and radiation resistance and a tendency for post-treatment relapse. Many of the signaling pathways involved in induction of EMT are involved in CSC generation and regulation. Here we are briefly reviewing the mechanism of TGF-β, Wnt, Notch, TNF-α, NF-κB, RTK signalling pathways which are involved in EMT as well as BCSCs maintenance. Therapeutic targeting or inhibition of the key/accessory players of these pathways could control growth of BCSCs and hence malignant cancer. Additionally several miRNAs are dysregulated in cancer stem cells indicating their roles as oncogenes or tumor suppressors. This review also lists the miRNA interactions identified in BCSCs and discusses on some newly identified targets in the BCSC regulatory pathways like SHIP2, nicastrin, Pin 1, IGF-1R, pro-inflammatory cytokines and syndecan which can be targeted for therapeutic achievements.« less

Detection of nuclei in 4D Nomarski DIC microscope images of early Caenorhabditis elegans embryos using local image entropy and object tracking

PubMed Central

Hamahashi, Shugo; Onami, Shuichi; Kitano, Hiroaki

2005-01-01

Background The ability to detect nuclei in embryos is essential for studying the development of multicellular organisms. A system of automated nuclear detection has already been tested on a set of four-dimensional (4D) Nomarski differential interference contrast (DIC) microscope images of Caenorhabditis elegans embryos. However, the system needed laborious hand-tuning of its parameters every time a new image set was used. It could not detect nuclei in the process of cell division, and could detect nuclei only from the two- to eight-cell stages. Results We developed a system that automates the detection of nuclei in a set of 4D DIC microscope images of C. elegans embryos. Local image entropy is used to produce regions of the images that have the image texture of the nucleus. From these regions, those that actually detect nuclei are manually selected at the first and last time points of the image set, and an object-tracking algorithm then selects regions that detect nuclei in between the first and last time points. The use of local image entropy makes the system applicable to multiple image sets without the need to change its parameter values. The use of an object-tracking algorithm enables the system to detect nuclei in the process of cell division. The system detected nuclei with high sensitivity and specificity from the one- to 24-cell stages. Conclusion A combination of local image entropy and an object-tracking algorithm enabled highly objective and productive detection of nuclei in a set of 4D DIC microscope images of C. elegans embryos. The system will facilitate genomic and computational analyses of C. elegans embryos. PMID:15910690

Single-Cell Imaging Using Radioluminescence Microscopy Reveals Unexpected Binding Target for [18F]HFB.

PubMed

Kiru, Louise; Kim, Tae Jin; Shen, Bin; Chin, Frederick T; Pratx, Guillem

2018-06-01

Cell-based therapies are showing great promise for a variety of diseases, but remain hindered by the limited information available regarding the biological fate, migration routes and differentiation patterns of infused cells in trials. Previous studies have demonstrated the feasibility of using positron emission tomography (PET) to track single cells utilising an approach known as positron emission particle tracking (PEPT). The radiolabel hexadecyl-4-[ 18 F]fluorobenzoate ([ 18 F]HFB) was identified as a promising candidate for PEPT, due to its efficient and long-lasting labelling capabilities. The purpose of this work was to characterise the labelling efficiency of [ 18 F]HFB in vitro at the single-cell level prior to in vivo studies. The binding efficiency of [ 18 F]HFB to MDA-MB-231 and Jurkat cells was verified in vitro using bulk gamma counting. The measurements were subsequently repeated in single cells using a new method known as radioluminescence microscopy (RLM) and binding of the radiolabel to the single cells was correlated with various fluorescent dyes. Similar to previous reports, bulk cell labelling was significantly higher with [ 18 F]HFB (18.75 ± 2.47 dpm/cell, n = 6) than 2-deoxy-2-[ 18 F]fluoro-D-glucose ([ 18 F]FDG) (7.59 ± 0.73 dpm/cell, n = 7; p ≤ 0.01). However, single-cell imaging using RLM revealed that [ 18 F]HFB accumulation in live cells (8.35 ± 1.48 cpm/cell, n = 9) was not significantly higher than background levels (4.83 ± 0.52 cpm/cell, n = 12; p > 0.05) and was 1.7-fold lower than [ 18 F]FDG uptake in the same cell line (14.09 ± 1.90 cpm/cell, n = 13; p < 0.01). Instead, [ 18 F]HFB was found to bind significantly to fragmented membranes associated with dead cell nuclei, suggesting an alternative binding target for [ 18 F]HFB. This study demonstrates that bulk analysis alone does not always accurately portray the labelling efficiency, therefore highlighting the need for more routine screening of

Plasmonic nanobubbles for target cell-specific gene and drug delivery and multifunctional processing of heterogeneous cell systems

NASA Astrophysics Data System (ADS)

Lukianova-Hleb, Ekaterina Y.; Huye, Leslie E.; Brenner, Malcolm K.; Lapotko, Dmitri O.

2014-03-01

Cell and gene cancer therapies require ex vivo cell processing of human grafts. Such processing requires at least three steps - cell enrichment, cell separation (destruction), and gene transfer - each of which requires the use of a separate technology. While these technologies may be satisfactory for research use, they are of limited usefulness in the clinical treatment setting because they have a low processing rate, as well as a low transfection and separation efficacy and specificity in heterogeneous human grafts. Most problematic, because current technologies are administered in multiple steps - rather than in a single, multifunctional, and simultaneous procedure - they lengthen treatment process and introduce an unnecessary level of complexity, labor, and resources into clinical treatment; all these limitations result in high losses of valuable cells. We report a universal, high-throughput, and multifunctional technology that simultaneously (1) inject free external cargo in target cells, (2) destroys unwanted cells, and (3) preserve valuable non-target cells in heterogeneous grafts. Each of these functions has single target cell specificity in heterogeneous cell system, processing rate > 45 mln cell/min, injection efficacy 90% under 96% viability of the injected cells, target cell destruction efficacy > 99%, viability of not-target cells >99% The developed technology employs novel cellular agents, called plasmonic nanobubbles (PNBs). PNBs are not particles, but transient, intracellular events, a vapor nanobubbles that expand and collapse in mere nanoseconds under optical excitation of gold nanoparticles with short picosecond laser pulses. PNBs of different, cell-specific, size (1) inject free external cargo with small PNBs, (2) Destroy other target cells mechanically with large PNBs and (3) Preserve non-target cells. The multi-functionality, precision, and high throughput of all-in-one PNB technology will tremendously impact cell and gene therapies and other

Cardiomyocyte cell cycle control and growth estimation in vivo--an analysis based on cardiomyocyte nuclei.

PubMed

Walsh, Stuart; Pontén, Annica; Fleischmann, Bernd K; Jovinge, Stefan

2010-06-01

Adult mammalian cardiomyocytes are traditionally viewed as being permanently withdrawn from the cell cycle. Whereas some groups have reported none, others have reported extensive mitosis in adult myocardium under steady-state conditions. Recently, a highly specific assay of 14C dating in humans has suggested a continuous generation of cardiomyocytes in the adult, albeit at a very low rate. Mice represent the most commonly used animal model for these studies, but their short lifespan makes them unsuitable for 14C studies. Herein, we investigate the cellular growth pattern for murine cardiomyocyte growth under steady-state conditions, addressed with new analytical and technical strategies, and we furthermore relate this to gene expression patterns. The observed levels of DNA synthesis in early life were associated with cardiomyocyte proliferation. Mitosis was prolonged into early life, longer than the most conservative previous estimates. DNA synthesis in neonatal life was attributable to bi-nucleation, therefore suggesting that cardiomyocytes withdraw from the cell cycle shortly after birth. No cell cycle activity was observed in adult cardiomyocytes and significant polyploidy was observed in cardiomyocyte nuclei. Gene analyses identified 32 genes whose expression was predicted to be particular to day 3-4 neonatal myocytes, compared with embryonic or adult cells. These cell cycle-associated genes are crucial to the understanding of the mechanisms of bi-nucleation and physiological cellular growth in the neonatal period.

Energetic Nuclei, Superdensity and Biomedicine

ERIC Educational Resources Information Center

Baldin, A. M.

1977-01-01

High-energy, relativistic nuclei were first observed in cosmic rays. Studing these nuclei has provided an opportunity for analyzing the composition of cosmic rays and for experimentally verifying principles governing the behavior of nuclear matter at high and super-high temperatures. Medical research using accelerated nuclei is suggested.…

The oncogenic transforming potential of the passage of single α particles through mammalian cell nuclei

PubMed Central

Miller, Richard C.; Randers-Pehrson, Gerhard; Geard, Charles R.; Hall, Eric J.; Brenner, David J.

1999-01-01

Domestic, low-level exposure to radon gas is considered a major environmental lung-cancer hazard involving DNA damage to bronchial cells by α particles from radon progeny. At domestic exposure levels, the relevant bronchial cells are very rarely traversed by more than one α particle, whereas at higher radon levels—at which epidemiological studies in uranium miners allow lung-cancer risks to be quantified with reasonable precision—these bronchial cells are frequently exposed to multiple α-particle traversals. Measuring the oncogenic transforming effects of exactly one α particle without the confounding effects of multiple traversals has hitherto been unfeasible, resulting in uncertainty in extrapolations of risk from high to domestic radon levels. A technique to assess the effects of single α particles uses a charged-particle microbeam, which irradiates individual cells or cell nuclei with predefined exact numbers of particles. Although previously too slow to assess the relevant small oncogenic risks, recent improvements in throughput now permit microbeam irradiation of large cell numbers, allowing the first oncogenic risk measurements for the traversal of exactly one α particle through a cell nucleus. Given positive controls to ensure that the dosimetry and biological controls were comparable, the measured oncogenicity from exactly one α particle was significantly lower than for a Poisson-distributed mean of one α particle, implying that cells traversed by multiple α particles contribute most of the risk. If this result applies generally, extrapolation from high-level radon risks (involving cellular traversal by multiple α particles) may overestimate low-level (involving only single α particles) radon risks. PMID:9874764

Acridine Orange Conjugated Polymersomes for Simultaneous Nuclear Delivery of Gemcitabine and Doxorubicin to Pancreatic Cancer Cells.

PubMed

Anajafi, Tayebeh; Scott, Michael D; You, Seungyong; Yang, Xiaoyu; Choi, Yongki; Qian, Steven Y; Mallik, Sanku

2016-03-16

Considering the systemic toxicity of chemotherapeutic agents, there is an urgent need to develop new targeted drug delivery systems. Herein, we have developed a new nuclear targeted, redox sensitive, drug delivery vehicle to simultaneously deliver the anticancer drugs gemcitabine and doxorubicin to the nuclei of pancreatic cancer cells. We prepared polymeric bilayer vesicles (polymersomes), and actively encapsulated the drug combination by the pH gradient method. A redox-sensitive polymer (PEG-S-S-PLA) was incorporated to sensitize the formulation to reducing agent concentration. Acridine orange (AO) was conjugated to the surface of the polymersomes imparting nuclear localizing property. The polymersomes' toxicity and efficacy were compared with those of a free drug combination using monolayer and three-dimensional spheroid cultures of pancreatic cancer cells. We observed that the redox sensitive, nuclear-targeted polymersomes released more than 60% of their encapsulated contents in response to 50 mM glutathione. The nanoparticles are nontoxic; however, the drug encapsulated vesicles have significant toxicity. The prepared formulation can increase the drug's therapeutic index by delivering the drugs directly to the cells' nuclei, one of the key organelles in the cells. This study is likely to initiate research in targeted nuclear delivery using other drug formulations in other types of cancers.

Commissioning a Rotating Target Wheel Assembly for Heavy Element Studies

NASA Astrophysics Data System (ADS)

Fields, L. D.; Bennett, M. E.; Mayorov, D. A.; Folden, C. M.

2013-10-01

The heaviest elements are produced artificially by fusing nuclei of light elements within an accelerator to form heavier nuclei. The most direct method to increase the production rate of nuclei is to increase the beam intensity, necessitating the use of a rotating target to minimize damage to the target by deposited heat. Such a target wheel was constructed for heavy element research at Texas A&M University, Cyclotron Institute, consisting of a wheel with three banana-shaped target cutouts. The target is designed to rotate at 1700 rpm, and a fiber optic cable provides a signal to trigger beam pulsing in order to avoid irradiating the spokes between target segments. Following minor mechanical modifications and construction of a dedicated electrical panel, the rotating target assembly was commissioned for a beam experiment. A 15 MeV/u beam of 20Ne was delivered from the K500 cyclotron and detected by a ruggedized silicon detector. The beam pulsing response time was characterized as a function of the rational frequency of the target wheel. Preliminary analysis suggests that the K500 is capable of pulsing at rates of up to 250 Hz, which is sufficient for planned future experiments. Funded by DOE and NSF-REU Program.

The Mechanism of Gene Targeting in Human Somatic Cells

PubMed Central

Kan, Yinan; Ruis, Brian; Lin, Sherry; Hendrickson, Eric A.

2014-01-01

Gene targeting in human somatic cells is of importance because it can be used to either delineate the loss-of-function phenotype of a gene or correct a mutated gene back to wild-type. Both of these outcomes require a form of DNA double-strand break (DSB) repair known as homologous recombination (HR). The mechanism of HR leading to gene targeting, however, is not well understood in human cells. Here, we demonstrate that a two-end, ends-out HR intermediate is valid for human gene targeting. Furthermore, the resolution step of this intermediate occurs via the classic DSB repair model of HR while synthesis-dependent strand annealing and Holliday Junction dissolution are, at best, minor pathways. Moreover, and in contrast to other systems, the positions of Holliday Junction resolution are evenly distributed along the homology arms of the targeting vector. Most unexpectedly, we demonstrate that when a meganuclease is used to introduce a chromosomal DSB to augment gene targeting, the mechanism of gene targeting is inverted to an ends-in process. Finally, we demonstrate that the anti-recombination activity of mismatch repair is a significant impediment to gene targeting. These observations significantly advance our understanding of HR and gene targeting in human cells. PMID:24699519

Synthesis, Decay Properties, and Identification of Superheavy Nuclei Produced in 48Ca-induced Reactions

NASA Astrophysics Data System (ADS)

Oganessian, Yu. Ts.; Utyonkov, V. K.; Lobanov, Yu. V.; Abdullin, F. Sh.; Polyakov, A. N.; Sagaidak, R. N.; Shirokovsky, I. V.; Tsyganov, Yu. S.; Voinov, A. A.; Iliev, S.; Subbotin, V. G.; Sukhov, A. M.; Gulbekian, G. G.; Bogomolov, S. L.; Gikal, B. N.; Mezentsev, A. N.; Subotic, K.; Zagrebaev, V. I.; Itkis, M. G.; Moody, K. J.; Henderson, R. A.; Patin, J. B.; Shaughnessy, D. A.; Stoyer, M. A.; Stoyer, N. J.; Wilk, P. A.; Kenneally, J. M.; Landrum, J. H.; Wild, J. F.; Lougheed, R. W.

2007-10-01

Thirty-four new nuclides with Z = 104-116, 118 and N = 161-177 have been synthesized in the complete-fusion reactions of 238U, 237Np, 242,244Pu, 243Am, 245,248Cm, and 249Cf targets with 48Ca beams. The masses of evaporation residues were identified through measurements of the excitation functions of the xn-evaporation channels and from cross bombardments. The decay properties of the new nuclei agree with those of previously known heavy nuclei and with predictions from different theoretical models. A discussion of self-consistent interpretations of all observed decay chains originating from the parent isotopes 282,283112, 282113, 286-289114, 287,288115, 290-293116, and 294118 is presented. Decay energies and lifetimes of the neutron-rich superheavy nuclei as well as their production cross sections indicate a considerable increase in the stability of nuclei with the approach to the theoretically predicted nuclear shells with N = 184 and Z = 114.

Synthesis, Decay Properties, and Identification of Superheavy Nuclei Produced in 48CA-INDUCED Reactions

NASA Astrophysics Data System (ADS)

Oganessian, Yu. Ts.; Utyonkov, V. K.; Lobanov, Yu. V.; Abdullin, F. Sh.; Polyakov, A. N.; Sagaidak, R. N.; Shirokovsky, I. V.; Tsyganov, Yu. S.; Voinov, A. A.; Iliev, S.; Subbotin, V. G.; Sukhov, A. M.; Gulbekian, G. G.; Bogomolov, S. L.; Gikal, B. N.; Mezentsev, A. N.; Subotic, K.; Zagrebaev, V. I.; Itkis, M. G.; Moody, K. J.; Henderson, R. A.; Patin, J. B.; Shaughnessy, D. A.; Stoyer, M. A.; Stoyer, N. J.; Wilk, P. A.; Kenneally, J. M.; Landrum, J. H.; Wild, J. F.; Lougheed, R. W.

2008-04-01

Thirty-four new nuclides with Z = 104-116, 118 and N = 161-177 have been synthesized in the complete-fusion reactions of 238U, 237Np, 242,244Pu, 243Am, 245,248Cm, and 249Cf targets with 48Ca beams. The masses of evaporation residues were identified through measurements of the excitation functions of the xn-evaporation channels and from cross bombardments. The decay properties of the new nuclei agree with those of previously known heavy nuclei and with predictions from different theoretical models. A discussion of self-consistent interpretations of all observed decay chains originating from the parent isotopes 282,283112, 282113, 286-289114, 287,288115, 290-293116, and 294118 is presented. Decay energies and lifetimes of the neutron-rich superheavy nuclei as well as their production cross sections indicate a considerable increase in the stability of nuclei with the approach to the theoretically predicted nuclear shells with N = 184 and Z = 114.

Theoretical study on production cross sections of exotic actinide nuclei in multinucleon transfer reactions

NASA Astrophysics Data System (ADS)

Zhu, Long

2017-12-01

Within the dinuclear system (DNS) model, the multinucleon transfer reactions 129,136Xe + 248Cm, 112Sn + 238U, and 144Xe + 248Cm are investigated. The production cross sections of primary fragments are calculated with the DNS model. By using a statistical model, we investigate the influence of charged particle evaporation channels on production cross sections of exotic nuclei. It is found that for excited neutron-deficient nuclei the charged particle evaporation competes with neutron emission and plays an important role in the cooling process. The production cross sections of several exotic actinide nuclei are predicted in the reactions 112Sn + 238U and 136,144Xe + 248Cm. Considering the beam intensities, the collisions of 136,144Xe projectiles with a 248Cm target for producing neutron-rich nuclei with Z=92-96 are investigated. Supported by National Natural Science Foundation of China (11605296) and Natural Science Foundation of Guangdong Province, China (2016A030310208)

Pro-Tumoral Inflammatory Myeloid Cells as Emerging Therapeutic Targets.

PubMed

Szebeni, Gabor J; Vizler, Csaba; Nagy, Lajos I; Kitajka, Klara; Puskas, Laszlo G

2016-11-23

Since the observation of Virchow, it has long been known that the tumor microenvironment constitutes the soil for the infiltration of inflammatory cells and for the release of inflammatory mediators. Under certain circumstances, inflammation remains unresolved and promotes cancer development. Here, we review some of these indisputable experimental and clinical evidences of cancer related smouldering inflammation. The most common myeloid infiltrate in solid tumors is composed of myeloid-derived suppressor cells (MDSCs) and tumor-associated macrophages (TAMs). These cells promote tumor growth by several mechanisms, including their inherent immunosuppressive activity, promotion of neoangiogenesis, mediation of epithelial-mesenchymal transition and alteration of cellular metabolism. The pro-tumoral functions of TAMs and MDSCs are further enhanced by their cross-talk offering a myriad of potential anti-cancer therapeutic targets. We highlight these main pro-tumoral mechanisms of myeloid cells and give a general overview of their phenotypical and functional diversity, offering examples of possible therapeutic targets. Pharmacological targeting of inflammatory cells and molecular mediators may result in therapies improving patient condition and prognosis. Here, we review experimental and clinical findings on cancer-related inflammation with a major focus on creating an inventory of current small molecule-based therapeutic interventions targeting cancer-related inflammatory cells: TAMs and MDSCs.

Cortical and subcortical afferents to the nucleus reticularis tegmenti pontis and basal pontine nuclei in the macaque monkey.

PubMed

Giolli, R A; Gregory, K M; Suzuki, D A; Blanks, R H; Lui, F; Betelak, K F

2001-01-01

Anatomical findings are presented that identify cortical and subcortical sources of afferents to the nucleus reticularis tegmenti pontis (NRTP) and basal pontine nuclei. Projections from the middle temporal visual area (MT), medial superior temporal visual area (MST), lateral intraparietal area (LIP), and areas 7a and 7b to the basal pontine nuclei were studied using 3H-leucine autoradiography. The results complemented a parallel study of retrograde neuronal labeling attributable to injecting WGA-HRP into NRTP and neighboring pontine nuclei. Small 3H-leucine injections confined to MT, MST, LIP, area 7a, or area 7b, produced multiple patches of pontine terminal label distributed as follows: (1) An injection within MT produced terminal label limited to the dorsolateral and lateral pontine nuclei. (2) Injections restricted to MST or LIP showed patches of terminal label in the dorsal, dorsolateral, lateral, and peduncular pontine nuclei. (3) Area 7a targets the dorsal, dorsolateral, lateral, peduncular, and ventral pontine nuclei, whereas area 7b projects, additionally, to the dorsomedial and paramedian pontine nuclei. Notably, no projections were seen to NRTP from any of these cortical areas. In contrast, injections made by other investigators into cortical areas anterior to the central sulcus revealed cerebrocortical afferents to NRTP, in addition to nuclei of the basal pontine gray. With our pontine WGA-HRP injections, retrograde neuronal labeling was observed over a large extent of the frontal cortex continuing onto the medial surface which included the lining of the cingulate sulcus and cingulate gyrus. Significant subcortical sources for afferents to the NRTP and basal pontine nuclei were the zona incerta, ventral mesencephalic tegmentum, dorsomedial hypothalamic area, rostral interstitial nucleus of the medial longitudinal fasciculus, red nucleus, and subthalamic nucleus. The combined anterograde and retrograde labeling data indicated that visuo-motor cortico

Prodrug strategy for cancer cell-specific targeting: A recent overview.

PubMed

Zhang, Xian; Li, Xiang; You, Qidong; Zhang, Xiaojin

2017-10-20

The increasing development of targeted cancer therapy provides extensive possibilities in clinical trials, and numerous strategies have been explored. The prodrug is one of the most promising strategies in targeted cancer therapy to improve the selectivity and efficacy of cytotoxic compounds. Compared with normal tissues, cancer cells are characterized by unique aberrant markers, thus inactive prodrugs targeting these markers are excellent therapeutics to release active drugs, killing cancer cells without damaging normal tissues. In this review, we explore an integrated view of potential prodrugs applied in targeted cancer therapy based on aberrant cancer specific markers and some examples are provided for inspiring new ideas of prodrug strategy for cancer cell-specific targeting. Copyright © 2017. Published by Elsevier Masson SAS.

DNA content of hepatocyte and erythrocyte nuclei of the spined loach (Cobitis taenia L.) and its polyploid forms.

PubMed

Juchno, Dorota; Lackowska, Bozena; Boron, Alicja; Kilarski, Wincenty

2010-09-01

We analyzed the DNA content of hepatocyte and erythrocyte nuclei of the spined loach Cobitis taenia (diploid) and its allopolyploid forms. Twenty triploid females and one tetraploid were used. At least 20,000 hepatocyte and erythrocyte nuclei were acquired and analyzed by flow cytometry. C. taenia erythrocyte nuclei contain 3.15 +/- 0.21 pg of DNA and the hepatocyte nuclei 4.45 +/- 0.46 pg of DNA. Triploid Cobitis have 5.08 +/- 0.41 pg of DNA in erythrocyte nuclei and 6.11 +/- 0.40 pg of DNA in hepatocyte nuclei, whereas the tetraploid erythrocyte and hepatocyte nuclei contained 6.60 and 7.40 pg of DNA, respectively. In general, the DNA contents correlate positively with the ploidy level of the fish investigated. The DNA content variation in the hepatocyte and erythrocyte nuclei may be due to differences in extent of chromatin condensation, which is more pronounced in the erythrocyte than hepatocyte nuclei, or to the several orders of ploidy that occur in the parenchymal liver cells.

Active galactic nuclei

PubMed Central

Fabian, Andrew C.

1999-01-01

Active galactic nuclei are the most powerful, long-lived objects in the Universe. Recent data confirm the theoretical idea that the power source is accretion into a massive black hole. The common occurrence of obscuration and outflows probably means that the contribution of active galactic nuclei to the power density of the Universe has been generally underestimated. PMID:10220363

Nuclei and the Unitary Limit

NASA Astrophysics Data System (ADS)

Hammer, H.-W.

2018-07-01

Few-body systems with large scattering length display universal properties which are independent of the details of short-distance dynamics. These features include universal correlations between few-body observables and a geometric spectrum of three- and higher-body bound states. They can be observed in a wide range of systems from ultracold atoms to hadrons and nuclei. In this contribution, we review universality in nuclei dominated by few-body physics. In particular, we discuss halo nuclei and the description of light nuclei in a strict expansion around the unitary limit of infinite scattering length.

Deuteron-induced nucleon transfer reactions within an ab initio framework: First application to p -shell nuclei

DOE PAGES

Raimondi, Francesco; Hupin, Guillaume; Navratil, Petr; ...

2016-05-10

Low-energy transfer reactions in which a proton is stripped from a deuteron projectile and dropped into a target play a crucial role in the formation of nuclei in both primordial and stellar nucleosynthesis, as well as in the study of exotic nuclei using radioactive beam facilities and inverse kinematics. Here, ab initio approaches have been successfully applied to describe the 3H(d,n) 4He and 3He(d,p) 4He fusion processes. An ab initio treatment of transfer reactions would also be desirable for heavier targets. In this work, we extend the ab initio description of (d,p) reactions to processes with light p-shell nuclei. Asmore » a first application, we study the elastic scattering of deuterium on 7Li and the 7Li(d,p) 8Li transfer reaction based on a two-body Hamiltonian. We use the no-core shell model to compute the wave functions of the nuclei involved in the reaction, and describe the dynamics between targets and projectiles with the help of microscopic-cluster states in the spirit of the resonating group method. The shapes of the excitation functions for deuterons impinging on 7Li are qualitatively reproduced up to the deuteron breakup energy. The interplay between d– 7Li and p– 8Li particle-decay channels determines some features of the 9Be spectrum above the d+ 7Li threshold. Our prediction for the parity of the 17.298 MeV resonance is at odds with the experimental assignment. Deuteron stripping reactions with p-shell targets can now be computed ab initio, but calculations are very demanding. Finally, a quantitative description of the 7Li(d,p) 8Li reaction will require further work to include the effect of three-nucleon forces and additional decay channels and to improve the convergence rate of our calculations.« less

Deuteron-induced nucleon transfer reactions within an ab initio framework: First application to p -shell nuclei

DOE Office of Scientific and Technical Information (OSTI.GOV)

Raimondi, Francesco; Hupin, Guillaume; Navratil, Petr

Low-energy transfer reactions in which a proton is stripped from a deuteron projectile and dropped into a target play a crucial role in the formation of nuclei in both primordial and stellar nucleosynthesis, as well as in the study of exotic nuclei using radioactive beam facilities and inverse kinematics. Here, ab initio approaches have been successfully applied to describe the 3H(d,n) 4He and 3He(d,p) 4He fusion processes. An ab initio treatment of transfer reactions would also be desirable for heavier targets. In this work, we extend the ab initio description of (d,p) reactions to processes with light p-shell nuclei. Asmore » a first application, we study the elastic scattering of deuterium on 7Li and the 7Li(d,p) 8Li transfer reaction based on a two-body Hamiltonian. We use the no-core shell model to compute the wave functions of the nuclei involved in the reaction, and describe the dynamics between targets and projectiles with the help of microscopic-cluster states in the spirit of the resonating group method. The shapes of the excitation functions for deuterons impinging on 7Li are qualitatively reproduced up to the deuteron breakup energy. The interplay between d– 7Li and p– 8Li particle-decay channels determines some features of the 9Be spectrum above the d+ 7Li threshold. Our prediction for the parity of the 17.298 MeV resonance is at odds with the experimental assignment. Deuteron stripping reactions with p-shell targets can now be computed ab initio, but calculations are very demanding. Finally, a quantitative description of the 7Li(d,p) 8Li reaction will require further work to include the effect of three-nucleon forces and additional decay channels and to improve the convergence rate of our calculations.« less

In Situ Target Engagement Studies in Adherent Cells.

PubMed

Axelsson, Hanna; Almqvist, Helena; Otrocka, Magdalena; Vallin, Michaela; Lundqvist, Sara; Hansson, Pia; Karlsson, Ulla; Lundbäck, Thomas; Seashore-Ludlow, Brinton

2018-04-20

A prerequisite for successful drugs is effective binding of the desired target protein in the complex environment of a living system. Drug-target engagement has typically been difficult to monitor in physiologically relevant models, and with current methods, especially, while maintaining spatial information. One recent technique for quantifying drug-target engagement is the cellular thermal shift assay (CETSA), in which ligand-induced protein stabilization is measured after a heat challenge. Here, we describe a CETSA protocol in live A431 cells for p38α (MAPK14), where remaining soluble protein is detected in situ, using high-content imaging in 384-well, microtiter plates. We validate this assay concept using a number of known p38α inhibitors and further demonstrate the potential of this technology for chemical probe and drug discovery purposes by performing a small pilot screen for novel p38α binders. Importantly, this protocol creates a workflow that is amenable to adherent cells in their native state and yields spatially resolved target engagement information measurable at the single-cell level.

Fragments of Target Cells are Internalized into Retroviral Envelope Protein-Expressing Cells during Cell-Cell Fusion by Endocytosis

PubMed Central

Izumida, Mai; Kamiyama, Haruka; Suematsu, Takashi; Honda, Eri; Koizumi, Yosuke; Yasui, Kiyoshi; Hayashi, Hideki; Ariyoshi, Koya; Kubo, Yoshinao

2016-01-01

Retroviruses enter into host cells by fusion between viral and host cell membranes. Retroviral envelope glycoprotein (Env) induces the membrane fusion, and also mediates cell-cell fusion. There are two types of cell-cell fusions induced by the Env protein. Fusion-from-within is induced by fusion between viral fusogenic Env protein-expressing cells and susceptible cells, and virions induce fusion-from-without by fusion between adjacent cells. Although entry of ecotropic murine leukemia virus (E-MLV) requires host cell endocytosis, the involvement of endocytosis in cell fusion is unclear. By fluorescent microscopic analysis of the fusion-from-within, we found that fragments of target cells are internalized into Env-expressing cells. Treatment of the Env-expressing cells with an endocytosis inhibitor more significantly inhibited the cell fusion than that of the target cells, indicating that endocytosis in Env-expressing cells is required for the cell fusion. The endocytosis inhibitor also attenuated the fusion-from-without. Electron microscopic analysis suggested that the membrane fusion resulting in fusion-from-within initiates in endocytic membrane dents. This study shows that two types of the viral cell fusion both require endocytosis, and provides the cascade of fusion-from-within. PMID:26834711

Target fragmentation in radiobiology

NASA Technical Reports Server (NTRS)

Wilson, John W.; Cucinotta, Francis A.; Shinn, Judy L.; Townsend, Lawrence W.

1993-01-01

Nuclear reactions in biological systems produce low-energy fragments of the target nuclei seen as local high events of linear energy transfer (LET). A nuclear-reaction formalism is used to evaluate the nuclear-induced fields within biosystems and their effects within several biological models. On the basis of direct ionization interaction, one anticipates high-energy protons to have a quality factor and relative biological effectiveness (RBE) of unity. Target fragmentation contributions raise the effective quality factor of 10 GeV protons to 3.3 in reasonable agreement with RBE values for induced micronuclei in bean sprouts. Application of the Katz model indicates that the relative increase in RBE with decreasing exposure observed in cell survival experiments with 160 MeV protons is related solely to target fragmentation events. Target fragment contributions to lens opacity given an RBE of 1.4 for 2 GeV protons in agreement with the work of Lett and Cox. Predictions are made for the effective RBE for Harderian gland tumors induced by high-energy protons. An exposure model for lifetime cancer risk is derived from NCRP 98 risk tables, and protraction effects are examined for proton and helium ion exposures. The implications of dose rate enhancement effects on space radiation protection are considered.

The therapeutic potential of cell cycle targeting in multiple myeloma.

PubMed

Maes, Anke; Menu, Eline; Veirman, Kim De; Maes, Ken; Vand Erkerken, Karin; De Bruyne, Elke

2017-10-27

Proper cell cycle progression through the interphase and mitosis is regulated by coordinated activation of important cell cycle proteins (including cyclin-dependent kinases and mitotic kinases) and several checkpoint pathways. Aberrant activity of these cell cycle proteins and checkpoint pathways results in deregulation of cell cycle progression, which is one of the key hallmarks of cancer. Consequently, intensive research on targeting these cell cycle regulatory proteins identified several candidate small molecule inhibitors that are able to induce cell cycle arrest and even apoptosis in cancer cells. Importantly, several of these cell cycle regulatory proteins have also been proposed as therapeutic targets in the plasma cell malignancy multiple myeloma (MM). Despite the enormous progress in the treatment of MM the past 5 years, MM still remains most often incurable due to the development of drug resistance. Deregulated expression of the cyclins D is observed in virtually all myeloma patients, emphasizing the potential therapeutic interest of cyclin-dependent kinase inhibitors in MM. Furthermore, other targets have also been identified in MM, such as microtubules, kinesin motor proteins, aurora kinases, polo-like kinases and the anaphase promoting complex/cyclosome. This review will provide an overview of the cell cycle proteins and checkpoint pathways deregulated in MM and discuss the therapeutic potential of targeting proteins or protein complexes involved in cell cycle control in MM.

Optokinetic and Vestibular Responsiveness in the Macaque Rostral Vestibular and Fastigial Nuclei

PubMed Central

Bryan, Ayanna S.; Angelaki, Dora E.

2009-01-01

We recorded from rostral vestibular (VN) and rostral fastigial nuclei (FN) neurons that did not respond to eye movements during three-dimensional (3D) vestibular and optokinetic stimulation (OKS). The majority of neurons in both areas (76 and 69% in VN and FN, respectively) responded during both rotational and translational motion. Preferred directions scattered throughout 3D space for translation but showed some preference for pitch/roll over yaw for rotation. VN/FN neurons were also tested during OKS while monkeys suppressed their optokinetic nystagmus by fixating a head-fixed target. Only a handful of cells (VN: 17%, FN: 6%) modulated during 0.5-Hz OKS suppression, but the number of responsive cells increased (VN: 40%, FN: 48%) during 0.02-Hz OKS. Preferred directions for rotation and OKS were not matched on individual neurons, and OKS gains were smaller than the respective gains during rotation. These results were generally similar for VN and FN neurons. We conclude that optokinetic-vestibular convergence might not be as prevalent as earlier studies have suggested. PMID:19073813

Cancer stem cell-targeted therapeutics and delivery strategies.

PubMed

Ahmad, Gulzar; Amiji, Mansoor M

2017-08-01

Cancer initiating or stem cells (CSCs) are a small population of cells in the tumor mass, which have been reported to be present in different types of cancers. CSCs usually reside within the tumor and are responsible for reoccurrence of cancer. The imprecise, inaccessible nature and increased efflux of conventional therapeutic drugs make these cells resistant to drugs. We discuss the specific markers for identification of these cells, role of CSCs in chemotherapy resistance and use of different therapeutic means to target them, including elucidation of specific cell markers, exploitation of different signaling pathways and use of nanotechnology. Area covered: This review covers cancer stem cell signaling which are used by these cells to maintain their quiescence, stemness and resistant phenotype, distinct cell surface markers, contribution of these cells in drug resistance, inevitability to cure cancer and use of nanotechnology to overcome this hurdle. Expert opinion: Cancer stem cells are the main culprit of our failure to cure cancer. In order to cure cancer along with other cells types in cancer, cancer stem cells need to be targeted in the tumor bed. Nanotechnology solutions can facilitate clinical translation of the therapeutics along with other emerging technologies to cure cancer.

β-decay spectroscopy of r-process nuclei with N = 126 at KISS

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hirayama, Y.; Watanabe, Y. X.; Imai, N.

2014-05-02

The β-decay properties of nuclei with N = 126, which are believed to act as progenitors in the rapid neutron capture (r-) process path forming the third peak (A ∼ 195) in the observed r-abundance element distribution, are considered critical for understanding the production of heavy elements such as gold and platinum at astrophysical sites. We have constructed the KEK Isotope Separation System (KISS), which consists of a gas cell based laser ion source (atomic number selection) and an isotope separation on-line (ISOL) (mass number selection), to produce pure low-energy beams of neutron-rich isotopes around N = 126 and tomore » study their β-decay properties, which are also of interest for astrophysics. The isotopes of interest will be produced by multi-nucleon transfer reactions in heavy ion collisions (e.g. {sup 136}Xe projectile on {sup 198}Pt target). KISS will allow us to study unknown isotopes produced in weak reaction channels under low background conditions. We successfully extracted the stable {sup 56}Fe beam from KISS at the last commissioning on-line experiment with the extraction efficiency of 0.25% and beam purity of more than 98%. We can access the nuclei with N = 126 and measure their half-lives using the KISS in the case of the extraction efficiency of 0.1%.« less

Pros and Cons of Antigen-Presenting Cell Targeted Tumor Vaccines.

PubMed

Goyvaerts, Cleo; Breckpot, Karine

2015-01-01

In therapeutic antitumor vaccination, dendritic cells play the leading role since they decide if, how, when, and where a potent antitumor immune response will take place. Since the disentanglement of the complexity and merit of different antigen-presenting cell subtypes, antitumor immunotherapeutic research started to investigate the potential benefit of targeting these subtypes in situ. This review will discuss which antigen-presenting cell subtypes are at play and how they have been targeted and finally question the true meaning of targeting antitumor-based vaccines.

Properties of true quaternary fission of nuclei with allowance for its multistep and sequential character

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kadmensky, S. G., E-mail: kadmensky@phys.vsu.ru; Titova, L. V.; Bulychev, A. O.

An analysis of basicmechanisms of binary and ternary fission of nuclei led to the conclusion that true ternary and quaternary fission of nuclei has a sequential two-step (three-step) character, where, at the first step, a fissile nucleus emits a third light particle (third and fourth light particles) under shakeup effects associated with a nonadiabatic character of its collective deformation motion, whereupon the residual nucleus undergoes fission to two fission fragments. Owing to this, the formulas derived earlier for the widths with respect to sequential two- and three-step decays of nuclei in constructing the theory of two-step twoproton decays and multistepmore » decays in chains of genetically related nuclei could be used to describe the relative yields and angular and energy distributions of third and fourth light particles emitted in (α, α), (t, t), and (α, t) pairs upon the true quaternary spontaneous fission of {sup 252}Cf and thermal-neutron-induced fission of {sup 235}U and {sup 233}U target nuclei. Mechanisms that explain a sharp decrease in the yield of particles appearing second in time and entering into the composition of light-particle pairs that originate from true quaternary fission of nuclei in relation to the yields of analogous particles in true ternary fission of nuclei are proposed.« less

Purification-Free, Target-Selective Immobilization of a Protein from Cell Lysates.

PubMed

Cha, Jaehyun; Kwon, Inchan

2018-02-27

Protein immobilization has been widely used for laboratory experiments and industrial processes. Preparation of a recombinant protein for immobilization usually requires laborious and expensive purification steps. Here, a novel purification-free, target-selective immobilization technique of a protein from cell lysates is reported. Purification steps are skipped by immobilizing a target protein containing a clickable non-natural amino acid (p-azidophenylalanine) in cell lysates onto alkyne-functionalized solid supports via bioorthogonal azide-alkyne cycloaddition. In order to achieve a target protein-selective immobilization, p-azidophenylalanine was introduced into an exogenous target protein, but not into endogenous non-target proteins using host cells with amber codon-free genomic DNAs. Immobilization of superfolder fluorescent protein (sfGFP) from cell lysates is as efficient as that of the purified sfGFP. Using two fluorescent proteins (sfGFP and mCherry), the authors also demonstrated that the target proteins are immobilized with a minimal immobilization of non-target proteins (target-selective immobilization). © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Methods to isolate a large amount of generative cells, sperm cells and vegetative nuclei from tomato pollen for "omics" analysis.

PubMed

Lu, Yunlong; Wei, Liqin; Wang, Tai

2015-01-01

The development of sperm cells (SCs) from microspores involves a set of finely regulated molecular and cellular events and the coordination of these events. The mechanisms underlying these events and their interconnections remain a major challenge. Systems analysis of genome-wide molecular networks and functional modules with high-throughput "omics" approaches is crucial for understanding the mechanisms; however, this study is hindered because of the difficulty in isolating a large amount of cells of different types, especially generative cells (GCs), from the pollen. Here, we optimized the conditions of tomato pollen germination and pollen tube growth to allow for long-term growth of pollen tubes in vitro with SCs generated in the tube. Using this culture system, we developed methods for isolating GCs, SCs and vegetative cell nuclei (VN) from just-germinated tomato pollen grains and growing pollen tubes and their purification by Percoll density gradient centrifugation. The purity and viability of isolated GCs and SCs were confirmed by microscopy examination and fluorescein diacetate staining, respectively, and the integrity of VN was confirmed by propidium iodide staining. We could obtain about 1.5 million GCs and 2.0 million SCs each from 180 mg initiated pollen grains, and 10 million VN from 270 mg initiated pollen grains germinated in vitro in each experiment. These methods provide the necessary preconditions for systematic biology studies of SC development and differentiation in higher plants.

Ion mediated targeting of cells with nanoparticles

NASA Astrophysics Data System (ADS)

Maheshwari, Vivek; Fu, Jinlong

2010-03-01

In eukaryotic cells, Ca^2+ ions are necessary for intracellular signaling, in activity of mitochondria and a variety of other cellular process that have been linked to cell apoptosis, proteins synthesis and cell-cycle regulation. Here we show that Ca^2+ ions, serving as the bio-compatible interface can be used to target Saccharomyces cerevisiae (SaC, baker's yeast), a model eukaryotic cell, with Au nanoparticles (10 nm). The Ca^2+ ions bind to the carboxylic acid groups in the citrate functionalized Au nanoparticles. This transforms the nanoparticles into micron long 1-D branched chain assemblies due to inter-particle dipole-dipole interaction and inter-particle bonding due to the divalent nature of the Ca^2+ ion. A similar transformation is observed with the use of divalent ions Mg^2+, Cd^2+ and Fe^2+. The 1-D assembly aids the interfacing of ion-nanoparticles on the cell by providing multiple contact points. Further monovalent ions such as Na^+ are also effective for the targeting of the cell with nanoparticles. However Na-Au nanoparticles are limited in their deposition as they exist in solution as single particles. The cells remain alive after the deposition process and their vitality is unaffected by the interfacing with ion-nanoparticles.

T-REX on-demand redox targeting in live cells.

PubMed

Parvez, Saba; Long, Marcus J C; Lin, Hong-Yu; Zhao, Yi; Haegele, Joseph A; Pham, Vanha N; Lee, Dustin K; Aye, Yimon

2016-12-01

This protocol describes targetable reactive electrophiles and oxidants (T-REX)-a live-cell-based tool designed to (i) interrogate the consequences of specific and time-resolved redox events, and (ii) screen for bona fide redox-sensor targets. A small-molecule toolset comprising photocaged precursors to specific reactive redox signals is constructed such that these inert precursors specifically and irreversibly tag any HaloTag-fused protein of interest (POI) in mammalian and Escherichia coli cells. Syntheses of the alkyne-functionalized endogenous reactive signal 4-hydroxynonenal (HNE(alkyne)) and the HaloTag-targetable photocaged precursor to HNE(alkyne) (also known as Ht-PreHNE or HtPHA) are described. Low-energy light prompts photo-uncaging (t 1/2 <1-2 min) and target-specific modification. The targeted modification of the POI enables precisely timed and spatially controlled redox events with no off-target modification. Two independent pathways are described, along with a simple setup to functionally validate known targets or discover novel sensors. T-REX sidesteps mixed responses caused by uncontrolled whole-cell swamping with reactive signals. Modification and downstream response can be analyzed by in-gel fluorescence, proteomics, qRT-PCR, immunofluorescence, fluorescence resonance energy transfer (FRET)-based and dual-luciferase reporters, or flow cytometry assays. T-REX targeting takes 4 h from initial probe treatment. Analysis of targeted redox responses takes an additional 4-24 h, depending on the nature of the pathway and the type of readouts used.

T-REX on-demand redox targeting in live cells

PubMed Central

Parvez, Saba; Long, Marcus J C; Lin, Hong-Yu; Zhao, Yi; Haegele, Joseph A; Pham, Vanha N; Lee, Dustin K; Aye, Yimon

2017-01-01

This protocol describes targetable reactive electrophiles and oxidants (T-REX)—a live-cell-based tool designed to (i) interrogate the consequences of specific and time-resolved redox events, and (ii) screen for bona fide redox-sensor targets. A small-molecule toolset comprising photocaged precursors to specific reactive redox signals is constructed such that these inert precursors specifically and irreversibly tag any HaloTag-fused protein of interest (POI) in mammalian and Escherichia coli cells. Syntheses of the alkyne-functionalized endogenous reactive signal 4-hydroxynonenal (HNE (alkyne)) and the HaloTag-targetable photocaged precursor to HNE (alkyne) (also known as Ht-PreHNE or HtPHA) are described. Low-energy light prompts photo-uncaging (t1/2 <1–2 min) and target-specific modification. The targeted modification of the POI enables precisely timed and spatially controlled redox events with no off-target modification. Two independent pathways are described, along with a simple setup to functionally validate known targets or discover novel sensors. T-REX sidesteps mixed responses caused by uncontrolled whole-cell swamping with reactive signals. Modification and downstream response can be analyzed by in-gel fluorescence, proteomics, qRT-PCR, immunofluorescence, fluorescence resonance energy transfer (FRET)-based and dual-luciferase reporters, or flow cytometry assays. T-REX targeting takes 4 h from initial probe treatment. Analysis of targeted redox responses takes an additional 4–24 h, depending on the nature of the pathway and the type of readouts used. PMID:27809314

Medium effects in λK+ pair production by 2.83 GeV protons on nuclei

NASA Astrophysics Data System (ADS)

Paryev, E. Ya.; Hartmann, M.; Kiselev, Yu. T.

2017-12-01

We study ΛK+ pair production in the interaction of protons of 2.83 GeV kinetic energy with C, Cu, Ag, and Au target nuclei in the framework of the nuclear spectral function approach for incoherent primary proton-nucleon and secondary pion-nucleon production processes, and processes associated with the creation of intermediate Σ0K+ pairs. The approach accounts for the initial proton and final Λ hyperon absorption, final K+ meson distortion in nuclei, target nucleon binding, and Fermi motion, as well as nuclear mean-field potential effects on these processes. We calculate the Λ momentum dependence of the absolute ΛK+ yield from the target nuclei considered, in the kinematical conditions of the ANKE experiment, performed at COSY, within the different scenarios for the Λ-nucleus effective scalar potential. We show that the above observable is appreciably sensitive to this potential in the low-momentum region. Therefore, direct comparison of the results of our calculations with the data from the ANKE-at-COSY experiment can help to determine the above potential at finite momenta. We also demonstrate that the two-step pion-nucleon production channels dominate in the low-momentum ΛK+ production in the chosen kinematics and, therefore, they have to be taken into account in the analysis of these data. Supported by the Ministry of Education and Science of the Russian Federation

NeuN+ Neuronal Nuclei in Non-Human Primate Prefrontal Cortex and Subcortical White Matter After Clozapine Exposure

PubMed Central

Halene, Tobias B.; Kozlenkov, Alexey; Jiang, Yan; Mitchell, Amanda; Javidfar, Behnam; Dincer, Aslihan; Park, Royce; Wiseman, Jennifer; Croxson, Paula; Giannaris, Eustathia Lela; Hof, Patrick R.; Roussos, Panos; Dracheva, Stella; Hemby, Scott E.; Akbarian, Schahram

2016-01-01

Increased neuronal densities in subcortical white matter have been reported for some cases with schizophrenia. The underlying cellular and molecular mechanisms remain unresolved. We exposed 26 young adult macaque monkeys for 6 months to either clozapine, haloperidol or placebo and measured by structural MRI frontal gray and white matter volumes before and after treatment, followed by observer-independent, flow-cytometry-based quantification of neuronal and non-neuronal nuclei and molecular fingerprinting of cell-type specific transcripts. After clozapine exposure, the proportion of nuclei expressing the neuronal marker NeuN increased by approximately 50% in subcortical white matter, in conjunction with a more subtle and non-significant increase in overlying gray matter. Numbers and proportions of nuclei expressing the oligodendrocyte lineage marker, OLIG2, and cell-type specific RNA expression patterns, were maintained after antipsychotic drug exposure. Frontal lobe gray and white matter volumes remained indistinguishable between antipsychotic-drug-exposed and control groups. Chronic clozapine exposure increases the proportion of NeuN+ nuclei in frontal subcortical white matter, without alterations in frontal lobe volumes or cell type-specific gene expression. Further exploration of neurochemical plasticity in non-human primate brain exposed to antipsychotic drugs is warranted. PMID:26776227

Landscape phages and their fusion proteins targeted to breast cancer cells

PubMed Central

Fagbohun, Olusegun A.; Bedi, Deepa; Grabchenko, Natalia I.; Deinnocentes, Patricia A.; Bird, Richard C.; Petrenko, Valery A.

2012-01-01

Breast cancer is a leading cause of death among women in the USA. The efficacy of existing anticancer therapeutics can be improved by targeting them through conjugation with ligands binding to cellular receptors. Recently, we developed a novel drug targeting strategy based on the use of pre-selected cancer-specific ‘fusion pVIII proteins’ (fpVIII), as targeting ligands. To study the efficiency of this approach in animal models, we developed a panel of breast cancer cell-binding phages as a source of targeted fpVIIIs. Two landscape phage peptide libraries (8-mer f8/8 and 9-mer f8/9) were screened to isolate 132 phage variants that recognize breast carcinoma cells MCF-7 and ZR-75-1 and internalize into the cells. When tested for their interaction with the breast cancer cells in comparison with liver cancer cells HepG2, human mammary cells MCF-10A cells and serum, 16 of the phage probes selectively interacted with the breast cancer cells whereas 32 bound both breast and liver cancer cells. The most prominent cancer-specific phage DMPGTVLP, demonstrating sub-nanomolar Kd in interaction with target cells, was used for affinity chromatography of cellular membrane molecules to reveal its potential binding receptor. The isolated protein was identified by direct sequencing as cellular surface nucleolin. This conclusion was confirmed by inhibition of the phage–cell interaction with nucleolin antibodies. Other prominent phage binders VPTDTDYS, VEEGGYIAA, and DWRGDSMDS demonstrate consensus motifs common to previously identified cancer-specific peptides. Isolated phage proteins exhibit inherent binding specificity towards cancer cells, demonstrating the functional activity of the selected fused peptides. The selected phages, their peptide inserts and intact fusion proteins can serve as promising ligands for the development of targeted nanomedicines and their study in model mice with xenograft of human cells MCF-7 and ZR-75-1. PMID:22490956

Modular cell-internalizing aptamer nanostructure enables targeted delivery of large functional RNAs in cancer cell lines.

PubMed

Porciani, David; Cardwell, Leah N; Tawiah, Kwaku D; Alam, Khalid K; Lange, Margaret J; Daniels, Mark A; Burke, Donald H

2018-06-11

Large RNAs and ribonucleoprotein complexes have powerful therapeutic potential, but effective cell-targeted delivery tools are limited. Aptamers that internalize into target cells can deliver siRNAs (<15 kDa, 19-21 nt/strand). We demonstrate a modular nanostructure for cellular delivery of large, functional RNA payloads (50-80 kDa, 175-250 nt) by aptamers that recognize multiple human B cell cancer lines and transferrin receptor-expressing cells. Fluorogenic RNA reporter payloads enable accelerated testing of platform designs and rapid evaluation of assembly and internalization. Modularity is demonstrated by swapping in different targeting and payload aptamers. Both modules internalize into leukemic B cell lines and remained colocalized within endosomes. Fluorescence from internalized RNA persists for ≥2 h, suggesting a sizable window for aptamer payloads to exert influence upon targeted cells. This demonstration of aptamer-mediated, cell-internalizing delivery of large RNAs with retention of functional structure raises the possibility of manipulating endosomes and cells by delivering large aptamers and regulatory RNAs.

The cancer cell adhesion resistome: mechanisms, targeting and translational approaches.

PubMed

Dickreuter, Ellen; Cordes, Nils

2017-06-27

Cell adhesion-mediated resistance limits the success of cancer therapies and is a great obstacle to overcome in the clinic. Since the 1990s, where it became clear that adhesion of tumor cells to the extracellular matrix is an important mediator of therapy resistance, a lot of work has been conducted to understand the fundamental underlying mechanisms and two paradigms were deduced: cell adhesion-mediated radioresistance (CAM-RR) and cell adhesion-mediated drug resistance (CAM-DR). Preclinical work has evidently demonstrated that targeting of integrins, adapter proteins and associated kinases comprising the cell adhesion resistome is a promising strategy to sensitize cancer cells to both radiotherapy and chemotherapy. Moreover, the cell adhesion resistome fundamentally contributes to adaptation mechanisms induced by radiochemotherapy as well as molecular drugs to secure a balanced homeostasis of cancer cells for survival and growth. Intriguingly, this phenomenon provides a basis for synthetic lethal targeted therapies simultaneously administered to standard radiochemotherapy. In this review, we summarize current knowledge about the cell adhesion resistome and highlight targeting strategies to override CAM-RR and CAM-DR.

Application of stem cells in targeted therapy of breast cancer: a systematic review.

PubMed

Madjd, Zahra; Gheytanchi, Elmira; Erfani, Elham; Asadi-Lari, Mohsen

2013-01-01

The aim of this systematic review was to investigate whether stem cells could be effectively applied in targeted therapy of breast cancer. A systematic literature search was performed for original articles published from January 2007 until May 2012. Nine studies met the inclusion criteria for phase I or II clinical trials, of which three used stem cells as vehicles, two trials used autologous hematopoetic stem cells and in four trials cancer stem cells were targeted. Mesenchymal stem cells (MSCs) were applied as cellular vehicles to transfer therapeutic agents. Cell therapy with MSC can successfully target resistant cancers. Cancer stem cells were selectively targeted via a proteasome-dependent suicide gene leading to tumor regression. Wnt/β-catenin signaling pathway has been also evidenced to be an attractive CSC-target. This systematic review focused on two different concepts of stem cells and breast cancer marking a turning point in the trials that applied stem cells as cellular vehicles for targeted delivery therapy as well as CSC-targeted therapies. Applying stem cells as targeted therapy could be an effective therapeutic approach for treatment of breast cancer in the clinic and in therapeutic marketing; however this needs to be confirmed with further clinical investigations.

Replication of pea enation mosaic virus RNA in isolated pea nuclei

PubMed Central

Powell, C. A.; Zoeten, G. A. de

1977-01-01

Isolated nuclei from healthy pea plants were primed with pea enation mosaic virus (PEMV), southern bean mosaic virus (SBMV), radish mosaic virus (RdMV), tobacco mosaic virus (TMV), PEMV RNA, SBMV RNA, RdMV RNA, or TMV RNA. RNA replication occurred only with PEMV RNA and not with intact PEMV or any of the other viruses or RNAs, as judged by ensuing actinomycin D-insensitive polymerase activity. Molecular hybridization experiments showed that some of the product of the polymerase was PEMV-specific (-)RNA. The substrate and ionic requirements of this polymerase were the same as those for the RNA-dependent RNA polymerase present in nuclei isolated from PEMV-infected pea plants. No virus particles could be recovered from nuclei primed with PEMV RNA. These results are discussed in relation to the possible mechanism for in vivo infection of pea cells. PMID:16592421

Monte Carlo calculations of the cellular S-values for α-particle-emitting radionuclides incorporated into the nuclei of cancer cells of the MDA-MB231, MCF7 and PC3 lines.

PubMed

Rojas-Calderón, E L; Ávila, O; Ferro-Flores, G

2018-05-01

S-values (dose per unit of cumulated activity) for alpha particle-emitting radionuclides and monoenergetic alpha sources placed in the nuclei of three cancer cell models (MCF7, MDA-MB231 breast cancer cells and PC3 prostate cancer cells) were obtained by Monte Carlo simulation. The MCNPX code was used to calculate the fraction of energy deposited in the subcellular compartments due to the alpha sources in order to obtain the S-values. A comparison with internationally accepted S-values reported by the MIRD Cellular Committee for alpha sources in three sizes of spherical cells was also performed leading to an agreement within 4% when an alpha extended source uniformly distributed in the nucleus is simulated. This result allowed to apply the Monte Carlo Methodology to evaluate S-values for alpha particles in cancer cells. The calculation of S-values for nucleus, cytoplasm and membrane of cancer cells considering their particular geometry, distribution of the radionuclide source and chemical composition by means of Monte Carlo simulation provides a good approach for dosimetry assessment of alpha emitters inside cancer cells. Results from this work provide information and tools that may help researchers in the selection of appropriate radiopharmaceuticals in alpha-targeted cancer therapy and improve its dosimetry evaluation. Copyright © 2018 Elsevier Ltd. All rights reserved.

Localization Microscopy Analyses of MRE11 Clusters in 3D-Conserved Cell Nuclei of Different Cell Lines.

PubMed

Eryilmaz, Marion; Schmitt, Eberhard; Krufczik, Matthias; Theda, Franziska; Lee, Jin-Ho; Cremer, Christoph; Bestvater, Felix; Schaufler, Wladimir; Hausmann, Michael; Hildenbrand, Georg

2018-01-22

In radiation biophysics, it is a subject of nowadays research to investigate DNA strand break repair in detail after damage induction by ionizing radiation. It is a subject of debate as to what makes up the cell's decision to use a certain repair pathway and how the repair machinery recruited in repair foci is spatially and temporarily organized. Single-molecule localization microscopy (SMLM) allows super-resolution analysis by precise localization of single fluorescent molecule tags, resulting in nuclear structure analysis with a spatial resolution in the 10 nm regime. Here, we used SMLM to study MRE11 foci. MRE11 is one of three proteins involved in the MRN-complex (MRE11-RAD50-NBS1 complex), a prominent DNA strand resection and broken end bridging component involved in homologous recombination repair (HRR) and alternative non-homologous end joining (a-NHEJ). We analyzed the spatial arrangements of antibody-labelled MRE11 proteins in the nuclei of a breast cancer and a skin fibroblast cell line along a time-course of repair (up to 48 h) after irradiation with a dose of 2 Gy. Different kinetics for cluster formation and relaxation were determined. Changes in the internal nano-scaled structure of the clusters were quantified and compared between the two cell types. The results indicate a cell type-dependent DNA damage response concerning MRE11 recruitment and cluster formation. The MRE11 data were compared to H2AX phosphorylation detected by γH2AX molecule distribution. These data suggested modulations of MRE11 signal frequencies that were not directly correlated to DNA damage induction. The application of SMLM in radiation biophysics offers new possibilities to investigate spatial foci organization after DNA damaging and during subsequent repair.

ELECTRON MICROSCOPY OF HELA CELLS INFECTED WITH ADENOVIRUSES

PubMed Central

Harford, Carl G.; Hamlin, Alice; Parker, Esther; van Ravenswaay, Theodore

1956-01-01

HeLa cells were infected with adenoviruses (types 1–4) and sectioned for electron microscopy after intervals of 20 to 48 hours. Clusters of virus-like particles were found within the nuclei of infected cultures but not in those of uninfected controls. The particles were often arranged in rows as if in crystalline formation. Maximal diameter of particles was approximately 65 mµ, and internal bodies were demonstrated. Lesions of infected cells included target-like structures of the nuclear membrane, large nuclear vacuoles (type 2), and increased numbers of large irregular electron-dense granules in the cytoplasm 48 hours after infection. Examination of infected cultures by light microscopy, using the Feulgen reaction, showed intranuclear inclusion bodies and a cytopathogenic effect consisting of clumping of cells without pyknosis of nuclei. A lipide stain showed numerous cytoplasmic granules that were not identical with the large, irregular, electron-dense granules of the cytoplasm. Practically all the cells showed the viral cytopathogenic effect, but only a minority of cells were found to contain virus-like particles or intranuclear inclusion bodies. PMID:13357696

Concise Review: Emerging Drugs Targeting Epithelial Cancer Stem-Like Cells.

PubMed

Ahmed, Mehreen; Chaudhari, Kritika; Babaei-Jadidi, Roya; Dekker, Lodewijk V; Shams Nateri, Abdolrahman

2017-04-01

Increasing evidence suggests that cancer cell populations contain a small proportion of cells that display stem-like cell properties and which may be responsible for overall tumor maintenance. These cancer stem-like cells (CSCs) appear to have unique tumor-initiating ability and innate survival mechanisms that allow them to resist cancer therapies, consequently promoting relapses. Selective targeting of CSCs may provide therapeutic benefit and several recent reports have indicated this may be possible. In this article, we review drugs targeting CSCs, in selected epithelial cell-derived cancers. Stem Cells 2017;35:839-850. © 2017 AlphaMed Press.

Scalar, Axial, and Tensor Interactions of Light Nuclei from Lattice QCD

NASA Astrophysics Data System (ADS)

Chang, Emmanuel; Davoudi, Zohreh; Detmold, William; Gambhir, Arjun S.; Orginos, Kostas; Savage, Martin J.; Shanahan, Phiala E.; Wagman, Michael L.; Winter, Frank; Nplqcd Collaboration

2018-04-01

Complete flavor decompositions of the matrix elements of the scalar, axial, and tensor currents in the proton, deuteron, diproton, and 3He at SU(3)-symmetric values of the quark masses corresponding to a pion mass mπ˜806 MeV are determined using lattice quantum chromodynamics. At the physical quark masses, the scalar interactions constrain mean-field models of nuclei and the low-energy interactions of nuclei with potential dark matter candidates. The axial and tensor interactions of nuclei constrain their spin content, integrated transversity, and the quark contributions to their electric dipole moments. External fields are used to directly access the quark-line connected matrix elements of quark bilinear operators, and a combination of stochastic estimation techniques is used to determine the disconnected sea-quark contributions. The calculated matrix elements differ from, and are typically smaller than, naive single-nucleon estimates. Given the particularly large, O (10 %), size of nuclear effects in the scalar matrix elements, contributions from correlated multinucleon effects should be quantified in the analysis of dark matter direct-detection experiments using nuclear targets.

Scalar, Axial, and Tensor Interactions of Light Nuclei from Lattice QCD

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chang, Emmanuel; Davoudi, Zohreh; Detmold, William

Complete flavor decompositions of the matrix elements of the scalar, axial, and tensor currents in the proton, deuteron, diproton, and 3He at SU(3)-symmetric values of the quark masses corresponding to a pion mass m π~806 MeV are determined using lattice quantum chromodynamics. At the physical quark masses, the scalar interactions constrain mean-field models of nuclei and the low-energy interactions of nuclei with potential dark matter candidates. The axial and tensor interactions of nuclei constrain their spin content, integrated transversity, and the quark contributions to their electric dipole moments. External fields are used to directly access the quark-line connected matrix elementsmore » of quark bilinear operators, and a combination of stochastic estimation techniques is used to determine the disconnected sea-quark contributions. The calculated matrix elements differ from, and are typically smaller than, naive single-nucleon estimates. Given the particularly large, O(10%), size of nuclear effects in the scalar matrix elements, contributions from correlated multinucleon effects should be quantified in the analysis of dark matter direct-detection experiments using nuclear targets.« less

Scalar, Axial, and Tensor Interactions of Light Nuclei from Lattice QCD

DOE PAGES

Chang, Emmanuel; Davoudi, Zohreh; Detmold, William; ...

2018-04-13

Complete flavor decompositions of the matrix elements of the scalar, axial, and tensor currents in the proton, deuteron, diproton, and 3He at SU(3)-symmetric values of the quark masses corresponding to a pion mass m π~806 MeV are determined using lattice quantum chromodynamics. At the physical quark masses, the scalar interactions constrain mean-field models of nuclei and the low-energy interactions of nuclei with potential dark matter candidates. The axial and tensor interactions of nuclei constrain their spin content, integrated transversity, and the quark contributions to their electric dipole moments. External fields are used to directly access the quark-line connected matrix elementsmore » of quark bilinear operators, and a combination of stochastic estimation techniques is used to determine the disconnected sea-quark contributions. The calculated matrix elements differ from, and are typically smaller than, naive single-nucleon estimates. Given the particularly large, O(10%), size of nuclear effects in the scalar matrix elements, contributions from correlated multinucleon effects should be quantified in the analysis of dark matter direct-detection experiments using nuclear targets.« less

Scalar, Axial, and Tensor Interactions of Light Nuclei from Lattice QCD.

PubMed

Chang, Emmanuel; Davoudi, Zohreh; Detmold, William; Gambhir, Arjun S; Orginos, Kostas; Savage, Martin J; Shanahan, Phiala E; Wagman, Michael L; Winter, Frank

2018-04-13

Complete flavor decompositions of the matrix elements of the scalar, axial, and tensor currents in the proton, deuteron, diproton, and ^{3}He at SU(3)-symmetric values of the quark masses corresponding to a pion mass m_{π}∼806  MeV are determined using lattice quantum chromodynamics. At the physical quark masses, the scalar interactions constrain mean-field models of nuclei and the low-energy interactions of nuclei with potential dark matter candidates. The axial and tensor interactions of nuclei constrain their spin content, integrated transversity, and the quark contributions to their electric dipole moments. External fields are used to directly access the quark-line connected matrix elements of quark bilinear operators, and a combination of stochastic estimation techniques is used to determine the disconnected sea-quark contributions. The calculated matrix elements differ from, and are typically smaller than, naive single-nucleon estimates. Given the particularly large, O(10%), size of nuclear effects in the scalar matrix elements, contributions from correlated multinucleon effects should be quantified in the analysis of dark matter direct-detection experiments using nuclear targets.

MicroRNA-944 Affects Cell Growth by Targeting EPHA7 in Non-Small Cell Lung Cancer.

PubMed

Liu, Minxia; Zhou, Kecheng; Cao, Yi

2016-09-26

MicroRNAs (miRNAs) have critical roles in lung tumorigenesis and development. To determine aberrantly expressed miRNAs involved in non-small cell lung cancer (NSCLC) and investigate pathophysiological functions and mechanisms, we firstly carried out small RNA deep sequencing in NSCLC cell lines (EPLC-32M1, A549 and 801D) and a human immortalized cell line 16HBE, we then studied miRNA function by cell proliferation and apoptosis. cDNA microarray, luciferase reporter assay and miRNA transfection were used to investigate interaction between the miRNA and target gene. miR-944 was significantly down-regulated in NSCLC and had many putative targets. Moreover, the forced expression of miR-944 significantly inhibited the proliferation of NSCLC cells in vitro. By integrating mRNA expression data and miR-944-target prediction, we disclosed that EPHA7 was a potential target of miR-944, which was further verified by luciferase reporter assay and microRNA transfection. Our data indicated that miR-944 targets EPHA7 in NSCLC and regulates NSCLC cell proliferation, which may offer a new mechanism underlying the development and progression of NSCLC.

New Strategies in Engineering T-cell Receptor Gene-Modified T cells to More Effectively Target Malignancies.

PubMed

Schmitt, Thomas M; Stromnes, Ingunn M; Chapuis, Aude G; Greenberg, Philip D

2015-12-01

The immune system, T cells in particular, have the ability to target and destroy malignant cells. However, antitumor immune responses induced from the endogenous T-cell repertoire are often insufficient for the eradication of established tumors, as illustrated by the failure of cancer vaccination strategies or checkpoint blockade for most tumors. Genetic modification of T cells to express a defined T-cell receptor (TCR) can provide the means to rapidly generate large numbers of tumor-reactive T cells capable of targeting tumor cells in vivo. However, cell-intrinsic factors as well as immunosuppressive factors in the tumor microenvironment can limit the function of such gene-modified T cells. New strategies currently being developed are refining and enhancing this approach, resulting in cellular therapies that more effectively target tumors and that are less susceptible to tumor immune evasion. ©2015 American Association for Cancer Research.

Probing Xist RNA Structure in Cells Using Targeted Structure-Seq

PubMed Central

Rutenberg-Schoenberg, Michael; Simon, Matthew D.

2015-01-01

The long non-coding RNA (lncRNA) Xist is a master regulator of X-chromosome inactivation in mammalian cells. Models for how Xist and other lncRNAs function depend on thermodynamically stable secondary and higher-order structures that RNAs can form in the context of a cell. Probing accessible RNA bases can provide data to build models of RNA conformation that provide insight into RNA function, molecular evolution, and modularity. To study the structure of Xist in cells, we built upon recent advances in RNA secondary structure mapping and modeling to develop Targeted Structure-Seq, which combines chemical probing of RNA structure in cells with target-specific massively parallel sequencing. By enriching for signals from the RNA of interest, Targeted Structure-Seq achieves high coverage of the target RNA with relatively few sequencing reads, thus providing a targeted and scalable approach to analyze RNA conformation in cells. We use this approach to probe the full-length Xist lncRNA to develop new models for functional elements within Xist, including the repeat A element in the 5’-end of Xist. This analysis also identified new structural elements in Xist that are evolutionarily conserved, including a new element proximal to the C repeats that is important for Xist function. PMID:26646615

Cell targeting peptides as smart ligands for targeting of therapeutic or diagnostic agents: a systematic review.

PubMed

Mousavizadeh, Ali; Jabbari, Ali; Akrami, Mohammad; Bardania, Hassan

2017-10-01

Cell targeting peptides (CTP) are small peptides which have high affinity and specificity to a cell or tissue targets. They are typically identified by using phage display and chemical synthetic peptide library methods. CTPs have attracted considerable attention as a new class of ligands to delivery specifically therapeutic and diagnostic agents, because of the fact they have several advantages including easy synthesis, smaller physical sizes, lower immunogenicity and cytotoxicity and their simple and better conjugation to nano-carriers and therapeutic or diagnostic agents compared to conventional antibodies. In this systematic review, we will focus on the basic concepts concerning the use of cell-targeting peptides (CTPs), following the approaches of selecting them from peptide libraries. We discuss several developed strategies for cell-specific delivery of different cargos by CTPs, which are designed for drug delivery and diagnostic applications. Copyright © 2017 Elsevier B.V. All rights reserved.

Tumor-targeting domains for chimeric antigen receptor T cells.

PubMed

Bezverbnaya, Ksenia; Mathews, Ashish; Sidhu, Jesse; Helsen, Christopher W; Bramson, Jonathan L

2017-01-01

Immunotherapy with chimeric antigen receptor (CAR) T cells has been advancing steadily in clinical trials. Since the ability of engineered T cells to recognize intended tumor-associated targets is crucial for the therapeutic success, antigen-binding domains play an important role in shaping T-cell responses. Single-chain antibody and T-cell receptor fragments, natural ligands, repeat proteins, combinations of the above and universal tag-specific domains have all been used in the antigen-binding moiety of chimeric receptors. Here we outline the advantages and disadvantages of different domains, discuss the concepts of affinity and specificity, and highlight the recent progress of each targeting strategy.

Optical cell monitoring system for underwater targets

NASA Astrophysics Data System (ADS)

Moon, SangJun; Manzur, Fahim; Manzur, Tariq; Demirci, Utkan

2008-10-01

We demonstrate a cell based detection system that could be used for monitoring an underwater target volume and environment using a microfluidic chip and charge-coupled-device (CCD). This technique allows us to capture specific cells and enumerate these cells on a large area on a microchip. The microfluidic chip and a lens-less imaging platform were then merged to monitor cell populations and morphologies as a system that may find use in distributed sensor networks. The chip, featuring surface chemistry and automatic cell imaging, was fabricated from a cover glass slide, double sided adhesive film and a transparent Polymethlymetacrylate (PMMA) slab. The optically clear chip allows detecting cells with a CCD sensor. These chips were fabricated with a laser cutter without the use of photolithography. We utilized CD4+ cells that are captured on the floor of a microfluidic chip due to the ability to address specific target cells using antibody-antigen binding. Captured CD4+ cells were imaged with a fluorescence microscope to verify the chip specificity and efficiency. We achieved 70.2 +/- 6.5% capturing efficiency and 88.8 +/- 5.4% specificity for CD4+ T lymphocytes (n = 9 devices). Bright field images of the captured cells in the 24 mm × 4 mm × 50 μm microfluidic chip were obtained with the CCD sensor in one second. We achieved an inexpensive system that rapidly captures cells and images them using a lens-less CCD system. This microfluidic device can be modified for use in single cell detection utilizing a cheap light-emitting diode (LED) chip instead of a wide range CCD system.

Biomarker evaluation of face transplant rejection: association of donor T cells with target cell injury.

PubMed

Lian, Christine Guo; Bueno, Ericka M; Granter, Scott R; Laga, Alvaro C; Saavedra, Arturo P; Lin, William M; Susa, Joseph S; Zhan, Qian; Chandraker, Anil K; Tullius, Stefan G; Pomahac, Bohdan; Murphy, George F

2014-06-01

This series of 113 sequential biopsies of full facial transplants provides findings of potential translational significance as well as biological insights that could prompt reexamination of conventional paradigms of effector pathways in skin allograft rejection. Serial biopsies before, during, and after rejection episodes were evaluated for clinicopathological assessment that in selected cases included specific biomarkers for donor-versus-recipient T cells. Histologic evidence of rejection included lymphocyte-associated injury to epidermal rete ridges, follicular infundibula, and dermal microvessels. Surprisingly, during active rejection, immune cells spatially associated with target cell injury consisted abundantly or predominantly of lymphocytes of donor origin with an immunophenotype typical of the resident memory T-cell subset. Current dogma assumes that skin allograft rejection is mediated by recipient T cells that attack epidermal targets, and the association of donor T cells with sites of target cell injury raises questions regarding the potential complexity of immune cell interactions in the rejection process. A more histopathologically refined and immune-based biomarker approach to assessment of rejection of facial transplants is now indicated.

Efficient Generation of Somatic Cell Nuclear Transfer-Competent Porcine Cells with Mutated Alleles at Multiple Target Loci by Using CRISPR/Cas9 Combined with Targeted Toxin-Based Selection System.

PubMed

Sato, Masahiro; Miyoshi, Kazuchika; Nakamura, Shingo; Ohtsuka, Masato; Sakurai, Takayuki; Watanabe, Satoshi; Kawaguchi, Hiroaki; Tanimoto, Akihide

2017-12-04

The recent advancement in genome editing such a CRISPR/Cas9 system has enabled isolation of cells with knocked multiple alleles through a one-step transfection. Somatic cell nuclear transfer (SCNT) has been frequently employed as one of the efficient tools for the production of genetically modified (GM) animals. To use GM cells as SCNT donor, efficient isolation of transfectants with mutations at multiple target loci is often required. The methods for the isolation of such GM cells largely rely on the use of drug selection-based approach using selectable genes; however, it is often difficult to isolate cells with mutations at multiple target loci. In this study, we used a novel approach for the efficient isolation of porcine cells with at least two target loci mutations by one-step introduction of CRISPR/Cas9-related components. A single guide (sg) RNA targeted to GGTA1 gene, involved in the synthesis of cell-surface α-Gal epitope (known as xenogenic antigen), is always a prerequisite. When the transfected cells were reacted with toxin-labeled BS-I-B₄ isolectin for 2 h at 37 C to eliminate α-Gal epitope-expressing cells, the surviving clones lacked α-Gal epitope expression and were highly expected to exhibit induced mutations at another target loci. Analysis of these α-Gal epitope-negative surviving cells demonstrated a 100% occurrence of genome editing at target loci. SCNT using these cells as donors resulted in the production of cloned blastocysts with the genotype similar to that of the donor cells used. Thus, this novel system will be useful for SCNT-mediated acquisition of GM cloned piglets, in which multiple target loci may be mutated.

Curcumin suppresses proliferation of colon cancer cells by targeting CDK2.

PubMed

Lim, Tae-Gyu; Lee, Sung-Young; Huang, Zunnan; Lim, Do Young; Chen, Hanyong; Jung, Sung Keun; Bode, Ann M; Lee, Ki Won; Dong, Zigang

2014-04-01

Curcumin, the yellow pigment of turmeric found in Southeast Indian food, is one of the most popular phytochemicals for cancer prevention. Numerous reports have demonstrated modulation of multiple cellular signaling pathways by curcumin and its molecular targets in various cancer cell lines. To identify a new molecular target of curcumin, we used shape screening and reverse docking to screen the Protein Data Bank against curcumin. Cyclin-dependent kinase 2 (CDK2), a major cell-cycle protein, was identified as a potential molecular target of curcumin. Indeed, in vitro and ex vivo kinase assay data revealed a dramatic suppressive effect of curcumin on CDK2 kinase activity. Furthermore, curcumin induced G1 cell-cycle arrest, which is regulated by CDK2 in HCT116 cells. Although the expression levels of CDK2 and its regulatory subunit, cyclin E, were not changed, the phosphorylation of retinoblastoma (Rb), a well-known CDK2 substrate, was reduced by curcumin. Because curcumin induced cell-cycle arrest, we investigated the antiproliferative effect of curcumin on HCT116 colon cancer cells. In this experiment, curcumin suppressed HCT116 cell proliferation effectively. To determine whether CDK2 is a direct target of curcumin, CDK2 expression was knocked down in HCT116 cells. As expected, HCT116 sh-CDK2 cells exhibited G1 arrest and reduced proliferation. Because of the low levels of CDK2 in HCT116 sh-CDK2 cells, the effects of curcumin on G1 arrest and cell proliferation were not substantially relative to HCT116 sh-control cells. From these results, we identified CDK2 as a direct target of curcumin in colon cancer cells.

Curcumin suppresses proliferation of colon cancer cells by targeting CDK2

PubMed Central

Lim, Tae-Gyu; Lee, Sung-Young; Huang, Zunnan; Lim, Do Young; Chen, Hanyong; Jung, Sung Keun; Bode, Ann M.; Lee, Ki Won; Dong, Zigang

2014-01-01

Curcumin, the yellow pigment of turmeric found in Southeast Indian food, is one of the most popular phytochemicals for cancer prevention. Numerous reports have demonstrated modulation of multiple cellular signaling pathways by curcumin and its molecular targets in various cancer cell lines. To identify a new molecular target of curcumin, we used shape screening and reverse docking to screen the protein data bank against curcumin. Cyclin dependent kinase 2 (CDK2), a major cell cycle protein, was identified as a potential molecular target of curcumin. Indeed, in vitro and ex vivo kinase assay data revealed a dramatic suppressive effect of curcumin on CDK2 kinase activity. Furthermore, curcumin induced G1 cell cycle arrest, which is regulated by CDK2 in HCT116 cells. Although the expression levels of CDK2 and its regulatory subunit, cyclin E, were not changed, the phosphorylation of Rb, a well-known CDK2 substrate, was reduced by curcumin. Because curcumin induced cell cycle arrest, we investigated the anti-proliferative effect of curcumin on HCT116 colon cancer cells. In this experiment, curcumin suppressed HCT116 cell proliferation effectively. To determine if CDK2 is a direct target of curcumin, CDK2 expression was knocked down in HCT116 cells. As expected, HCT116 sh-CDK2 cells exhibited G1 arrest and reduced proliferation. Because of the low levels of CDK2 in HCT116 sh-CDK2 cells, the effects of curcumin on G1 arrest and cell proliferation were not substantial relative to HCT116 sh-control cells. From these results, we identified CDK2 as a direct target of curcumin in colon cancer cells. PMID:24550143

Enhancing Oral Vaccine Potency by Targeting Intestinal M Cells

PubMed Central

Azizi, Ali; Kumar, Ashok; Diaz-Mitoma, Francisco; Mestecky, Jiri

2010-01-01

The immune system in the gastrointestinal tract plays a crucial role in the control of infection, as it constitutes the first line of defense against mucosal pathogens. The attractive features of oral immunization have led to the exploration of a variety of oral delivery systems. However, none of these oral delivery systems have been applied to existing commercial vaccines. To overcome this, a new generation of oral vaccine delivery systems that target antigens to gut-associated lymphoid tissue is required. One promising approach is to exploit the potential of microfold (M) cells by mimicking the entry of pathogens into these cells. Targeting specific receptors on the apical surface of M cells might enhance the entry of antigens, initiating the immune response and consequently leading to protection against mucosal pathogens. In this article, we briefly review the challenges associated with current oral vaccine delivery systems and discuss strategies that might potentially target mouse and human intestinal M cells. PMID:21085599

Microcinematographic and electron microscopic analysis of target cell lysis induced by cytotoxic T lymphocytes.

PubMed Central

Matter, A

1979-01-01

A study was carried out to determine the sequence of events of T-cell mediated target cell lysis in microcinematography and electron microscopy. Highly efficient cytotoxic T lymphocytes (CTL) were generated in vivo and in vitro using preimmunized spleen cells and purification procedures. Such CTL were highly specific. This specificity correlated well with the number of adhesions formed between CTL and targets and this criterion was used to study killer-target cell interaction. Microcinematography showed that target cell lysis at the single cell level, despite time variations, could be clearly separated into three phases: (a) a recognition phase, visible by random crawling of CTL over the target cell surface until firm contact was established; (b) a post-recognition phase, during which firm contact between CTL and target was maintained without gross modification of either cell; (c) a phase of target cell disintegration, mainly characterized by vigorous blebbing of the cell membrane resulting in a motionless carcass of the target cell but not in its total dissolution. Only later this carcass decayed and formed a necrotic ghost. Electron microscopic observations were put into sequence according to microcinematography. Post-recognition phase was characterized by a tight apposition of the membranes of CTL and target cell. No gap junctions could be observed. During target cell disintegration, profound cytoplasmic and nuclear changes occurred simultaneous with surface blebbing. Most noticeable were extensive internal vacuolization, mitochondrial swelling, nuclear pycnosis and dissolution of the nucleolus. These observations suggested that target cell lysis does not start with a surface phenomenon similar to complement lysis, but a process involving practically the whole cell simultaneously. It is conceivable, therefore, that the signal from the CTL is transmitted across the target cell, and that the switch to sudden cell death is manipulated deep inside the cell. Images

Colon-targeted delivery of live bacterial cell biotherapeutics including microencapsulated live bacterial cells

PubMed Central

Prakash, Satya; Malgorzata Urbanska, Aleksandra

2008-01-01

There has been an ample interest in delivery of therapeutic molecules using live cells. Oral delivery has been stipulated as best way to deliver live cells to humans for therapy. Colon, in particular, is a part of gastrointestinal (GI) tract that has been proposed to be an oral targeted site. The main objective of these oral therapy procedures is to deliver live cells not only to treat diseases like colorectal cancer, inflammatory bowel disease, and other GI tract diseases like intestinal obstruction and gastritis, but also to deliver therapeutic molecules for overall therapy in various diseases such as renal failure, coronary heart disease, hypertension, and others. This review provides a comprehensive summary of recent advancement in colon targeted live bacterial cell biotherapeutics. Current status of bacterial cell therapy, principles of artificial cells and its potentials in oral delivery of live bacterial cell biotherapeutics for clinical applications as well as biotherapeutic future perspectives are also discussed in our review. PMID:19707368

Star formation around active galactic nuclei

NASA Technical Reports Server (NTRS)

Keel, William C.

1987-01-01

Active galactic nuclei (Seyfert nuclei and their relatives) and intense star formation can both deliver substantial amounts of energy to the vicinity of a galactic nucleus. Many luminous nuclei have energetics dominated by one of these mechanisms, but detailed observations show that some have a mixture. Seeing both phenomena at once raises several interesting questions: (1) Is this a general property of some kinds of nuclei? How many AGNs have surround starbursts, and vice versa? (2) As in 1, how many undiscovered AGNs or starbursts are hidden by a more luminous instance of the other? (3) Does one cause the other, and by what means, or do both reflect common influences such as potential well shape or level of gas flow? (4) Can surrounding star formation tell us anything about the central active nuclei, such as lifetimes, kinetic energy output, or mechanical disturbance of the ISM? These are important points in the understanding of activity and star formation in galactic nuclei. Unfortunately, the observational ways of addressing them are as yet not well formulated. Some preliminary studies are reported, aimed at clarifying the issues involved in study of the relationships between stellar and nonstellar excitement in galactic nuclei.

Targeting Notch signalling pathway of cancer stem cells.

PubMed

Venkatesh, Vandana; Nataraj, Raghu; Thangaraj, Gopenath S; Karthikeyan, Murugesan; Gnanasekaran, Ashok; Kaginelli, Shanmukhappa B; Kuppanna, Gobianand; Kallappa, Chandrashekrappa Gowdru; Basalingappa, Kanthesh M

2018-01-01

Cancer stem cells (CSCs) have been defined as cells within tumor that possess the capacity to self-renew and to cause the heterogeneous lineages of cancer cells that comprise the tumor. CSCs have been increasingly identified in blood cancer, prostate, ovarian, lung, melanoma, pancreatic, colon, brain and many more malignancies. CSCs have slow growth rate and are resistant to chemotherapy and radiotherapy that lead to the failure of traditional current therapy. Eradicating the CSCs and recurrence, is promising aspect for the cure of cancer. The CSCs like any other stem cells activate the signal transduction pathways that involve the development and tissue homeostasis, which include Notch signaling pathway. The new treatment targets these pathway that control stem-cell replication, survival and differentiation that are under development. Notch inhibitors either single or in combination with chemotherapy drugs have been developed to treat cancer and its recurrence. This approach of targeting signaling pathway of CSCs represents a promising future direction for the therapeutic strategy to cure cancer.

Application of JAERI quantum molecular dynamics model for collisions of heavy nuclei

NASA Astrophysics Data System (ADS)

Ogawa, Tatsuhiko; Hashimoto, Shintaro; Sato, Tatsuhiko; Niita, Koji

2016-06-01

The quantum molecular dynamics (QMD) model incorporated into the general-purpose radiation transport code PHITS was revised for accurate prediction of fragment yields in peripheral collisions. For more accurate simulation of peripheral collisions, stability of the nuclei at their ground state was improved and the algorithm to reject invalid events was modified. In-medium correction on nucleon-nucleon cross sections was also considered. To clarify the effect of this improvement on fragmentation of heavy nuclei, the new QMD model coupled with a statistical decay model was used to calculate fragment production cross sections of Ag and Au targets and compared with the data of earlier measurement. It is shown that the revised version can predict cross section more accurately.

The nucleus is irreversibly shaped by motion of cell boundaries in cancer and non-cancer cells.

PubMed

Tocco, Vincent J; Li, Yuan; Christopher, Keith G; Matthews, James H; Aggarwal, Varun; Paschall, Lauren; Luesch, Hendrik; Licht, Jonathan D; Dickinson, Richard B; Lele, Tanmay P

2018-02-01

Actomyosin stress fibers impinge on the nucleus and can exert compressive forces on it. These compressive forces have been proposed to elongate nuclei in fibroblasts, and lead to abnormally shaped nuclei in cancer cells. In these models, the elongated or flattened nuclear shape is proposed to store elastic energy. However, we found that deformed shapes of nuclei are unchanged even after removal of the cell with micro-dissection, both for smooth, elongated nuclei in fibroblasts and abnormally shaped nuclei in breast cancer cells. The lack of shape relaxation implies that the nuclear shape in spread cells does not store any elastic energy, and the cellular stresses that deform the nucleus are dissipative, not static. During cell spreading, the deviation of the nucleus from a convex shape increased in MDA-MB-231 cancer cells, but decreased in MCF-10A cells. Tracking changes of nuclear and cellular shape on micropatterned substrata revealed that fibroblast nuclei deform only during deformations in cell shape and only in the direction of nearby moving cell boundaries. We propose that motion of cell boundaries exert a stress on the nucleus, which allows the nucleus to mimic cell shape. The lack of elastic energy in the nuclear shape suggests that nuclear shape changes in cells occur at constant surface area and volume. © 2017 Wiley Periodicals, Inc.

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

PubMed

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

2012-12-01

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

Glioblastoma Stem Cells as a New Therapeutic Target for Glioblastoma.

PubMed

Kalkan, Rasime

2015-01-01

Primary and secondary glioblastomas (GBMs) are two distinct diseases. The genetic and epigenetic background of these tumors is highly variable. The treatment procedure for these tumors is often unsuccessful because of the cellular heterogeneity and intrinsic ability of the tumor cells to invade healthy tissues. The fatal outcome of these tumors promotes researchers to find out new markers associated with the prognosis and treatment planning. In this communication, the role of glioblastoma stem cells in tumor progression and the malignant behavior of GBMs are summarized with attention to the signaling pathways and molecular regulators that are involved in maintaining the glioblastoma stem cell phenotype. A better understanding of these stem cell-like cells is necessary for designing new effective treatments and developing novel molecular strategies to target glioblastoma stem cells. We discuss hypoxia as a new therapeutic target for GBM. We focus on the inhibition of signaling pathways, which are associated with the hypoxia-mediated maintenance of glioblastoma stem cells, and the knockdown of hypoxia-inducible factors, which could be identified as attractive molecular target approaches for GBM therapeutics.

Monoclonal antibodies targeting non-small cell lung cancer stem-like cells by multipotent cancer stem cell monoclonal antibody library.

PubMed

Cao, Kaiyue; Pan, Yunzhi; Yu, Long; Shu, Xiong; Yang, Jing; Sun, Linxin; Sun, Lichao; Yang, Zhihua; Ran, Yuliang

2017-02-01

Cancer stem cells (CSCs) are a rare subset of cancer cells that play a significant role in cancer initiation, spreading, and recurrence. In this study, a subpopulation of lung cancer stem-like cells (LCSLCs) was identified from non-small cell lung carcinoma cell lines, SPCA-1 and A549, using serum-free suspension sphere-forming culture method. A monoclonal antibody library was constructed using immunized BLAB/c mice with the multipotent CSC cell line T3A-A3. Flow cytometry analysis showed that 33 mAbs targeted antigens can be enriched in sphere cells compared with the parental cells of SPCA-1 and A549 cell lines. Then, we performed functional antibody screening including sphere-forming inhibiting and invasion inhibiting assay. The results showed that two antibodies, 12C7 and 9B8, notably suppressed the self-renewal and invasion of LCSLCs. Fluorescence-activated cell sorting (FACs) found that the positive cells recognized by mAbs, 12C7 or 9B8, displayed features of LCSLCs. Interestingly, we found that these two antibodies recognized different subsets of cells and their combination effect was superior to the individual effect both in vitro and in vivo. Tissue microarrays were applied to detect the expression of the antigens targeted by these two antibodies. The positive expression of 12C7 and 9B8 targeted antigen was 84.4 and 82.5%, respectively, which was significantly higher than that in the non-tumor lung tissues. In conclusion, we screened two potential therapeutic antibodies that target different subsets of LCSLCs.

Tracking of cell nuclei for assessment of in vitro uptake kinetics in ultrasound-mediated drug delivery using fibered confocal fluorescence microscopy.

PubMed

Derieppe, Marc; de Senneville, Baudouin Denis; Kuijf, Hugo; Moonen, Chrit; Bos, Clemens

2014-10-01

Previously, we demonstrated the feasibility to monitor ultrasound-mediated uptake of a cell-impermeable model drug in real time with fibered confocal fluorescence microscopy. Here, we present a complete post-processing methodology, which corrects for cell displacements, to improve the accuracy of pharmacokinetic parameter estimation. Nucleus detection was performed based on the radial symmetry transform algorithm. Cell tracking used an iterative closest point approach. Pharmacokinetic parameters were calculated by fitting a two-compartment model to the time-intensity curves of individual cells. Cells were tracked successfully, improving time-intensity curve accuracy and pharmacokinetic parameter estimation. With tracking, 93 % of the 370 nuclei showed a fluorescence signal variation that was well-described by a two-compartment model. In addition, parameter distributions were narrower, thus increasing precision. Dedicated image analysis was implemented and enabled studying ultrasound-mediated model drug uptake kinetics in hundreds of cells per experiment, using fiber-based confocal fluorescence microscopy.

HLA-targeted flow cytometric sorting of blood cells allows separation of pure and viable microchimeric cell populations.

PubMed

Drabbels, Jos J M; van de Keur, Carin; Kemps, Berit M; Mulder, Arend; Scherjon, Sicco A; Claas, Frans H J; Eikmans, Michael

2011-11-10

Microchimerism is defined by the presence of low levels of nonhost cells in a person. We developed a reliable method for separating viable microchimeric cells from the host environment. For flow cytometric cell sorting, HLA antigens were targeted with human monoclonal HLA antibodies (mAbs). Optimal separation of microchimeric cells (present at a proportion as low as 0.01% in artificial mixtures) was obtained with 2 different HLA mAbs, one targeting the chimeric cells and the other the background cells. To verify purity of separated cell populations, flow-sorted fractions of 1000 cells were processed for DNA analysis by HLA-allele-specific and Y-chromosome-directed real-time quantitative PCR assays. After sorting, PCR signals of chimeric DNA markers in the positive fractions were significantly enhanced compared with those in the presort samples, and they were similar to those in 100% chimeric control samples. Next, we demonstrate applicability of HLA-targeted FACS sorting after pregnancy by separating chimeric maternal cells from child umbilical cord mononuclear cells. Targeting allelic differences with anti-HLA mAbs with FACS sorting allows maximal enrichment of viable microchimeric cells from a background cell population. The current methodology enables reliable microchimeric cell detection and separation in clinical specimens.

Zn(II)-curc targets p53 in thyroid cancer cells.

PubMed

Garufi, Alessia; D'Orazi, Valerio; Crispini, Alessandra; D'Orazi, Gabriella

2015-10-01

TP53 mutation is a common event in many cancers, including thyroid carcinoma. Defective p53 activity promotes cancer resistance to therapies and a more malignant phenotype, acquiring oncogenic functions. Rescuing the function of mutant p53 (mutp53) protein is an attractive anticancer therapeutic strategy. Zn(II)-curc is a novel small molecule that has been shown to target mutp53 protein in several cancer cells, but its effect in thyroid cancer cells remains unclear. Here, we investigated whether Zn(II)-curc could affect p53 in thyroid cancer cells with both p53 mutation (R273H) and wild-type p53. Zn(II)-curc induced mutp53H273 downregulation and reactivation of wild-type functions, such as binding to canonical target promoters and target gene transactivation. This latter effect was similar to that induced by PRIMA-1. In addition, Zn(II)-curc triggered p53 target gene expression in wild-type p53-carrying cells. In combination treatments, Zn(II)-curc enhanced the antitumor activity of chemotherapeutic drugs, in both mutant and wild-type-carrying cancer cells. Taken together, our data indicate that Zn(II)-curc promotes the reactivation of p53 in thyroid cancer cells, providing in vitro evidence for a potential therapeutic approach in thyroid cancers.

[Effect of Biejiajian Pills on Wnt signal pathway molecules β-catenin and GSK-3β and the target genes CD44v6 and VEGF in hepatocellular carcinoma cells].

PubMed

Sun, Haitao; He, Songqi; Wen, Bin; Jia, Wenyan; Fan, Eryan; Zheng, Yan

2014-10-01

To investigate the effect of Biejiajian Pills on the expressions of the signal molecules and target genes of Wnt signal pathway in HepG2 cells and explore the mechanisms by which Biejiajian pills suppress the invasiveness of hepatocellular carcinoma. HepG2 cells were cultured for 48 h in the presence of serum collected from rats fed with Biejiajian Pills. The expressions of β-catenin, GSK-3β and P-GSK-3β in the cultured cells were assessed by Western blotting and the expressions of CD44v6 and VEGF were detected using immunohistochemistry. HepG2 cells cultured with the serum of rats fed with Biejiajian Pills showed lowered expressions of β-catenin protein both in the cytoplasm and the nuclei with also inhibition of phosphorylation of GSK-3β and reduced expression of CD44v6 and VEGF. Biejiajian Pills can significantly reduce the expression of β-catenin by decreasing the phosphorylation of GSK-3β and blocking the Wnt/β-catenin signaling pathway to cause down-regulation of the target genes CD44v6 and VEGF, which may be one of the molecular mechanisms by which Biejiajian Pills suppress the proliferation and invasiveness of hepatocellular carcinoma.

ErbB-targeted CAR T-cell immunotherapy of cancer.

PubMed

Whilding, Lynsey M; Maher, John

2015-01-01

Chimeric antigen receptor (CAR) based immunotherapy has been under development for the last 25 years and is now a promising new treatment modality in the field of cancer immunotherapy. The approach involves genetically engineering T cells to target malignant cells through expression of a bespoke fusion receptor that couples an HLA-independent antigen recognition domain to one or more intracellular T-cell activating modules. Multiple clinical trials are now underway in several centers to investigate CAR T-cell immunotherapy of diverse hematologic and solid tumor types. The most successful results have been achieved in the treatment of patients with B-cell malignancies, in whom several complete and durable responses have been achieved. This review focuses on the preclinical and clinical development of CAR T-cell immunotherapy of solid cancers, targeted against members of the ErbB family.

Thalamic atrophy in antero-medial and dorsal nuclei correlates with six-month outcome after severe brain injury☆

PubMed Central

Lutkenhoff, Evan S.; McArthur, David L.; Hua, Xue; Thompson, Paul M.; Vespa, Paul M.; Monti, Martin M.

2013-01-01

The primary and secondary damage to neural tissue inflicted by traumatic brain injury is a leading cause of death and disability. The secondary processes, in particular, are of great clinical interest because of their potential susceptibility to intervention. We address the dynamics of tissue degeneration in cortico-subcortical circuits after severe brain injury by assessing volume change in individual thalamic nuclei over the first six-months post-injury in a sample of 25 moderate to severe traumatic brain injury patients. Using tensor-based morphometry, we observed significant localized thalamic atrophy over the six-month period in antero-dorsal limbic nuclei as well as in medio-dorsal association nuclei. Importantly, the degree of atrophy in these nuclei was predictive, even after controlling for full-brain volume change, of behavioral outcome at six-months post-injury. Furthermore, employing a data-driven decision tree model, we found that physiological measures, namely the extent of atrophy in the anterior thalamic nucleus, were the most predictive variables of whether patients had regained consciousness by six-months, followed by behavioral measures. Overall, these findings suggest that the secondary non-mechanical degenerative processes triggered by severe brain injury are still ongoing after the first week post-trauma and target specifically antero-medial and dorsal thalamic nuclei. This result therefore offers a potential window of intervention, and a specific target region, in agreement with the view that specific cortico-thalamo-cortical circuits are crucial to the maintenance of large-scale network neural activity and thereby the restoration of cognitive function after severe brain injury. PMID:24273723

Investigation of the structure of light exotic nuclei by proton elastic scattering in inverse kinematics

DOE Office of Scientific and Technical Information (OSTI.GOV)

Alkhazov, G. D.; Vorobyov, A. A.; Dobrovolsky, A. V., E-mail: dobrov@pnpi.spb.ru

2015-05-15

In order to study the spatial structure of exotic nuclei, it was proposed at the Petersburg Nuclear Physics Institute (PNPI) to measure the differential cross section for small-angle proton elastic scattering in inverse kinematics. Several experiments in beams of 0.7-GeV/nucleon exotic nuclei were performed at the heavy-ion accelerator facility of GSI (Gesellschaft für Schwerionenforschung, Darmstadt, Germany) by using the IKAR ionization spectrometer developed at PNPI. The IKAR ionization chamber filled with hydrogen at a pressure of 10 bar served simultaneously as a target and as a recoil-proton detector, which measured the recoil-proton energy. The beam-particle scattering angle was also measured.more » The results obtained for the cross sections in question were analyzed on the basis of the Glauber-Sitenko theory using phenomenological nuclear-density distributions with two free parameters. Nuclear-matter distributions and root-mean-square radii were found for the nuclei under investigation. The size of the halo in the {sup 6}He, {sup 8}He, {sup 11}Li, and {sup 14}Be nuclei was determined among other things. Information about neutron distributions in nuclei was deduced by combining the data obtained here with the known values of the radii of proton distributions. A sizable neutron skin was revealed in the {sup 8}Li, {sup 9}Li, and {sup 12}Be nuclei.« less

GEM-loaded magnetic albumin nanospheres modified with cetuximab for simultaneous targeting, magnetic resonance imaging, and double-targeted thermochemotherapy of pancreatic cancer cells.

PubMed

Wang, Ling; An, Yanli; Yuan, Chenyan; Zhang, Hao; Liang, Chen; Ding, Fengan; Gao, Qi; Zhang, Dongsheng

2015-01-01

Targeted delivery is a promising strategy to improve the diagnostic imaging and therapeutic effect of cancers. In this paper, novel cetuximab (C225)-conjugated, gemcitabine (GEM)-containing magnetic albumin nanospheres (C225-GEM/MANs) were fabricated and applied as a theranostic nanocarrier to conduct simultaneous targeting, magnetic resonance imaging (MRI), and double-targeted thermochemotherapy against pancreatic cancer cells. Fe3O4 nanoparticles (NPs) and GEM co-loaded albumin nanospheres (GEM/MANs) were prepared, and then C225 was further conjugated to synthesize C225-GEM/MANs. Their morphology, mean particle size, GEM encapsulation ratio, specific cell-binding ability, and thermal dynamic profiles were characterized. The effects of discriminating different EGFR-expressing pancreatic cancer cells (AsPC-1 and MIA PaCa-2) and monitoring cellular targeting effects were assessed by targeted MRI. Lastly, the antitumor efficiency of double/C225/magnetic-targeted and nontargeted thermochemotherapy was compared with chemotherapy alone using 3-(4, 5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) and flow cytometry (FCM) assay. When treated with targeted nanospheres, AsPC-1 cells showed a significantly less intense MRI T2 signal than MIA PaCa-2 cells, while both cells had similar signal strength when incubated with nontargeted nanospheres. T2 signal intensity was significantly lower when magnetic and C225 targeting were combined, rather than used alone. The inhibitory and apoptotic rates of each thermochemotherapy group were significantly higher than those of the chemotherapy-alone groups. Additionally, both MTT and FCM analysis verified that double-targeted thermochemotherapy had the highest targeted killing efficiency among all groups. The C225-GEM/MANs can distinguish various EGFR-expressing live pancreatic cancer cells, monitor diverse cellular targeting effects using targeted MRI imaging, and efficiently mediate double-targeted thermochemotherapy

Replication labeling patterns and chromosome territories typical of mammalian nuclei are conserved in the early metazoan Hydra.

PubMed

Alexandrova, Olga; Solovei, Irina; Cremer, Thomas; David, Charles N

2003-12-01

To investigate the evolutionary conservation of higher order nuclear architecture previously described for mammalian cells we have analyzed the nuclear architecture of the simple polyp Hydra. These diploblastic organisms have large nuclei (8-10 microm) containing about 3x10(9) bp of DNA organized in 15 chromosome pairs. They belong to the earliest metazoan phylum and are separated from mammals by at least 600 million years. Single and double pulse labeling with halogenated nucleotides (bromodeoxyuridine, iododeoxyuridine and chlorodeoxyuridine) revealed striking similarities to the known sequence of replication labeling patterns in mammalian nuclei. These patterns reflect a persistent nuclear arrangement of early, mid-, and late replicating chromatin foci that could be identified during all stages of interphase over at least 5-10 cell generations. Segregation of labeled chromatids led after several cell divisions to nuclei with single or a few labeled chromosome territories. In such nuclei distinct clusters of labeled chromatin foci were separated by extended nuclear areas with non-labeled chromatin, which is typical of a territorial arrangement of interphase chromosomes. Our results indicate the conservation of fundamental features of higher order chromatin arrangements throughout the evolution of metazoan animals and suggest the existence of conserved mechanism(s) controlling this architecture.

Proflavine sensitivity of RNA processing in isolated nuclei.

PubMed Central

Yannarell, A; Niemann, M; Schumm, D E; Webb, T E

1977-01-01

The intercalating agent proflavine inhibits the processing and subsequent release of preformed messenger RNA and ribosomal RNA from isolated liver nuclei to surrogate cytoplasm. The direct effect of proflavine on these processes, as monitored in a reconstituted cell-free system, supports the theory that base-paired segments (i.e. hairpin loops) in the precursor RNA's are involved as recognition sites in nuclear RNA processing. PMID:866181

Phenotypic high-throughput screening elucidates target pathway in breast cancer stem cell-like cells.

PubMed

Carmody, Leigh C; Germain, Andrew R; VerPlank, Lynn; Nag, Partha P; Muñoz, Benito; Perez, Jose R; Palmer, Michelle A J

2012-10-01

Cancer stem cells (CSCs) are resistant to standard cancer treatments and are likely responsible for cancer recurrence, but few therapies target this subpopulation. Due to the difficulty in propagating CSCs outside of the tumor environment, previous work identified CSC-like cells by inducing human breast epithelial cells into an epithelial-to-mesenchymal transdifferentiated state (HMLE_sh_ECad). A phenotypic screen was conducted against HMLE_sh_ECad with 300 718 compounds from the Molecular Libraries Small Molecule Repository to identify selective inhibitors of CSC growth. The screen yielded 2244 hits that were evaluated for toxicity and selectivity toward an isogenic control cell line. An acyl hydrazone scaffold emerged as a potent and selective scaffold targeting HMLE_sh_ECad. Fifty-three analogues were acquired and tested; compounds ranged in potency from 790 nM to inactive against HMLE_sh_ECad. Of the analogues, ML239 was best-in-class with an IC(50)= 1.18 µM against HMLE_sh_ECad, demonstrated a >23-fold selectivity over the control line, and was toxic to another CSC-like line, HMLE_shTwist, and a breast carcinoma cell line, MDA-MB-231. Gene expression studies conducted with ML239-treated cells showed altered gene expression in the NF-κB pathway in the HMLE_sh_ECad line but not in the isogenic control line. Future studies will be directed toward the identification of ML239 target(s).

Detecting drug-target binding in cells using fluorescence-activated cell sorting coupled with mass spectrometry analysis.

PubMed

Wilson, Kris; Webster, Scott P; Iredale, John P; Zheng, Xiaozhong; Homer, Natalie Z; Pham, Nhan T; Auer, Manfred; Mole, Damian J

2017-12-15

The assessment of drug-target engagement for determining the efficacy of a compound inside cells remains challenging, particularly for difficult target proteins. Existing techniques are more suited to soluble protein targets. Difficult target proteins include those with challenging in vitro solubility, stability or purification properties that preclude target isolation. Here, we report a novel technique that measures intracellular compound-target complex formation, as well as cellular permeability, specificity and cytotoxicity-the toxicity-affinity-permeability-selectivity (TAPS) technique. The TAPS assay is exemplified here using human kynurenine 3-monooxygenase (KMO), a challenging intracellular membrane protein target of significant current interest. TAPS confirmed target binding of known KMO inhibitors inside cells. We conclude that the TAPS assay can be used to facilitate intracellular hit validation on most, if not all intracellular drug targets.

Detecting drug-target binding in cells using fluorescence-activated cell sorting coupled with mass spectrometry analysis

NASA Astrophysics Data System (ADS)

Wilson, Kris; Webster, Scott P.; Iredale, John P.; Zheng, Xiaozhong; Homer, Natalie Z.; Pham, Nhan T.; Auer, Manfred; Mole, Damian J.

2018-01-01

The assessment of drug-target engagement for determining the efficacy of a compound inside cells remains challenging, particularly for difficult target proteins. Existing techniques are more suited to soluble protein targets. Difficult target proteins include those with challenging in vitro solubility, stability or purification properties that preclude target isolation. Here, we report a novel technique that measures intracellular compound-target complex formation, as well as cellular permeability, specificity and cytotoxicity-the toxicity-affinity-permeability-selectivity (TAPS) technique. The TAPS assay is exemplified here using human kynurenine 3-monooxygenase (KMO), a challenging intracellular membrane protein target of significant current interest. TAPS confirmed target binding of known KMO inhibitors inside cells. We conclude that the TAPS assay can be used to facilitate intracellular hit validation on most, if not all intracellular drug targets.

Targeting myeloid-derived suppressor cells for cancer immunotherapy.

PubMed

Liu, Yijun; Wei, Guowei; Cheng, Wesley A; Dong, Zhenyuan; Sun, Han; Lee, Vincent Y; Cha, Soung-Chul; Smith, D Lynne; Kwak, Larry W; Qin, Hong

2018-05-31

Myeloid-derived suppressor cells (MDSCs) are a heterogeneous population of immature myeloid cells with an immune suppressive phenotype. They represent a critical component of the immune suppressive niche described in cancer, where they support immune escape and tumor progression through direct effects on both the innate and adaptive immune responses, largely by contributing to maintenance of a high oxidative stress environment. The number of MDSCs positively correlates with protumoral activity, and often diminishes the effectiveness of immunotherapies, which is particularly problematic with the emergence of personalized medicine. Approaches targeting MDSCs showed promising results in preclinical studies and are under active investigation in clinical trials in combination with various immune checkpoint inhibitors. In this review, we discuss MDSC targets and therapeutic approaches targeting MDSC that have the aim of enhancing the existing tumor therapies.

Many si/shRNAs can kill cancer cells by targeting multiple survival genes through an off-target mechanism

PubMed Central

van Dongen, Stijn; Haluck-Kangas, Ashley; Sarshad, Aishe A; Bartom, Elizabeth T; Kim, Kwang-Youn A; Scholtens, Denise M; Hafner, Markus; Zhao, Jonathan C; Murmann, Andrea E

2017-01-01

Over 80% of multiple-tested siRNAs and shRNAs targeting CD95 or CD95 ligand (CD95L) induce a form of cell death characterized by simultaneous activation of multiple cell death pathways preferentially killing transformed and cancer stem cells. We now show these si/shRNAs kill cancer cells through canonical RNAi by targeting the 3’UTR of critical survival genes in a unique form of off-target effect we call DISE (death induced by survival gene elimination). Drosha and Dicer-deficient cells, devoid of most miRNAs, are hypersensitive to DISE, suggesting cellular miRNAs protect cells from this form of cell death. By testing 4666 shRNAs derived from the CD95 and CD95L mRNA sequences and an unrelated control gene, Venus, we have identified many toxic sequences - most of them located in the open reading frame of CD95L. We propose that specific toxic RNAi-active sequences present in the genome can kill cancer cells. PMID:29063830

Nonserotonergic projection neurons in the midbrain raphe nuclei contain the vesicular glutamate transporter VGLUT3.

PubMed

Jackson, Jesse; Bland, Brian H; Antle, Michael C

2009-01-01

The brainstem raphe nuclei are typically assigned a role in serotonergic brain function. However, numerous studies have reported that a large proportion of raphe projection cells are nonserotonergic. The identity of these projection cells is unknown. Recent studies have reported that the vesicular glutamate transporter VGLUT3 is found in both serotonergic and nonserotonergic neurons in both the median raphe (MR) and dorsal raphe (DR) nuclei. We injected the retrograde tracer cholera toxin subunit B into either the dorsal hippocampus or the medial septum (MS) and used triple labeled immunofluorescence to determine if nonserotonergic raphe cells projecting to these structures contained VGLUT3. Consistent with previous studies, only about half of retrogradely labeled MR neurons projecting to the hippocampus contained serotonin, whereas a majority of the retrogradely labeled nonserotonergic cells contained VGLUT3. Similar patterns were observed for MR cells projecting to the MS. About half of retrogradely labeled nonserotonergic neurons in the DR contained VGLUT3. Additionally, a large number of retrogradely labeled cells in the caudal linear and interpeduncular nuclei projecting to the MS were found to contain VGLUT3. These data suggest the enigmatic nonserotonergic projection from the MR to forebrain regions may be glutamatergic. In addition, these results demonstrate a dissociation between glutamatergic and serotonergic MR afferent inputs to the MS and hippocampus suggesting divergent and/or complementary roles of these pathways in modulating cellular activity within the septohippocampal network.

Compton scattering from nuclei and photo-absorption sum rules

NASA Astrophysics Data System (ADS)

Gorchtein, Mikhail; Hobbs, Timothy; Londergan, J. Timothy; Szczepaniak, Adam P.

2011-12-01

We revisit the photo-absorption sum rule for real Compton scattering from the proton and from nuclear targets. In analogy with the Thomas-Reiche-Kuhn sum rule appropriate at low energies, we propose a new “constituent quark model” sum rule that relates the integrated strength of hadronic resonances to the scattering amplitude on constituent quarks. We study the constituent quark model sum rule for several nuclear targets. In addition, we extract the α=0 pole contribution for both proton and nuclei. Using the modern high-energy proton data, we find that the α=0 pole contribution differs significantly from the Thomson term, in contrast with the original findings by Damashek and Gilman.

Light scattering microscopy measurements of single nuclei compared with GPU-accelerated FDTD simulations

NASA Astrophysics Data System (ADS)

Stark, Julian; Rothe, Thomas; Kieß, Steffen; Simon, Sven; Kienle, Alwin

2016-04-01

Single cell nuclei were investigated using two-dimensional angularly and spectrally resolved scattering microscopy. We show that even for a qualitative comparison of experimental and theoretical data, the standard Mie model of a homogeneous sphere proves to be insufficient. Hence, an accelerated finite-difference time-domain method using a graphics processor unit and domain decomposition was implemented to analyze the experimental scattering patterns. The measured cell nuclei were modeled as single spheres with randomly distributed spherical inclusions of different size and refractive index representing the nucleoli and clumps of chromatin. Taking into account the nuclear heterogeneity of a large number of inclusions yields a qualitative agreement between experimental and theoretical spectra and illustrates the impact of the nuclear micro- and nanostructure on the scattering patterns.

Light scattering microscopy measurements of single nuclei compared with GPU-accelerated FDTD simulations.

PubMed

Stark, Julian; Rothe, Thomas; Kieß, Steffen; Simon, Sven; Kienle, Alwin

2016-04-07

Single cell nuclei were investigated using two-dimensional angularly and spectrally resolved scattering microscopy. We show that even for a qualitative comparison of experimental and theoretical data, the standard Mie model of a homogeneous sphere proves to be insufficient. Hence, an accelerated finite-difference time-domain method using a graphics processor unit and domain decomposition was implemented to analyze the experimental scattering patterns. The measured cell nuclei were modeled as single spheres with randomly distributed spherical inclusions of different size and refractive index representing the nucleoli and clumps of chromatin. Taking into account the nuclear heterogeneity of a large number of inclusions yields a qualitative agreement between experimental and theoretical spectra and illustrates the impact of the nuclear micro- and nanostructure on the scattering patterns.

Activation of muscarinic receptors in rat parotid acinar cells induces AQP5 trafficking to nuclei and apical plasma membrane.

PubMed

Cho, Gota; Bragiel, Aneta M; Wang, Di; Pieczonka, Tomasz D; Skowronski, Mariusz T; Shono, Masayuki; Nielsen, Søren; Ishikawa, Yasuko

2015-04-01

The subcellular distribution of aquaporin-5 (AQP5) in rat parotid acinar cells in response to muscarinic acetylcholine receptor (mAChR) activation remains unclear. Immunoconfocal and immunoelectron microscopy were used to visualize the distribution of AQP5 in parotid acinar cells. Western blotting was used to analyze AQP5 levels in membranes. To clarify the characteristics of membrane domains associated with AQP5, detergent solubility and sucrose-density flotation experiments were performed. Under control conditions, AQP5 was diffusely distributed on the apical plasma membrane (APM) and apical plasmalemmal region and throughout the cytoplasm. Upon mAChR activation, AQP5 was predominantly located in the nucleus, APM and lateral plasma membrane (LPM). Subsequently, localization of AQP5 in the nucleus, APM and LPM was decreased. Prolonged atropine treatment inhibited mAChR agonist-induced translocation of AQP5 to the nucleus, APM and LPM. AQP5 levels were enhanced in isolated nuclei and nuclear membranes prepared from parotid tissues incubated with mAChR agonist. mAChR agonist induced AQP5 levels in both soluble and insoluble nuclear fractions solubilized with Triton X-100 or Lubrol WX. Small amounts of AQP5 in nuclei were detected using low-density sucrose gradient. When AQP5 was present in the nuclear membrane, nuclear size decreased. The activation of mAChR induced AQP5 translocation to the nucleus, APM and LPM, and AQP5 may trigger water transport across the nuclear membrane and plasma membrane in rat parotid acinar cells. AQP5 translocates to the nuclear membrane and may trigger the movement of water, inducing shrinkage of the nucleus and the start of nuclear functions. Copyright © 2015 Elsevier B.V. All rights reserved.

Determination of the accuracy for targeted irradiations of cellular substructures at SNAKE

NASA Astrophysics Data System (ADS)

Siebenwirth, C.; Greubel, C.; Drexler, S. E.; Girst, S.; Reindl, J.; Walsh, D. W. M.; Dollinger, G.; Friedl, A. A.; Schmid, T. E.; Drexler, G. A.

2015-04-01

In the last 10 years the ion microbeam SNAKE, installed at the Munich 14 MV tandem accelerator, has been successfully used for radiobiological experiments by utilizing pattern irradiation without targeting single cells. Now for targeted irradiation of cellular substructures a precise irradiation device was added to the live cell irradiation setup at SNAKE. It combines a sub-micrometer single ion irradiation facility with a high resolution optical fluorescence microscope. Most systematic errors can be reduced or avoided by using the same light path in the microscope for beam spot verification as well as for and target recognition. In addition online observation of the induced cellular responses is possible. The optical microscope and the beam delivering system are controlled by an in-house developed software which integrates the open-source image analysis software, CellProfiler, for semi-automatic target recognition. In this work the targeting accuracy was determined by irradiation of a cross pattern with 55 MeV carbon ions on nucleoli in U2OS and HeLa cells stably expressing a GFP-tagged repair protein MDC1. For target recognition, nuclei were stained with Draq5 and nucleoli were stained with Syto80 or Syto83. The damage response was determined by live-cell imaging of MDC1-GFP accumulation directly after irradiation. No systematic displacement and a random distribution of about 0.7 μm (SD) in x-direction and 0.8 μm (SD) in y-direction were observed. An independent analysis after immunofluorescence staining of the DNA damage marker yH2AX yielded similar results. With this performance a target with a size similar to that of nucleoli (i.e. a diameter of about 3 μm) is hit with a probability of more than 80%, which enables the investigation of the radiation response of cellular subcompartments after targeted ion irradiation in the future.

EGFR-targeted granzyme B expressed in NK cells enhances natural cytotoxicity and mediates specific killing of tumor cells.

PubMed

Oberoi, Pranav; Jabulowsky, Robert A; Bähr-Mahmud, Hayat; Wels, Winfried S

2013-01-01

Natural killer (NK) cells are highly specialized effectors of the innate immune system that hold promise for adoptive cancer immunotherapy. Their cell killing activity is primarily mediated by the pro-apoptotic serine protease granzyme B (GrB), which enters targets cells with the help of the pore-forming protein perforin. We investigated expression of a chimeric GrB fusion protein in NK cells as a means to augment their antitumoral activity. For selective targeting to tumor cells, we fused the epidermal growth factor receptor (EGFR) peptide ligand transforming growth factor α (TGFα) to human pre-pro-GrB. Established human NKL natural killer cells transduced with a lentiviral vector expressed this GrB-TGFα (GrB-T) molecule in amounts comparable to endogenous wildtype GrB. Activation of the genetically modified NK cells by cognate target cells resulted in the release of GrB-T together with endogenous granzymes and perforin, which augmented the effector cells' natural cytotoxicity against NK-sensitive tumor cells. Likewise, GrB-T was released into the extracellular space upon induction of degranulation with PMA and ionomycin. Secreted GrB-T fusion protein displayed specific binding to EGFR-overexpressing tumor cells, enzymatic activity, and selective target cell killing in the presence of an endosomolytic activity. Our data demonstrate that ectopic expression of a targeted GrB fusion protein in NK cells is feasible and can enhance antitumoral activity of the effector cells.

Nuclear Shell Structure and Beta Decay I. Odd A Nuclei II. Even A Nuclei

DOE R&D Accomplishments Database

Mayer, M.G.; Moszkowski, S.A.; Nordheim, L.W.

1951-05-01

In Part I a systematics is given of all transitions for odd A nuclei for which sufficiently reliable data are available. The allowed or forbidden characters of the transitions are correlated with the positions of the initial and final odd nucleon groups in the nuclear shell scheme. The nuclear shells show definite characteristics with respect to parity of the ground states. The latter is the same as the one obtained from known spins and magnetic moments in a one-particle interpretation. In Part II a systematics of the beta transitions of even-A nuclei is given. An interpretation of the character of the transitions in terms of nuclear shell structure is achieved on the hypothesis that the odd nucleon groups have the same structure as in odd-A nuclei, together with a simple coupling rule between the neutron and proton groups in odd-odd nuclei.

Reversible changes in size of cell nuclei isolated from Amoeba proteus: role of the cytoskeleton.

PubMed

Pomorski, P; Grebecka, L; Grebecki, A; Makuch, R

2000-01-01

Micrurgically isolated interphasal nuclei of Amoeba proteus, which preserve F-actin cytoskeletal shells on their surface, shrink after perfusion with imidazole buffer without ATP, and expand to about 200% of their cross-sectional area upon addition of pyrophosphate. These changes in size may be reproduced several times with the same nucleus. The shrunken nuclei are insensitive to the osmotic effects of sugars and distilled water, whereas the expanded ones react only to the distilled water, showing further swelling. The shrinking-expansion cycles are partially inhibited by cytochalasins. They are attributed to the state of actomyosin complex in the perinuclear cytoskeleton, which is supposed to be in the rigor state in the imidazole buffer without ATP, and to dissociate in the presence of pyrophosphate. Inflow of external medium to the nuclei during dissociation of the myosin from the perinuclear F-actin may be due to colloidal osmosis depending on other macromolecular components of the karyoplasm.

Multi-tissue and multi-scale approach for nuclei segmentation in H&E stained images.

PubMed

Salvi, Massimo; Molinari, Filippo

2018-06-20

Accurate nuclei detection and segmentation in histological images is essential for many clinical purposes. While manual annotations are time-consuming and operator-dependent, full automated segmentation remains a challenging task due to the high variability of cells intensity, size and morphology. Most of the proposed algorithms for the automated segmentation of nuclei were designed for specific organ or tissues. The aim of this study was to develop and validate a fully multiscale method, named MANA (Multiscale Adaptive Nuclei Analysis), for nuclei segmentation in different tissues and magnifications. MANA was tested on a dataset of H&E stained tissue images with more than 59,000 annotated nuclei, taken from six organs (colon, liver, bone, prostate, adrenal gland and thyroid) and three magnifications (10×, 20×, 40×). Automatic results were compared with manual segmentations and three open-source software designed for nuclei detection. For each organ, MANA obtained always an F1-score higher than 0.91, with an average F1 of 0.9305 ± 0.0161. The average computational time was about 20 s independently of the number of nuclei to be detected (anyway, higher than 1000), indicating the efficiency of the proposed technique. To the best of our knowledge, MANA is the first fully automated multi-scale and multi-tissue algorithm for nuclei detection. Overall, the robustness and versatility of MANA allowed to achieve, on different organs and magnifications, performances in line or better than those of state-of-art algorithms optimized for single tissues.

Adoptive therapy with CAR redirected T cells: the challenges in targeting solid tumors.

PubMed

Abken, Hinrich

2015-01-01

Recent spectacular success in the adoptive cell therapy of leukemia and lymphoma with chimeric antigen receptor (CAR)-modified T cells raised the expectations that this therapy may be efficacious in a wide range of cancer entities. The expectations are based on the predefined specificity of CAR T cells by an antibody-derived binding domain that acts independently of the natural T-cell receptor, recognizes targets independently of presentation by the major histocompatibility complex and allows targeting toward virtually any cell surface antigen. We here discuss that targeting CAR T cells toward solid tumors faces certain circumstances critical for the therapeutic success. Targeting tumor stroma and taking advantage of TRUCK cells, in other words, CAR T cells with inducible release of a transgenic payload, are some strategies envisaged to overcome current limitations in the near future.

Chaotic dynamics around cometary nuclei

NASA Astrophysics Data System (ADS)

Lages, José; Shevchenko, Ivan I.; Rollin, Guillaume

2018-06-01

We apply a generalized Kepler map theory to describe the qualitative chaotic dynamics around cometary nuclei, based on accessible observational data for five comets whose nuclei are well-documented to resemble dumb-bells. The sizes of chaotic zones around the nuclei and the Lyapunov times of the motion inside these zones are estimated. In the case of Comet 1P/Halley, the circumnuclear chaotic zone seems to engulf an essential part of the Hill sphere, at least for orbits of moderate to high eccentricity.

Adduct-specific monoclonal antibodies for the measurement of cisplatin-induced DNA lesions in individual cell nuclei

PubMed Central

Liedert, Bernd; Pluim, Dick; Schellens, Jan; Thomale, Jürgen

2006-01-01

The anticancer drug cisplatin executes its cytotoxic activity via formation of intra- and interstrand crosslinks in DNA. The relative contribution of structurally defined cisplatin adducts to induce apoptosis and the cellular processing of these lesions is still poorly understood mostly due to the lack of sensitive analytical tools for in vivo studies. Here we describe a new method to establish and characterize monoclonal antibodies (Mab) for structurally defined DNA adducts. The two major reaction products of cisplatin, the guanine–guanine (Pt-[GG]) and adenine–guanine (Pt-[AG]) intrastrand crosslinks are recognized by Mab R-C18 and R-B3, respectively. Both antibodies were employed in an immuno-cytological assay allowing the quantification of drug-induced lesions in individual cell nuclei at clinically relevant doses. Analyzing various tissues of cisplatin-treated C57Bl/6 mice the accumulation of Pt-(GG) was highest in kidney tubular cells compared with 30, 50 and 90% lower levels in kidney stroma, liver and peripheral blood cells, respectively. Adduct kinetics revealed that wild type mouse cells remove up to 80% of the crosslinks in contrast to their complete persistence in nucleotide excision repair-deficient (XPC−/−) mice. The aptitude of the immunoassay for human molecular dosimetry studies was demonstrated by measuring adduct levels in tumor biopsies from patients treated with cisplatin. PMID:16571898

Personalized targeted therapy for esophageal squamous cell carcinoma

PubMed Central

Kang, Xiaozheng; Chen, Keneng; Li, Yicheng; Li, Jianying; D'Amico, Thomas A; Chen, Xiaoxin

2015-01-01

Esophageal squamous cell carcinoma continues to heavily burden clinicians worldwide. Researchers have discovered the genomic landscape of esophageal squamous cell carcinoma, which holds promise for an era of personalized oncology care. One of the most pressing problems facing this issue is to improve the understanding of the newly available genomic data, and identify the driver-gene mutations, pathways, and networks. The emergence of a legion of novel targeted agents has generated much hope and hype regarding more potent treatment regimens, but the accuracy of drug selection is still arguable. Other problems, such as cancer heterogeneity, drug resistance, exceptional responders, and side effects, have to be surmounted. Evolving topics in personalized oncology, such as interpretation of genomics data, issues in targeted therapy, research approaches for targeted therapy, and future perspectives, will be discussed in this editorial. PMID:26167067

[Properties and localization of Mg- and Ca-ATpase activities in wheat embryo cell nuclei].

PubMed

Vasil'eva, N A; Belkina, G G; Stepanenko, S Y; Atalykova, F I; Oparin, A I

1978-05-01

The isolated nuclei of wheat embryo possess the ATPase activity. The addition of Mg2+ and Ca2+ significantly increases the activities of nuclear ATPases, whereas Hg2+, Cu2+ and Mn2+ inhibit the activity. The activating effect of Mg2+ is enhanced by an addition of Na and K ions. The activity of wheat embryo nuclear Mg-ATPase is higher than its Ca-ATPase activity; both ATPases also differ in their pH optima. Separation of total nuclear protein according to the solubility of its individual protein components in wheat and strong salt solutions, using the detergents, as well as ammonium sulfate precipitation and dialysis do not result in separation of Mg-activated and Ca-activated ATPases, although their levels of activities and ratios change in the course of fractionation. The Mg- and Ca-ATPase activities of the wheat embryo nuclei were found in the nuclear fraction of albumin, in nonhistone proteins and nuclear membranes. In the albumin nuclear fraction and subfractions of non-histone proteins the higher level of activity is observed in Ca-ATPase, whereas in the nuclei and soluble fractions of residual proteins in Mg-ATPase.

Systematic Identification of Combinatorial Drivers and Targets in Cancer Cell Lines

PubMed Central

Tabchy, Adel; Eltonsy, Nevine; Housman, David E.; Mills, Gordon B.

2013-01-01

There is an urgent need to elicit and validate highly efficacious targets for combinatorial intervention from large scale ongoing molecular characterization efforts of tumors. We established an in silico bioinformatic platform in concert with a high throughput screening platform evaluating 37 novel targeted agents in 669 extensively characterized cancer cell lines reflecting the genomic and tissue-type diversity of human cancers, to systematically identify combinatorial biomarkers of response and co-actionable targets in cancer. Genomic biomarkers discovered in a 141 cell line training set were validated in an independent 359 cell line test set. We identified co-occurring and mutually exclusive genomic events that represent potential drivers and combinatorial targets in cancer. We demonstrate multiple cooperating genomic events that predict sensitivity to drug intervention independent of tumor lineage. The coupling of scalable in silico and biologic high throughput cancer cell line platforms for the identification of co-events in cancer delivers rational combinatorial targets for synthetic lethal approaches with a high potential to pre-empt the emergence of resistance. PMID:23577104

Systematic identification of combinatorial drivers and targets in cancer cell lines.

PubMed

Tabchy, Adel; Eltonsy, Nevine; Housman, David E; Mills, Gordon B

2013-01-01

There is an urgent need to elicit and validate highly efficacious targets for combinatorial intervention from large scale ongoing molecular characterization efforts of tumors. We established an in silico bioinformatic platform in concert with a high throughput screening platform evaluating 37 novel targeted agents in 669 extensively characterized cancer cell lines reflecting the genomic and tissue-type diversity of human cancers, to systematically identify combinatorial biomarkers of response and co-actionable targets in cancer. Genomic biomarkers discovered in a 141 cell line training set were validated in an independent 359 cell line test set. We identified co-occurring and mutually exclusive genomic events that represent potential drivers and combinatorial targets in cancer. We demonstrate multiple cooperating genomic events that predict sensitivity to drug intervention independent of tumor lineage. The coupling of scalable in silico and biologic high throughput cancer cell line platforms for the identification of co-events in cancer delivers rational combinatorial targets for synthetic lethal approaches with a high potential to pre-empt the emergence of resistance.

Interactions of relativistic 36Ar and 40Ar nuclei in hydrogen: Isotopic production cross sections

NASA Astrophysics Data System (ADS)

Knott, C. N.; Albergo, S.; Caccia, Z.; Chen, C.-X.; Costa, S.; Crawford, H. J.; Cronqvist, M.; Engelage, J.; Greiner, L.; Guzik, T. G.; Insolia, A.; Lindstrom, P. J.; Mitchell, J. W.; Potenza, R.; Russo, G. V.; Soutoul, A.; Testard, O.; Tull, C. E.; Tuvé, C.; Waddington, C. J.; Webber, W. R.; Wefel, J. P.

1997-07-01

The interactions of 36Ar projectile nuclei with energies of 361, 546, and 765 MeV/nucleon and 40Ar nuclei with 352 MeV/nucleon, have been studied in a liquid-hydrogen target as part of a program to study interactions of relevance to the problem of cosmic-ray propagation in the interstellar medium. We have measured the cross sections for the production of isotopic fragments of the projectile nuclei in these interactions. The variations of these cross sections with mass, charge, and energy, are examined for insights into any systematic features of this type of fragmentation reaction that might aid predictions of other, unmeasured cross sections. These cross sections are also compared with the values derived from the most commonly used prediction techniques. It is suggested that these techniques could be improved by taking account of the systematic features identified here.

Cavitation inception from bubble nuclei

PubMed Central

Mørch, K. A.

2015-01-01

The tensile strength of ordinary water such as tap water or seawater is typically well below 1 bar. It is governed by cavitation nuclei in the water, not by the tensile strength of the water itself, which is extremely high. Different models of the nuclei have been suggested over the years, and experimental investigations of bubbles and cavitation inception have been presented. These results suggest that cavitation nuclei in equilibrium are gaseous voids in the water, stabilized by a skin which allows diffusion balance between gas inside the void and gas in solution in the surrounding liquid. The cavitation nuclei may be free gas bubbles in the bulk of water, or interfacial gaseous voids located on the surface of particles in the water, or on bounding walls. The tensile strength of these nuclei depends not only on the water quality but also on the pressure–time history of the water. A recent model and associated experiments throw new light on the effects of transient pressures on the tensile strength of water, which may be notably reduced or increased by such pressure changes. PMID:26442138

Interference of fission amplitudes of neutron resonances and T-odd asymmetry for various prescission third particles in the ternary fission of nuclei

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kadmensky, S. G., E-mail: kadmensky@phys.vsu.ru; Bunakov, V. E.; Kadmensky, S. S.

Differential cross sections for reactions of the true ternary fission of nuclei that was induced by cold polarized neutrons were constructed with allowance of the effect that Coriolis interaction and the interference between fission amplitudes of neutron resonances excited in fissile nuclei upon incidentneutron capture by target nuclei exerted on angular distributions of prescission third particles (alpha particles, neutrons, or photons). It is shown that T -odd TRI- and ROT-type asymmetries for prescission alpha particles are associated with, respectively, the odd and even components of the Coriolis interaction-perturbed amplitude of angular distributions of particles belonging to the types indicated above.more » These asymmetries have angular distributions differing from each other and stemming from a nontrivial dependence of these components on the neutron-resonance spins J{sub s} and their projections K{sub s} onto the symmetry axis of the nucleus involved. It is shown that angular distributions of prescission photons and neutrons from reactions of the ternary fission of nuclei that is induced by cold polarized neutrons are determined by the effect of Coriolis forces exclusively. Therefore, the emerging T-odd asymmetries have a character of a ROT-type asymmetry and are universal for all target nuclei.« less

Microchimeric cells in systemic lupus erythematosus: targets or innocent bystanders?

PubMed

Stevens, A M

2006-01-01

During pregnancy maternal and fetal cells commute back and forth leading to fetal microchimerism in the mother and maternal microchimerism in the child that can persist for years after the birth. Chimeric fetal and maternal cells can be hematopoietic or can differentiate into somatic cells in multiple organs, potentially acting as targets for 'autoimmunity' and so have been implicated in the pathogenesis of autoimmune diseases that resemble graft-versus-host disease after stem cell transplantation. Fetal cells have been found in women with systemic lupus erythematosus, both in the blood and a target organ, the kidney, suggesting that they may be involved in pathogenesis. Future studies will address how the host immune system normally tolerates maternal and fetal cells or how the balance may change during autoimmunity.

Free Extracellular miRNA Functionally Targets Cells by Transfecting Exosomes from Their Companion Cells.

PubMed

Bryniarski, Krzysztof; Ptak, Wlodzimierz; Martin, Emilia; Nazimek, Katarzyna; Szczepanik, Marian; Sanak, Marek; Askenase, Philip W

2015-01-01

Lymph node and spleen cells of mice doubly immunized by epicutaneous and intravenous hapten application produce a suppressive component that inhibits the action of the effector T cells that mediate contact sensitivity reactions. We recently re-investigated this phenomenon in an immunological system. CD8+ T lymphocyte-derived exosomes transferred suppressive miR-150 to the effector T cells antigen-specifically due to exosome surface coat of antibody light chains made by B1a lymphocytes. Extracellular RNA (exRNA) is protected from plasma RNases by carriage in exosomes or by chaperones. Exosome transfer of functional RNA to target cells is well described, whereas the mechanism of transfer of exRNA free of exosomes remains unclear. In the current study we describe extracellular miR-150, extracted from exosomes, yet still able to mediate antigen-specific suppression. We have determined that this was due to miR-150 association with antibody-coated exosomes produced by B1a cell companions of the effector T cells, which resulted in antigen-specific suppression of their function. Thus functional cell targeting by free exRNA can proceed by transfecting companion cell exosomes that then transfer RNA cargo to the acceptor cells. This contrasts with the classical view on release of RNA-containing exosomes from the multivesicular bodies for subsequent intercellular targeting. This new alternate pathway for transfer of exRNA between cells has distinct biological and immunological significance, and since most human blood exRNA is not in exosomes may be relevant to evaluation and treatment of diseases.

Free Extracellular miRNA Functionally Targets Cells by Transfecting Exosomes from Their Companion Cells

PubMed Central

Bryniarski, Krzysztof; Ptak, Wlodzimierz; Martin, Emilia; Nazimek, Katarzyna; Szczepanik, Marian; Sanak, Marek; Askenase, Philip W.

2015-01-01

Lymph node and spleen cells of mice doubly immunized by epicutaneous and intravenous hapten application produce a suppressive component that inhibits the action of the effector T cells that mediate contact sensitivity reactions. We recently re-investigated this phenomenon in an immunological system. CD8+ T lymphocyte-derived exosomes transferred suppressive miR-150 to the effector T cells antigen-specifically due to exosome surface coat of antibody light chains made by B1a lymphocytes. Extracellular RNA (exRNA) is protected from plasma RNases by carriage in exosomes or by chaperones. Exosome transfer of functional RNA to target cells is well described, whereas the mechanism of transfer of exRNA free of exosomes remains unclear. In the current study we describe extracellular miR-150, extracted from exosomes, yet still able to mediate antigen-specific suppression. We have determined that this was due to miR-150 association with antibody-coated exosomes produced by B1a cell companions of the effector T cells, which resulted in antigen-specific suppression of their function. Thus functional cell targeting by free exRNA can proceed by transfecting companion cell exosomes that then transfer RNA cargo to the acceptor cells. This contrasts with the classical view on release of RNA-containing exosomes from the multivesicular bodies for subsequent intercellular targeting. This new alternate pathway for transfer of exRNA between cells has distinct biological and immunological significance, and since most human blood exRNA is not in exosomes may be relevant to evaluation and treatment of diseases. PMID:25923429

Adoptive immunotherapy for B-cell malignancies with autologous chimeric antigen receptor modified tumor targeted T cells.

PubMed

Park, Jae H; Brentjens, Renier J

2010-04-01

Chemotherapy-resistant B-cell hematologic malignancies may be cured with allogeneic hematopoietic stem cell transplantation (HSCT), demonstrating the potential susceptibility of these tumors to donor T-cell mediated immune responses. However, high rates of transplant-related morbidity and mortality limit this approach. For this reason, there is an urgent need for less-toxic forms of immune-based cellular therapy to treat these malignancies. Adoptive transfer of autologous T cells genetically modified to express chimeric antigen receptors (CARs) targeted to specific tumor-associated antigens represents an attractive means of overcoming the limitations of conventional HSCT. To this end, investigators have generated CARs targeted to various antigens expressed by B-cell malignancies, optimized the design of these CARs to enhance receptor mediated T cell signaling, and demonstrated significant anti-tumor efficacy of the resulting CAR modified T cells both in vitro and in vivo mouse tumor models. These encouraging preclinical data have justified the translation of this approach to the clinical setting with currently 12 open clinical trials and one completed clinical trial treating various B-cell malignancies utilizing CAR modified T cells targeted to either the CD19 or CD20 B-cell specific antigens.

Vesicle-associated membrane protein 7 (VAMP-7) is essential for target cell killing in a natural killer cell line

DOE Office of Scientific and Technical Information (OSTI.GOV)

Marcet-Palacios, Marcelo; Odemuyiwa, Solomon O.; Coughlin, Jason J.

2008-02-15

Natural killer cells recognize and induce apoptosis in foreign, transformed or virus-infected cells through the release of perforin and granzymes from secretory lysosomes. Clinically, NK-cell mediated killing is a major limitation to successful allo- and xenotransplantation. The molecular mechanisms that regulate the fusion of granzyme B-containing secretory lysosomes to the plasma membrane in activated NK cells, prior to target cell killing, are not fully understood. Using the NK cell line YT-Indy as a model, we have investigated the expression of SNAP REceptors (SNAREs), both target (t-) and vesicular (v-) SNAREs, and their function in granzyme B-mediated target cell killing. Ourmore » data showed that YT-Indy cells express VAMP-7 and SNAP-23, but not VAMP-2. VAMP-7 was associated with granzyme B-containing lysosomal granules. Using VAMP-7 small interfering RNA (siRNA), we successfully knocked down the expression of VAMP-7 protein in YT-Indy to less than 10% of untreated cells in 24 h. VAMP7-deficient YT-Indy cells activated via co-culture with Jurkat cells released <1 ng/mL of granzyme B, compared to 1.5-2.5 {mu}g/mL from controls. Using Jurkat cells as targets, we showed a 7-fold reduction in NK cell-mediated killing by VAMP-7 deficient YT-Indy cells. Our results show that VAMP-7 is a crucial component of granzyme B release and target cell killing in the NK cell line YT-Indy. Thus, targeting VAMP-7 expression specifically with siRNA, following transplantation, may be a viable strategy for preventing NK cell-mediated transplant rejection, in vivo.« less

Tumor-targeting delivery of herb-based drugs with cell-penetrating/tumor-targeting peptide-modified nanocarriers

PubMed Central

Kebebe, Dereje; Liu, Yuanyuan; Wu, Yumei; Vilakhamxay, Maikhone; Liu, Zhidong; Li, Jiawei

2018-01-01

Cancer has become one of the leading causes of mortality globally. The major challenges of conventional cancer therapy are the failure of most chemotherapeutic agents to accumulate selectively in tumor cells and their severe systemic side effects. In the past three decades, a number of drug delivery approaches have been discovered to overwhelm the obstacles. Among these, nanocarriers have gained much attention for their excellent and efficient drug delivery systems to improve specific tissue/organ/cell targeting. In order to enhance targeting efficiency further and reduce limitations of nanocarriers, nanoparticle surfaces are functionalized with different ligands. Several kinds of ligand-modified nanomedicines have been reported. Cell-penetrating peptides (CPPs) are promising ligands, attracting the attention of researchers due to their efficiency to transport bioactive molecules intracellularly. However, their lack of specificity and in vivo degradation led to the development of newer types of CPP. Currently, activable CPP and tumor-targeting peptide (TTP)-modified nanocarriers have shown dramatically superior cellular specific uptake, cytotoxicity, and tumor growth inhibition. In this review, we discuss recent advances in tumor-targeting strategies using CPPs and their limitations in tumor delivery systems. Special emphasis is given to activable CPPs and TTPs. Finally, we address the application of CPPs and/or TTPs in the delivery of plant-derived chemotherapeutic agents. PMID:29563797

Deep magnetic capture of magnetically loaded cells for spatially targeted therapeutics.

PubMed

Huang, Zheyong; Pei, Ning; Wang, Yanyan; Xie, Xinxing; Sun, Aijun; Shen, Li; Zhang, Shuning; Liu, Xuebo; Zou, Yunzeng; Qian, Juying; Ge, Junbo

2010-03-01

Magnetic targeting has recently demonstrated potential in promoting magnetically loaded cell delivery to target lesion, but its application is limited by magnetic attenuation. For deep magnetic capture of cells for spatial targeting therapeutics, we designed a magnetic pole, in which the magnetic field density can be focused at a distance from the pole. As flowing through a tube served as a model of blood vessels, the magnetically loaded mesenchymal stem cells (MagMSCs) were highly enriched at the site distance from the magnetic pole. The cell capture efficiency was positively influenced by the magnetic flux density, and inversely influenced by the flow velocity, and well-fitted with the deductive value by theoretical considerations. It appeared to us that the spatially-focused property of the magnetic apparatus promises a new deep targeting strategy to promote homing and engraftment for cellular therapy. Copyright (c) 2009 Elsevier Ltd. All rights reserved.

Black-sphere approximation to nuclei and its application to reactions with neutron-rich nuclei

NASA Astrophysics Data System (ADS)

Kohama, Akihisa; Iida, Kei; Oyamatsu, Kazuhiro

2013-09-01

We briefly review our formula for a proton-nucleus total reaction cross section, σR, constructed in the black-sphere approximation of nuclei, in which a nucleus is viewed as a "black" sphere of radius "a". An extension to reactions involving neutron-rich nuclei is also reported.

Controversies in cancer stem cells: targeting embryonic signaling pathways.

PubMed

Takebe, Naoko; Ivy, S Percy

2010-06-15

Selectively targeting cancer stem cells (CSC) or tumor-initiating cells (TIC; from this point onward referred to as CSCs) with novel agents is a rapidly emerging field of oncology. Our knowledge of CSCs and their niche microenvironments remains a nascent field. CSC's critical dependence upon self-renewal makes these regulatory signaling pathways ripe for the development of experimental therapeutic agents. Investigational agents targeting the Notch, Hedgehog, and Wnt pathways are currently in late preclinical development stages, with some early phase 1-2 testing in human subjects. This series of articles will provide an overview and summary of the current state of knowledge of CSCs, their interactive microenvironment, and how they may serve as important targets for antitumor therapies. We also examine the scope and stage of development of early experimental agents that specifically target these highly conserved embryonic signaling pathways. (c) 2010 AACR.

MPN estimation of qPCR target sequence recoveries from whole cell calibrator samples.

PubMed

Sivaganesan, Mano; Siefring, Shawn; Varma, Manju; Haugland, Richard A

2011-12-01

DNA extracts from enumerated target organism cells (calibrator samples) have been used for estimating Enterococcus cell equivalent densities in surface waters by a comparative cycle threshold (Ct) qPCR analysis method. To compare surface water Enterococcus density estimates from different studies by this approach, either a consistent source of calibrator cells must be used or the estimates must account for any differences in target sequence recoveries from different sources of calibrator cells. In this report we describe two methods for estimating target sequence recoveries from whole cell calibrator samples based on qPCR analyses of their serially diluted DNA extracts and most probable number (MPN) calculation. The first method employed a traditional MPN calculation approach. The second method employed a Bayesian hierarchical statistical modeling approach and a Monte Carlo Markov Chain (MCMC) simulation method to account for the uncertainty in these estimates associated with different individual samples of the cell preparations, different dilutions of the DNA extracts and different qPCR analytical runs. The two methods were applied to estimate mean target sequence recoveries per cell from two different lots of a commercially available source of enumerated Enterococcus cell preparations. The mean target sequence recovery estimates (and standard errors) per cell from Lot A and B cell preparations by the Bayesian method were 22.73 (3.4) and 11.76 (2.4), respectively, when the data were adjusted for potential false positive results. Means were similar for the traditional MPN approach which cannot comparably assess uncertainty in the estimates. Cell numbers and estimates of recoverable target sequences in calibrator samples prepared from the two cell sources were also used to estimate cell equivalent and target sequence quantities recovered from surface water samples in a comparative Ct method. Our results illustrate the utility of the Bayesian method in accounting for

A Filtration-based Method of Preparing High-quality Nuclei from Cross-linked Skeletal Muscle for Chromatin Immunoprecipitation.

PubMed

Nohara, Kazunari; Chen, Zheng; Yoo, Seung-Hee

2017-07-06

Chromatin immunoprecipitation (ChIP) is a powerful method to determine protein binding to chromatin DNA. Fiber-rich skeletal muscle, however, has been a challenge for ChIP due to technical difficulty in isolation of high-quality nuclei with minimal contamination of myofibrils. Previous protocols have attempted to purify nuclei before cross-linking, which incurs the risk of altered DNA-protein interaction during the prolonged nuclei preparation process. In the current protocol, we first cross-linked the skeletal muscle tissue collected from mice, and the tissues were minced and sonicated. Since we found that ultracentrifugation was not able to separate nuclei from myofibrils using cross-linked muscle tissue, we devised a sequential filtration procedure to obtain high-quality nuclei devoid of significant myofibril contamination. We subsequently prepared chromatin by using an ultrasonicator, and ChIP assays with anti-BMAL1 antibody revealed robust circadian binding pattern of BMAL1 to target gene promoters. This filtration protocol constitutes an easily applicable method to isolate high-quality nuclei from cross-linked skeletal muscle tissue, allowing consistent sample processing for circadian and other time-sensitive studies. In combination with next-generation sequencing (NGS), our method can be deployed for various mechanistic and genomic studies focusing on skeletal muscle function.

Basic Proteins of Plant Nuclei during Normal and Pathological Cell Growth

PubMed Central

Rasch, Ellen; Woodard, John W.

1959-01-01

Histone proteins were studied by microphotometry of plant tissue sections stained with fast green at pH 8.1. For comparative purposes the Feulgen reaction was used for deoxyribose nuclei acid (DNA); the Sakaguchi reaction for arginine; and the Millon reaction for estimates of total protein. Analysis of Tradescantia tissues indicated that amounts of nuclear histone fell into approximate multiples of the gametic (egg or sperm) quantity except in dividing tissues, where amounts intermediate between multiples were found. In differentiated tissues of lily, corn, onion, and broad bean, histones occurred in constant amounts per nucleus, characteristic of the species, as was found also for DNA. Unlike the condition in several animal species, the basic proteins of sperm nuclei in these higher plants were of the histone type; no evidence of protamine was found. In a plant neoplasm, crown gall of broad bean, behavior of the basic nuclear proteins closely paralleled that of DNA. Thus, alterations of DNA levels in tumor tissues were accompanied by quantitatively similar changes in histone levels to maintain the same Feulgen/fast green ratios found in homologous normal tissues. PMID:14436319

Liver cell-targeted delivery of therapeutic molecules.

PubMed

Kang, Jeong-Hun; Toita, Riki; Murata, Masaharu

2016-01-01

The liver is the largest internal organ in mammals and is involved in metabolism, detoxification, synthesis of proteins and lipids, secretion of cytokines and growth factors and immune/inflammatory responses. Hepatitis, alcoholic or non-alcoholic liver disease, hepatocellular carcinoma, hepatic veno-occlusive disease, and liver fibrosis and cirrhosis are the most common liver diseases. Safe and efficient delivery of therapeutic molecules (drugs, genes or proteins) into the liver is very important to increase the clinical efficacy of these molecules and to reduce their side effects in other organs. Several liver cell-targeted delivery systems have been developed and tested in vivo or ex vivo/in vitro. In this review, we discuss the literature concerning liver cell-targeted delivery systems, with a particular emphasis on the results of in vivo studies.

Metabolic and structural integrity of magnetic nanoparticle-loaded primary endothelial cells for targeted cell therapy.

PubMed

Orynbayeva, Zulfiya; Sensenig, Richard; Polyak, Boris

2015-05-01

To successfully translate magnetically mediated cell targeting from bench to bedside, there is a need to systematically assess the potential adverse effects of magnetic nanoparticles (MNPs) interacting with 'therapeutic' cells. Here, we examined in detail the effects of internalized polymeric MNPs on primary rat endothelial cells' structural intactness, metabolic integrity and proliferation potential. The intactness of cytoskeleton and organelles was studied by fluorescent confocal microscopy, flow cytometry and high-resolution respirometry. MNP-loaded primary endothelial cells preserve intact cytoskeleton and organelles, maintain normal rate of proliferation, calcium signaling and mitochondria energy metabolism. This study provides supportive evidence that MNPs at doses necessary for targeting did not induce significant adverse effects on structural integrity and functionality of primary endothelial cells - potential cell therapy vectors.

Precision measurement of the mass difference between light nuclei and anti-nuclei

NASA Astrophysics Data System (ADS)

Alice Collaboration; Adam, J.; Adamová, D.; Aggarwal, M. M.; Aglieri Rinella, G.; Agnello, M.; Agrawal, N.; Ahammed, Z.; Ahmed, I.; Ahn, S. U.; Aimo, I.; Aiola, S.; Ajaz, M.; Akindinov, A.; Alam, S. N.; Aleksandrov, D.; Alessandro, B.; Alexandre, D.; Alfaro Molina, R.; Alici, A.; Alkin, A.; Alme, J.; Alt, T.; Altinpinar, S.; Altsybeev, I.; Alves Garcia Prado, C.; Andrei, C.; Andronic, A.; Anguelov, V.; Anielski, J.; Antičić, T.; Antinori, F.; Antonioli, P.; Aphecetche, L.; Appelshäuser, H.; Arcelli, S.; Armesto, N.; Arnaldi, R.; Aronsson, T.; Arsene, I. C.; Arslandok, M.; Augustinus, A.; Averbeck, R.; Azmi, M. D.; Bach, M.; Badalà, A.; Baek, Y. W.; Bagnasco, S.; Bailhache, R.; Bala, R.; Baldisseri, A.; Ball, M.; Baltasar Dos Santos Pedrosa, F.; Baral, R. C.; Barbano, A. M.; Barbera, R.; Barile, F.; Barnaföldi, G. G.; Barnby, L. S.; Barret, V.; Bartalini, P.; Bartke, J.; Bartsch, E.; Basile, M.; Bastid, N.; Basu, S.; Bathen, B.; Batigne, G.; Batista Camejo, A.; Batyunya, B.; Batzing, P. C.; Bearden, I. G.; Beck, H.; Bedda, C.; Behera, N. K.; Belikov, I.; Bellini, F.; Bello Martinez, H.; Bellwied, R.; Belmont, R.; Belmont-Moreno, E.; Belyaev, V.; Bencedi, G.; Beole, S.; Berceanu, I.; Bercuci, A.; Berdnikov, Y.; Berenyi, D.; Bertens, R. A.; Berzano, D.; Betev, L.; Bhasin, A.; Bhat, I. R.; Bhati, A. K.; Bhattacharjee, B.; Bhom, J.; Bianchi, L.; Bianchi, N.; Bianchin, C.; Bielčík, J.; Bielčíková, J.; Bilandzic, A.; Biswas, S.; Bjelogrlic, S.; Blanco, F.; Blau, D.; Blume, C.; Bock, F.; Bogdanov, A.; Bøggild, H.; Boldizsár, L.; Bombara, M.; Book, J.; Borel, H.; Borissov, A.; Borri, M.; Bossú, F.; Botje, M.; Botta, E.; Böttger, S.; Braun-Munzinger, P.; Bregant, M.; Breitner, T.; Broker, T. A.; Browning, T. A.; Broz, M.; Brucken, E. J.; Bruna, E.; Bruno, G. E.; Budnikov, D.; Buesching, H.; Bufalino, S.; Buncic, P.; Busch, O.; Buthelezi, Z.; Buxton, J. T.; Caffarri, D.; Cai, X.; Caines, H.; Calero Diaz, L.; Caliva, A.; Calvo Villar, E.; Camerini, P.; Carena, F.; Carena, W.; Castillo Castellanos, J.; Castro, A. J.; Casula, E. A. R.; Cavicchioli, C.; Ceballos Sanchez, C.; Cepila, J.; Cerello, P.; Chang, B.; Chapeland, S.; Chartier, M.; Charvet, J. L.; Chattopadhyay, Subhasis; Chattopadhyay, Sukalyan; Chelnokov, V.; Cherney, M.; Cheshkov, C.; Cheynis, B.; Chibante Barroso, V.; Chinellato, D. D.; Chochula, P.; Choi, K.; Chojnacki, M.; Choudhury, S.; Christakoglou, P.; Christensen, C. H.; Christiansen, P.; Chujo, T.; Chung, S. U.; Cicalo, C.; Cifarelli, L.; Cindolo, F.; Cleymans, J.; Colamaria, F.; Colella, D.; Collu, A.; Colocci, M.; Conesa Balbastre, G.; Conesa Del Valle, Z.; Connors, M. E.; Contreras, J. G.; Cormier, T. M.; Corrales Morales, Y.; Cortés Maldonado, I.; Cortese, P.; Cosentino, M. R.; Costa, F.; Crochet, P.; Cruz Albino, R.; Cuautle, E.; Cunqueiro, L.; Dahms, T.; Dainese, A.; Danu, A.; Das, D.; Das, I.; Das, S.; Dash, A.; Dash, S.; de, S.; de Caro, A.; de Cataldo, G.; de Cuveland, J.; de Falco, A.; de Gruttola, D.; De Marco, N.; de Pasquale, S.; Deisting, A.; Deloff, A.; Dénes, E.; D'Erasmo, G.; di Bari, D.; di Mauro, A.; di Nezza, P.; Diaz Corchero, M. A.; Dietel, T.; Dillenseger, P.; Divià, R.; Djuvsland, Ø.; Dobrin, A.; Dobrowolski, T.; Domenicis Gimenez, D.; Dönigus, B.; Dordic, O.; Dubey, A. K.; Dubla, A.; Ducroux, L.; Dupieux, P.; Ehlers, R. J.; Elia, D.; Engel, H.; Erazmus, B.; Erhardt, F.; Eschweiler, D.; Espagnon, B.; Estienne, M.; Esumi, S.; Evans, D.; Evdokimov, S.; Eyyubova, G.; Fabbietti, L.; Fabris, D.; Faivre, J.; Fantoni, A.; Fasel, M.; Feldkamp, L.; Felea, D.; Feliciello, A.; Feofilov, G.; Ferencei, J.; Fernández Téllez, A.; Ferreiro, E. G.; Ferretti, A.; Festanti, A.; Figiel, J.; Figueredo, M. A. S.; Filchagin, S.; Finogeev, D.; Fionda, F. M.; Fiore, E. M.; Fleck, M. G.; Floris, M.; Foertsch, S.; Foka, P.; Fokin, S.; Fragiacomo, E.; Francescon, A.; Frankenfeld, U.; Fuchs, U.; Furget, C.; Furs, A.; Fusco Girard, M.; Gaardhøje, J. J.; Gagliardi, M.; Gago, A. M.; Gallio, M.; Gangadharan, D. R.; Ganoti, P.; Gao, C.; Garabatos, C.; Garcia-Solis, E.; Gargiulo, C.; Gasik, P.; Germain, M.; Gheata, A.; Gheata, M.; Ghosh, P.; Ghosh, S. K.; Gianotti, P.; Giubellino, P.; Giubilato, P.; Gladysz-Dziadus, E.; Glässel, P.; Goméz Coral, D. M.; Gomez Ramirez, A.; González-Zamora, P.; Gorbunov, S.; Görlich, L.; Gotovac, S.; Grabski, V.; Graczykowski, L. K.; Grelli, A.; Grigoras, A.; Grigoras, C.; Grigoriev, V.; Grigoryan, A.; Grigoryan, S.; Grinyov, B.; Grion, N.; Grosse-Oetringhaus, J. F.; Grossiord, J.-Y.; Grosso, R.; Guber, F.; Guernane, R.; Guerzoni, B.; Gulbrandsen, K.; Gulkanyan, H.; Gunji, T.; Gupta, A.; Gupta, R.; Haake, R.; Haaland, Ø.; Hadjidakis, C.; Haiduc, M.; Hamagaki, H.; Hamar, G.; Hanratty, L. D.; Hansen, A.; Harris, J. W.; Hartmann, H.; Harton, A.; Hatzifotiadou, D.; Hayashi, S.; Heckel, S. T.; Heide, M.; Helstrup, H.; Herghelegiu, A.; Herrera Corral, G.; Hess, B. A.; Hetland, K. F.; Hilden, T. E.; Hillemanns, H.; Hippolyte, B.; Hristov, P.; Huang, M.; Humanic, T. J.; Hussain, N.; Hussain, T.; Hutter, D.; Hwang, D. S.; Ilkaev, R.; Ilkiv, I.; Inaba, M.; Ionita, C.; Ippolitov, M.; Irfan, M.; Ivanov, M.; Ivanov, V.; Izucheev, V.; Jacobs, P. M.; Jahnke, C.; Jang, H. J.; Janik, M. A.; Jayarathna, P. H. S. Y.; Jena, C.; Jena, S.; Jimenez Bustamante, R. T.; Jones, P. G.; Jung, H.; Jusko, A.; Kalinak, P.; Kalweit, A.; Kamin, J.; Kang, J. H.; Kaplin, V.; Kar, S.; Karasu Uysal, A.; Karavichev, O.; Karavicheva, T.; Karpechev, E.; Kebschull, U.; Keidel, R.; Keijdener, D. L. D.; Keil, M.; Khan, K. H.; Khan, M. Mohisin; Khan, P.; Khan, S. A.; Khanzadeev, A.; Kharlov, Y.; Kileng, B.; Kim, B.; Kim, D. W.; Kim, D. J.; Kim, H.; Kim, J. S.; Kim, Mimae.; Kim, Minwoo; Kim, S.; Kim, T.; Kirsch, S.; Kisel, I.; Kiselev, S.; Kisiel, A.; Kiss, G.; Klay, J. L.; Klein, C.; Klein, J.; Klein-Bösing, C.; Kluge, A.; Knichel, M. L.; Knospe, A. G.; Kobayashi, T.; Kobdaj, C.; Kofarago, M.; Köhler, M. K.; Kollegger, T.; Kolojvari, A.; Kondratiev, V.; Kondratyeva, N.; Kondratyuk, E.; Konevskikh, A.; Kour, M.; Kouzinopoulos, C.; Kovalenko, V.; Kowalski, M.; Kox, S.; Koyithatta Meethaleveedu, G.; Kral, J.; Králik, I.; Kravčáková, A.; Krelina, M.; Kretz, M.; Krivda, M.; Krizek, F.; Kryshen, E.; Krzewicki, M.; Kubera, A. M.; Kučera, V.; Kucheriaev, Y.; Kugathasan, T.; Kuhn, C.; Kuijer, P. G.; Kulakov, I.; Kumar, A.; Kumar, J.; Kumar, L.; Kurashvili, P.; Kurepin, A.; Kurepin, A. B.; Kuryakin, A.; Kushpil, S.; Kweon, M. J.; Kwon, Y.; La Pointe, S. L.; La Rocca, P.; Lagana Fernandes, C.; Lakomov, I.; Langoy, R.; Lara, C.; Lardeux, A.; Lattuca, A.; Laudi, E.; Lea, R.; Leardini, L.; Lee, G. R.; Lee, S.; Legrand, I.; Lehnert, J.; Lemmon, R. C.; Lenti, V.; Leogrande, E.; León Monzón, I.; Leoncino, M.; Lévai, P.; Li, S.; Li, X.; Lien, J.; Lietava, R.; Lindal, S.; Lindenstruth, V.; Lippmann, C.; Lisa, M. A.; Ljunggren, H. M.; Lodato, D. F.; Loenne, P. I.; Loggins, V. R.; Loginov, V.; Loizides, C.; Lopez, X.; López Torres, E.; Lowe, A.; Lu, X.-G.; Luettig, P.; Lunardon, M.; Luparello, G.; Maevskaya, A.; Mager, M.; Mahajan, S.; Mahmood, S. M.; Maire, A.; Majka, R. D.; Malaev, M.; Maldonado Cervantes, I.; Malinina, L.; Mal'Kevich, D.; Malzacher, P.; Mamonov, A.; Manceau, L.; Manko, V.; Manso, F.; Manzari, V.; Marchisone, M.; Mareš, J.; Margagliotti, G. V.; Margotti, A.; Margutti, J.; Marín, A.; Markert, C.; Marquard, M.; Martashvili, I.; Martin, N. A.; Martin Blanco, J.; Martinengo, P.; Martínez, M. I.; Martínez García, G.; Martinez Pedreira, M.; Martynov, Y.; Mas, A.; Masciocchi, S.; Masera, M.; Masoni, A.; Massacrier, L.; Mastroserio, A.; Matyja, A.; Mayer, C.; Mazer, J.; Mazzoni, M. A.; McDonald, D.; Meddi, F.; Menchaca-Rocha, A.; Meninno, E.; Mercado Pérez, J.; Meres, M.; Miake, Y.; Mieskolainen, M. M.; Mikhaylov, K.; Milano, L.; Milosevic, J.; Minervini, L. M.; Mischke, A.; Mishra, A. N.; Miśkowiec, D.; Mitra, J.; Mitu, C. M.; Mohammadi, N.; Mohanty, B.; Molnar, L.; Montaño Zetina, L.; Montes, E.; Morando, M.; Moreira de Godoy, D. A.; Moreno, L. A. P.; Moretto, S.; Morreale, A.; Morsch, A.; Muccifora, V.; Mudnic, E.; Mühlheim, D.; Muhuri, S.; Mukherjee, M.; Müller, H.; Mulligan, J. D.; Munhoz, M. G.; Murray, S.; Musa, L.; Musinsky, J.; Nandi, B. K.; Nania, R.; Nappi, E.; Naru, M. U.; Nattrass, C.; Nayak, K.; Nayak, T. K.; Nazarenko, S.; Nedosekin, A.; Nellen, L.; Ng, F.; Nicassio, M.; Niculescu, M.; Niedziela, J.; Nielsen, B. S.; Nikolaev, S.; Nikulin, S.; Nikulin, V.; Noferini, F.; Nomokonov, P.; Nooren, G.; Norman, J.; Nyanin, A.; Nystrand, J.; Oeschler, H.; Oh, S.; Oh, S. K.; Ohlson, A.; Okatan, A.; Okubo, T.; Olah, L.; Oleniacz, J.; Oliveira da Silva, A. C.; Oliver, M. H.; Onderwaater, J.; Oppedisano, C.; Ortiz Velasquez, A.; Oskarsson, A.; Otwinowski, J.; Oyama, K.; Ozdemir, M.; Pachmayer, Y.; Pagano, P.; Paić, G.; Pajares, C.; Pal, S. K.; Pan, J.; Pandey, A. K.; Pant, D.; Papikyan, V.; Pappalardo, G. S.; Pareek, P.; Park, W. J.; Parmar, S.; Passfeld, A.; Paticchio, V.; Paul, B.; Pawlak, T.; Peitzmann, T.; Pereira da Costa, H.; Pereira de Oliveira Filho, E.; Peresunko, D.; Pérez Lara, C. E.; Peskov, V.; Pestov, Y.; Petráček, V.; Petrov, V.; Petrovici, M.; Petta, C.; Piano, S.; Pikna, M.; Pillot, P.; Pinazza, O.; Pinsky, L.; Piyarathna, D. B.; Płoskoń, M.; Planinic, M.; Pluta, J.; Pochybova, S.; Podesta-Lerma, P. L. M.; Poghosyan, M. G.; Polichtchouk, B.; Poljak, N.; Poonsawat, W.; Pop, A.; Porteboeuf-Houssais, S.; Porter, J.; Pospisil, J.; Prasad, S. K.; Preghenella, R.; Prino, F.; Pruneau, C. A.; Pshenichnov, I.; Puccio, M.; Puddu, G.; Pujahari, P.; Punin, V.; Putschke, J.; Qvigstad, H.; Rachevski, A.; Raha, S.; Rajput, S.; Rak, J.; Rakotozafindrabe, A.; Ramello, L.; Raniwala, R.; Raniwala, S.; Räsänen, S. S.; Rascanu, B. T.; Rathee, D.; Razazi, V.; Read, K. F.; Real, J. S.; Redlich, K.; Reed, R. J.; Rehman, A.; Reichelt, P.; Reicher, M.; Reidt, F.; Ren, X.; Renfordt, R.; Reolon, A. R.; Reshetin, A.; Rettig, F.; Revol, J.-P.; Reygers, K.; Riabov, V.; Ricci, R. A.; Richert, T.; Richter, M.; Riedler, P.; Riegler, W.; Riggi, F.; Ristea, C.; Rivetti, A.; Rocco, E.; Rodríguez Cahuantzi, M.; Rodriguez Manso, A.; Røed, K.; Rogochaya, E.; Rohr, D.; Röhrich, D.; Romita, R.; Ronchetti, F.; Ronflette, L.; Rosnet, P.; Rossi, A.; Roukoutakis, F.; Roy, A.; Roy, C.; Roy, P.; Rubio Montero, A. J.; Rui, R.; Russo, R.; Ryabinkin, E.; Ryabov, Y.; Rybicki, A.; Sadovsky, S.; Šafařík, K.; Sahlmuller, B.; Sahoo, P.; Sahoo, R.; Sahoo, S.; Sahu, P. K.; Saini, J.; Sakai, S.; Saleh, M. A.; Salgado, C. A.; Salzwedel, J.; Sambyal, S.; Samsonov, V.; Sanchez Castro, X.; Šándor, L.; Sandoval, A.; Sano, M.; Santagati, G.; Sarkar, D.; Scapparone, E.; Scarlassara, F.; Scharenberg, R. P.; Schiaua, C.; Schicker, R.; Schmidt, C.; Schmidt, H. R.; Schuchmann, S.; Schukraft, J.; Schulc, M.; Schuster, T.; Schutz, Y.; Schwarz, K.; Schweda, K.; Scioli, G.; Scomparin, E.; Scott, R.; Seeder, K. S.; Seger, J. E.; Sekiguchi, Y.; Selyuzhenkov, I.; Senosi, K.; Seo, J.; Serradilla, E.; Sevcenco, A.; Shabanov, A.; Shabetai, A.; Shadura, O.; Shahoyan, R.; Shangaraev, A.; Sharma, A.; Sharma, M.; Sharma, N.; Shigaki, K.; Shtejer, K.; Sibiriak, Y.; Siddhanta, S.; Sielewicz, K. M.; Siemiarczuk, T.; Silvermyr, D.; Silvestre, C.; Simatovic, G.; Simonetti, G.; Singaraju, R.; Singh, R.; Singha, S.; Singhal, V.; Sinha, B. C.; Sinha, T.; Sitar, B.; Sitta, M.; Skaali, T. B.; Slupecki, M.; Smirnov, N.; Snellings, R. J. M.; Snellman, T. W.; Søgaard, C.; Soltz, R.; Song, J.; Song, M.; Song, Z.; Soramel, F.; Sorensen, S.; Spacek, M.; Spiriti, E.; Sputowska, I.; Spyropoulou-Stassinaki, M.; Srivastava, B. K.; Stachel, J.; Stan, I.; Stefanek, G.; Steinpreis, M.; Stenlund, E.; Steyn, G.; Stiller, J. H.; Stocco, D.; Strmen, P.; Suaide, A. A. P.; Sugitate, T.; Suire, C.; Suleymanov, M.; Sultanov, R.; Šumbera, M.; Symons, T. J. M.; Szabo, A.; Szanto de Toledo, A.; Szarka, I.; Szczepankiewicz, A.; Szymanski, M.; Takahashi, J.; Tanaka, N.; Tangaro, M. A.; Tapia Takaki, J. D.; Tarantola Peloni, A.; Tariq, M.; Tarzila, M. G.; Tauro, A.; Tejeda Muñoz, G.; Telesca, A.; Terasaki, K.; Terrevoli, C.; Teyssier, B.; Thäder, J.; Thomas, D.; Tieulent, R.; Timmins, A. R.; Toia, A.; Trogolo, S.; Trubnikov, V.; Trzaska, W. H.; Tsuji, T.; Tumkin, A.; Turrisi, R.; Tveter, T. S.; Ullaland, K.; Uras, A.; Usai, G. L.; Utrobicic, A.; Vajzer, M.; Vala, M.; Valencia Palomo, L.; Vallero, S.; van der Maarel, J.; van Hoorne, J. W.; van Leeuwen, M.; Vanat, T.; Vande Vyvre, P.; Varga, D.; Vargas, A.; Vargyas, M.; Varma, R.; Vasileiou, M.; Vasiliev, A.; Vauthier, A.; Vechernin, V.; Veen, A. M.; Veldhoen, M.; Velure, A.; Venaruzzo, M.; Vercellin, E.; Vergara Limón, S.; Vernet, R.; Verweij, M.; Vickovic, L.; Viesti, G.; Viinikainen, J.; Vilakazi, Z.; Villalobos Baillie, O.; Villatoro Tello, A.; Vinogradov, A.; Vinogradov, L.; Vinogradov, Y.; Virgili, T.; Vislavicius, V.; Viyogi, Y. P.; Vodopyanov, A.; Völkl, M. A.; Voloshin, K.; Voloshin, S. A.; Volpe, G.; von Haller, B.; Vorobyev, I.; Vranic, D.; Vrláková, J.; Vulpescu, B.; Vyushin, A.; Wagner, B.; Wagner, J.; Wang, H.; Wang, M.; Wang, Y.; Watanabe, D.; Weber, M.; Weber, S. G.; Wessels, J. P.; Westerhoff, U.; Wiechula, J.; Wikne, J.; Wilde, M.; Wilk, G.; Wilkinson, J.; Williams, M. C. S.; Windelband, B.; Winn, M.; Yaldo, C. G.; Yamaguchi, Y.; Yang, H.; Yang, P.; Yano, S.; Yasnopolskiy, S.; Yin, Z.; Yokoyama, H.; Yoo, I.-K.; Yurchenko, V.; Yushmanov, I.; Zaborowska, A.; Zaccolo, V.; Zaman, A.; Zampolli, C.; Zanoli, H. J. C.; Zaporozhets, S.; Zarochentsev, A.; Závada, P.; Zaviyalov, N.; Zbroszczyk, H.; Zgura, I. S.; Zhalov, M.; Zhang, H.; Zhang, X.; Zhang, Y.; Zhao, C.; Zhigareva, N.; Zhou, D.; Zhou, Y.; Zhou, Z.; Zhu, H.; Zhu, J.; Zhu, X.; Zichichi, A.; Zimmermann, A.; Zimmermann, M. B.; Zinovjev, G.; Zyzak, M.

2015-10-01

The measurement of the mass differences for systems bound by the strong force has reached a very high precision with protons and anti-protons. The extension of such measurement from (anti-)baryons to (anti-)nuclei allows one to probe any difference in the interactions between nucleons and anti-nucleons encoded in the (anti-)nuclei masses. This force is a remnant of the underlying strong interaction among quarks and gluons and can be described by effective theories, but cannot yet be directly derived from quantum chromodynamics. Here we report a measurement of the difference between the ratios of the mass and charge of deuterons (d) and anti-deuterons (), and 3He and nuclei carried out with the ALICE (A Large Ion Collider Experiment) detector in Pb-Pb collisions at a centre-of-mass energy per nucleon pair of 2.76 TeV. Our direct measurement of the mass-over-charge differences confirms CPT invariance to an unprecedented precision in the sector of light nuclei. This fundamental symmetry of nature, which exchanges particles with anti-particles, implies that all physics laws are the same under the simultaneous reversal of charge(s) (charge conjugation C), reflection of spatial coordinates (parity transformation P) and time inversion (T).

Precision measurement of the mass difference between light nuclei and anti-nuclei

DOE Office of Scientific and Technical Information (OSTI.GOV)

Adam, J.

The measurement of the mass differences for systems bound by the strong force has reached a very high precision with protons and anti-protons. The extension of such measurement from (anti-)baryons to (anti-)nuclei allows one to probe any difference in the interactions between nucleons and anti-nucleons encoded in the (anti-)nuclei masses. Also, this force is a remnant of the underlying strong interaction among quarks and gluons and can be described by effective theories, but cannot yet be directly derived from quantum chromodynamics. Here we report a measurement of the difference between the ratios of the mass and charge of deuterons (d) and anti-deuterons (more » $$-\\atop{d}$$), and 3He and 3$$-\\atop{He}$$nuclei carried out with the ALICE (A Large Ion Collider Experiment) detector in Pb–Pb collisions at a centre-of-mass energy per nucleon pair of 2.76 TeV. Our direct measurement of the mass-over-charge differences confirms CPT invariance to an unprecedented precision in the sector of light nuclei. This fundamental symmetry of nature, which exchanges particles with anti-particles, implies that all physics laws are the same under the simultaneous reversal of charge(s) (charge conjugation C), reflection of spatial coordinates (parity transformation P) and time inversion (T).« less

Precision measurement of the mass difference between light nuclei and anti-nuclei

DOE PAGES

Adam, J.

2015-08-17

The measurement of the mass differences for systems bound by the strong force has reached a very high precision with protons and anti-protons. The extension of such measurement from (anti-)baryons to (anti-)nuclei allows one to probe any difference in the interactions between nucleons and anti-nucleons encoded in the (anti-)nuclei masses. Also, this force is a remnant of the underlying strong interaction among quarks and gluons and can be described by effective theories, but cannot yet be directly derived from quantum chromodynamics. Here we report a measurement of the difference between the ratios of the mass and charge of deuterons (d) and anti-deuterons (more » $$-\\atop{d}$$), and 3He and 3$$-\\atop{He}$$nuclei carried out with the ALICE (A Large Ion Collider Experiment) detector in Pb–Pb collisions at a centre-of-mass energy per nucleon pair of 2.76 TeV. Our direct measurement of the mass-over-charge differences confirms CPT invariance to an unprecedented precision in the sector of light nuclei. This fundamental symmetry of nature, which exchanges particles with anti-particles, implies that all physics laws are the same under the simultaneous reversal of charge(s) (charge conjugation C), reflection of spatial coordinates (parity transformation P) and time inversion (T).« less

Localization Microscopy Analyses of MRE11 Clusters in 3D-Conserved Cell Nuclei of Different Cell Lines

PubMed Central

Eryilmaz, Marion; Schmitt, Eberhard; Krufczik, Matthias; Theda, Franziska; Lee, Jin-Ho; Cremer, Christoph; Bestvater, Felix; Schaufler, Wladimir; Hildenbrand, Georg

2018-01-01

In radiation biophysics, it is a subject of nowadays research to investigate DNA strand break repair in detail after damage induction by ionizing radiation. It is a subject of debate as to what makes up the cell’s decision to use a certain repair pathway and how the repair machinery recruited in repair foci is spatially and temporarily organized. Single-molecule localization microscopy (SMLM) allows super-resolution analysis by precise localization of single fluorescent molecule tags, resulting in nuclear structure analysis with a spatial resolution in the 10 nm regime. Here, we used SMLM to study MRE11 foci. MRE11 is one of three proteins involved in the MRN-complex (MRE11-RAD50-NBS1 complex), a prominent DNA strand resection and broken end bridging component involved in homologous recombination repair (HRR) and alternative non-homologous end joining (a-NHEJ). We analyzed the spatial arrangements of antibody-labelled MRE11 proteins in the nuclei of a breast cancer and a skin fibroblast cell line along a time-course of repair (up to 48 h) after irradiation with a dose of 2 Gy. Different kinetics for cluster formation and relaxation were determined. Changes in the internal nano-scaled structure of the clusters were quantified and compared between the two cell types. The results indicate a cell type-dependent DNA damage response concerning MRE11 recruitment and cluster formation. The MRE11 data were compared to H2AX phosphorylation detected by γH2AX molecule distribution. These data suggested modulations of MRE11 signal frequencies that were not directly correlated to DNA damage induction. The application of SMLM in radiation biophysics offers new possibilities to investigate spatial foci organization after DNA damaging and during subsequent repair. PMID:29361783

A 20-Amino Acid Module of Protein Kinase Cϵ Involved in Translocation and Selective Targeting at Cell-Cell Contacts*

PubMed Central

Diouf, Barthélémy; Collazos, Alejandra; Labesse, Gilles; Macari, Françoise; Choquet, Armelle; Clair, Philippe; Gauthier-Rouvière, Cécile; Guérineau, Nathalie C.; Jay, Philippe; Hollande, Frédéric; Joubert, Dominique

2009-01-01

In the pituitary gland, activated protein kinase C (PKC) isoforms accumulate either selectively at the cell-cell contact (α and ϵ) or at the entire plasma membrane (β1 and δ). The molecular mechanisms underlying these various subcellular locations are not known. Here, we demonstrate the existence within PKCϵ of a cell-cell contact targeting sequence (3CTS) that, upon stimulation, is capable of targeting PKCδ, chimerin-α1, and the PKCϵ C1 domain to the cell-cell contact. We show that this selective targeting of PKCϵ is lost upon overexpression of 3CTS fused to a (R-Ahx-R)4 (where Ahx is 6-aminohexanoic acid) vectorization peptide, reflecting a dominant-negative effect of the overexpressed 3CTS on targeting selectivity. 3CTS contains a putative amphipathic α-helix, a 14-3-3-binding site, and the Glu-374 amino acid, involved in targeting selectivity. We show that the integrity of the α-helix is important for translocation but that 14-3-3 is not involved in targeting selectivity. However, PKCϵ translocation is increased when PKCϵ/14-3-3 interaction is abolished, suggesting that phorbol 12-myristate 13-acetate activation may initiate two sets of PKCϵ functions, those depending on 14-3-3 and those depending on translocation to cell-cell contacts. Thus, 3CTS is involved in the modulation of translocation via its 14-3-3-binding site, in cytoplasmic desequestration via the α-helix, and in selective PKCϵ targeting at the cell-cell contact via Glu-374. PMID:19429675

E-selectin liposomal and nanotube-targeted delivery of doxorubicin to circulating tumor cells

PubMed Central

Mitchell, Michael J.; Chen, Christina S.; Ponmudi, Varun; Hughes, Andrew D.; King, Michael R.

2012-01-01

The presence of circulating tumor cells (CTCs) is believed to lead to the formation of secondary tumors via an adhesion cascade involving interaction between adhesion receptors of endothelial cells and ligands on CTCs. Many CTCs express sialylated carbohydrate ligands on their surfaces that adhere to selectin protein found on inflamed endothelial cells. We have investigated the feasibility of using immobilized selectin proteins as a targeting mechanism for CTCs under flow. Herein, targeted liposomal doxorubicin (L-DXR) was functionalized with recombinant human E-selectin (ES) and polyethylene glycol (PEG) to target and kill cancer cells under shear flow, both when immobilized along a microtube device or sheared in a cone-and-plate viscometer in a dilute suspension. Healthy circulating cells such as red blood cells were not targeted by this mechanism and were left to freely circulate, and minimal leukocyte death was observed. Halloysite nanotube (HNT)-coated microtube devices immobilized with nanoscale liposomes significantly enhanced the targeting, capture, and killing of cancer cells. This work demonstrates that E-selectin functionalized L-DXR, sheared in suspension or immobilized onto microtube devices, provides a novel approach to selectively target and deliver chemotherapeutics to CTCs in the bloodstream. PMID:22421423

3D segmentations of neuronal nuclei from confocal microscope image stacks

PubMed Central

LaTorre, Antonio; Alonso-Nanclares, Lidia; Muelas, Santiago; Peña, José-María; DeFelipe, Javier

2013-01-01

In this paper, we present an algorithm to create 3D segmentations of neuronal cells from stacks of previously segmented 2D images. The idea behind this proposal is to provide a general method to reconstruct 3D structures from 2D stacks, regardless of how these 2D stacks have been obtained. The algorithm not only reuses the information obtained in the 2D segmentation, but also attempts to correct some typical mistakes made by the 2D segmentation algorithms (for example, under segmentation of tightly-coupled clusters of cells). We have tested our algorithm in a real scenario—the segmentation of the neuronal nuclei in different layers of the rat cerebral cortex. Several representative images from different layers of the cerebral cortex have been considered and several 2D segmentation algorithms have been compared. Furthermore, the algorithm has also been compared with the traditional 3D Watershed algorithm and the results obtained here show better performance in terms of correctly identified neuronal nuclei. PMID:24409123

3D segmentations of neuronal nuclei from confocal microscope image stacks.

PubMed

Latorre, Antonio; Alonso-Nanclares, Lidia; Muelas, Santiago; Peña, José-María; Defelipe, Javier

2013-01-01

In this paper, we present an algorithm to create 3D segmentations of neuronal cells from stacks of previously segmented 2D images. The idea behind this proposal is to provide a general method to reconstruct 3D structures from 2D stacks, regardless of how these 2D stacks have been obtained. The algorithm not only reuses the information obtained in the 2D segmentation, but also attempts to correct some typical mistakes made by the 2D segmentation algorithms (for example, under segmentation of tightly-coupled clusters of cells). We have tested our algorithm in a real scenario-the segmentation of the neuronal nuclei in different layers of the rat cerebral cortex. Several representative images from different layers of the cerebral cortex have been considered and several 2D segmentation algorithms have been compared. Furthermore, the algorithm has also been compared with the traditional 3D Watershed algorithm and the results obtained here show better performance in terms of correctly identified neuronal nuclei.

Improved segmentation of abnormal cervical nuclei using a graph-search based approach

NASA Astrophysics Data System (ADS)

Zhang, Ling; Liu, Shaoxiong; Wang, Tianfu; Chen, Siping; Sonka, Milan

2015-03-01

Reliable segmentation of abnormal nuclei in cervical cytology is of paramount importance in automation-assisted screening techniques. This paper presents a general method for improving the segmentation of abnormal nuclei using a graph-search based approach. More specifically, the proposed method focuses on the improvement of coarse (initial) segmentation. The improvement relies on a transform that maps round-like border in the Cartesian coordinate system into lines in the polar coordinate system. The costs consisting of nucleus-specific edge and region information are assigned to the nodes. The globally optimal path in the constructed graph is then identified by dynamic programming. We have tested the proposed method on abnormal nuclei from two cervical cell image datasets, Herlev and H and E stained liquid-based cytology (HELBC), and the comparative experiments with recent state-of-the-art approaches demonstrate the superior performance of the proposed method.

Concise Review: Cell Surface N-Linked Glycoproteins as Potential Stem Cell Markers and Drug Targets.

PubMed

Boheler, Kenneth R; Gundry, Rebekah L

2017-01-01

Stem cells and their derivatives hold great promise to advance regenerative medicine. Critical to the progression of this field is the identification and utilization of antibody-accessible cell-surface proteins for immunophenotyping and cell sorting-techniques essential for assessment and isolation of defined cell populations with known functional and therapeutic properties. Beyond their utility for cell identification and selection, cell-surface proteins are also major targets for pharmacological intervention. Although comprehensive cell-surface protein maps are highly valuable, they have been difficult to define until recently. In this review, we discuss the application of a contemporary targeted chemoproteomic-based technique for defining the cell-surface proteomes of stem and progenitor cells. In applying this approach to pluripotent stem cells (PSCs), these studies have improved the biological understanding of these cells, led to the enhanced use and development of antibodies suitable for immunophenotyping and sorting, and contributed to the repurposing of existing drugs without the need for high-throughput screening. The utility of this latter approach was first demonstrated with human PSCs (hPSCs) through the identification of small molecules that are selectively toxic to hPSCs and have the potential for eliminating confounding and tumorigenic cells in hPSC-derived progeny destined for research and transplantation. Overall, the cutting-edge technologies reviewed here will accelerate the development of novel cell-surface protein targets for immunophenotyping, new reagents to improve the isolation of therapeutically qualified cells, and pharmacological studies to advance the treatment of intractable diseases amenable to cell-replacement therapies. Stem Cells Translational Medicine 2017;6:131-138. © 2016 The Authors Stem Cells Translational Medicine published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.