High-frame-rate imaging of biological samples with optoacoustic micro-tomography

NASA Astrophysics Data System (ADS)

Deán-Ben, X. Luís.; López-Schier, Hernán.; Razansky, Daniel

2018-02-01

Optical microscopy remains a major workhorse in biological discovery despite the fact that light scattering limits its applicability to depths of ˜ 1 mm in scattering tissues. Optoacoustic imaging has been shown to overcome this barrier by resolving optical absorption with microscopic resolution in significantly deeper regions. Yet, the time domain is paramount for the observation of biological dynamics in living systems that exhibit fast motion. Commonly, acquisition of microscopy data involves raster scanning across the imaged volume, which significantly limits temporal resolution in 3D. To overcome these limitations, we have devised a fast optoacoustic micro-tomography (OMT) approach based on simultaneous acquisition of 3D image data with a high-density hemispherical ultrasound array having effective detection bandwidth around 25 MHz. We performed experiments by imaging tissue-mimicking phantoms and zebrafish larvae, demonstrating that OMT can provide nearly cellular resolution and imaging speed of 100 volumetric frames per second. As opposed to other optical microscopy techniques, OMT is a hybrid method that resolves optical absorption contrast acoustically using unfocused light excitation. Thus, no penetration barriers are imposed by light scattering in deep tissues, suggesting it as a powerful approach for multi-scale functional and molecular imaging applications.

Axial tomography in live cell laser microscopy

NASA Astrophysics Data System (ADS)

Richter, Verena; Bruns, Sarah; Bruns, Thomas; Weber, Petra; Wagner, Michael; Cremer, Christoph; Schneckenburger, Herbert

2017-09-01

Single cell microscopy in a three-dimensional (3-D) environment is reported. Cells are grown in an agarose culture gel, located within microcapillaries and observed from different sides after adaptation of an innovative device for sample rotation. Thus, z-stacks can be recorded by confocal microscopy in different directions and used for illustration in 3-D. This gives additional information, since cells or organelles that appear superimposed in one direction, may be well resolved in another one. The method is tested and validated with single cells expressing a membrane or a mitochondrially associated green fluorescent protein, or cells accumulating fluorescent quantum dots. In addition, axial tomography supports measurements of cellular uptake and distribution of the anticancer drug doxorubicin in the nucleus (2 to 6 h after incubation) or the cytoplasm (24 h). This paper discusses that upon cell rotation an enhanced optical resolution in lateral direction compared to axial direction can be utilized to obtain an improved effective 3-D resolution, which represents an important step toward super-resolution microscopy of living cells.

Noninvasive metabolic imaging of engineered 3D human adipose tissue in a perfusion bioreactor.

PubMed

Ward, Andrew; Quinn, Kyle P; Bellas, Evangelia; Georgakoudi, Irene; Kaplan, David L

2013-01-01

The efficacy and economy of most in vitro human models used in research is limited by the lack of a physiologically-relevant three-dimensional perfused environment and the inability to noninvasively quantify the structural and biochemical characteristics of the tissue. The goal of this project was to develop a perfusion bioreactor system compatible with two-photon imaging to noninvasively assess tissue engineered human adipose tissue structure and function in vitro. Three-dimensional (3D) vascularized human adipose tissues were engineered in vitro, before being introduced to a perfusion environment and tracked over time by automated quantification of endogenous markers of metabolism using two-photon excited fluorescence (TPEF). Depth-resolved image stacks were analyzed for redox ratio metabolic profiling and compared to prior analyses performed on 3D engineered adipose tissue in static culture. Traditional assessments with H&E staining were used to qualitatively measure extracellular matrix generation and cell density with respect to location within the tissue. The distribution of cells within the tissue and average cellular redox ratios were different between static and perfusion cultures, while the trends of decreased redox ratio and increased cellular proliferation with time in both static and perfusion cultures were similar. These results establish a basis for noninvasive optical tracking of tissue structure and function in vitro, which can be applied to future studies to assess tissue development or drug toxicity screening and disease progression.

Micro-Mirrors for Nanoscale Three-Dimensional Microscopy

PubMed Central

Seale, Kevin; Janetopoulos, Chris; Wikswo, John

2013-01-01

A research-grade optical microscope is capable of resolving fine structures in two-dimensional images. However, three-dimensional resolution, or the ability of the microscope to distinguish between objects lying above or below the focal plane from in-focus objects, is not nearly as good as in-plane resolution. In this issue of ACS Nano, McMahon et al. report the use of mirrored pyramidal wells with a conventional microscope for rapid, 3D localization and tracking of nanoparticles. Mirrors have been used in microscopy before, but recent work with MPWs is unique because it enables the rapid determination of the x-, y-, and z-position of freely diffusing nanoparticles and cellular nanostructures with unprecedented speed and spatial accuracy. As inexpensive tools for 3D visualization, mirrored pyramidal wells may prove to be invaluable aids in nanotechnology and engineering of nanomaterials. PMID:19309167

Comparative studies of cellular viability levels on 2D and 3D in vitro culture matrices.

PubMed

Gargotti, M; Lopez-Gonzalez, U; Byrne, H J; Casey, A

2018-02-01

In this study, the cellular viability and function of immortalized human cervical and dermal cells are monitored and compared in conventional 2D and two commercial 3D membranes, Collagen and Geltrex, of varying working concentration and volume. Viability was monitored with the aid of the Alamar Blue assay, cellular morphology was monitored with confocal microscopy, and cell cycle studies and cell death mechanism studies were performed with flow cytometry. The viability studies showed apparent differences between the 2D and 3D culture systems, the differences attributed in part to the physical transition from 2D to 3D environment causing alterations to effective resazurin concentration, uptake and conversion rates, which was dependent on exposure time, but also due to the effect of the membrane itself on cellular function. These effects were verified by flow cytometry, in which no significant differences in viable cell numbers between 2D and 3D systems were observed after 24 h culture. The results showed the observed effect was different after shorter exposure periods, was also dependent on working concentration of the 3D system and could be mediated by altering the culture vessel size. Cell cycle analysis revealed cellular function could be altered by growth on the 3D substrates and the alterations were noted to be dependent on 3D membrane concentration. The use of 3D culture matrices has been widely interpreted to result in "improved viability levels" or "reduced" toxicity or cellular "resistance" compared to cells cultured on traditional 2D systems. The results of this study show that cellular health and viability levels are not altered by culture in 3D environments, but their normal cycle can be altered as indicated in the cell cycle studies performed and such variations must be accounted for in studies employing 3D membranes for in vitro cellular screening.

Aspirin-triggered resolvin D1 is produced during self-resolving gram-negative bacterial pneumonia and regulates host immune responses for the resolution of lung inflammation.

PubMed

Abdulnour, R E; Sham, H P; Douda, D N; Colas, R A; Dalli, J; Bai, Y; Ai, X; Serhan, C N; Levy, B D

2016-09-01

Bacterial pneumonia is a leading cause of morbidity and mortality worldwide. Host responses to contain infection and mitigate pathogen-mediated lung inflammation are critical for pneumonia resolution. Aspirin-triggered resolvin D1 (AT-RvD1; 7S,8R,17R-trihydroxy-4Z,9E,11E,13Z,15E,19Z-docosahexaenoic acid) is a lipid mediator (LM) that displays organ-protective actions in sterile lung inflammation, and regulates pathogen-initiated cellular responses. Here, in a self-resolving murine model of Escherichia coli pneumonia, LM metabololipidomics performed on lungs obtained at baseline, 24, and 72 h after infection uncovered temporal regulation of endogenous AT-RvD1 production. Early treatment with exogenous AT-RvD1 (1 h post infection) enhanced clearance of E. coli and Pseudomonas aeruginosa in vivo, and lung macrophage phagocytosis of fluorescent bacterial particles ex vivo. Characterization of macrophage subsets in the alveolar compartment during pneumonia identified efferocytosis by infiltrating macrophages (CD11b(Hi) CD11c(Low)) and exudative macrophages (CD11b(Hi) CD11c(Hi)). AT-RvD1 increased efferocytosis by these cells ex vivo, and accelerated neutrophil clearance during pneumonia in vivo. These anti-bacterial and pro-resolving actions of AT-RvD1 were additive to antibiotic therapy. Taken together, these findings suggest that the pro-resolving actions of AT-RvD1 during pneumonia represent a novel host-directed therapeutic strategy to complement the current antibiotic-centered approach for combatting infections.

Resolvin D3 Is Dysregulated in Arthritis and Reduces Arthritic Inflammation.

PubMed

Arnardottir, Hildur H; Dalli, Jesmond; Norling, Lucy V; Colas, Romain A; Perretti, Mauro; Serhan, Charles N

2016-09-15

Uncontrolled inflammation is a unifying component of many chronic inflammatory diseases, such as arthritis. Resolvins (Rvs) are a new family from the endogenous specialized proresolving mediators (SPMs) that actively stimulate resolution of inflammation. In this study, using lipid mediator metabololipidomics with murine joints we found a temporal regulation of endogenous SPMs during self-resolving inflammatory arthritis. The SPMs present in self-resolving arthritic joints include the D-series Rvs, for example, RvD1, RvD2, RvD3, and RvD4. Of note, RvD3 levels were reduced in inflamed joints from mice with delayed-resolving arthritis when compared with self-resolving inflammatory arthritis. RvD3 was also reduced in serum from rheumatoid arthritis patients compared with healthy controls. RvD3 administration reduced joint leukocytes as well as paw joint eicosanoids, clinical scores, and edema. Taken together, these findings provide evidence for dysregulated endogenous RvD3 levels in inflamed paw joints and its potent actions in reducing murine arthritis. Copyright © 2016 by The American Association of Immunologists, Inc.

A New Approach to Time-Resolved 3D-PTV

NASA Astrophysics Data System (ADS)

Boomsma, Aaron; Troolin, Dan; Bjorkquist, Dan; TSI Inc Team

2017-11-01

Volumetric three-component velocimetry via particle tracking is a powerful alternative to TomoPIV. It has been thoroughly documented that compared to TomoPIV, particle tracking velocimetry (PTV) methods (whether 2D or 3D) better resolve regions of high velocity gradient, identify fewer ghost particles, and are less computationally demanding, which results in shorter processing times. Recently, 3D-PTV has seen renewed interest in the PIV community with the availability of time-resolved data. Of course, advances in hardware are partly to thank for that availability-higher speed cameras, more effective memory management, and higher speed lasers. But in software, algorithms that utilize time resolved data to improve 3D particle reconstruction and particle tracking are also under development and advancing (e.g. shake-the-box, neighbor tracking reconstruction, etc.). .In the current study, we present a new 3D-PTV method that incorporates time-resolved data. We detail the method, its performance in terms of particle identification and reconstruction error and their relation to varying seeding densities, as well as computational performance.

Hard X-ray submicrometer tomography of human brain tissue at Diamond Light Source

NASA Astrophysics Data System (ADS)

Khimchenko, A.; Bikis, C.; Schulz, G.; Zdora, M.-C.; Zanette, I.; Vila-Comamala, J.; Schweighauser, G.; Hench, J.; Hieber, S. E.; Deyhle, H.; Thalmann, P.; Müller, B.

2017-06-01

There is a lack of the necessary methodology for three-dimensional (3D) investigation of soft tissues with cellular resolution without staining or tissue transformation. Synchrotron radiation based hard X-ray in-line phase contrast tomography using single-distance phase reconstruction (SDPR) provides high spatial resolution and density contrast for the visualization of individual cells using a standard specimen preparation and data reconstruction. In this study, we demonstrate the 3D characterization of a formalin-fixed paraffin-embedded (FFPE) human cerebellum specimen by SDPR at the Diamond-Manchester Imaging Branchline I13-2 (Diamond Light Source, UK) at pixel sizes down to 0.45 μm. The approach enables visualization of cerebellar layers (Stratum moleculare and Stratum granulosum), the 3D characterization of individual cells (Purkinje, stellate and granule cells) and can even resolve some subcellular structures (nucleus and nucleolus of Purkinje cells). The tomographic results are qualitatively compared to hematoxylin and eosin (H&E) stained histological sections. We demonstrate the potential benefits of hard X-ray microtomography for the investigations of biological tissues in comparison to conventional histology.

Structured illumination multimodal 3D-resolved quantitative phase and fluorescence sub-diffraction microscopy

PubMed Central

Chowdhury, Shwetadwip; Eldridge, Will J.; Wax, Adam; Izatt, Joseph A.

2017-01-01

Sub-diffraction resolution imaging has played a pivotal role in biological research by visualizing key, but previously unresolvable, sub-cellular structures. Unfortunately, applications of far-field sub-diffraction resolution are currently divided between fluorescent and coherent-diffraction regimes, and a multimodal sub-diffraction technique that bridges this gap has not yet been demonstrated. Here we report that structured illumination (SI) allows multimodal sub-diffraction imaging of both coherent quantitative-phase (QP) and fluorescence. Due to SI’s conventionally fluorescent applications, we first demonstrate the principle of SI-enabled three-dimensional (3D) QP sub-diffraction imaging with calibration microspheres. Image analysis confirmed enhanced lateral and axial resolutions over diffraction-limited QP imaging, and established striking parallels between coherent SI and conventional optical diffraction tomography. We next introduce an optical system utilizing SI to achieve 3D sub-diffraction, multimodal QP/fluorescent visualization of A549 biological cells fluorescently tagged for F-actin. Our results suggest that SI has a unique utility in studying biological phenomena with significant molecular, biophysical, and biochemical components. PMID:28663887

Decoding the dynamics of cellular metabolism and the action of 3-bromopyruvate and 2-deoxyglucose using pulsed stable isotope-resolved metabolomics.

PubMed

Pietzke, Matthias; Zasada, Christin; Mudrich, Susann; Kempa, Stefan

2014-01-01

Cellular metabolism is highly dynamic and continuously adjusts to the physiological program of the cell. The regulation of metabolism appears at all biological levels: (post-) transcriptional, (post-) translational, and allosteric. This regulatory information is expressed in the metabolome, but in a complex manner. To decode such complex information, new methods are needed in order to facilitate dynamic metabolic characterization at high resolution. Here, we describe pulsed stable isotope-resolved metabolomics (pSIRM) as a tool for the dynamic metabolic characterization of cellular metabolism. We have adapted gas chromatography-coupled mass spectrometric methods for metabolomic profiling and stable isotope-resolved metabolomics. In addition, we have improved robustness and reproducibility and implemented a strategy for the absolute quantification of metabolites. By way of examples, we have applied this methodology to characterize central carbon metabolism of a panel of cancer cell lines and to determine the mode of metabolic inhibition of glycolytic inhibitors in times ranging from minutes to hours. Using pSIRM, we observed that 2-deoxyglucose is a metabolic inhibitor, but does not directly act on the glycolytic cascade.

Interwoven Aligned Conductive Nanofiber Yarn/Hydrogel Composite Scaffolds for Engineered 3D Cardiac Anisotropy.

PubMed

Wu, Yaobin; Wang, Ling; Guo, Baolin; Ma, Peter X

2017-06-27

Mimicking the anisotropic cardiac structure and guiding 3D cellular orientation play a critical role in designing scaffolds for cardiac tissue regeneration. Significant advances have been achieved to control cellular alignment and elongation, but it remains an ongoing challenge for engineering 3D cardiac anisotropy using these approaches. Here, we present a 3D hybrid scaffold based on aligned conductive nanofiber yarns network (NFYs-NET, composition: polycaprolactone, silk fibroin, and carbon nanotubes) within a hydrogel shell for mimicking the native cardiac tissue structure, and further demonstrate their great potential for engineering 3D cardiac anisotropy for cardiac tissue engineering. The NFYs-NET structures are shown to control cellular orientation and enhance cardiomyocytes (CMs) maturation. 3D hybrid scaffolds were then fabricated by encapsulating NFYs-NET layers within hydrogel shell, and these 3D scaffolds performed the ability to promote aligned and elongated CMs maturation on each layer and individually control cellular orientation on different layers in a 3D environment. Furthermore, endothelialized myocardium was constructed by using this hybrid strategy via the coculture of CMs on NFYs-NET layer and endothelial cells within hydrogel shell. Therefore, these 3D hybrid scaffolds, containing NFYs-NET layer inducing cellular orientation, maturation, and anisotropy and hydrogel shell providing a suitable 3D environment for endothelialization, has great potential in engineering 3D cardiac anisotropy.

Enhanced labeling density and whole-cell 3D dSTORM imaging by repetitive labeling of target proteins.

PubMed

Venkataramani, Varun; Kardorff, Markus; Herrmannsdörfer, Frank; Wieneke, Ralph; Klein, Alina; Tampé, Robert; Heilemann, Mike; Kuner, Thomas

2018-04-03

With continuing advances in the resolving power of super-resolution microscopy, the inefficient labeling of proteins with suitable fluorophores becomes a limiting factor. For example, the low labeling density achieved with antibodies or small molecule tags limits attempts to reveal local protein nano-architecture of cellular compartments. On the other hand, high laser intensities cause photobleaching within and nearby an imaged region, thereby further reducing labeling density and impairing multi-plane whole-cell 3D super-resolution imaging. Here, we show that both labeling density and photobleaching can be addressed by repetitive application of trisNTA-fluorophore conjugates reversibly binding to a histidine-tagged protein by a novel approach called single-epitope repetitive imaging (SERI). For single-plane super-resolution microscopy, we demonstrate that, after multiple rounds of labeling and imaging, the signal density is increased. Using the same approach of repetitive imaging, washing and re-labeling, we demonstrate whole-cell 3D super-resolution imaging compensated for photobleaching above or below the imaging plane. This proof-of-principle study demonstrates that repetitive labeling of histidine-tagged proteins provides a versatile solution to break the 'labeling barrier' and to bypass photobleaching in multi-plane, whole-cell 3D experiments.

Phononic Band Gaps in 2D Quadratic and 3D Cubic Cellular Structures

PubMed Central

Warmuth, Franziska; Körner, Carolin

2015-01-01

The static and dynamic mechanical behaviour of cellular materials can be designed by the architecture of the underlying unit cell. In this paper, the phononic band structure of 2D and 3D cellular structures is investigated. It is shown how the geometry of the unit cell influences the band structure and eventually leads to full band gaps. The mechanism leading to full band gaps is elucidated. Based on this knowledge, a 3D cellular structure with a broad full band gap is identified. Furthermore, the dependence of the width of the gap on the geometry parameters of the unit cell is presented. PMID:28793713

Phononic Band Gaps in 2D Quadratic and 3D Cubic Cellular Structures.

PubMed

Warmuth, Franziska; Körner, Carolin

2015-12-02

The static and dynamic mechanical behaviour of cellular materials can be designed by the architecture of the underlying unit cell. In this paper, the phononic band structure of 2D and 3D cellular structures is investigated. It is shown how the geometry of the unit cell influences the band structure and eventually leads to full band gaps. The mechanism leading to full band gaps is elucidated. Based on this knowledge, a 3D cellular structure with a broad full band gap is identified. Furthermore, the dependence of the width of the gap on the geometry parameters of the unit cell is presented.

Quantification of substrate and cellular strains in stretchable 3D cell cultures: an experimental and computational framework.

PubMed

González-Avalos, P; Mürnseer, M; Deeg, J; Bachmann, A; Spatz, J; Dooley, S; Eils, R; Gladilin, E

2017-05-01

The mechanical cell environment is a key regulator of biological processes . In living tissues, cells are embedded into the 3D extracellular matrix and permanently exposed to mechanical forces. Quantification of the cellular strain state in a 3D matrix is therefore the first step towards understanding how physical cues determine single cell and multicellular behaviour. The majority of cell assays are, however, based on 2D cell cultures that lack many essential features of the in vivo cellular environment. Furthermore, nondestructive measurement of substrate and cellular mechanics requires appropriate computational tools for microscopic image analysis and interpretation. Here, we present an experimental and computational framework for generation and quantification of the cellular strain state in 3D cell cultures using a combination of 3D substrate stretcher, multichannel microscopic imaging and computational image analysis. The 3D substrate stretcher enables deformation of living cells embedded in bead-labelled 3D collagen hydrogels. Local substrate and cell deformations are determined by tracking displacement of fluorescent beads with subsequent finite element interpolation of cell strains over a tetrahedral tessellation. In this feasibility study, we debate diverse aspects of deformable 3D culture construction, quantification and evaluation, and present an example of its application for quantitative analysis of a cellular model system based on primary mouse hepatocytes undergoing transforming growth factor (TGF-β) induced epithelial-to-mesenchymal transition. © 2017 The Authors. Journal of Microscopy published by JohnWiley & Sons Ltd on behalf of Royal Microscopical Society.

Ombuin-3-O-β-D-glucopyranoside from Gynostemma pentaphyllum is a dual agonistic ligand of peroxisome proliferator-activated receptors α and δ/β

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

Malek, Mastura Abd; Hoang, Minh-Hien; Jia, Yaoyao

Highlights: ► Ombuin-3-O-β-D-glucopyranoside is a dual ligand for PPARα and δ/β. ► Ombuin-3-O-β-D-glucopyranoside reduces cellular lipid levels in multiple cell types. ► Cells stimulated with ombuine up-regulated target genes in cholesterol efflux. ► Cells stimulated with ombuine regulated target fatty acid β-oxidation and synthesis. ► Ombuin-3-O-β-D-glucopyranoside could ameliorate hyperlipidemia and hepatic steatosis. -- Abstract: We demonstrated that ombuin-3-O-β-D-glucopyranoside (ombuine), a flavonoid from Gynostemma pentaphyllum, is a dual agonist for peroxisome proliferator-activated receptors (PPARs) α and δ/β. Using surface plasmon resonance (SPR), time-resolved fluorescence resonance energy transfer (FRET) analyses, and reporter gene assays, we showed that ombuine bound directly to PPARαmore » and δ/β but not to PPARγ or liver X receptors (LXRs). Cultured HepG2 hepatocytes stimulated with ombuine significantly reduced intracellular concentrations of triglyceride and cholesterol and downregulated the expression of lipogenic genes, including sterol regulatory element binding protein-1c (SREBP1c) and stearoyl-CoA desaturase-1 (SCD-1), with activation of PPARα and δ/β. Activation of LXRs by ombuine was confirmed by reporter gene assays, however, SPR and cell-based FRET assays showed no direct binding of ombuine to either of the LXRs suggesting LXR activation by ombuine may be operated via PPARα stimulation. Ombuine-stimulated macrophages showed significantly induced transcription of ATP binding cassette cholesterol transporter A1 (ABCA1) and G1 (ABCG1), the key genes in reverse cholesterol transport, which led to reduced cellular cholesterol concentrations. These results suggest that ombuine is a dual PPAR ligand for PPARα and δ/β with the ability to decrease lipid concentrations by reducing lipogenic gene expression in hepatocytes and inducing genes involved in cholesterol efflux in macrophages.« less

Adipose-derived stem cells and BMP-2 delivery in chitosan-based 3D constructs to enhance bone regeneration in a rat mandibular defect model.

PubMed

Fan, Jiabing; Park, Hyejin; Lee, Matthew K; Bezouglaia, Olga; Fartash, Armita; Kim, Jinku; Aghaloo, Tara; Lee, Min

2014-08-01

Reconstructing segmental mandiblular defects remains a challenge in the clinic. Tissue engineering strategies provide an alternative option to resolve this problem. The objective of the present study was to determine the effects of adipose-derived stem cells (ASCs) and bone morphogenetic proteins-2 (BMP-2) in three-dimensional (3D) scaffolds on mandibular repair in a small animal model. Noggin expression levels in ASCs were downregulated by a lentiviral short hairpin RNA strategy to enhance ASC osteogenesis (ASCs(Nog-)). Chitosan (CH) and chondroitin sulfate (CS), natural polysaccharides, were fabricated into 3D porous scaffolds, which were further modified with apatite coatings for enhanced cellular responses and efficient delivery of BMP-2. The efficacy of 3D apatite-coated CH/CS scaffolds supplemented with ASCs(Nog-) and BMP-2 were evaluated in a rat critical-sized mandibular defect model. After 8 weeks postimplantation, the scaffolds treated with ASCs(Nog-) and BMP-2 significantly promoted rat mandibular regeneration as demonstrated by micro-computerized tomography, histology, and immunohistochemistry, compared with the groups treated with ASCs(Nog-) or BMP-2 alone. These results suggest that our combinatorial strategy of ASCs(Nog-)+BMP-2 in 3D apatite microenvironments can significantly promote mandibular regeneration, and these may provide a potential tissue engineering approach to repair large bony defects.

Optimal MRI sequence for identifying occlusion location in acute stroke: which value of time-resolved contrast-enhanced MRA?

PubMed

Le Bras, A; Raoult, H; Ferré, J-C; Ronzière, T; Gauvrit, J-Y

2015-06-01

Identifying occlusion location is crucial for determining the optimal therapeutic strategy during the acute phase of ischemic stroke. The purpose of this study was to assess the diagnostic efficacy of MR imaging, including conventional sequences plus time-resolved contrast-enhanced MRA in comparison with DSA for identifying arterial occlusion location. Thirty-two patients with 34 occlusion levels referred for thrombectomy during acute cerebral stroke events were consecutively included from August 2010 to December 2012. Before thrombectomy, we performed 3T MR imaging, including conventional 3D-TOF and gradient-echo T2 sequences, along with time-resolved contrast-enhanced MRA of the extra- and intracranial arteries. The 3D-TOF, gradient-echo T2, and time-resolved contrast-enhanced MRA results were consensually assessed by 2 neuroradiologists and compared with prethrombectomy DSA results in terms of occlusion location. The Wilcoxon test was used for statistical analysis to compare MR imaging sequences with DSA, and the κ coefficient was used to determine intermodality agreement. The occlusion level on the 3D-TOF and gradient-echo T2 images differed significantly from that of DSA (P < .001 and P = .002, respectively), while no significant difference was observed between DSA and time-resolved contrast-enhanced MRA (P = .125). κ coefficients for intermodality agreement with DSA (95% CI, percentage agreement) were 0.43 (0.3%-0.6; 62%), 0.32 (0.2%-0.5; 56%), and 0.81 (0.6%-1.0; 88%) for 3D-TOF, gradient-echo T2, and time-resolved contrast-enhanced MRA, respectively. The time-resolved contrast-enhanced MRA sequence proved reliable for identifying occlusion location in acute stroke with performance superior to that of 3D-TOF and gradient-echo T2 sequences. © 2015 by American Journal of Neuroradiology.

Advanced 3D Printers for Cellular Solids

DTIC Science & Technology

2016-06-30

2211 3d printing , cellular solids REPORT DOCUMENTATION PAGE 11. SPONSOR/MONITOR’S REPORT NUMBER(S) 10. SPONSOR/MONITOR’S ACRONYM(S) ARO 8...quality 3D printing and rapid prototyping in a fraction of the time taken by traditional 3D printers, using eco-friendly, inexpensive office paper and...STL file, which then can be used in printing the 3D model. Mechanical performance using compressive crushing of the 3D printed part will be studied

FIB-SEM tomography of human skin telocytes and their extracellular vesicles.

PubMed

Cretoiu, Dragos; Gherghiceanu, Mihaela; Hummel, Eric; Zimmermann, Hans; Simionescu, Olga; Popescu, Laurentiu M

2015-04-01

We have shown in 2012 the existence of telocytes (TCs) in human dermis. TCs were described by transmission electron microscopy (TEM) as interstitial cells located in non-epithelial spaces (stroma) of many organs (see www.telocytes.com). TCs have very long prolongations (tens to hundreds micrometers) named Telopodes (Tps). These Tps have a special conformation with dilated portions named podoms (containing mitochondria, endoplasmic reticulum and caveolae) and very thin segments (below resolving power of light microscopy), called podomers. To show the real 3D architecture of TC network, we used the most advanced available electron microscope technology: focused ion beam scanning electron microscopy (FIB-SEM) tomography. Generally, 3D reconstruction of dermal TCs by FIB-SEM tomography revealed the existence of Tps with various conformations: (i) long, flattened irregular veils (ribbon-like segments) with knobs, corresponding to podoms, and (ii) tubular structures (podomers) with uneven calibre because of irregular dilations (knobs) - the podoms. FIB-SEM tomography also showed numerous extracellular vesicles (diameter 438.6 ± 149.1 nm, n = 30) released by a human dermal TC. Our data might be useful for understanding the role(s) of TCs in intercellular signalling and communication, as well as for comprehension of pathologies like scleroderma, multiple sclerosis, psoriasis, etc. © 2015 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

Cellular Factors Shape 3D Genome Landscape

Cancer.gov

Researchers, using novel large-scale imaging technology, have mapped the spatial location of individual genes in the nucleus of human cells and identified 50 cellular factors required for the proper 3D positioning of genes. These spatial locations play important roles in gene expression, DNA repair, genome stability, and other cellular activities.

Can Positron 2D-ACAR Resolve the Electronic Structure of HIGH-Tc Superconductors?

NASA Astrophysics Data System (ADS)

Chan, L. P.; Lynn, K. G.; Harshman, D. R.

We examine the ability of the positron Two-Dimensional Angular Correlation Annihilation Radiation (2D-ACAR) technique to resolve the electronic structures of high-Tc cuprate superconductors. Following a short description of the technique, discussions of the theoretical assumptions, data analysis and experimental considerations, in relation to the high-Tc superconductors, are given. We briefly review recent 2D-ACAR experiments on YBa2Cu3O7-x, Bi2Sr2CaCuO8+δ and La2-xSrxCuO4. The 2D-ACAR technique is useful in resolving the band crossings associated with the layers of the superconductors that are preferentially sampled by the positrons. Together with other Fermi surface measurements (namely angle-resolved photoemission), 2D-ACAR can resolve some of the electronic structures of high-Tc cuprate superconductors. In addition, 2D-ACAR measurements of YBa2Cu3O7-x and Bi2Sr2CaCuO8+δ also reveal an interesting temperature dependence in the fine structures, and a change in the positron lifetime in the former.

3D Encoding of Musical Score Information and the Playback Method Used by the Cellular Phone

NASA Astrophysics Data System (ADS)

Kubo, Hitoshi; Sugiura, Akihiko

Recently, 3G cellular phone that can take a movie has spread by improving the digital camera function. And, 2Dcode has accurate readout and high operability. And it has spread as an information transmission means. However, the symbol is expanded and complicated when information of 2D codes increases. To solve these, 3D code was proposed. But it need the special equipment for readout, and specializes in the enhancing reality feeling technology. Therefore, it is difficult to apply it to the cellular phone. And so, we propose 3D code that can be recognized by the movie shooting function of the cellular phone. And, score information was encoded. We apply Gray Code to the property of music, and encode it. And the effectiveness was verified.

Time-resolved spectroscopic imaging reveals the fundamentals of cellular NADH fluorescence.

PubMed

Li, Dong; Zheng, Wei; Qu, Jianan Y

2008-10-15

A time-resolved spectroscopic imaging system is built to study the fluorescence characteristics of nicotinamide adenine dinucleotide (NADH), an important metabolic coenzyme and endogenous fluorophore in cells. The system provides a unique approach to measure fluorescence signals in different cellular organelles and cytoplasm. The ratios of free over protein-bound NADH signals in cytosol and nucleus are slightly higher than those in mitochondria. The mitochondrial fluorescence contributes about 70% of overall cellular fluorescence and is not a completely dominant signal. Furthermore, NADH signals in mitochondria, cytosol, and the nucleus respond to the changes of cellular activity differently, suggesting that cytosolic and nuclear fluorescence may complicate the well-known relationship between mitochondrial fluorescence and cellular metabolism.

Contrast-enhanced time-resolved MRA for follow-up of intracranial aneurysms treated with the pipeline embolization device.

PubMed

Boddu, S R; Tong, F C; Dehkharghani, S; Dion, J E; Saindane, A M

2014-01-01

Endovascular reconstruction and flow diversion by using the Pipeline Embolization Device is an effective treatment for complex cerebral aneurysms. Accurate noninvasive alternatives to DSA for follow-up after Pipeline Embolization Device treatment are desirable. This study evaluated the accuracy of contrast-enhanced time-resolved MRA for this purpose, hypothesizing that contrast-enhanced time-resolved MRA will be comparable with DSA and superior to 3D-TOF MRA. During a 24-month period, 37 Pipeline Embolization Device-treated intracranial aneurysms in 26 patients underwent initial follow-up by using 3D-TOF MRA, contrast-enhanced time-resolved MRA, and DSA. MRA was performed on a 1.5T unit by using 3D-TOF and time-resolved imaging of contrast kinetics. All patients underwent DSA a median of 0 days (range, 0-68) after MRA. Studies were evaluated for aneurysm occlusion, quality of visualization of the reconstructed artery, and measurable luminal diameter of the Pipeline Embolization Device, with DSA used as the reference standard. The sensitivity, specificity, and positive and negative predictive values of contrast-enhanced time-resolved MRA relative to DSA for posttreatment aneurysm occlusion were 96%, 85%, 92%, and 92%. Contrast-enhanced time-resolved MRA demonstrated superior quality of visualization (P = .0001) and a higher measurable luminal diameter (P = .0001) of the reconstructed artery compared with 3D-TOF MRA but no significant difference compared with DSA. Contrast-enhanced time-resolved MRA underestimated the luminal diameter of the reconstructed artery by 0.965 ± 0.497 mm (27% ± 13%) relative to DSA. Contrast-enhanced time-resolved MRA is a reliable noninvasive method for monitoring intracranial aneurysms following flow diversion and vessel reconstruction by using the Pipeline Embolization Device. © 2014 by American Journal of Neuroradiology.

Magnetic field effects on spectrally resolved lifetime of on-line oxygen monitoring using magneto-optic probes

NASA Astrophysics Data System (ADS)

Mermut, O.; Gallant, P.; Le Bouch, N.; Leclair, S.; Noiseux, I.; Vernon, M.; Morin, J.-F.; Diamond, K.; Patterson, M. S.; Samkoe, K.; Pogue, B.

2009-02-01

Multimodal agents that serve as both probes for contrast and light-activated effectors of cellular processes in diseased tissue were developed. These agents were introduced into multicellular tumor spheroids (3D tissue models) and in the chorioallantoic membrane (CAM) of a chicken embryo. The luminescence decay was examined using a novel technique involving a spectrally-resolved fluorescence lifetime apparatus integrated with a weak electromagnet. A spectrallyresolved lifetime setup was used to identify magneto-optic species sensitive to magnetic field effects and distinguish from background emissions. We demonstrate that the applied magnetic fields can alter reaction rates and product distribution of some dyes detected by time- and spectrally-resolved luminescence changes. We will discuss the use of exogenous magneto-optical probes taken up in tumors to both induce phototoxicity, a process that is governed by complex and dynamically evolving mechanisms involving reactive oxygen species, and monitor treatment progress. The magnetic field enhancement, measured over a range of weak fields (0-300 mT) is correlated to oxygenation and may be used to monitor dynamic changes occurring due to oxygen consumption over the course of photodynamic therapy. Such online measurements provide the possibility to derive real-time information about response to treatment via monitoring magnetic field enhancement/suppression of the time-resolved, spectrally-resolved luminescence of the probe at the site of the treatment directly. Magnetic perturbation of lifetime can serve as a status reporter, providing optical feedback of oxygen-mediated treatments in situ and allowing for real-time adjustment of a phototherapy treatment plan.

Precipitation Processes developed during ARM (1997), TOGA COARE (1992), GATE (1974), SCSMEX (1998), and KWAJEX (1999), Consistent 2D, semi-3D and 3D Cloud Resolving Model Simulations

NASA Technical Reports Server (NTRS)

Tao, Wei-Kuo; Hou, A.; Atlas, R.; Starr, D.; Sud, Y.

2003-01-01

Real clouds and cloud systems are inherently three-dimensional (3D). Because of the limitations in computer resources, however, most cloud-resolving models (CRMs) today are still two-dimensional (2D). A few 3D CRMs have been used to study the response of clouds to large-scale forcing. In these 3D simulations, the model domain was small, and the integration time was 6 hours. The major objectives of this paper are: (1) to assess the performance of the super-parameterization technique (i.e. is 2D or semi-3D CRM appropriate for the super-parameterization?); (2) calculate and examine the surface energy (especially radiation) and water budgets; (3) identify the differences and similarities in the organization and entrainment rates of convection between simulated 2D and 3D cloud systems.

Structure and Reversibility of 2D von Neumann Cellular Automata Over Triangular Lattice

NASA Astrophysics Data System (ADS)

Uguz, Selman; Redjepov, Shovkat; Acar, Ecem; Akin, Hasan

2017-06-01

Even though the fundamental main structure of cellular automata (CA) is a discrete special model, the global behaviors at many iterative times and on big scales could be a close, nearly a continuous, model system. CA theory is a very rich and useful phenomena of dynamical model that focuses on the local information being relayed to the neighboring cells to produce CA global behaviors. The mathematical points of the basic model imply the computable values of the mathematical structure of CA. After modeling the CA structure, an important problem is to be able to move forwards and backwards on CA to understand their behaviors in more elegant ways. A possible case is when CA is to be a reversible one. In this paper, we investigate the structure and the reversibility of two-dimensional (2D) finite, linear, triangular von Neumann CA with null boundary case. It is considered on ternary field ℤ3 (i.e. 3-state). We obtain their transition rule matrices for each special case. For given special triangular information (transition) rule matrices, we prove which triangular linear 2D von Neumann CAs are reversible or not. It is known that the reversibility cases of 2D CA are generally a much challenged problem. In the present study, the reversibility problem of 2D triangular, linear von Neumann CA with null boundary is resolved completely over ternary field. As far as we know, there is no structure and reversibility study of von Neumann 2D linear CA on triangular lattice in the literature. Due to the main CA structures being sufficiently simple to investigate in mathematical ways, and also very complex to obtain in chaotic systems, it is believed that the present construction can be applied to many areas related to these CA using any other transition rules.

Accelerated time-resolved three-dimensional MR velocity mapping of blood flow patterns in the aorta using SENSE and k-t BLAST.

PubMed

Stadlbauer, Andreas; van der Riet, Wilma; Crelier, Gerard; Salomonowitz, Erich

2010-07-01

To assess the feasibility and potential limitations of the acceleration techniques SENSE and k-t BLAST for time-resolved three-dimensional (3D) velocity mapping of aortic blood flow. Furthermore, to quantify differences in peak velocity versus heart phase curves. Time-resolved 3D blood flow patterns were investigated in eleven volunteers and two patients suffering from aortic diseases with accelerated PC-MR sequences either in combination with SENSE (R=2) or k-t BLAST (6-fold). Both sequences showed similar data acquisition times and hence acceleration efficiency. Flow-field streamlines were calculated and visualized using the GTFlow software tool in order to reconstruct 3D aortic blood flow patterns. Differences between the peak velocities from single-slice PC-MRI experiments using SENSE 2 and k-t BLAST 6 were calculated for the whole cardiac cycle and averaged for all volunteers. Reconstruction of 3D flow patterns in volunteers revealed attenuations in blood flow dynamics for k-t BLAST 6 compared to SENSE 2 in terms of 3D streamlines showing fewer and less distinct vortices and reduction in peak velocity, which is caused by temporal blurring. Solely by time-resolved 3D MR velocity mapping in combination with SENSE detected pathologic blood flow patterns in patients with aortic diseases. For volunteers, we found a broadening and flattering of the peak velocity versus heart phase diagram between the two acceleration techniques, which is an evidence for the temporal blurring of the k-t BLAST approach. We demonstrated the feasibility of SENSE and detected potential limitations of k-t BLAST when used for time-resolved 3D velocity mapping. The effects of higher k-t BLAST acceleration factors have to be considered for application in 3D velocity mapping. Copyright 2009 Elsevier Ireland Ltd. All rights reserved.

3D Spatially Resolved Models of the Intracellular Dynamics of the Hepatitis C Genome Replication Cycle

PubMed Central

Reiter, Sebastian; Grillo, Alfio; Herrmann, Eva; Wittum, Gabriel

2017-01-01

Mathematical models of virus dynamics have not previously acknowledged spatial resolution at the intracellular level despite substantial arguments that favor the consideration of intracellular spatial dependence. The replication of the hepatitis C virus (HCV) viral RNA (vRNA) occurs within special replication complexes formed from membranes derived from endoplasmatic reticulum (ER). These regions, termed membranous webs, are generated primarily through specific interactions between nonstructural virus-encoded proteins (NSPs) and host cellular factors. The NSPs are responsible for the replication of the vRNA and their movement is restricted to the ER surface. Therefore, in this study we developed fully spatio-temporal resolved models of the vRNA replication cycle of HCV. Our simulations are performed upon realistic reconstructed cell structures—namely the ER surface and the membranous webs—based on data derived from immunostained cells replicating HCV vRNA. We visualized 3D simulations that reproduced dynamics resulting from interplay of the different components of our models (vRNA, NSPs, and a host factor), and we present an evaluation of the concentrations for the components within different regions of the cell. Thus far, our model is restricted to an internal portion of a hepatocyte and is qualitative more than quantitative. For a quantitative adaption to complete cells, various additional parameters will have to be determined through further in vitro cell biology experiments, which can be stimulated by the results described in the present study. PMID:28973992

Current State-of-the-Art 3D Tissue Models and Their Compatibility with Live Cell Imaging.

PubMed

Bardsley, Katie; Deegan, Anthony J; El Haj, Alicia; Yang, Ying

2017-01-01

Mammalian cells grow within a complex three-dimensional (3D) microenvironment where multiple cells are organized and surrounded by extracellular matrix (ECM). The quantity and types of ECM components, alongside cell-to-cell and cell-to-matrix interactions dictate cellular differentiation, proliferation and function in vivo. To mimic natural cellular activities, various 3D tissue culture models have been established to replace conventional two dimensional (2D) culture environments. Allowing for both characterization and visualization of cellular activities within possibly bulky 3D tissue models presents considerable challenges due to the increased thickness and subsequent light scattering features of such 3D models. In this chapter, state-of-the-art methodologies used to establish 3D tissue models are discussed, first with a focus on both scaffold-free and scaffold-based 3D tissue model formation. Following on, multiple 3D live cell imaging systems, mainly optical imaging modalities, are introduced. Their advantages and disadvantages are discussed, with the aim of stimulating more research in this highly demanding research area.

Transient inter-cellular polymeric linker.

PubMed

Ong, Siew-Min; He, Lijuan; Thuy Linh, Nguyen Thi; Tee, Yee-Han; Arooz, Talha; Tang, Guping; Tan, Choon-Hong; Yu, Hanry

2007-09-01

Three-dimensional (3D) tissue-engineered constructs with bio-mimicry cell-cell and cell-matrix interactions are useful in regenerative medicine. In cell-dense and matrix-poor tissues of the internal organs, cells support one another via cell-cell interactions, supplemented by small amount of the extra-cellular matrices (ECM) secreted by the cells. Here we connect HepG2 cells directly but transiently with inter-cellular polymeric linker to facilitate cell-cell interaction and aggregation. The linker consists of a non-toxic low molecular-weight polyethyleneimine (PEI) backbone conjugated with multiple hydrazide groups that can aggregate cells within 30 min by reacting with the aldehyde handles on the chemically modified cell-surface glycoproteins. The cells in the cellular aggregates proliferated; and maintained the cortical actin distribution of the 3D cell morphology while non-aggregated cells died over 7 days of suspension culture. The aggregates lost distinguishable cell-cell boundaries within 3 days; and the ECM fibers became visible around cells from day 3 onwards while the inter-cellular polymeric linker disappeared from the cell surfaces over time. The transient inter-cellular polymeric linker can be useful for forming 3D cellular and tissue constructs without bulk biomaterials or extensive network of engineered ECM for various applications.

A new detection scheme for ultrafast 2D J-resolved spectroscopy

NASA Astrophysics Data System (ADS)

Giraudeau, Patrick; Akoka, Serge

2007-06-01

Recent ultrafast techniques enable 2D NMR spectra to be obtained in a single scan. A modification of the detection scheme involved in this technique is proposed, permitting the achievement of 2D 1H J-resolved spectra in 500 ms. The detection gradient echoes are substituted by spin echoes to obtain spectra where the coupling constants are encoded along the direct ν2 domain. The use of this new J-resolved detection block after continuous phase-encoding excitation schemes is discussed in terms of resolution and sensitivity. J-resolved spectra obtained on cinnamic acid and 3-ethyl bromopropionate are presented, revealing the expected 2D J-patterns with coupling constants as small as 2 Hz.

On the star partition dimension of comb product of cycle and path

NASA Astrophysics Data System (ADS)

Alfarisi, Ridho; Darmaji

2017-08-01

Let G = (V, E) be a connected graphs with vertex set V(G), edge set E(G) and S ⊆ V(G). Given an ordered partition Π = {S1, S2, S3, …, Sk} of the vertex set V of G, the representation of a vertex v ∈ V with respect to Π is the vector r(v|Π) = (d(v, S1), d(v, S2), …, d(v, Sk)), where d(v, Sk) represents the distance between the vertex v and the set Sk and d(v, Sk) = min{d(v, x)|x ∈ Sk }. A partition Π of V(G) is a resolving partition if different vertices of G have distinct representations, i.e., for every pair of vertices u, v ∈ V(G), r(u|Π) ≠ r(v|Π). The minimum k of Π resolving partition is a partition dimension of G, denoted by pd(G). The resolving partition Π = {S1, S2, S3, …, Sk } is called a star resolving partition for G if it is a resolving partition and each subgraph induced by Si, 1 ≤ i ≤ k, is a star. The minimum k for which there exists a star resolving partition of V(G) is the star partition dimension of G, denoted by spd(G). Finding the star partition dimension of G is classified to be a NP-Hard problem. In this paper, we will show that the partition dimension of comb product of cycle and path namely Cm⊳Pn and Pn⊳Cm for n ≥ 2 and m ≥ 3.

3D printed phantoms of retinal photoreceptor cells for evaluating adaptive optics imaging modalities

NASA Astrophysics Data System (ADS)

Kedia, Nikita; Liu, Zhuolin; Sochol, Ryan; Hammer, Daniel X.; Agrawal, Anant

2018-02-01

Adaptive optics-enabled optical coherence tomography (AO-OCT) and scanning laser ophthalmoscopy (AO-SLO) devices can resolve retinal cones and rods in three dimensions. To evaluate the improved resolution of AO-OCT and AO-SLO, a phantom that mimics retinal anatomy at the cellular level is required. We used a two-photon polymerization approach to fabricate three-dimensional (3D) photoreceptor phantoms modeled on the central foveal cones. By using a femtosecond laser to selectively photocure precise locations within a liquid-based photoresist via two-photon absorption, we produced high-resolution phantoms with μm-level dimensions similar to true anatomy. In this work, we present two phantoms to evaluate the resolution limits of an AO imaging system: one that models only the outer segments of the photoreceptor cells at varying retinal eccentricities and another that contains anatomically relevant features of the full-length photoreceptor. With these phantoms we are able to quantitatively estimate transverse resolution of an AO system and produce images that are comparable to those found in the human retina.

Live Cell Imaging of Viscosity in 3D Tumour Cell Models.

PubMed

Shirmanova, Marina V; Shimolina, Lubov' E; Lukina, Maria M; Zagaynova, Elena V; Kuimova, Marina K

2017-01-01

Abnormal levels of viscosity in tissues and cells are known to be associated with disease and malfunction. While methods to measure bulk macroscopic viscosity of bio-tissues are well developed, imaging viscosity at the microscopic scale remains a challenge, especially in vivo. Molecular rotors are small synthetic viscosity-sensitive fluorophores in which fluorescence parameters are strongly correlated to the microviscosity of their immediate environment. Hence, molecular rotors represent a promising instrument for mapping of viscosity in living cells and tissues at the microscopic level. Quantitative measurements of viscosity can be achieved by recording time-resolved fluorescence decays of molecular rotor using fluorescence lifetime imaging microscopy (FLIM), which is also suitable for dynamic viscosity mapping, both in cellulo and in vivo. Among tools of experimental oncology, 3D tumour cultures, or spheroids, are considered a more adequate in vitro model compared to a cellular monolayer, and represent a less labour-intensive and more unified approach compared to animal tumour models. This chapter describes a methodology for microviscosity imaging in tumour spheroids using BODIPY-based molecular rotors and two photon-excited FLIM.

Configuration of a high-content imaging platform for hit identification and pharmacological assessment of JMJD3 demethylase enzyme inhibitors.

PubMed

Mulji, Alpa; Haslam, Carl; Brown, Fiona; Randle, Rebecca; Karamshi, Bhumika; Smith, Julia; Eagle, Robert; Munoz-Muriedas, Jordi; Taylor, Joanna; Sheikh, Arshad; Bridges, Angela; Gill, Kirsty; Jepras, Rob; Smee, Penny; Barker, Mike; Woodrow, Mike; Liddle, John; Thomas, Pamela; Jones, Emma; Gordon, Laurie; Tanner, Rob; Leveridge, Melanie; Hutchinson, Sue; Martin, Margaret; Brown, Murray; Kruidenier, Laurens; Katso, Roy

2012-01-01

The biological complexity associated with the regulation of histone demethylases makes it desirable to configure a cellular mechanistic assay format that simultaneously encompasses as many of the relevant cellular processes as possible. In this report, the authors describe the configuration of a JMJD3 high-content cellular mechanistic imaging assay that uses single-cell multiparameter measurements to accurately assess cellular viability and the enzyme-dependent demethylation of the H3K27(Me)3 mark by exogenously expressed JMJD3. This approach couples robust statistical analyses with the spatial resolving power of cellular imaging. This enables segregation of expressing and nonexpressing cells into discrete subpopulations and consequently pharmacological quantification of compounds of interest in the expressing population at varying JMJD3 expression levels. Moreover, the authors demonstrate the utility of this hit identification strategy through the successful prosecution of a medium-throughput focused campaign of an 87 500-compound file, which has enabled the identification of JMJD3 cellular-active chemotypes. This study represents the first report of a demethylase high-content imaging assay with the ability to capture a repertoire of pharmacological tools, which are likely both to inform our mechanistic understanding of how JMJD3 is modulated and, more important, to contribute to the identification of novel therapeutic modalities for this demethylase enzyme.

Development of 3D woven cellular structures for adaptive composites based on thermoplastic hybrid yarns

NASA Astrophysics Data System (ADS)

Sennewald, C.; Vorhof, M.; Schegner, P.; Hoffmann, G.; Cherif, C.; Boblenz, J.; Sinapius, M.; Hühne, C.

2018-05-01

Flexible cellular 3D structures with structure-inherent compliance made of fiber-reinforced composites have repeatedly aroused the interest of international research groups. Such structures offer the possibility to meet the increasing demand for flexible and adaptive structures. The aim of this paper is the development of cellular 3D structures based on weaving technology. Considering the desired geometry of the 3D structure, algorithms are developed for the formation of geometry through tissue sub-areas. Subsequently, these sub-areas are unwound into the weaving level and appropriate weave patterns are developed. A particular challenge is the realization of compliant mechanisms in the woven fabric. This can be achieved either by combining different materials or, in particular, by implementing large stiffness gradients by means of varying the woven fabrics thickness, whereas differences in wall thickness have to be realized with a factor of 1:10. A manufacturing technology based on the weaving process is developed for the realization of the developed 3D cellular structures. To this end, solutions for the processing of hybrid thermoplastic materials (e.g. tapes), solutions for the integration of inlays in the weaving process (thickening of partial areas), and solutions for tissue retraction, as well as for the fabric pull-off (linear pull-off system) are being developed. In this way, woven cellular 3D structures with woven outer layers and woven joint areas (compliance) can be realized in a single process step and are subsequently characterized.

Quantitative analyses of the 3D nuclear landscape recorded with super-resolved fluorescence microscopy.

PubMed

Schmid, Volker J; Cremer, Marion; Cremer, Thomas

2017-07-01

Recent advancements of super-resolved fluorescence microscopy have revolutionized microscopic studies of cells, including the exceedingly complex structural organization of cell nuclei in space and time. In this paper we describe and discuss tools for (semi-) automated, quantitative 3D analyses of the spatial nuclear organization. These tools allow the quantitative assessment of highly resolved different chromatin compaction levels in individual cell nuclei, which reflect functionally different regions or sub-compartments of the 3D nuclear landscape, and measurements of absolute distances between sites of different chromatin compaction. In addition, these tools allow 3D mapping of specific DNA/RNA sequences and nuclear proteins relative to the 3D chromatin compaction maps and comparisons of multiple cell nuclei. The tools are available in the free and open source R packages nucim and bioimagetools. We discuss the use of masks for the segmentation of nuclei and the use of DNA stains, such as DAPI, as a proxy for local differences in chromatin compaction. We further discuss the limitations of 3D maps of the nuclear landscape as well as problems of the biological interpretation of such data. Copyright © 2017 Elsevier Inc. All rights reserved.

Improving the accuracy and efficiency of time-resolved electronic spectra calculations: cellular dephasing representation with a prefactor.

PubMed

Zambrano, Eduardo; Šulc, Miroslav; Vaníček, Jiří

2013-08-07

Time-resolved electronic spectra can be obtained as the Fourier transform of a special type of time correlation function known as fidelity amplitude, which, in turn, can be evaluated approximately and efficiently with the dephasing representation. Here we improve both the accuracy of this approximation-with an amplitude correction derived from the phase-space propagator-and its efficiency-with an improved cellular scheme employing inverse Weierstrass transform and optimal scaling of the cell size. We demonstrate the advantages of the new methodology by computing dispersed time-resolved stimulated emission spectra in the harmonic potential, pyrazine, and the NCO molecule. In contrast, we show that in strongly chaotic systems such as the quartic oscillator the original dephasing representation is more appropriate than either the cellular or prefactor-corrected methods.

4D blood flow mapping using SPIM-microPIV in the developing zebrafish heart

NASA Astrophysics Data System (ADS)

Zickus, Vytautas; Taylor, Jonathan M.

2018-02-01

Fluid-structure interaction in the developing heart is an active area of research in developmental biology. However, investigation of heart dynamics is mostly limited to computational uid dynamics simulations using heart wall structure information only, or single plane blood ow information - so there is a need for 3D + time resolved data to fully understand cardiac function. We present an imaging platform combining selective plane illumination microscopy (SPIM) with micro particle image velocimetry (μPIV) to enable 3D-resolved flow mapping in a microscopic environment, free from many of the sources of error and bias present in traditional epi uorescence-based μPIV systems. By using our new system in conjunction with optical heart beat synchronization, we demonstrate the ability obtain non-invasive 3D + time resolved blood flow measurements in the heart of a living zebrafish embryo.

An x-ray fluorescence imaging system for gold nanoparticle detection.

PubMed

Ricketts, K; Guazzoni, C; Castoldi, A; Gibson, A P; Royle, G J

2013-11-07

Gold nanoparticles (GNPs) may be used as a contrast agent to identify tumour location and can be modified to target and image specific tumour biological parameters. There are currently no imaging systems in the literature that have sufficient sensitivity to GNP concentration and distribution measurement at sufficient tissue depth for use in in vivo and in vitro studies. We have demonstrated that high detecting sensitivity of GNPs can be achieved using x-ray fluorescence; furthermore this technique enables greater depth imaging in comparison to optical modalities. Two x-ray fluorescence systems were developed and used to image a range of GNP imaging phantoms. The first system consisted of a 10 mm(2) silicon drift detector coupled to a slightly focusing polycapillary optic which allowed 2D energy resolved imaging in step and scan mode. The system has sensitivity to GNP concentrations as low as 1 ppm. GNP concentrations different by a factor of 5 could be resolved, offering potential to distinguish tumour from non-tumour. The second system was designed to avoid slow step and scan image acquisition; the feasibility of excitation of the whole specimen with a wide beam and detection of the fluorescent x-rays with a pixellated controlled drift energy resolving detector without scanning was investigated. A parallel polycapillary optic coupled to the detector was successfully used to ascertain the position where fluorescence was emitted. The tissue penetration of the technique was demonstrated to be sufficient for near-surface small-animal studies, and for imaging 3D in vitro cellular constructs. Previous work demonstrates strong potential for both imaging systems to form quantitative images of GNP concentration.

3D-resolved fluorescence and phosphorescence lifetime imaging using temporal focusing wide-field two-photon excitation

PubMed Central

Choi, Heejin; Tzeranis, Dimitrios S.; Cha, Jae Won; Clémenceau, Philippe; de Jong, Sander J. G.; van Geest, Lambertus K.; Moon, Joong Ho; Yannas, Ioannis V.; So, Peter T. C.

2012-01-01

Fluorescence and phosphorescence lifetime imaging are powerful techniques for studying intracellular protein interactions and for diagnosing tissue pathophysiology. While lifetime-resolved microscopy has long been in the repertoire of the biophotonics community, current implementations fall short in terms of simultaneously providing 3D resolution, high throughput, and good tissue penetration. This report describes a new highly efficient lifetime-resolved imaging method that combines temporal focusing wide-field multiphoton excitation and simultaneous acquisition of lifetime information in frequency domain using a nanosecond gated imager from a 3D-resolved plane. This approach is scalable allowing fast volumetric imaging limited only by the available laser peak power. The accuracy and performance of the proposed method is demonstrated in several imaging studies important for understanding peripheral nerve regeneration processes. Most importantly, the parallelism of this approach may enhance the imaging speed of long lifetime processes such as phosphorescence by several orders of magnitude. PMID:23187477

An image encryption algorithm based on 3D cellular automata and chaotic maps

NASA Astrophysics Data System (ADS)

Del Rey, A. Martín; Sánchez, G. Rodríguez

2015-05-01

A novel encryption algorithm to cipher digital images is presented in this work. The digital image is rendering into a three-dimensional (3D) lattice and the protocol consists of two phases: the confusion phase where 24 chaotic Cat maps are applied and the diffusion phase where a 3D cellular automata is evolved. The encryption method is shown to be secure against the most important cryptanalytic attacks.

Optical computed tomography for spatially isotropic four-dimensional imaging of live single cells

PubMed Central

Kelbauskas, Laimonas; Shetty, Rishabh; Cao, Bin; Wang, Kuo-Chen; Smith, Dean; Wang, Hong; Chao, Shi-Hui; Gangaraju, Sandhya; Ashcroft, Brian; Kritzer, Margaret; Glenn, Honor; Johnson, Roger H.; Meldrum, Deirdre R.

2017-01-01

Quantitative three-dimensional (3D) computed tomography (CT) imaging of living single cells enables orientation-independent morphometric analysis of the intricacies of cellular physiology. Since its invention, x-ray CT has become indispensable in the clinic for diagnostic and prognostic purposes due to its quantitative absorption-based imaging in true 3D that allows objects of interest to be viewed and measured from any orientation. However, x-ray CT has not been useful at the level of single cells because there is insufficient contrast to form an image. Recently, optical CT has been developed successfully for fixed cells, but this technology called Cell-CT is incompatible with live-cell imaging due to the use of stains, such as hematoxylin, that are not compatible with cell viability. We present a novel development of optical CT for quantitative, multispectral functional 4D (three spatial + one spectral dimension) imaging of living single cells. The method applied to immune system cells offers truly isotropic 3D spatial resolution and enables time-resolved imaging studies of cells suspended in aqueous medium. Using live-cell optical CT, we found a heterogeneous response to mitochondrial fission inhibition in mouse macrophages and differential basal remodeling of small (0.1 to 1 fl) and large (1 to 20 fl) nuclear and mitochondrial structures on a 20- to 30-s time scale in human myelogenous leukemia cells. Because of its robust 3D measurement capabilities, live-cell optical CT represents a powerful new tool in the biomedical research field. PMID:29226240

State-resolved three-dimensional electron-momentum correlation in nonsequential double ionization of benzene

NASA Astrophysics Data System (ADS)

Winney, Alexander H.; Lin, Yun Fei; Lee, Suk Kyoung; Adhikari, Pradip; Li, Wen

2016-03-01

We report state-resolved electron-momentum correlation measurement of strong-field nonsequential double ionization in benzene. With a novel coincidence detection apparatus, highly efficient triple coincidence (electron-electron dication) and quadruple coincidence (electron-electron-cation-cation) are used to resolve the final ionic states and to characterize three-dimensional (3D) electron-momentum correlation. The primary states associated with dissociative and nondissociative dications are assigned. A 3D momentum anticorrelation is observed for the electrons in coincidence with dissociative benzene dication states whereas such a correlation is absent for nondissociative dication states.

Multiplex, quantitative cellular analysis in large tissue volumes with clearing-enhanced 3D microscopy (Ce3D)

PubMed Central

Li, Weizhe; Germain, Ronald N.

2017-01-01

Organ homeostasis, cellular differentiation, signal relay, and in situ function all depend on the spatial organization of cells in complex tissues. For this reason, comprehensive, high-resolution mapping of cell positioning, phenotypic identity, and functional state in the context of macroscale tissue structure is critical to a deeper understanding of diverse biological processes. Here we report an easy to use method, clearing-enhanced 3D (Ce3D), which generates excellent tissue transparency for most organs, preserves cellular morphology and protein fluorescence, and is robustly compatible with antibody-based immunolabeling. This enhanced signal quality and capacity for extensive probe multiplexing permits quantitative analysis of distinct, highly intermixed cell populations in intact Ce3D-treated tissues via 3D histo-cytometry. We use this technology to demonstrate large-volume, high-resolution microscopy of diverse cell types in lymphoid and nonlymphoid organs, as well as to perform quantitative analysis of the composition and tissue distribution of multiple cell populations in lymphoid tissues. Combined with histo-cytometry, Ce3D provides a comprehensive strategy for volumetric quantitative imaging and analysis that bridges the gap between conventional section imaging and disassociation-based techniques. PMID:28808033

A Novel Water-Soluble Fluorescence Probe with Wash-Free Cellular Imaging Capacity Based on AIE Characteristics.

PubMed

Qian, Yunxia; Liu, Hongmei; Tan, Haijian; Yang, Qingmin; Zhang, Shuchen; Han, Lingui; Yi, Xuegang; Huo, Li; Zhao, Hongchi; Wu, Yonggang; Bai, Libin; Ba, Xinwu

2017-05-01

A potential real-time imaging water-soluble fluorescent polymer (P3) is facilely prepared via one-pot method. For P3, tetraphenylethene unit serves as the fluorescent unit, poly(acryloyl ethylene diamine) (a kind of polyelectrolyte) with specific degree of polymerization acts as water-soluble part. 1 H-NMR, gel permeation chromatography (GPC), UV-vis spectroscopy, photoluminescence (PL), and confocal laser scanning microscopy are undertaken to characterize the structure and property of P3. The results of wash-free cellular imaging show that the signal-to-noise ratio is high as the concentration of P3 is 50 μg mL -1 . In addition, the pH-responsive and Cd 2+ -responsive are also investigated in this paper. The results coming from pH-responsive show that P3 solution displays significant fluorescence under near neutral. And the result from the cellular imaging shows that intracellular fluorescence intensity enhances with the augment of concentration of Cd 2+ , which reveals that P3 can give a hint to resolve the dilemma of traditional fluorescent dyes used as living cellular fluorescent probe. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Robust and highly performant ring detection algorithm for 3d particle tracking using 2d microscope imaging

NASA Astrophysics Data System (ADS)

Afik, Eldad

2015-09-01

Three-dimensional particle tracking is an essential tool in studying dynamics under the microscope, namely, fluid dynamics in microfluidic devices, bacteria taxis, cellular trafficking. The 3d position can be determined using 2d imaging alone by measuring the diffraction rings generated by an out-of-focus fluorescent particle, imaged on a single camera. Here I present a ring detection algorithm exhibiting a high detection rate, which is robust to the challenges arising from ring occlusion, inclusions and overlaps, and allows resolving particles even when near to each other. It is capable of real time analysis thanks to its high performance and low memory footprint. The proposed algorithm, an offspring of the circle Hough transform, addresses the need to efficiently trace the trajectories of many particles concurrently, when their number in not necessarily fixed, by solving a classification problem, and overcomes the challenges of finding local maxima in the complex parameter space which results from ring clusters and noise. Several algorithmic concepts introduced here can be advantageous in other cases, particularly when dealing with noisy and sparse data. The implementation is based on open-source and cross-platform software packages only, making it easy to distribute and modify. It is implemented in a microfluidic experiment allowing real-time multi-particle tracking at 70 Hz, achieving a detection rate which exceeds 94% and only 1% false-detection.

5D-intravital tomography as a novel tool for non-invasive in-vivo analysis of human skin

NASA Astrophysics Data System (ADS)

König, Karsten; Weinigel, Martin; Breunig, Hans G.; Gregory, Axel; Fischer, Peter; Kellner-Höfer, Marcel; Bückle, Rainer; Schwarz, Martin; Riemann, Iris; Stracke, Frank; Huck, Volker; Gorzelanny, Christian; Schneider, Stefan W.

2010-02-01

Some years ago, CE-marked clinical multiphoton systems for 3D imaging of human skin with subcellular resolution have been launched. These tomographs provide optical biopsies with submicron resolution based on two-photon excited autofluorescence (NAD(P)H, flavoproteins, keratin, elastin, melanin, porphyrins) and second harmonic generation by collagen. The 3D tomograph was now transferred into a 5D imaging system by the additional detection of the emission spectrum and the fluorescence lifetime based on spatially and spectrally resolved time-resolved single photon counting. The novel 5D intravital tomograph (5D-IVT) was employed for the early detection of atopic dermatitis and the analysis of treatment effects.

Optimized respiratory-resolved motion-compensated 3D Cartesian coronary MR angiography.

PubMed

Correia, Teresa; Ginami, Giulia; Cruz, Gastão; Neji, Radhouene; Rashid, Imran; Botnar, René M; Prieto, Claudia

2018-04-22

To develop a robust and efficient reconstruction framework that provides high-quality motion-compensated respiratory-resolved images from free-breathing 3D whole-heart Cartesian coronary magnetic resonance angiography (CMRA) acquisitions. Recently, XD-GRASP (eXtra-Dimensional Golden-angle RAdial Sparse Parallel MRI) was proposed to achieve 100% scan efficiency and provide respiratory-resolved 3D radial CMRA images by exploiting sparsity in the respiratory dimension. Here, a reconstruction framework for Cartesian CMRA imaging is proposed, which provides respiratory-resolved motion-compensated images by incorporating 2D beat-to-beat translational motion information to increase sparsity in the respiratory dimension. The motion information is extracted from interleaved image navigators and is also used to compensate for 2D translational motion within each respiratory phase. The proposed Optimized Respiratory-resolved Cartesian Coronary MR Angiography (XD-ORCCA) method was tested on 10 healthy subjects and 2 patients with cardiovascular disease, and compared against XD-GRASP. The proposed XD-ORCCA provides high-quality respiratory-resolved images, allowing clear visualization of the right and left coronary arteries, even for irregular breathing patterns. Compared with XD-GRASP, the proposed method improves the visibility and sharpness of both coronaries. Significant differences (p < .05) in visible vessel length and proximal vessel sharpness were found between the 2 methods. The XD-GRASP method provides good-quality images in the absence of intraphase motion. However, motion blurring is observed in XD-GRASP images for respiratory phases with larger motion amplitudes and subjects with irregular breathing patterns. A robust respiratory-resolved motion-compensated framework for Cartesian CMRA has been proposed and tested in healthy subjects and patients. The proposed XD-ORCCA provides high-quality images for all respiratory phases, independently of the regularity of the breathing pattern. © 2018 The Authors Magnetic Resonance in Medicine published by Wiley Periodicals, Inc. on behalf of International Society for Magnetic Resonance in Medicine.

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

PubMed

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

2018-02-27

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

Docosahexaenoic Acid Signalolipidomics in Nutrition: Significance in Aging, Neuroinflammation, Macular Degeneration, Alzheimer’s, and Other Neurodegenerative Diseases

PubMed Central

Bazan, Nicolas G.; Molina, Miguel F.; Gordon, William C.

2012-01-01

Essential polyunsaturated fatty acids (PUFAs) are critical nutritional lipids that must be obtained from the diet to sustain homeostasis. Omega-3 and -6 PUFAs are key components of biomembranes and play important roles in cell integrity, development, maintenance, and function. The essential omega-3 fatty acid family member docosahexaenoic acid (DHA) is avidly retained and uniquely concentrated in the nervous system, particularly in photoreceptors and synaptic membranes. DHA plays a key role in vision, neuroprotection, successful aging, memory, and other functions. In addition, DHA displays anti-inflammatory and inflammatory resolving properties in contrast to the proinflammatory actions of several members of the omega-6 PUFAs family. This review discusses DHA signalolipidomics, comprising the cellular/tissue organization of DHA uptake, its distribution among cellular compartments, the organization and function of membrane domains rich in DHA-containing phospholipids, and the cellular and molecular events revealed by the uncovering of signaling pathways regulated by DHA and docosanoids, the DHA-derived bioactive lipids, which include neuroprotectin D1 (NPD1), a novel DHA-derived stereoselective mediator. NPD1 synthesis agonists include neurotrophins and oxidative stress; NPD1 elicits potent anti-inflammatory actions and prohomeostatic bioactivity, is anti-angiogenic, promotes corneal nerve regeneration, and induces cell survival. In the context of DHA signalolipidomics, this review highlights aging and the evolving studies on the significance of DHA in Alzheimer’s disease, macular degeneration, Parkinson’s disease, and other brain disorders. DHA signalolipidomics in the nervous system offers emerging targets for pharmaceutical intervention and clinical translation. PMID:21756134

Contrast-enhanced time-resolved 4D MRA of congenital heart and vessel anomalies: image quality and diagnostic value compared with 3D MRA.

PubMed

Vogt, Florian M; Theysohn, Jens M; Michna, Dariusz; Hunold, Peter; Neudorf, Ulrich; Kinner, Sonja; Barkhausen, Jörg; Quick, Harald H

2013-09-01

To evaluate time-resolved interleaved stochastic trajectories (TWIST) contrast-enhanced 4D magnetic resonance angiography (MRA) and compare it with 3D FLASH MRA in patients with congenital heart and vessel anomalies. Twenty-six patients with congenital heart and vessel anomalies underwent contrast-enhanced MRA with both 3D FLASH and 4D TWIST MRA. Images were subjectively evaluated regarding total image quality, artefacts, diagnostic value and added diagnostic value of 4D dynamic imaging. Quantitative comparison included signal-to-noise ratio (SNR), contrast-to-noise ratio (CNR) and vessel sharpness measurements. Three-dimensional FLASH MRA was judged to be significantly better in terms of image quality (4.0 ± 0.6 vs 3.4 ± 0.6, P < 0.05) and artefacts (3.8 ± 0.4 vs 3.3 ± 0.5, P < 0.05); no difference in diagnostic value was found (4.2 ± 0.4 vs 4.0 ± 0.4); important additional functional information was found in 21/26 patients. SNR and CNR were higher in the pulmonary trunk in 4D TWIST, but slightly higher in the systemic arteries in 3D FLASH. No difference in vessel sharpness delineation was found. Although image quality was inferior compared with 3D FLASH MRA, 4D TWIST MRA yields robust images and added diagnostic value through dynamic acquisition was found. Thus, 4D TWIST MRA is an attractive alternative to 3D FLASH MRA. • New magnetic resonance angiography (MRA) techniques are increasingly introduced for congenital cardiovascular problems. • Time-resolved angiography with interleaved stochastic trajectories (TWIST) is an example. • Four-dimensional TWIST MRA provided inferior image quality compared to 3D FLASH MRA but without significant difference in vessel sharpness. • Four-dimensional TWIST MRA gave added diagnostic value.

Recent Advances in 3D Time-Resolved Contrast-Enhanced MR Angiography

PubMed Central

Riederer, Stephen J.; Haider, Clifton R.; Borisch, Eric A.; Weavers, Paul T.; Young, Phillip M.

2015-01-01

Contrast-enhanced MR angiography (CE-MRA) was first introduced for clinical studies approximately 20 years ago. Early work provided 3 to 4 mm spatial resolution with acquisition times in the 30 sec range. Since that time there has been continuing effort to provide improved spatial resolution with reduced acquisition time, allowing high resolution three-dimensional (3D) time-resolved studies. The purpose of this work is to describe how this has been accomplished. Specific technical enablers have been: improved gradients allowing reduced repetition times, improved k-space sampling and reconstruction methods, parallel acquisition particularly in two directions, and improved and higher count receiver coil arrays. These have collectively made high resolution time-resolved studies readily available for many anatomic regions. Depending on the application, approximate 1 mm isotropic resolution is now possible with frame times of several seconds. Clinical applications of time-resolved CE-MRA are briefly reviewed. PMID:26032598

The third dimension bridges the gap between cell culture and live tissue.

PubMed

Pampaloni, Francesco; Reynaud, Emmanuel G; Stelzer, Ernst H K

2007-10-01

Moving from cell monolayers to three-dimensional (3D) cultures is motivated by the need to work with cellular models that mimic the functions of living tissues. Essential cellular functions that are present in tissues are missed by 'petri dish'-based cell cultures. This limits their potential to predict the cellular responses of real organisms. However, establishing 3D cultures as a mainstream approach requires the development of standard protocols, new cell lines and quantitative analysis methods, which include well-suited three-dimensional imaging techniques. We believe that 3D cultures will have a strong impact on drug screening and will also decrease the use of laboratory animals, for example, in the context of toxicity assays.

The 3-dimensional cellular automata for HIV infection

NASA Astrophysics Data System (ADS)

Mo, Youbin; Ren, Bin; Yang, Wencao; Shuai, Jianwei

2014-04-01

The HIV infection dynamics is discussed in detail with a 3-dimensional cellular automata model in this paper. The model can reproduce the three-phase development, i.e., the acute period, the asymptotic period and the AIDS period, observed in the HIV-infected patients in a clinic. We show that the 3D HIV model performs a better robustness on the model parameters than the 2D cellular automata. Furthermore, we reveal that the occurrence of a perpetual source to successively generate infectious waves to spread to the whole system drives the model from the asymptotic state to the AIDS state.

Computational Model of Secondary Palate Fusion and Disruption

EPA Science Inventory

Morphogenetic events are driven by cell-generated physical forces and complex cellular dynamics. To improve our capacity to predict developmental effects from cellular alterations, we built a multi-cellular agent-based model in CompuCell3D that recapitulates the cellular networks...

Smooth muscle-like tissue constructs with circumferentially oriented cells formed by the cell fiber technology.

PubMed

Hsiao, Amy Y; Okitsu, Teru; Onoe, Hiroaki; Kiyosawa, Mahiro; Teramae, Hiroki; Iwanaga, Shintaroh; Kazama, Tomohiko; Matsumoto, Taro; Takeuchi, Shoji

2015-01-01

The proper functioning of many organs and tissues containing smooth muscles greatly depends on the intricate organization of the smooth muscle cells oriented in appropriate directions. Consequently controlling the cellular orientation in three-dimensional (3D) cellular constructs is an important issue in engineering tissues of smooth muscles. However, the ability to precisely control the cellular orientation at the microscale cannot be achieved by various commonly used 3D tissue engineering building blocks such as spheroids. This paper presents the formation of coiled spring-shaped 3D cellular constructs containing circumferentially oriented smooth muscle-like cells differentiated from dedifferentiated fat (DFAT) cells. By using the cell fiber technology, DFAT cells suspended in a mixture of extracellular proteins possessing an optimized stiffness were encapsulated in the core region of alginate shell microfibers and uniformly aligned to the longitudinal direction. Upon differentiation induction to the smooth muscle lineage, DFAT cell fibers self-assembled to coiled spring structures where the cells became circumferentially oriented. By changing the initial core-shell microfiber diameter, we demonstrated that the spring pitch and diameter could be controlled. 21 days after differentiation induction, the cell fibers contained high percentages of ASMA-positive and calponin-positive cells. Our technology to create these smooth muscle-like spring constructs enabled precise control of cellular alignment and orientation in 3D. These constructs can further serve as tissue engineering building blocks for larger organs and cellular implants used in clinical treatments.

Smooth Muscle-Like Tissue Constructs with Circumferentially Oriented Cells Formed by the Cell Fiber Technology

PubMed Central

Hsiao, Amy Y.; Okitsu, Teru; Onoe, Hiroaki; Kiyosawa, Mahiro; Teramae, Hiroki; Iwanaga, Shintaroh; Kazama, Tomohiko; Matsumoto, Taro; Takeuchi, Shoji

2015-01-01

The proper functioning of many organs and tissues containing smooth muscles greatly depends on the intricate organization of the smooth muscle cells oriented in appropriate directions. Consequently controlling the cellular orientation in three-dimensional (3D) cellular constructs is an important issue in engineering tissues of smooth muscles. However, the ability to precisely control the cellular orientation at the microscale cannot be achieved by various commonly used 3D tissue engineering building blocks such as spheroids. This paper presents the formation of coiled spring-shaped 3D cellular constructs containing circumferentially oriented smooth muscle-like cells differentiated from dedifferentiated fat (DFAT) cells. By using the cell fiber technology, DFAT cells suspended in a mixture of extracellular proteins possessing an optimized stiffness were encapsulated in the core region of alginate shell microfibers and uniformly aligned to the longitudinal direction. Upon differentiation induction to the smooth muscle lineage, DFAT cell fibers self-assembled to coiled spring structures where the cells became circumferentially oriented. By changing the initial core-shell microfiber diameter, we demonstrated that the spring pitch and diameter could be controlled. 21 days after differentiation induction, the cell fibers contained high percentages of ASMA-positive and calponin-positive cells. Our technology to create these smooth muscle-like spring constructs enabled precise control of cellular alignment and orientation in 3D. These constructs can further serve as tissue engineering building blocks for larger organs and cellular implants used in clinical treatments. PMID:25734774

Identification of a flavonoid C-glycoside as potent antioxidant.

PubMed

Wen, Lingrong; Zhao, Yupeng; Jiang, Yueming; Yu, Limei; Zeng, Xiaofang; Yang, Jiali; Tian, Miaomiao; Liu, Huiling; Yang, Bao

2017-09-01

Flavonoids have been documented to have good antioxidant activities in vitro. However, reports on the cellular antioxidant activities of flavonoid C-glycosides are very limited. In this work, an apigenin C-glycoside was purified from Artocarpus heterophyllus by column chromatography and was identified to be 2″-O-β-D-xylosylvitexin by nuclear magnetic resonance spectroscopy. The cellular antioxidant activity and anticancer activity of 2″-O-β-D-xylosylvitexin were evaluated for the first time. The quantitative structure-activity relationship was analysed by molecular modeling. Apigenin presented an unexpected cellular antioxidation behaviour. It had an antioxidant activity at low concentration and a prooxidant activity at high concentration, whereas 2″-O-β-D-xylosylvitexin showed a dose-dependent cellular antioxidant activity. It indicated that C-glycosidation improved the cellular antioxidation performance of apigenin and eliminated the prooxidant effect. The ortho-dihydroxyl at C-3'/C-4' and C-3 hydroxyl in the flavonoid skeleton play important roles in the antioxidation behaviour. The cell proliferation assay revealed a low cytotoxicity of 2″-O-β-D-xylosylvitexin. Copyright © 2017 Elsevier Inc. All rights reserved.

Time-Resolved 3D Quantitative Flow MRI of the Major Intracranial Vessels: Initial Experience and Comparative Evaluation at 1.5T and 3.0T in Combination With Parallel Imaging

PubMed Central

Bammer, Roland; Hope, Thomas A.; Aksoy, Murat; Alley, Marcus T.

2012-01-01

Exact knowledge of blood flow characteristics in the major cerebral vessels is of great relevance for diagnosing cerebrovascular abnormalities. This involves the assessment of hemodynamically critical areas as well as the derivation of biomechanical parameters such as wall shear stress and pressure gradients. A time-resolved, 3D phase-contrast (PC) MRI method using parallel imaging was implemented to measure blood flow in three dimensions at multiple instances over the cardiac cycle. The 4D velocity data obtained from 14 healthy volunteers were used to investigate dynamic blood flow with the use of multiplanar reformatting, 3D streamlines, and 4D particle tracing. In addition, the effects of magnetic field strength, parallel imaging, and temporal resolution on the data were investigated in a comparative evaluation at 1.5T and 3T using three different parallel imaging reduction factors and three different temporal resolutions in eight of the 14 subjects. Studies were consistently performed faster at 3T than at 1.5T because of better parallel imaging performance. A high temporal resolution (65 ms) was required to follow dynamic processes in the intracranial vessels. The 4D flow measurements provided a high degree of vascular conspicuity. Time-resolved streamline analysis provided features that have not been reported previously for the intracranial vasculature. PMID:17195166

On the star partition dimension of comb product of cycle and complete graph

NASA Astrophysics Data System (ADS)

Alfarisi, Ridho; Darmaji; Dafik

2017-06-01

Let G = (V, E) be a connected graphs with vertex set V (G), edge set E(G) and S ⊆ V (G). For an ordered partition Π = {S 1, S 2, S 3, …, Sk } of V (G), the representation of a vertex v ∈ V (G) with respect to Π is the k-vectors r(v|Π) = (d(v, S 1), d(v, S 2), …, d(v, Sk )), where d(v, Sk ) represents the distance between the vertex v and the set Sk , defined by d(v, Sk ) = min{d(v, x)|x ∈ Sk}. The partition Π of V (G) is a resolving partition if the k-vektors r(v|Π), v ∈ V (G) are distinct. The minimum resolving partition Π is a partition dimension of G, denoted by pd(G). The resolving partition Π = {S 1, S 2, S 3, …, Sk} is called a star resolving partition for G if it is a resolving partition and each subgraph induced by Si , 1 ≤ i ≤ k, is a star. The minimum k for which there exists a star resolving partition of V (G) is the star partition dimension of G, denoted by spd(G). Finding a star partition dimension of G is classified to be a NP-Hard problem. Furthermore, the comb product between G and H, denoted by G ⊲ H, is a graph obtained by taking one copy of G and |V (G)| copies of H and grafting the i-th copy of H at the vertex o to the i-th vertex of G. By definition of comb product, we can say that V (G ⊲ H) = {(a, u)|a ∈ V (G), u ∈ V (H)} and (a, u)(b, v) ∈ E(G ⊲ H) whenever a = b and uv ∈ E(H), or ab ∈ E(G) and u = v = o. In this paper, we will study the star partition dimension of comb product of cycle and complete graph, namely Cn ⊲ Km and Km ⊲ Cn for n ≥ 3 and m ≥ 3.

Ethanol-induced disruption of Golgi apparatus morphology, primary neurite number and cellular orientation in developing cortical neurons

PubMed Central

Powrozek, Teresa A.; Olson, Eric C.

2012-01-01

Prenatal ethanol exposure disrupts cortical neurite initiation and outgrowth, but prior studies have reported both ethanol-dependent growth promotion and inhibition. To resolve this ambiguity and better approximate in vivo conditions, we quantitatively analyzed neuronal morphology using a new, whole hemisphere explant model. In this model, Layer 6 (L6) cortical neurons migrate, laminate and extend neurites in an organotypic fashion. To selectively label L6 neurons we performed ex utero electroporation of a GFP expression construct at embryonic day 13 and allowed the explants to develop for 2 days in vitro. Explants were exposed to (400mg/dL) ethanol for either 4 or 24 hrs prior to fixation. Complete 3-D reconstructions were made of >80 GFP-positive neurons in each experimental condition. Acute responses to ethanol exposure included compaction of the Golgi apparatus accompanied by elaboration of supernumerary primary apical neurites, as well as a modest (~15%) increase in higher order apical neurite length. With longer exposure time, ethanol exposure leads to a consistent, significant disorientation of the cell (cell body, primary apical neurite, and Golgi) with respect to the pial surface. The effects on cellular orientation were accompanied by decreased expression of cytoskeletal elements, microtubule associated protein 2 and F-actin. These findings indicate that upon exposure to ethanol, developing L6 neurons manifest disruptions in Golgi apparatus and cytoskeletal elements which may in turn trigger selective and significant perturbations to primary neurite formation and neuronal polarity. PMID:22840816

Autofluorescence imaging captures heterogeneous drug response differences between 2D and 3D breast cancer cultures

PubMed Central

Cannon, T. M.; Shah, A. T.; Skala, M. C.

2017-01-01

Two-photon microscopy of cellular autofluorescence intensity and lifetime (optical metabolic imaging, or OMI) is a promising tool for preclinical drug development. OMI, which exploits the endogenous fluorescence from the metabolic coenzymes NAD(P)H and FAD, is sensitive to changes in cell metabolism produced by drug treatment. Previous studies have shown that drug response, genetic expression, cell-cell communication, and cell signaling in 3D culture match those of the original in vivo tumor, but not those of 2D culture. The goal of this study is to use OMI to quantify dynamic cell-level metabolic differences in drug response in 2D cell lines vs. 3D organoids generated from xenograft tumors of the same cell origin. BT474 cells and Herceptin-resistant BT474 (HR6) cells were tested. Cells were treated with vehicle control, Herceptin, XL147 (PI3K inhibitor), and the combination. The OMI index was used to quantify response, and is a linear combination of the redox ratio (intensity of NAD(P)H divided by FAD), mean NADH lifetime, and mean FAD lifetime. The results confirm that the OMI index resolves significant differences (p<0.05) in drug response for 2D vs. 3D cultures, specifically for BT474 cells 24 hours after Herceptin treatment, for HR6 cells 24 and 72 hours after combination treatment, and for HR6 cells 72 hours after XL147 treatment. Cell-level analysis of the OMI index also reveals differences in the number of cell sub-populations in 2D vs. 3D culture at 24, 48, and 72 hours post-treatment in control and treated groups. Finally, significant increases (p<0.05) in the mean lifetime of NADH and FAD were measured in 2D vs. 3D for both cell lines at 72 hours post-treatment in control and all treatment groups. These whole-population differences in the mean NADH and FAD lifetimes are supported by differences in the number of cell sub-populations in 2D vs. 3D. Overall, these studies confirm that OMI is sensitive to differences in drug response in 2D vs. 3D, and provides further information on dynamic changes in the relative abundance of metabolic cell sub-populations that contribute to this difference. PMID:28663873

Immunogold labeling reveals subcellular localisation of silica nanoparticles in a human blood-brain barrier model

NASA Astrophysics Data System (ADS)

Ye, Dong; Anguissola, Sergio; O'Neill, Tiina; Dawson, Kenneth A.

2015-05-01

Subcellular location of nanoparticles has been widely investigated with fluorescence microscopy, via fluorescently labeled antibodies to visualise target antigens in cells. However, fluorescence microscopy, such as confocal or live cell imaging, has generally limited 3D spatial resolution. Conventional electron microscopy can be useful in bridging resolution gap, but still not ideal in resolving subcellular organelle identities. Using the pre-embedding immunogold electron microscopic imaging, we performed accurate examination of the intracellular trafficking and gathered further evidence of transport mechanisms of silica nanoparticles across a human in vitro blood-brain barrier model. Our approach can effectively immunolocalise a variety of intracellular compartments and provide new insights into the uptake and subcellular transport of nanoparticles.Subcellular location of nanoparticles has been widely investigated with fluorescence microscopy, via fluorescently labeled antibodies to visualise target antigens in cells. However, fluorescence microscopy, such as confocal or live cell imaging, has generally limited 3D spatial resolution. Conventional electron microscopy can be useful in bridging resolution gap, but still not ideal in resolving subcellular organelle identities. Using the pre-embedding immunogold electron microscopic imaging, we performed accurate examination of the intracellular trafficking and gathered further evidence of transport mechanisms of silica nanoparticles across a human in vitro blood-brain barrier model. Our approach can effectively immunolocalise a variety of intracellular compartments and provide new insights into the uptake and subcellular transport of nanoparticles. Electronic supplementary information (ESI) available: Nanoparticle characterisation data, preservation of cellular structures, staining controls, optimisation of size amplification via the silver enhancement, and more imaging results from anti-clathrin and anti-caveolin 1 immunolabeling. See DOI: 10.1039/c5nr01539a

Antibody-Mediated Rejection of the Kidney after Simultaneous Pancreas-Kidney Transplantation

PubMed Central

Pascual, Julio; Samaniego, Milagros D.; Torrealba, José R.; Odorico, Jon S.; Djamali, Arjang; Becker, Yolanda T.; Voss, Barbara; Leverson, Glen E.; Knechtle, Stuart J.; Sollinger, Hans W.; Pirsch, John D.

2008-01-01

The prevalence, risk factors, and outcome of antibody-mediated rejection (AMR) of the kidney after simultaneous pancreas-kidney transplantation are unknown. In 136 simultaneous pancreas-kidney recipients who were followed for an average of 3.1 yr, 21 episodes of AMR of the kidney allograft were identified. Eight episodes occurred early (≤90 d) after transplantation, and 13 occurred later. Histologic evidence of concomitant acute cellular rejection was noted in 12 cases; the other nine had evidence only of humoral rejection. In 13 cases, clinical rejection of the pancreas was diagnosed simultaneously, and two of these were biopsy proven and were positive for C4d immunostaining. Multivariate analysis identified only one significant risk factor: Female patients were three times more likely to experience AMR. Nearly all early episodes resolved with treatment and did not predict graft loss, but multivariate Cox models revealed that late AMR episodes more than tripled the risk for kidney and pancreas graft loss; therefore, new strategies are needed to prevent and to treat late AMR in simultaneous pancreas-kidney transplant recipients. PMID:18235091

3D Freeze-Casting of Cellular Graphene Films for Ultrahigh-Power-Density Supercapacitors.

PubMed

Shao, Yuanlong; El-Kady, Maher F; Lin, Cheng-Wei; Zhu, Guanzhou; Marsh, Kristofer L; Hwang, Jee Youn; Zhang, Qinghong; Li, Yaogang; Wang, Hongzhi; Kaner, Richard B

2016-08-01

3D cellular graphene films with open porosity, high electrical conductivity, and good tensile strength, can be synthesized by a method combining freeze-casting and filtration. The resulting supercapacitors based on 3D porous reduced graphene oxide (RGO) film exhibit extremely high specific power densities and high energy densities. The fabrication process provides an effective means for controlling the pore size, electronic conductivity, and loading mass of the electrode materials, toward devices with high energy-storage performance. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Three-dimensional super-resolved live cell imaging through polarized multi-angle TIRF.

PubMed

Zheng, Cheng; Zhao, Guangyuan; Liu, Wenjie; Chen, Youhua; Zhang, Zhimin; Jin, Luhong; Xu, Yingke; Kuang, Cuifang; Liu, Xu

2018-04-01

Measuring three-dimensional nanoscale cellular structures is challenging, especially when the structure is dynamic. Owing to the informative total internal reflection fluorescence (TIRF) imaging under varied illumination angles, multi-angle (MA) TIRF has been examined to offer a nanoscale axial and a subsecond temporal resolution. However, conventional MA-TIRF still performs badly in lateral resolution and fails to characterize the depth image in densely distributed regions. Here, we emphasize the lateral super-resolution in the MA-TIRF, exampled by simply introducing polarization modulation into the illumination procedure. Equipped with a sparsity and accelerated proximal algorithm, we examine a more precise 3D sample structure compared with previous methods, enabling live cell imaging with a temporal resolution of 2 s and recovering high-resolution mitochondria fission and fusion processes. We also shared the recovery program, which is the first open-source recovery code for MA-TIRF, to the best of our knowledge.

Two-dimensional and three-dimensional viability measurements of adult stem cells with optical coherence phase microscopy

NASA Astrophysics Data System (ADS)

Bagnaninchi, Pierre O.; Holmes, Christina; Drummond, Nicola; Daoud, Jamal; Tabrizian, Maryam

2011-08-01

Cell viability assays are essential tools for cell biology. They assess healthy cells in a sample and enable the quantification of cellular responses to reagents of interest. Noninvasive and label-free assays are desirable in two-dimensional (2D) and three-dimensional (3D) cell culture to facilitate time-course viability studies. Cellular micromotion, emanating from cell to substrate distance variations, has been demonstrated as a marker of cell viability with electric cell-substrate impedance sensing (ECIS). In this study we investigated if optical coherence phase microscopy (OCPM) was able to report phase fluctuations of adult stem cells in 2D and 3D that could be associated with cellular micromotion. An OCPM has been developed around a Thorlabs engine (λo = 930 nm) and integrated in an inverted microscope with a custom scanning head. Human adipose derived stem cells (ADSCs, Invitrogen) were cultured in Mesenpro RS medium and seeded either on ECIS arrays, 2D cell culture dishes, or in 3D highly porous microplotted polymeric scaffolds. ADSC micromotion was confirmed by ECIS analysis. Live and fixed ADSCs were then investigated in 2D and 3D with OCPM. Significant differences were found in phase fluctuations between the different conditions. This study indicated that OCPM could potentially assess cell vitality in 2D and in 3D microstructures.

Bioprinting of 3D hydrogels.

PubMed

Stanton, M M; Samitier, J; Sánchez, S

2015-08-07

Three-dimensional (3D) bioprinting has recently emerged as an extension of 3D material printing, by using biocompatible or cellular components to build structures in an additive, layer-by-layer methodology for encapsulation and culture of cells. These 3D systems allow for cell culture in a suspension for formation of highly organized tissue or controlled spatial orientation of cell environments. The in vitro 3D cellular environments simulate the complexity of an in vivo environment and natural extracellular matrices (ECM). This paper will focus on bioprinting utilizing hydrogels as 3D scaffolds. Hydrogels are advantageous for cell culture as they are highly permeable to cell culture media, nutrients, and waste products generated during metabolic cell processes. They have the ability to be fabricated in customized shapes with various material properties with dimensions at the micron scale. 3D hydrogels are a reliable method for biocompatible 3D printing and have applications in tissue engineering, drug screening, and organ on a chip models.

Low-dose AgNPs reduce lung mechanical function and innate immune defense in the absence of cellular toxicity.

PubMed

Botelho, Danielle J; Leo, Bey Fen; Massa, Christopher B; Sarkar, Srijata; Tetley, Terry D; Chung, Kian Fan; Chen, Shu; Ryan, Mary P; Porter, Alexandra E; Zhang, Junfeng; Schwander, Stephan K; Gow, Andrew J

2016-01-01

Multiple studies have examined the direct cellular toxicity of silver nanoparticles (AgNPs). However, the lung is a complex biological system with multiple cell types and a lipid-rich surface fluid; therefore, organ level responses may not depend on direct cellular toxicity. We hypothesized that interaction with the lung lining is a critical determinant of organ level responses. Here, we have examined the effects of low dose intratracheal instillation of AgNPs (0.05 μg/g body weight) 20 and 110 nm diameter in size, and functionalized with citrate or polyvinylpyrrolidone. Both size and functionalization were significant factors in particle aggregation and lipid interaction in vitro. One day post-intratracheal instillation lung function was assessed, and bronchoalveolar lavage (BAL) and lung tissue collected. There were no signs of overt inflammation. There was no change in surfactant protein-B content in the BAL but there was loss of surfactant protein-D with polyvinylpyrrolidone (PVP)-stabilized particles. Mechanical impedance data demonstrated a significant increase in pulmonary elastance as compared to control, greatest with 110 nm PVP-stabilized particles. Seven days post-instillation of PVP-stabilized particles increased BAL cell counts, and reduced lung function was observed. These changes resolved by 21 days. Hence, AgNP-mediated alterations in the lung lining and mechanical function resolve by 21 days. Larger particles and PVP stabilization produce the largest disruptions. These studies demonstrate that low dose AgNPs elicit deficits in both mechanical and innate immune defense function, suggesting that organ level toxicity should be considered.

Three-dimensional printing of Hela cells for cervical tumor model in vitro.

PubMed

Zhao, Yu; Yao, Rui; Ouyang, Liliang; Ding, Hongxu; Zhang, Ting; Zhang, Kaitai; Cheng, Shujun; Sun, Wei

2014-09-01

Advances in three-dimensional (3D) printing have enabled the direct assembly of cells and extracellular matrix materials to form in vitro cellular models for 3D biology, the study of disease pathogenesis and new drug discovery. In this study, we report a method of 3D printing for Hela cells and gelatin/alginate/fibrinogen hydrogels to construct in vitro cervical tumor models. Cell proliferation, matrix metalloproteinase (MMP) protein expression and chemoresistance were measured in the printed 3D cervical tumor models and compared with conventional 2D planar culture models. Over 90% cell viability was observed using the defined printing process. Comparisons of 3D and 2D results revealed that Hela cells showed a higher proliferation rate in the printed 3D environment and tended to form cellular spheroids, but formed monolayer cell sheets in 2D culture. Hela cells in 3D printed models also showed higher MMP protein expression and higher chemoresistance than those in 2D culture. These new biological characteristics from the printed 3D tumor models in vitro as well as the novel 3D cell printing technology may help the evolution of 3D cancer study.

3D Printing of Tissue Engineered Constructs for In Vitro Modeling of Disease Progression and Drug Screening.

PubMed

Vanderburgh, Joseph; Sterling, Julie A; Guelcher, Scott A

2017-01-01

2D cell culture and preclinical animal models have traditionally been implemented for investigating the underlying cellular mechanisms of human disease progression. However, the increasing significance of 3D vs. 2D cell culture has initiated a new era in cell culture research in which 3D in vitro models are emerging as a bridge between traditional 2D cell culture and in vivo animal models. Additive manufacturing (AM, also known as 3D printing), defined as the layer-by-layer fabrication of parts directed by digital information from a 3D computer-aided design file, offers the advantages of simultaneous rapid prototyping and biofunctionalization as well as the precise placement of cells and extracellular matrix with high resolution. In this review, we highlight recent advances in 3D printing of tissue engineered constructs that recapitulate the physical and cellular properties of the tissue microenvironment for investigating mechanisms of disease progression and for screening drugs.

3D Printing of Tissue Engineered Constructs for in vitro Modeling of Disease Progression and Drug Screening

PubMed Central

Vanderburgh, Joseph; Sterling, Julie A.

2016-01-01

2D cell culture and preclinical animal models have traditionally been implemented for investigating the underlying cellular mechanisms of human disease progression. However, the increasing significance of 3D versus 2D cell culture has initiated a new era in cell culture research in which 3D in vitro models are emerging as a bridge between traditional 2D cell culture and in vivo animal models. Additive manufacturing (AM, also known as 3D printing), defined as the layer-by-layer fabrication of parts directed by digital information from a 3D computer-aided design (CAD) file, offers the advantages of simultaneous rapid prototyping and biofunctionalization as well as the precise placement of cells and extracellular matrix with high resolution. In this review, we highlight recent advances in 3D printing of tissue engineered constructs (TECs) that recapitulate the physical and cellular properties of the tissue microenvironment for investigating mechanisms of disease progression and for screening drugs. PMID:27169894

Effect of 3D Cultivation Conditions on the Differentiation of Endodermal Cells

PubMed Central

Petrakova, O. S.; Ashapkin, V. V.; Voroteliak, E. A.; Bragin, E. Y.; Shtratnikova, V. Y.; Chernioglo, E. S.; Sukhanov, Y. V.; Terskikh, V. V.; Vasiliev, A. V.

2012-01-01

Cellular therapy of endodermal organs is one of the most important issues in modern cellular biology and biotechnology. One of the most promising directions in this field is the study of the transdifferentiation abilities of cells within the same germ layer. A method for anin vitroinvestigation of the cell differentiation potential (the cell culture in a three-dimensional matrix) is described in this article. Cell cultures of postnatal salivary gland cells and postnatal liver progenitor cells were obtained; their comparative analysis under 2D and 3D cultivation conditions was carried out. Both cell types have high proliferative abilities and can be cultivated for more than 20 passages. Under 2D cultivation conditions, the cells remain in an undifferentiated state. Under 3D conditions, they undergo differentiation, which was confirmed by a lower cell proliferation and by an increase in the differentiation marker expression. Salivary gland cells can undergo hepatic and pancreatic differentiation under 3D cultivation conditions. Liver progenitor cells also acquire a pancreatic differentiation capability under conditions of 3D cultivation. Thus, postnatal salivary gland cells exhibit a considerable differentiation potential within the endodermal germ layer and can be used as a promising source of endodermal cells for the cellular therapy of liver pathologies. Cultivation of cells under 3D conditions is a useful model for thein vitroanalysis of the cell differentiation potential. PMID:23346379

Local metric dimension of circulant graph c i r c (n :1 ,2 ,…,n/+1 2 )

NASA Astrophysics Data System (ADS)

Rimadhany, Ruzika; Darmaji

2017-08-01

Let G be a connected graph with two vertices u and v. The distance between u and v, denoted by d(u, v), is defined as length of the shortest path from u to v in G. For an ordered set W = {w1, w2, w3, … , wk} of k distinct vertices in a nontrivial connected graph G, the representation of a vertex v of V(G) respect to W is r(v|W) = (d(v, w1), d(v, w2), … , d(v, wk)). The set W is a resolving set of G if r(v|W) for each vertex v ∈ V(G) is distinct. A resolving set of minimum cardinality is a metric dimension and denoted by dim(G). The set W is a local resolving set of G if r(v|W) for every two adjacent vertices of V(G) is distinct. The minimum cardinality of local resolving set of G is a local metric dimension and denoted by ldim(G). In this research, we determine local metric dimension of circulant graph c i r c (n :1 ,2 ,3 ,…,n/+1 2 ) .

Stereological estimation of cell wall density of DR12 tomato mutant using three-dimensional confocal imaging

PubMed Central

Legland, David; Guillon, Fabienne; Kiêu, Kiên; Bouchet, Brigitte; Devaux, Marie-Françoise

2010-01-01

Background and Aims The cellular structure of fleshy fruits is of interest to study fruit shape, size, mechanical behaviour or sensory texture. The cellular structure is usually not observed in the whole fruit but, instead, in a sample of limited size and volume. It is therefore difficult to extend measurements to the whole fruit and/or to a specific genotype, or to describe the cellular structure heterogeneity within the fruit. Methods An integrated method is presented to describe the cellular structure of the whole fruit from partial three-dimensional (3D) observations, involving the following steps: (1) fruit sampling, (2) 3D image acquisition and processing and (3) measurement and estimation of relevant 3D morphological parameters. This method was applied to characterize DR12 mutant and wild-type tomatoes (Solanum lycopersicum). Key Results The cellular structure was described using the total volume of the pericarp, the surface area of the cell walls and the ratio of cell-wall surface area to pericarp volume, referred to as the cell-wall surface density. The heterogeneity of cellular structure within the fruit was investigated by estimating variations in the cell-wall surface density with distance to the epidermis. Conclusions The DR12 mutant presents a greater pericarp volume and an increase of cell-wall surface density under the epidermis. PMID:19952012

Regulation of cellular growth by the Drosophila target of rapamycin dTOR

PubMed Central

Zhang, Hongbing; Stallock, James P.; Ng, Joyce C.; Reinhard, Christoph; Neufeld, Thomas P.

2000-01-01

The TOR protein kinases (TOR1 and TOR2 in yeast; mTOR/FRAP/RAFT1 in mammals) promote cellular proliferation in response to nutrients and growth factors, but their role in development is poorly understood. Here, we show that the Drosophila TOR homolog dTOR is required cell autonomously for normal growth and proliferation during larval development, and for increases in cellular growth caused by activation of the phosphoinositide 3-kinase (PI3K) signaling pathway. As in mammalian cells, the kinase activity of dTOR is required for growth factor-dependent phosphorylation of p70 S6 kinase (p70S6K) in vitro, and we demonstrate that overexpression of p70S6K in vivo can rescue dTOR mutant animals to viability. Loss of dTOR also results in cellular phenotypes characteristic of amino acid deprivation, including reduced nucleolar size, lipid vesicle aggregation in the larval fat body, and a cell type-specific pattern of cell cycle arrest that can be bypassed by overexpression of the S-phase regulator cyclin E. Our results suggest that dTOR regulates growth during animal development by coupling growth factor signaling to nutrient availability. PMID:11069888

Label free cell tracking in 3D tissue engineering constructs with high resolution imaging

NASA Astrophysics Data System (ADS)

Smith, W. A.; Lam, K.-P.; Dempsey, K. P.; Mazzocchi-Jones, D.; Richardson, J. B.; Yang, Y.

2014-02-01

Within the field of tissue engineering there is an emphasis on studying 3-D live tissue structures. Consequently, to investigate and identify cellular activities and phenotypes in a 3-D environment for all in vitro experiments, including shape, migration/proliferation and axon projection, it is necessary to adopt an optical imaging system that enables monitoring 3-D cellular activities and morphology through the thickness of the construct for an extended culture period without cell labeling. This paper describes a new 3-D tracking algorithm developed for Cell-IQ®, an automated cell imaging platform, which has been equipped with an environmental chamber optimized to enable capturing time-lapse sequences of live cell images over a long-term period without cell labeling. As an integral part of the algorithm, a novel auto-focusing procedure was developed for phase contrast microscopy equipped with 20x and 40x objectives, to provide a more accurate estimation of cell growth/trajectories by allowing 3-D voxels to be computed at high spatiotemporal resolution and cell density. A pilot study was carried out in a phantom system consisting of horizontally aligned nanofiber layers (with precise spacing between them), to mimic features well exemplified in cellular activities of neuronal growth in a 3-D environment. This was followed by detailed investigations concerning axonal projections and dendritic circuitry formation in a 3-D tissue engineering construct. Preliminary work on primary animal neuronal cells in response to chemoattractant and topographic cue within the scaffolds has produced encouraging results.

Three-dimensional morphogenesis of MDCK cells induced by cellular contractile forces on a viscous substrate

PubMed Central

Imai, Misako; Furusawa, Kazuya; Mizutani, Takeomi; Kawabata, Kazushige; Haga, Hisashi

2015-01-01

Substrate physical properties are essential for many physiological events such as embryonic development and 3D tissue formation. Physical properties of the extracellular matrix such as viscoelasticity and geometrical constraints are understood as factors that affect cell behaviour. In this study, we focused on the relationship between epithelial cell 3D morphogenesis and the substrate viscosity. We observed that Madin-Darby Canine Kidney (MDCK) cells formed 3D structures on a viscous substrate (Matrigel). The structures appear as a tulip hat. We then changed the substrate viscosity by genipin (GP) treatment. GP is a cross-linker of amino groups. Cells cultured on GP-treated-matrigel changed their 3D morphology in a substrate viscosity-dependent manner. Furthermore, to elucidate the spatial distribution of the cellular contractile force, localization of mono-phosphorylated and di-phosphorylated myosin regulatory light chain (P-MRLCs) was visualized by immunofluorescence. P-MRLCs localized along the periphery of epithelial sheets. Treatment with Y-27632, a Rho-kinase inhibitor, blocked the P-MRLCs localization at the edge of epithelial sheets and halted 3D morphogenesis. Our results indicate that the substrate viscosity, the substrate deformation, and the cellular contractile forces induced by P-MRLCs play crucial roles in 3D morphogenesis. PMID:26374384

Three-dimensional morphogenesis of MDCK cells induced by cellular contractile forces on a viscous substrate.

PubMed

Imai, Misako; Furusawa, Kazuya; Mizutani, Takeomi; Kawabata, Kazushige; Haga, Hisashi

2015-09-16

Substrate physical properties are essential for many physiological events such as embryonic development and 3D tissue formation. Physical properties of the extracellular matrix such as viscoelasticity and geometrical constraints are understood as factors that affect cell behaviour. In this study, we focused on the relationship between epithelial cell 3D morphogenesis and the substrate viscosity. We observed that Madin-Darby Canine Kidney (MDCK) cells formed 3D structures on a viscous substrate (Matrigel). The structures appear as a tulip hat. We then changed the substrate viscosity by genipin (GP) treatment. GP is a cross-linker of amino groups. Cells cultured on GP-treated-matrigel changed their 3D morphology in a substrate viscosity-dependent manner. Furthermore, to elucidate the spatial distribution of the cellular contractile force, localization of mono-phosphorylated and di-phosphorylated myosin regulatory light chain (P-MRLCs) was visualized by immunofluorescence. P-MRLCs localized along the periphery of epithelial sheets. Treatment with Y-27632, a Rho-kinase inhibitor, blocked the P-MRLCs localization at the edge of epithelial sheets and halted 3D morphogenesis. Our results indicate that the substrate viscosity, the substrate deformation, and the cellular contractile forces induced by P-MRLCs play crucial roles in 3D morphogenesis.

Luminescent single-walled carbon nanotube-sensitized europium nanoprobes for cellular imaging

PubMed Central

Avti, Pramod K; Sitharaman, Balaji

2012-01-01

Lanthanoid-based optical probes with excitation wavelengths in the ultra-violet (UV) range (300–325 nm) have been widely developed as imaging probes. Efficient cellular imaging requires that lanthanoid optical probes be excited at visible wavelengths, to avoid UV damage to cells. The efficacy of europium-catalyzed single-walled carbon nanotubes (Eu-SWCNTs), as visible nanoprobes for cellular imaging, is reported in this study. Confocal fluorescence microscopy images of breast cancer cells (SK-BR-3 and MCF-7) and normal cells (NIH 3T3), treated with Eu-SWCNT at 0.2 μg/mL concentration, showed bright red luminescence after excitation at 365 nm and 458 nm wavelengths. Cell viability analysis showed no cytotoxic effects after the incubation of cells with Eu-SWCNTs at this concentration. Eu-SWCNT uptake is via the endocytosis mechanism. Labeling efficiency, defined as the percentage of incubated cells that uptake Eu-SWCNT, was 95%–100% for all cell types. The average cellular uptake concentration was 6.68 ng Eu per cell. Intracellular localization was further corroborated by transmission electron microscopy and Raman microscopy. The results indicate that Eu-SWCNT shows potential as a novel cellular imaging probe, wherein SWCNT sensitizes Eu3+ ions to allow excitation at visible wavelengths, and stable time-resolved red emission. The ability to functionalize biomolecules on the exterior surface of Eu-SWCNT makes it an excellent candidate for targeted cellular imaging. PMID:22619533

Decoupling diffusional from dimensional control of signaling in 3D culture reveals a role for myosin in tubulogenesis

PubMed Central

Raghavan, Srivatsan; Shen, Colette J.; Desai, Ravi A.; Sniadecki, Nathan J.; Nelson, Celeste M.; Chen, Christopher S.

2010-01-01

We present a novel microfabricated platform to culture cells within arrays of micrometer-scale three-dimensional (3D) extracellular matrix scaffolds (microgels). These microscale cultures eliminate diffusion barriers that are intrinsic to traditional 3D culture systems (macrogels) and enable uniform cytokine stimulation of the entire culture population, as well as allow immunolabeling, imaging and population-based biochemical assays across the relatively coplanar microgels. Examining early signaling associated with hepatocyte growth factor (HGF)-mediated scattering and tubulogenesis of MDCK cells revealed that 3D culture modulates cellular responses both through dimensionality and altered stimulation rates. Comparing responses in 2D culture, microgels and macrogels demonstrated that HGF-induced ERK signaling was driven by the dynamics of stimulation and not by whether cells were in a 2D or 3D environment, and that this ERK signaling was equally important for HGF-induced cell scattering on 2D substrates and tubulogenesis in 3D. By contrast, we discovered a specific HGF-induced increase in myosin expression leading to sustained downregulation of myosin activity that occurred only within 3D contexts and was required for 3D tubulogenesis but not 2D scattering. Interestingly, although absent in cells on collagen-coated plates, downregulation of myosin activity also occurred for cells on collagen gels, but was transient and mediated by a combination of myosin dephosphorylation and enhanced myosin expression. Furthermore, upregulating myosin activity via siRNA targeted to a myosin phosphatase did not attenuate scattering in 2D but did inhibit tubulogenesis in 3D. Together, these results demonstrate that cellular responses to soluble cues in 3D culture are regulated by both rates of stimulation and by matrix dimensionality, and highlight the importance of decoupling these effects to identify early signals relevant to cellular function in 3D environments. PMID:20682635

Efficient non-hydrostatic modelling of 3D wave-induced currents using a subgrid approach

NASA Astrophysics Data System (ADS)

Rijnsdorp, Dirk P.; Smit, Pieter B.; Zijlema, Marcel; Reniers, Ad J. H. M.

2017-08-01

Wave-induced currents are an ubiquitous feature in coastal waters that can spread material over the surf zone and the inner shelf. These currents are typically under resolved in non-hydrostatic wave-flow models due to computational constraints. Specifically, the low vertical resolutions adequate to describe the wave dynamics - and required to feasibly compute at the scales of a field site - are too coarse to account for the relevant details of the three-dimensional (3D) flow field. To describe the relevant dynamics of both wave and currents, while retaining a model framework that can be applied at field scales, we propose a two grid approach to solve the governing equations. With this approach, the vertical accelerations and non-hydrostatic pressures are resolved on a relatively coarse vertical grid (which is sufficient to accurately resolve the wave dynamics), whereas the horizontal velocities and turbulent stresses are resolved on a much finer subgrid (of which the resolution is dictated by the vertical scale of the mean flows). This approach ensures that the discrete pressure Poisson equation - the solution of which dominates the computational effort - is evaluated on the coarse grid scale, thereby greatly improving efficiency, while providing a fine vertical resolution to resolve the vertical variation of the mean flow. This work presents the general methodology, and discusses the numerical implementation in the SWASH wave-flow model. Model predictions are compared with observations of three flume experiments to demonstrate that the subgrid approach captures both the nearshore evolution of the waves, and the wave-induced flows like the undertow profile and longshore current. The accuracy of the subgrid predictions is comparable to fully resolved 3D simulations - but at much reduced computational costs. The findings of this work thereby demonstrate that the subgrid approach has the potential to make 3D non-hydrostatic simulations feasible at the scale of a realistic coastal region.

Generalized thick strip modelling for vortex-induced vibration of long flexible cylinders

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

Bao, Y., E-mail: ybao@sjtu.edu.cn; Department of Aeronautics, Imperial College London, South Kensington Campus, London; Palacios, R., E-mail: r.palacios@imperial.ac.uk

2016-09-15

We propose a generalized strip modelling method that is computationally efficient for the VIV prediction of long flexible cylinders in three-dimensional incompressible flow. In order to overcome the shortcomings of conventional strip-theory-based 2D models, the fluid domain is divided into “thick” strips, which are sufficiently thick to locally resolve the small scale turbulence effects and three dimensionality of the flow around the cylinder. An attractive feature of the model is that we independently construct a three-dimensional scale resolving model for individual strips, which have local spanwise scale along the cylinder's axial direction and are only coupled through the structural modelmore » of the cylinder. Therefore, this approach is able to cover the full spectrum for fully resolved 3D modelling to 2D strip theory. The connection between these strips is achieved through the calculation of a tensioned beam equation, which is used to represent the dynamics of the flexible body. In the limit, however, a single “thick” strip would fill the full 3D domain. A parallel Fourier spectral/hp element method is employed to solve the 3D flow dynamics in the strip-domain, and then the VIV response prediction is achieved through the strip–structure interactions. Numerical tests on both laminar and turbulent flows as well as the comparison against the fully resolved DNS are presented to demonstrate the applicability of this approach.« less

Cellular Microcultures: Programming Mechanical and Physicochemical Properties of 3D Hydrogel Cellular Microcultures via Direct Ink Writing (Adv. Healthcare Mater. 9/2016).

PubMed

McCracken, Joselle M; Badea, Adina; Kandel, Mikhail E; Gladman, A Sydney; Wetzel, David J; Popescu, Gabriel; Lewis, Jennifer A; Nuzzo, Ralph G

2016-05-01

R. Nuzzo and co-workers show on page 1025 how compositional differences in hydrogels are used to tune their cellular compliance by controlling their polymer mesh properties and subsequent uptake of the protein poly-l-lysine (green spheres in circled inset). The cover image shows pyramid micro-scaffolds prepared using direct ink writing (DIW) that differentially direct fibroblast and preosteoblast growth in 3D, depending on cell motility and surface treatment. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Precipitation Processes Developed During ARM (1997), TOGA COARE (1992) GATE (1974), SCSMEX (1998), and KWAJEX (1999): Consistent 3D, Semi-3D and 3D Cloud Resolving Model Simulations

NASA Technical Reports Server (NTRS)

Tao, W.-K.; Hou, A.; Atlas, R.; Starr, D.; Sud, Y.

2003-01-01

Real clouds and cloud systems are inherently three-dimensional (3D). Because of the limitations in computer resources, however, most cloud-resolving models (CRMs) today are still two-dimensional (2D) have been used to study the response of clouds to large-scale forcing. IN these 3D simulators, the model domain was small, and the integration time was 6 hours. Only recently have 3D experiments been performed for multi-day periods for tropical clouds systems with large horizontal domains at the National Center of Atmospheric Research (NCAR) and at NASA Goddard Space Center. At Goddard, a 3D cumulus Ensemble (GCE) model was used to simulate periods during TOGA COARE, GATE, SCSMEX, ARM, and KWAJEX using a 512 by 512 km domain (with 2-km resolution). The result indicate that surface precipitation and latent heating profiles are very similar between the 2D and 3D GCE model simulation. The major objective of this paper are: (1) to assess the performance of the super-parametrization technique, (2) calculate and examine the surface energy (especially radiation) and water budget, and (3) identify the differences and similarities in the organization and entrainment rates of convection between simulated 2D and 3D cloud systems.

Magnetic Resonance Angiography in the Diagnosis of Cerebral Arteriovenous Malformation and Dural Arteriovenous Fistulas: Comparison of Time-Resolved Magnetic Resonance Angiography and Three Dimensional Time-of-Flight Magnetic Resonance Angiography

PubMed Central

Cheng, Yu-Ching; Chen, Hung-Chieh; Wu, Chen-Hao; Wu, Yi-Ying; Sun, Ming-His; Chen, Wen-Hsien; Chai, Jyh-Wen; Chi-Chang Chen, Clayton

2016-01-01

Background Traditional digital subtraction angiography (DSA) is currently the gold standard diagnostic method for the diagnosis and evaluation of cerebral arteriovenous malformation (AVM) and dural arteriovenous fistulas (dAVF). Objectives The aim of this study was to analyze different less invasive magnetic resonance angiography (MRA) images, time-resolved MRA (TR-MRA) and three-dimensional time-of-flight MRA (3D TOF MRA) to identify their diagnostic accuracy and to determine which approach is most similar to DSA. Patients and Methods A total of 41 patients with AVM and dAVF at their initial evaluation or follow-up after treatment were recruited in this study. We applied time-resolved angiography using keyhole (4D-TRAK) MRA to perform TR-MRA and 3D TOF MRA examinations simultaneously followed by DSA, which was considered as a standard reference. Two experienced neuroradiologists reviewed the images to compare the diagnostic accuracy, arterial feeder and venous drainage between these two MRA images. Inter-observer agreement for different MRA images was assessed by Kappa coefficient and the differences of diagnostic accuracy between MRA images were evaluated by the Wilcoxon rank sum test. Results Almost all vascular lesions (92.68%) were correctly diagnosed using 4D-TRAK MRA. However, 3D TOF MRA only diagnosed 26 patients (63.41%) accurately. There were statistically significant differences regarding lesion diagnostic accuracy (P = 0.008) and venous drainage identification (P < 0.0001) between 4D-TRAK MRA and 3D TOF MRA. The results indicate that 4D-TRAK MRA is superior to 3D TOF MRA in the assessment of lesions. Conclusion Compared with 3D TOF MRA, 4D-TRAK MRA proved to be a more reliable screening modality and follow-up method for the diagnosis of cerebral AVM and dAVF. PMID:27679690

Display dimensionality and conflict geometry effects on maneuver preferences for resolving in-flight conflicts.

PubMed

Thomas, Lisa C; Wickens, Christopher D

2008-08-01

Two experiments explored the effects of display dimensionality, conflict geometry, and time pressure on pilot maneuvering preferences for resolving en route conflicts. With the presence of a cockpit display of traffic information (CDTI) that provides graphical airspace information, pilots can use a variety of conflict resolution maneuvers in response to how they perceive the conflict. Inconsistent preference findings from previous research on conflict resolution using CDTIs may be attributable to inherent ambiguities in 3-D perspective displays and/or a limited range of conflict geometries. Pilots resolved predicted conflicts using CDTIs with three levels of display dimensionality; the first had two 2-D orthogonal views, the second depicted the airspace in two alternating 3-D perspective views, and the third had a pilot-controlled swiveling viewpoint. Pilots demonstrated the same preferences that have been observed in previous research for vertical over lateral maneuvers in low workload and climbs over descents for level-flight conflicts. With increasing workload the two 3-D perspective displays, but not the 2-D displays, resulted in an increased preference for lateral over vertical maneuvers. Increased time pressure resulted in increased vertical maneuvers, an effect again limited to the two 3-D perspective displays. Resolution preferences were more affected by workload and time pressure when the 3-D perspective displays were used, as compared with the 2-D displays, although overall preferences were milder than in previous studies. Investigating maneuver preferences using the strategic flight planning paradigm employed in this study may be the key to better ensure pilot acceptance of computer-generated resolution maneuvers.

High Resolution, Large Deformation 3D Traction Force Microscopy

PubMed Central

López-Fagundo, Cristina; Reichner, Jonathan; Hoffman-Kim, Diane; Franck, Christian

2014-01-01

Traction Force Microscopy (TFM) is a powerful approach for quantifying cell-material interactions that over the last two decades has contributed significantly to our understanding of cellular mechanosensing and mechanotransduction. In addition, recent advances in three-dimensional (3D) imaging and traction force analysis (3D TFM) have highlighted the significance of the third dimension in influencing various cellular processes. Yet irrespective of dimensionality, almost all TFM approaches have relied on a linear elastic theory framework to calculate cell surface tractions. Here we present a new high resolution 3D TFM algorithm which utilizes a large deformation formulation to quantify cellular displacement fields with unprecedented resolution. The results feature some of the first experimental evidence that cells are indeed capable of exerting large material deformations, which require the formulation of a new theoretical TFM framework to accurately calculate the traction forces. Based on our previous 3D TFM technique, we reformulate our approach to accurately account for large material deformation and quantitatively contrast and compare both linear and large deformation frameworks as a function of the applied cell deformation. Particular attention is paid in estimating the accuracy penalty associated with utilizing a traditional linear elastic approach in the presence of large deformation gradients. PMID:24740435

Respiratory motion resolved, self-gated 4D-MRI using Rotating Cartesian K-space (ROCK)

PubMed Central

Han, Fei; Zhou, Ziwu; Cao, Minsong; Yang, Yingli; Sheng, Ke; Hu, Peng

2017-01-01

Purpose To propose and validate a respiratory motion resolved, self-gated (SG) 4D-MRI technique to assess patient-specific breathing motion of abdominal organs for radiation treatment planning. Methods The proposed 4D-MRI technique was based on the balanced steady-state free-precession (bSSFP) technique and 3D k-space encoding. A novel ROtating Cartesian K-space (ROCK) reordering method was designed that incorporates repeatedly sampled k-space centerline as the SG motion surrogate and allows for retrospective k-space data binning into different respiratory positions based on the amplitude of the surrogate. The multiple respiratory-resolved 3D k-space data were subsequently reconstructed using a joint parallel imaging and compressed sensing method with spatial and temporal regularization. The proposed 4D-MRI technique was validated using a custom-made dynamic motion phantom and was tested in 6 healthy volunteers, in whom quantitative diaphragm and kidney motion measurements based on 4D-MRI images were compared with those based on 2D-CINE images. Results The 5-minute 4D-MRI scan offers high-quality volumetric images in 1.2×1.2×1.6mm3 and 8 respiratory positions, with good soft-tissue contrast. In phantom experiments with triangular motion waveform, the motion amplitude measurements based on 4D-MRI were 11.89% smaller than the ground truth, whereas a −12.5% difference was expected due to data binning effects. In healthy volunteers, the difference between the measurements based on 4D-MRI and the ones based on 2D-CINE were 6.2±4.5% for the diaphragm, 8.2±4.9% and 8.9±5.1% for the right and left kidney. Conclusion The proposed 4D-MRI technique could provide high resolution, high quality, respiratory motion resolved 4D images with good soft-tissue contrast and are free of the “stitching” artifacts usually seen on 4D-CT and 4D-MRI based on resorting 2D-CINE. It could be used to visualize and quantify abdominal organ motion for MRI-based radiation treatment planning. PMID:28133752

Respiratory motion-resolved, self-gated 4D-MRI using rotating cartesian k-space (ROCK).

PubMed

Han, Fei; Zhou, Ziwu; Cao, Minsong; Yang, Yingli; Sheng, Ke; Hu, Peng

2017-04-01

To propose and validate a respiratory motion resolved, self-gated (SG) 4D-MRI technique to assess patient-specific breathing motion of abdominal organs for radiation treatment planning. The proposed 4D-MRI technique was based on the balanced steady-state free-precession (bSSFP) technique and 3D k-space encoding. A novel rotating cartesian k-space (ROCK) reordering method was designed which incorporates repeatedly sampled k-space centerline as the SG motion surrogate and allows for retrospective k-space data binning into different respiratory positions based on the amplitude of the surrogate. The multiple respiratory-resolved 3D k-space data were subsequently reconstructed using a joint parallel imaging and compressed sensing method with spatial and temporal regularization. The proposed 4D-MRI technique was validated using a custom-made dynamic motion phantom and was tested in six healthy volunteers, in whom quantitative diaphragm and kidney motion measurements based on 4D-MRI images were compared with those based on 2D-CINE images. The 5-minute 4D-MRI scan offers high-quality volumetric images in 1.2 × 1.2 × 1.6 mm 3 and eight respiratory positions, with good soft-tissue contrast. In phantom experiments with triangular motion waveform, the motion amplitude measurements based on 4D-MRI were 11.89% smaller than the ground truth, whereas a -12.5% difference was expected due to data binning effects. In healthy volunteers, the difference between the measurements based on 4D-MRI and the ones based on 2D-CINE were 6.2 ± 4.5% for the diaphragm, 8.2 ± 4.9% and 8.9 ± 5.1% for the right and left kidney. The proposed 4D-MRI technique could provide high-resolution, high-quality, respiratory motion-resolved 4D images with good soft-tissue contrast and are free of the "stitching" artifacts usually seen on 4D-CT and 4D-MRI based on resorting 2D-CINE. It could be used to visualize and quantify abdominal organ motion for MRI-based radiation treatment planning. © 2017 American Association of Physicists in Medicine.

Uterine flushing proteome of the tammar wallaby after reactivation from diapause.

PubMed

Martin, Florine C; Ang, Ching-Seng; Gardner, David K; Renfree, Marilyn B; Shaw, Geoff

2016-11-01

The marsupial tammar wallaby has the longest period of embryonic diapause of any mammal, up to 11 months, during which there is no cell division or blastocyst growth. Since the blastocyst in diapause is surrounded by acellular coats, the signals that maintain or terminate diapause involve factors that reside in uterine secretions. The nature of such factors remains to be resolved. In this study, uterine flushings (UFs) were used to assess changes in uterine secretions of tammars using liquid chromatography-mass spectrometry (LC-MS/MS) during diapause (day 0 and 3) and reactivation days (d) 4, 5, 6, 8, 9, 11 and 24 after removal of pouch young (RPY), which initiates embryonic development. This study supports earlier suggestions that the presence of specific factors stimulate reactivation, early embryonic growth and cell proliferation. A mitogen, hepatoma-derived growth factor and soluble epidermal growth factor receptors were observed from d3 until at least d11 RPY when these secreted proteins constituted 21% of the UF proteome. Binding of these factors to specific cellular receptors or growth factors may directly stimulate DNA synthesis and division in endometrial gland cells. Proteins involved in the p53/CDKN1A (p21) cell cycle inhibition pathway were also observed in the diapause samples. Progesterone and most of the oestrogen-regulated proteins were present in the UF after d3, which is concomitant with the start of blastocyst mitoses at d4. We propose that once the p21 inhibition of the cell cycle is lost, growth factors including HDGF and EGFR are responsible for reactivation of the diapausing blastocyst via the uterine secretions. © 2016 Society for Reproduction and Fertility.

Quantifying the correlation between spatially defined oxygen gradients and cell fate in an engineered three-dimensional culture model.

PubMed

Ardakani, Amir G; Cheema, Umber; Brown, Robert A; Shipley, Rebecca J

2014-09-06

A challenge in three-dimensional tissue culture remains the lack of quantitative information linking nutrient delivery and cellular distribution. Both in vivo and in vitro, oxygen is delivered by diffusion from its source (blood vessel or the construct margins). The oxygen level at a defined distance from its source depends critically on the balance of diffusion and cellular metabolism. Cells may respond to this oxygen environment through proliferation, death and chemotaxis, resulting in spatially resolved gradients in cellular density. This study extracts novel spatially resolved and simultaneous data on tissue oxygenation, cellular proliferation, viability and chemotaxis in three-dimensional spiralled, cellular collagen constructs. Oxygen concentration gradients drove preferential cellular proliferation rates and viability in the higher oxygen zones and induced chemotaxis along the spiral of the collagen construct; an oxygen gradient of 1.03 mmHg mm(-1) in the spiral direction induced a mean migratory speed of 1015 μm day(-1). Although this movement was modest, it was effective in balancing the system to a stable cell density distribution, and provided insights into the natural cell mechanism for adapting cell number and activity to a prevailing oxygen regime.

Geometric phase effects in the ultracold D + HD $$ \\rightarrow $$ D + HD and D + HD $$\\leftrightarrow $$ H + D 2 reactions

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

Kendrick, Brian Kent; Hazra, Jisha; Balakrishnan, Naduvaluth

The results of accurate quantum reactive scattering calculations for the D + HD(v = 4, j = 0)more » $$\\to $$ D + HD($$v^{\\prime} $$, $$j^{\\prime} $$), D + HD(v = 4, j = 0) $$\\to $$ H + D2($$v^{\\prime} $$, $$j^{\\prime} $$) and H + D2(v = 4, j = 0) $$\\to $$ D + HD($$v^{\\prime} $$, $$j^{\\prime} $$) reactions are presented for collision energies between $$1\\,\\mu {\\rm{K}}$$ and $$100\\,{\\rm{K}}$$. The ab initio BKMP2 PES for the ground electronic state of H3 is used and all values of total angular momentum between $J=0-4$ are included. The general vector potential approach is used to include the geometric phase. The rotationally resolved, vibrationally resolved, and total reaction rate coefficients are reported as a function of collision energy. Rotationally resolved differential cross sections are also reported as a function of collision energy and scattering angle. Large geometric phase effects appear in the ultracold reaction rate coefficients which result in a significant enhancement or suppression of the rate coefficient (up to 3 orders of magnitude) relative to calculations which ignore the geometric phase. The results are interpreted using a new quantum interference mechanism which is unique to ultracold collisions. Significant effects of the geometric phase also appear in the rotationally resolved differential cross sections which lead to a very different oscillatory structure in both energy and scattering angle. Several shape resonances occur in the 1–$$10\\,{\\rm{K}}$$ energy range and the geometric phase is shown to significantly alter the predicted resonance spectrum. The geometric phase effects and ultracold rate coefficients depend sensitively on the nuclear spin. Furthermore, experimentalists may be able to control the reaction by the selection of a particular nuclear spin state.« less

Geometric phase effects in the ultracold D + HD $$ \\rightarrow $$ D + HD and D + HD $$\\leftrightarrow $$ H + D 2 reactions

DOE PAGES

Kendrick, Brian Kent; Hazra, Jisha; Balakrishnan, Naduvaluth

2016-12-15

The results of accurate quantum reactive scattering calculations for the D + HD(v = 4, j = 0)more » $$\\to $$ D + HD($$v^{\\prime} $$, $$j^{\\prime} $$), D + HD(v = 4, j = 0) $$\\to $$ H + D2($$v^{\\prime} $$, $$j^{\\prime} $$) and H + D2(v = 4, j = 0) $$\\to $$ D + HD($$v^{\\prime} $$, $$j^{\\prime} $$) reactions are presented for collision energies between $$1\\,\\mu {\\rm{K}}$$ and $$100\\,{\\rm{K}}$$. The ab initio BKMP2 PES for the ground electronic state of H3 is used and all values of total angular momentum between $J=0-4$ are included. The general vector potential approach is used to include the geometric phase. The rotationally resolved, vibrationally resolved, and total reaction rate coefficients are reported as a function of collision energy. Rotationally resolved differential cross sections are also reported as a function of collision energy and scattering angle. Large geometric phase effects appear in the ultracold reaction rate coefficients which result in a significant enhancement or suppression of the rate coefficient (up to 3 orders of magnitude) relative to calculations which ignore the geometric phase. The results are interpreted using a new quantum interference mechanism which is unique to ultracold collisions. Significant effects of the geometric phase also appear in the rotationally resolved differential cross sections which lead to a very different oscillatory structure in both energy and scattering angle. Several shape resonances occur in the 1–$$10\\,{\\rm{K}}$$ energy range and the geometric phase is shown to significantly alter the predicted resonance spectrum. The geometric phase effects and ultracold rate coefficients depend sensitively on the nuclear spin. Furthermore, experimentalists may be able to control the reaction by the selection of a particular nuclear spin state.« less

Detection of anatomical changes in lung cancer patients with 2D time-integrated, 2D time-resolved and 3D time-integrated portal dosimetry: a simulation study

NASA Astrophysics Data System (ADS)

Wolfs, Cecile J. A.; Brás, Mariana G.; Schyns, Lotte E. J. R.; Nijsten, Sebastiaan M. J. J. G.; van Elmpt, Wouter; Scheib, Stefan G.; Baltes, Christof; Podesta, Mark; Verhaegen, Frank

2017-08-01

The aim of this work is to assess the performance of 2D time-integrated (2D-TI), 2D time-resolved (2D-TR) and 3D time-integrated (3D-TI) portal dosimetry in detecting dose discrepancies between the planned and (simulated) delivered dose caused by simulated changes in the anatomy of lung cancer patients. For six lung cancer patients, tumor shift, tumor regression and pleural effusion are simulated by modifying their CT images. Based on the modified CT images, time-integrated (TI) and time-resolved (TR) portal dose images (PDIs) are simulated and 3D-TI doses are calculated. The modified and original PDIs and 3D doses are compared by a gamma analysis with various gamma criteria. Furthermore, the difference in the D 95% (ΔD 95%) of the GTV is calculated and used as a gold standard. The correlation between the gamma fail rate and the ΔD 95% is investigated, as well the sensitivity and specificity of all combinations of portal dosimetry method, gamma criteria and gamma fail rate threshold. On the individual patient level, there is a correlation between the gamma fail rate and the ΔD 95%, which cannot be found at the group level. The sensitivity and specificity analysis showed that there is not one combination of portal dosimetry method, gamma criteria and gamma fail rate threshold that can detect all simulated anatomical changes. This work shows that it will be more beneficial to relate portal dosimetry and DVH analysis on the patient level, rather than trying to quantify a relationship for a group of patients. With regards to optimizing sensitivity and specificity, different combinations of portal dosimetry method, gamma criteria and gamma fail rate should be used to optimally detect certain types of anatomical changes.

Detection of anatomical changes in lung cancer patients with 2D time-integrated, 2D time-resolved and 3D time-integrated portal dosimetry: a simulation study.

PubMed

Wolfs, Cecile J A; Brás, Mariana G; Schyns, Lotte E J R; Nijsten, Sebastiaan M J J G; van Elmpt, Wouter; Scheib, Stefan G; Baltes, Christof; Podesta, Mark; Verhaegen, Frank

2017-07-12

The aim of this work is to assess the performance of 2D time-integrated (2D-TI), 2D time-resolved (2D-TR) and 3D time-integrated (3D-TI) portal dosimetry in detecting dose discrepancies between the planned and (simulated) delivered dose caused by simulated changes in the anatomy of lung cancer patients. For six lung cancer patients, tumor shift, tumor regression and pleural effusion are simulated by modifying their CT images. Based on the modified CT images, time-integrated (TI) and time-resolved (TR) portal dose images (PDIs) are simulated and 3D-TI doses are calculated. The modified and original PDIs and 3D doses are compared by a gamma analysis with various gamma criteria. Furthermore, the difference in the D 95% (ΔD 95% ) of the GTV is calculated and used as a gold standard. The correlation between the gamma fail rate and the ΔD 95% is investigated, as well the sensitivity and specificity of all combinations of portal dosimetry method, gamma criteria and gamma fail rate threshold. On the individual patient level, there is a correlation between the gamma fail rate and the ΔD 95% , which cannot be found at the group level. The sensitivity and specificity analysis showed that there is not one combination of portal dosimetry method, gamma criteria and gamma fail rate threshold that can detect all simulated anatomical changes. This work shows that it will be more beneficial to relate portal dosimetry and DVH analysis on the patient level, rather than trying to quantify a relationship for a group of patients. With regards to optimizing sensitivity and specificity, different combinations of portal dosimetry method, gamma criteria and gamma fail rate should be used to optimally detect certain types of anatomical changes.

Mapping the contact surfaces in the Lamin A:AIMP3 complex by hydrogen/deuterium exchange FT-ICR mass spectrometry.

PubMed

Tao, Yeqing; Fang, Pengfei; Kim, Sunghoon; Guo, Min; Young, Nicolas L; Marshall, Alan G

2017-01-01

Aminoacyl-tRNA synthetases-interacting multifunctional protein3 (AIMP3/p18) is involved in the macromolecular tRNA synthetase complex via its interaction with several aminoacyl-tRNA synthetases. Recent reports reveal a novel function of AIMP3 as a tumor suppressor by accelerating cellular senescence and causing defects in nuclear morphology. AIMP3 specifically mediates degradation of mature Lamin A (LmnA), a major component of the nuclear envelope matrix; however, the mechanism of how AIMP3 interacts with LmnA is unclear. Here we report solution-phase hydrogen/deuterium exchange (HDX) for AIMP3, LmnA, and AIMP3 in association with the LmnA C-terminus. Reversed-phase LC coupled with LTQ 14.5 T Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) results in high mass accuracy and resolving power for comparing the D-uptake profiles for AIMP3, LmnA, and their complex. The results show that the AIMP3-LmnA interaction involves one of the two putative binding sites and an adjacent novel interface on AIMP3. LmnA binds AIMP3 via its extreme C-terminus. Together these findings provide a structural insight for understanding the interaction between AIMP3 and LmnA in AIMP3 degradation.

MPACT Theory Manual, Version 2.2.0

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

Downar, Thomas; Collins, Benjamin S.; Gehin, Jess C.

2016-06-09

This theory manual describes the three-dimensional (3-D) whole-core, pin-resolved transport calculation methodology employed in the MPACT code. To provide sub-pin level power distributions with sufficient accuracy, MPACT employs the method of characteristics (MOC) solutions in the framework of a 3-D coarse mesh finite difference (CMFD) formulation. MPACT provides a 3D MOC solution, but also a 2D/1D solution in which the 2D planar solution is provided by MOC and the axial coupling is resolved by one-dimensional (1-D) lower order (diffusion or P3) solutions. In Chapter 2 of the manual, the MOC methodology is described for calculating the regional angular and scalarmore » fluxes from the Boltzmann transport equation. In Chapter 3, the 2D/1D methodology is described, together with the description of the CMFD iteration process involving dynamic homogenization and solution of the multigroup CMFD linear system. A description of the MPACT depletion algorithm is given in Chapter 4, followed by a discussion of the subgroup and ESSM resonance processing methods in Chapter 5. The final Chapter 6 describes a simplified thermal hydraulics model in MPACT.« less

Three-dimensional shape perception from chromatic orientation flows

PubMed Central

Zaidi, Qasim; Li, Andrea

2010-01-01

The role of chromatic information in 3-D shape perception is controversial. We resolve this controversy by showing that chromatic orientation flows are sufficient for accurate perception of 3-D shape. Chromatic flows required less cone contrast to convey shape than did achromatic flows, thus ruling out luminance artifacts as a problem. Luminance artifacts were also ruled out by a protanope’s inability to see 3-D shape from chromatic flows. Since chromatic orientation flows can only be extracted from retinal images by neurons that are responsive to color modulations and selective for orientation, the psychophysical results also resolve the controversy over the existence of such neurons. In addition, we show that identification of 3-D shapes from chromatic flows can be masked by luminance modulations, indicating that it is subserved by orientation-tuned neurons sensitive to both chromatic and luminance modulations. PMID:16961963

Resolving power for the diffusion orientation distribution function.

PubMed

Jensen, Jens H; Helpern, Joseph A

2016-08-01

The diffusion orientation distribution function (dODF) is primarily used for white matter fiber tractography. Here the resolving power of the dODF is investigated for a simple diffusion model of two intersecting axonal fiber bundles. The resolving power for the dODF is evaluated using the Sparrow criterion. This is determined for the exact dODF and also for q-space imaging (QSI), q-ball, and kurtosis approximations. Based on theoretical and numerical calculations, the resolving power is found to depend on the eigenvalues of the diffusion model and on the degree of radial weighting for the dODF. The resolving powers of the QSI and q-ball dODFs improve with increased b-value. The kurtosis dODF has a resolving power similar to that of the exact dODF. The dODFs, whether exact or approximate, have finite resolving powers that limit their sensitivity to fiber crossings. The resolving powers for the different dODFs considered here provide convenient benchmarks for assessing and comparing their performance. Magn Reson Med 76:679-688, 2016. © 2015 Wiley Periodicals, Inc. © 2015 Wiley Periodicals, Inc.

Real-time measurement of hyperpolarized lactate production and efflux as a biomarker of tumor aggressiveness in an MR compatible 3D cell culture bioreactor.

PubMed

Sriram, Renuka; Van Criekinge, Mark; Hansen, Ailin; Wang, Zhen J; Vigneron, Daniel B; Wilson, David M; Keshari, Kayvan R; Kurhanewicz, John

2015-09-01

We have developed a 3D cell/tissue culture bioreactor compatible with hyperpolarized (HP) (13)C MR and interrogated HP [1-(13)C]lactate production and efflux in human renal cell carcinoma (RCC) cells. This platform is capable of resolving intracellular and extracellular HP lactate pools, allowing the kinetic measurement of lactate production and efflux in the context of cancer aggressiveness and response to therapy. HP (13)C MR studies were performed on three immortalized human renal cell lines: HK2, a normal renal proximal tubule cell line from which a majority of RCCs arise, UMRC6, a cell line derived from a localized RCC, and UOK262, an aggressive and metastatic RCC. The intra- (Lacin ) and extracellular (Lacex ) HP lactate signals were robustly resolved in dynamic (13)C spectra of the cell lines due to a very small but reproducible chemical shift difference (0.031 ± 0.0005 ppm). Following HP [1-(13)C]pyruvate delivery, the ratio of HP Lacin /Lacex was significantly lower for UOK262 cells compared with both UMRC6 and HK2 cells due to a significant (p < 0.05) increase in the Lacex pool size. Lacin /Lacex correlated with the MCT4 mRNA expression of the cell lines, and inhibition of MCT4 transport using DIDS resulted in a significant reduction in the HP Lacex pool size. The extension of these studies to living patient-derived RCC tissue slices using HP [1,2-(13)C2]pyruvate demonstrated a similarly split lactate doublet with a high Lacex pool fraction; in contrast, only a single NMR resonance is noted for HP [5-(13)C]glutamate, consistent with intracellular localization. These studies support the importance of lactate efflux as a biomarker of cancer aggressiveness and metastatic potential, and the utility of the MR compatible 3D cell/tissue culture bioreactor to study not only cellular metabolism but also transport. Additionally, this platform offers a sophisticated way to follow therapeutic interventions and screen novel therapies that target lactate export. Copyright © 2015 John Wiley & Sons, Ltd.

Resolvin D1 protects the liver from ischemia/reperfusion injury by enhancing M2 macrophage polarization and efferocytosis.

PubMed

Kang, Jung-Woo; Lee, Sun-Mee

2016-09-01

Resolution of inflammation is an active process involving a novel category of lipid factors known as specialized pro-resolving lipid mediators, which includes Resolvin D1 (RvD1). While accumulating evidence suggests that RvD1 counteracts proinflammatory signaling and promotes resolution, the specific cellular targets and mechanisms of action of RvD1 remain largely unknown. In the present study, we investigated the role and molecular mechanisms of RvD1 in ischemia/reperfusion (IR)-induced sterile liver inflammation. Male C57BL/6 mice underwent 70% hepatic ischemia for 60min, followed by reperfusion. RvD1 (5, 10, and 15μg/kg, i.p.) was administered to the mice 1h before ischemia and then immediately prior to reperfusion. RvD1 attenuated IR-induced hepatocellular damage and the proinflammatory response. In purified Kupffer cells (KCs) from mice exposed to IR, the levels of M1 marker genes (Nos2a and Cd40) increased, while those of M2 marker genes (Arg1, Cd206, and Mst1r) decreased, demonstrating a proinflammatory shift. RvD1 markedly attenuated these changes. Depletion of KCs by liposome clodronate abrogated the effects of RvD1 on proinflammatory mediators and macrophage polarization. In addition, RvD1 attenuated increases in myeloperoxidase activity and Cxcl1 and Cxcl2 mRNA expression. RvD1 markedly augmented the efferocytic activity of KCs, as indicated by increases in F4/80(+)Gr-1(+) cells in the liver. However, antagonist pretreatment or gene silencing of the RvD1 receptor, ALX/FPR2, abrogated the anti-inflammatory and pro-resolving actions of RvD1. These data indicate that RvD1 ameliorates IR-induced liver injury, and this protection is associated with enhancement of M2 polarization and efferocytosis via ALX/FPR2 activation. Copyright © 2016 Elsevier B.V. All rights reserved.

Giant Thermal Expansion in 2D and 3D Cellular Materials.

PubMed

Zhu, Hanxing; Fan, Tongxiang; Peng, Qing; Zhang, Di

2018-05-01

When temperature increases, the volume of an object changes. This property was quantified as the coefficient of thermal expansion only a few hundred years ago. Part of the reason is that the change of volume due to the variation of temperature is in general extremely small and imperceptible. Here, abnormal giant linear thermal expansions in different types of two-ingredient microstructured hierarchical and self-similar cellular materials are reported. The cellular materials can be 2D or 3D, and isotropic or anisotropic, with a positive or negative thermal expansion due to the convex or/and concave shape in their representative volume elements respectively. The magnitude of the thermal expansion coefficient can be several times larger than the highest value reported in the literature. This study suggests an innovative approach to develop temperature-sensitive functional materials and devices. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Appropriate Minimal Dose of Gadobutrol for 3D Time-Resolved MRA of the Supra-Aortic Arteries: Comparison with Conventional Single-Phase High-Resolution 3D Contrast-Enhanced MRA.

PubMed

Bak, S H; Roh, H G; Moon, W-J; Choi, J W; An, H S

2017-07-01

The development of nephrogenic systemic fibrosis and neural tissue deposition is gadolinium dose-dependent. The purpose of this study was to determine the appropriate minimal dose of gadobutrol with time-resolved MRA to assess supra-aortic arterial stenosis with contrast-enhanced MRA as a reference standard. Four hundred sixty-two consecutive patients underwent both standard-dose contrast-enhanced MRA and low-dose time-resolved MRA and were classified into 3 groups; group A (a constant dose of 1 mL for time-resolved MRA), group B (2 mL), or group C (3 mL). All studies were independently evaluated by 2 radiologists for image quality by using a 5-point scale (from 0 = failure to 4 = excellent), grading of arterial stenosis (0 = normal, 1 = mild [<30%], 2 = moderate [30%-69%], 3 = severe to occlusion [≥70%]), and signal-to-noise ratio. The image quality of time-resolved MRA was similar to that of contrast-enhanced MRA in groups B and C, but it was inferior to contrast-enhanced MRA in group A. For the grading of arterial stenosis, there was an excellent correlation between contrast-enhanced MRA and time-resolved MRA ( R = 0.957 for group A, R = 0.988 for group B, R = 0.991 for group C). The SNR of time-resolved MRA tended to be lower than that of contrast-enhanced MRA in groups A and B. However, SNR was higher for time-resolved MRA compared with contrast-enhanced MRA in group C. Low-dose time-resolved MRA is feasible in the evaluation of supra-aortic stenosis and could be used as an alternative to contrast-enhanced MRA for a diagnostic technique in high-risk populations. © 2017 by American Journal of Neuroradiology.

Human cardiac telocytes: 3D imaging by FIB-SEM tomography.

PubMed

Cretoiu, D; Hummel, E; Zimmermann, H; Gherghiceanu, M; Popescu, L M

2014-11-01

Telocyte (TC) is a newly identified type of cell in the cardiac interstitium (www.telocytes.com). TCs are described by classical transmission electron microscopy as cells with very thin and long telopodes (Tps; cellular prolongations) having podoms (dilations) and podomers (very thin segments). TCs' three-dimensional (3D) morphology is still unknown. Cardiac TCs seem to be particularly involved in long and short distance intercellular signalling and, therefore, their 3D architecture is important for understanding their spatial connections. Using focused ion beam scanning electron microscopy (FIB-SEM) we show, for the first time, the whole ultrastructural anatomy of cardiac TCs. 3D reconstruction of cardiac TCs by FIB-SEM tomography confirms that they have long, narrow but flattened (ribbon-like) telopodes, with humps generated by the podoms. FIB-SEM tomography also confirms the network made by TCs in the cardiac interstitium through adherens junctions. This study provides the first FIB-SEM tomography of a human cell type. © 2014 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

Building blocks for the synthesis of glycosyl-myo-inositols involved in the insulin intracellular signalling process.

PubMed

Zapata, A; Martín-Lomas, M

1992-10-09

Glycosylation of (+/- )-1-O-benzyl-2,3:5,6-di-O-isopropylidene-myo-inositol (4) with 6-O-acetyl-4-O-allyl-2-azido-3-O-benzyl-2-deoxy-beta-D-glucopyranosyl trichloroacetimidate (6) gave the 4-O-(2-amino-2-deoxy-alpha-D-glucopyranosyl)- myo-inositol derivative (9) as a mixture of diastereoisomers which could be resolved by chromatography. Likewise alpha-glycosylation of 4 with 6-O-acetyl-2-azido-3-O-benzoyl-2-deoxy-4-O-(2,3,4,6-tetra-O-acetyl-beta- D- galactopyranosyl)-D-glucopyranosyl trichloroacetimidate (10) gave the corresponding pseudotrisaccharide derivative 16 as a mixture of diastereomers which could be resolved partially by chromatography. alpha-Glycosylation of enantiomerically pure 2,3:5,6- (18) and 2,3:4,5-di-O-isopropylidene-1-O-menthoxycarbonyl-myo-inositol (19) with 3,4,6-tri-O-acetyl-2-azido-2-deoxy-D-glucopyranosyl trichloroacetimidate (20) gave the pseudodisaccharide derivatives 21 and 22, respectively. Likewise, alpha-glycosylation of 18 with 10 afforded a pseudotrisaccharide derivative (23).

Around the macrolide - Impact of 3D structure of macrocycles on lipophilicity and cellular accumulation.

PubMed

Koštrun, Sanja; Munic Kos, Vesna; Matanović Škugor, Maja; Palej Jakopović, Ivana; Malnar, Ivica; Dragojević, Snježana; Ralić, Jovica; Alihodžić, Sulejman

2017-06-16

The aim of this study was to investigate lipophilicity and cellular accumulation of rationally designed azithromycin and clarithromycin derivatives at the molecular level. The effect of substitution site and substituent properties on a global physico-chemical profile and cellular accumulation of investigated compounds was studied using calculated structural parameters as well as experimentally determined lipophilicity. In silico models based on the 3D structure of molecules were generated to investigate conformational effect on studied properties and to enable prediction of lipophilicity and cellular accumulation for this class of molecules based on non-empirical parameters. The applicability of developed models was explored on a validation and test sets and compared with previously developed empirical models. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

New approaches for the reliable in vitro assessment of binding affinity based on high-resolution real-time data acquisition of radioligand-receptor binding kinetics.

PubMed

Zeilinger, Markus; Pichler, Florian; Nics, Lukas; Wadsak, Wolfgang; Spreitzer, Helmut; Hacker, Marcus; Mitterhauser, Markus

2017-12-01

Resolving the kinetic mechanisms of biomolecular interactions have become increasingly important in early-phase drug development. Since traditional in vitro methods belong to dose-dependent assessments, binding kinetics is usually overlooked. The present study aimed at the establishment of two novel experimental approaches for the assessment of binding affinity of both, radiolabelled and non-labelled compounds targeting the A 3 R, based on high-resolution real-time data acquisition of radioligand-receptor binding kinetics. A novel time-resolved competition assay was developed and applied to determine the K i of eight different A 3 R antagonists, using CHO-K1 cells stably expressing the hA 3 R. In addition, a new kinetic real-time cell-binding approach was established to quantify the rate constants k on and k off , as well as the dedicated K d of the A 3 R agonist [ 125 I]-AB-MECA. Furthermore, lipophilicity measurements were conducted to control influences due to physicochemical properties of the used compounds. Two novel real-time cell-binding approaches were successfully developed and established. Both experimental procedures were found to visualize the kinetic binding characteristics with high spatial and temporal resolution, resulting in reliable affinity values, which are in good agreement with values previously reported with traditional methods. Taking into account the lipophilicity of the A 3 R antagonists, no influences on the experimental performance and the resulting affinity were investigated. Both kinetic binding approaches comprise tracer administration and subsequent binding to living cells, expressing the dedicated target protein. Therefore, the experiments resemble better the true in vivo physiological conditions and provide important markers of cellular feedback and biological response.

An approach to estimate spatial distribution of analyte within cells using spectrally-resolved fluorescence microscopy.

PubMed

Sharma, Dharmendar Kumar; Irfanullah, Mir; Basu, Santanu Kumar; Madhu, Sheri; De, Suman; Jadhav, Sameer; Ravikanth, Mangalampalli; Chowdhury, Arindam

2017-01-18

While fluorescence microscopy has become an essential tool amongst chemists and biologists for the detection of various analyte within cellular environments, non-uniform spatial distribution of sensors within cells often restricts extraction of reliable information on relative abundance of analytes in different subcellular regions. As an alternative to existing sensing methodologies such as ratiometric or FRET imaging, where relative proportion of analyte with respect to the sensor can be obtained within cells, we propose a methodology using spectrally-resolved fluorescence microscopy, via which both the relative abundance of sensor as well as their relative proportion with respect to the analyte can be simultaneously extracted for local subcellular regions. This method is exemplified using a BODIPY sensor, capable of detecting mercury ions within cellular environments, characterized by spectral blue-shift and concurrent enhancement of emission intensity. Spectral emission envelopes collected from sub-microscopic regions allowed us to compare the shift in transition energies as well as integrated emission intensities within various intracellular regions. Construction of a 2D scatter plot using spectral shifts and emission intensities, which depend on the relative amount of analyte with respect to sensor and the approximate local amounts of the probe, respectively, enabled qualitative extraction of relative abundance of analyte in various local regions within a single cell as well as amongst different cells. Although the comparisons remain semi-quantitative, this approach involving analysis of multiple spectral parameters opens up an alternative way to extract spatial distribution of analyte in heterogeneous systems. The proposed method would be especially relevant for fluorescent probes that undergo relatively nominal shift in transition energies compared to their emission bandwidths, which often restricts their usage for quantitative ratiometric imaging in cellular media due to strong cross-talk between energetically separated detection channels.

An approach to estimate spatial distribution of analyte within cells using spectrally-resolved fluorescence microscopy

NASA Astrophysics Data System (ADS)

Sharma, Dharmendar Kumar; Irfanullah, Mir; Basu, Santanu Kumar; Madhu, Sheri; De, Suman; Jadhav, Sameer; Ravikanth, Mangalampalli; Chowdhury, Arindam

2017-03-01

While fluorescence microscopy has become an essential tool amongst chemists and biologists for the detection of various analyte within cellular environments, non-uniform spatial distribution of sensors within cells often restricts extraction of reliable information on relative abundance of analytes in different subcellular regions. As an alternative to existing sensing methodologies such as ratiometric or FRET imaging, where relative proportion of analyte with respect to the sensor can be obtained within cells, we propose a methodology using spectrally-resolved fluorescence microscopy, via which both the relative abundance of sensor as well as their relative proportion with respect to the analyte can be simultaneously extracted for local subcellular regions. This method is exemplified using a BODIPY sensor, capable of detecting mercury ions within cellular environments, characterized by spectral blue-shift and concurrent enhancement of emission intensity. Spectral emission envelopes collected from sub-microscopic regions allowed us to compare the shift in transition energies as well as integrated emission intensities within various intracellular regions. Construction of a 2D scatter plot using spectral shifts and emission intensities, which depend on the relative amount of analyte with respect to sensor and the approximate local amounts of the probe, respectively, enabled qualitative extraction of relative abundance of analyte in various local regions within a single cell as well as amongst different cells. Although the comparisons remain semi-quantitative, this approach involving analysis of multiple spectral parameters opens up an alternative way to extract spatial distribution of analyte in heterogeneous systems. The proposed method would be especially relevant for fluorescent probes that undergo relatively nominal shift in transition energies compared to their emission bandwidths, which often restricts their usage for quantitative ratiometric imaging in cellular media due to strong cross-talk between energetically separated detection channels. Dedicated to Professor Kankan Bhattacharyya.

Interplay between self-assembled structure of bone morphogenetic protein-2 (BMP-2) and osteoblast functions in three-dimensional titanium alloy scaffolds: Stimulation of osteogenic activity.

PubMed

Nune, K C; Kumar, A; Murr, L E; Misra, R D K

2016-02-01

Three-dimensional cellular scaffolds are receiving significant attention in bone tissue engineering to treat segmental bone defects. However, there are indications of lack of significant osteoinductive ability of three-dimensional cellular scaffolds. In this regard, the objective of the study is to elucidate the interplay between bone morphogenetic protein (BMP-2) and osteoblast functions on 3D mesh structures with different porosities and pore size that were fabricated by electron beam melting. Self-assembled dendritic microstructure with interconnected cellular-type morphology of BMP-2 on 3D scaffolds stimulated osteoblast functions including adhesion, proliferation, and mineralization, with prominent effect on 2-mm mesh. Furthermore, immunofluorescence studies demonstrated higher density and viability of osteoblasts on lower porosity mesh structure (2 mm) as compared to 3- and 4-mm mesh structures. Enhanced filopodia cellular extensions with extensive cell spreading was observed on BMP-2 treated mesh structures, a behavior that is attributed to the unique self-assembled structure of BMP-2 that effectively communicates with the cells. The study underscores the potential of BMP-2 in imparting osteoinductive capability to the 3D printed scaffolds. © 2015 Wiley Periodicals, Inc.

3D surface reconstruction and visualization of the Drosophila wing imaginal disc at cellular resolution

NASA Astrophysics Data System (ADS)

Bai, Linge; Widmann, Thomas; Jülicher, Frank; Dahmann, Christian; Breen, David

2013-01-01

Quantifying and visualizing the shape of developing biological tissues provide information about the morphogenetic processes in multicellular organisms. The size and shape of biological tissues depend on the number, size, shape, and arrangement of the constituting cells. To better understand the mechanisms that guide tissues into their final shape, it is important to investigate the cellular arrangement within tissues. Here we present a data processing pipeline to generate 3D volumetric surface models of epithelial tissues, as well as geometric descriptions of the tissues' apical cell cross-sections. The data processing pipeline includes image acquisition, editing, processing and analysis, 2D cell mesh generation, 3D contourbased surface reconstruction, cell mesh projection, followed by geometric calculations and color-based visualization of morphological parameters. In their first utilization we have applied these procedures to construct a 3D volumetric surface model at cellular resolution of the wing imaginal disc of Drosophila melanogaster. The ultimate goal of the reported effort is to produce tools for the creation of detailed 3D geometric models of the individual cells in epithelial tissues. To date, 3D volumetric surface models of the whole wing imaginal disc have been created, and the apicolateral cell boundaries have been identified, allowing for the calculation and visualization of cell parameters, e.g. apical cross-sectional area of cells. The calculation and visualization of morphological parameters show position-dependent patterns of cell shape in the wing imaginal disc. Our procedures should offer a general data processing pipeline for the construction of 3D volumetric surface models of a wide variety of epithelial tissues.

Advances in 4D Treatment Planning for Scanned Particle Beam Therapy — Report of Dedicated Workshops

PubMed Central

Bert, Christoph; Graeff, Christian; Riboldi, Marco; Nill, Simeon; Baroni, Guido; Knopf, Antje-Christin

2014-01-01

We report on recent progress in the field of mobile tumor treatment with scanned particle beams, as discussed in the latest editions of the 4D treatment planning workshop. The workshop series started in 2009, with about 20 people from 4 research institutes involved, all actively working on particle therapy delivery and development. The first workshop resulted in a summary of recommendations for the treatment of mobile targets, along with a list of requirements to apply these guidelines clinically. The increased interest in the treatment of mobile tumors led to a continuously growing number of attendees: the 2012 edition counted more than 60 participants from 20 institutions and commercial vendors. The focus of research discussions among workshop participants progressively moved from 4D treatment planning to complete 4D treatments, aiming at effective and safe treatment delivery. Current research perspectives on 4D treatments include all critical aspects of time resolved delivery, such as in-room imaging, motion detection, beam application, and quality assurance techniques. This was motivated by the start of first clinical treatments of hepato cellular tumors with a scanned particle beam, relying on gating or abdominal compression for motion mitigation. Up to date research activities emphasize significant efforts in investigating advanced motion mitigation techniques, with a specific interest in the development of dedicated tools for experimental validation. Potential improvements will be made possible in the near future through 4D optimized treatment plans that require upgrades of the currently established therapy control systems for time resolved delivery. But since also these novel optimization techniques rely on the validity of the 4DCT, research focusing on alternative 4D imaging technique, such as MRI based 4DCT generation will continue. PMID:24354749

Cancer cells growing on perfused 3D collagen model produced higher reactive oxygen species level and were more resistant to cisplatin compared to the 2D model.

PubMed

Liu, Qingxi; Zhang, Zijiang; Liu, Yupeng; Cui, Zhanfeng; Zhang, Tongcun; Li, Zhaohui; Ma, Wenjian

2018-03-01

Three-dimensional (3D) collagen scaffold models, due to their ability to mimic the tissue and organ structure in vivo, have received increasing interest in drug discovery and toxicity evaluation. In this study, we developed a perfused 3D model and studied cellular response to cytotoxic drugs in comparison with traditional 2D cell cultures as evaluated by cancer drug cisplatin. Cancer cells grown in perfused 3D environments showed increased levels of reactive oxygen species (ROS) production compared to the 2D culture. As determined by growth analysis, cells in the 3D culture, after forming a spheroid, were more resistant to the cancer drug cisplatin compared to that of the 2D cell culture. In addition, 3D culturing cells showed elevated level of ROS, indicating a physiological change or the formation of a microenvironment that resembles tumor cells in vivo. These data revealed that cellular response to drugs for cells growing in 3D environments are dramatically different from that of 2D cultured cells. Thus, the perfused 3D collagen scaffold model we report here might be a potentially very useful tool for drug analysis.

Simulation of orographic effects with a Quasi-3-D Multiscale Modeling Framework: Basic algorithm and preliminary results

DOE PAGES

Jung, Joon -Hee

2016-10-11

Here, the global atmospheric models based on the Multi-scale Modeling Framework (MMF) are able to explicitly resolve subgrid-scale processes by using embedded 2-D Cloud-Resolving Models (CRMs). Up to now, however, those models do not include the orographic effects on the CRM grid scale. This study shows that the effects of CRM grid-scale orography can be simulated reasonably well by the Quasi-3-D MMF (Q3D MMF), which has been developed as a second-generation MMF. In the Q3D framework, the surface topography can be included in the CRM component by using a block representation of the mountains, so that no smoothing of themore » topographic height is necessary. To demonstrate the performance of such a model, the orographic effects over a steep mountain are simulated in an idealized experimental setup with each of the Q3D MMF and the full 3-D CRM. The latter is used as a benchmark. Comparison of the results shows that the Q3D MMF is able to reproduce the horizontal distribution of orographic precipitation and the flow changes around mountains as simulated by the 3-D CRM, even though the embedded CRMs of the Q3D MMF recognize only some aspects of the complex 3-D topography. It is also shown that the use of 3-D CRMs in the Q3D framework, rather than 2-D CRMs, has positive impacts on the simulation of wind fields but does not substantially change the simulated precipitation.« less

Simulation of orographic effects with a Quasi-3-D Multiscale Modeling Framework: Basic algorithm and preliminary results

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

Jung, Joon -Hee

Here, the global atmospheric models based on the Multi-scale Modeling Framework (MMF) are able to explicitly resolve subgrid-scale processes by using embedded 2-D Cloud-Resolving Models (CRMs). Up to now, however, those models do not include the orographic effects on the CRM grid scale. This study shows that the effects of CRM grid-scale orography can be simulated reasonably well by the Quasi-3-D MMF (Q3D MMF), which has been developed as a second-generation MMF. In the Q3D framework, the surface topography can be included in the CRM component by using a block representation of the mountains, so that no smoothing of themore » topographic height is necessary. To demonstrate the performance of such a model, the orographic effects over a steep mountain are simulated in an idealized experimental setup with each of the Q3D MMF and the full 3-D CRM. The latter is used as a benchmark. Comparison of the results shows that the Q3D MMF is able to reproduce the horizontal distribution of orographic precipitation and the flow changes around mountains as simulated by the 3-D CRM, even though the embedded CRMs of the Q3D MMF recognize only some aspects of the complex 3-D topography. It is also shown that the use of 3-D CRMs in the Q3D framework, rather than 2-D CRMs, has positive impacts on the simulation of wind fields but does not substantially change the simulated precipitation.« less

Simulation of orographic effects with a Quasi-3-D Multiscale Modeling Framework: Basic algorithm and preliminary results

NASA Astrophysics Data System (ADS)

Jung, Joon-Hee

2016-12-01

The global atmospheric models based on the Multi-scale Modeling Framework (MMF) are able to explicitly resolve subgrid-scale processes by using embedded 2-D Cloud-Resolving Models (CRMs). Up to now, however, those models do not include the orographic effects on the CRM grid scale. This study shows that the effects of CRM grid-scale orography can be simulated reasonably well by the Quasi-3-D MMF (Q3D MMF), which has been developed as a second-generation MMF. In the Q3D framework, the surface topography can be included in the CRM component by using a block representation of the mountains, so that no smoothing of the topographic height is necessary. To demonstrate the performance of such a model, the orographic effects over a steep mountain are simulated in an idealized experimental setup with each of the Q3D MMF and the full 3-D CRM. The latter is used as a benchmark. Comparison of the results shows that the Q3D MMF is able to reproduce the horizontal distribution of orographic precipitation and the flow changes around mountains as simulated by the 3-D CRM, even though the embedded CRMs of the Q3D MMF recognize only some aspects of the complex 3-D topography. It is also shown that the use of 3-D CRMs in the Q3D framework, rather than 2-D CRMs, has positive impacts on the simulation of wind fields but does not substantially change the simulated precipitation.

TH-EF-BRA-11: Feasibility of Super-Resolution Time-Resolved 4DMRI for Multi-Breath Volumetric Motion Simulation in Radiotherapy Planning

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

Li, G; Zakian, K; Deasy, J

Purpose: To develop a novel super-resolution time-resolved 4DMRI technique to evaluate multi-breath, irregular and complex organ motion without respiratory surrogate for radiotherapy planning. Methods: The super-resolution time-resolved (TR) 4DMRI approach combines a series of low-resolution 3D cine MRI images acquired during free breathing (FB) with a high-resolution breath-hold (BH) 3DMRI via deformable image registration (DIR). Five volunteers participated in the study under an IRB-approved protocol. The 3D cine images with voxel size of 5×5×5 mm{sup 3} at two volumes per second (2Hz) were acquired coronally using a T1 fast field echo sequence, half-scan (0.8) acceleration, and SENSE (3) parallel imaging.more » Phase-encoding was set in the lateral direction to minimize motion artifacts. The BH image with voxel size of 2×2×2 mm{sup 3} was acquired using the same sequence within 10 seconds. A demons-based DIR program was employed to produce super-resolution 2Hz 4DMRI. Registration quality was visually assessed using difference images between TR 4DMRI and 3D cine and quantitatively assessed using average voxel correlation. The fidelity of the 3D cine images was assessed using a gel phantom and a 1D motion platform by comparing mobile and static images. Results: Owing to voxel intensity similarity using the same MRI scanning sequence, accurate DIR between FB and BH images is achieved. The voxel correlations between 3D cine and TR 4DMRI are greater than 0.92 in all cases and the difference images illustrate minimal residual error with little systematic patterns. The 3D cine images of the mobile gel phantom preserve object geometry with minimal scanning artifacts. Conclusion: The super-resolution time-resolved 4DMRI technique has been achieved via DIR, providing a potential solution for multi-breath motion assessment. Accurate DIR mapping has been achieved to map high-resolution BH images to low-resolution FB images, producing 2Hz volumetric high-resolution 4DMRI. Further validation and improvement are still required prior to clinical applications. This study is in part supported by the NIH (U54CA137788/U54CA132378).« less

An analytical tool that quantifies cellular morphology changes from three-dimensional fluorescence images.

PubMed

Haass-Koffler, Carolina L; Naeemuddin, Mohammad; Bartlett, Selena E

2012-08-31

The most common software analysis tools available for measuring fluorescence images are for two-dimensional (2D) data that rely on manual settings for inclusion and exclusion of data points, and computer-aided pattern recognition to support the interpretation and findings of the analysis. It has become increasingly important to be able to measure fluorescence images constructed from three-dimensional (3D) datasets in order to be able to capture the complexity of cellular dynamics and understand the basis of cellular plasticity within biological systems. Sophisticated microscopy instruments have permitted the visualization of 3D fluorescence images through the acquisition of multispectral fluorescence images and powerful analytical software that reconstructs the images from confocal stacks that then provide a 3D representation of the collected 2D images. Advanced design-based stereology methods have progressed from the approximation and assumptions of the original model-based stereology even in complex tissue sections. Despite these scientific advances in microscopy, a need remains for an automated analytic method that fully exploits the intrinsic 3D data to allow for the analysis and quantification of the complex changes in cell morphology, protein localization and receptor trafficking. Current techniques available to quantify fluorescence images include Meta-Morph (Molecular Devices, Sunnyvale, CA) and Image J (NIH) which provide manual analysis. Imaris (Andor Technology, Belfast, Northern Ireland) software provides the feature MeasurementPro, which allows the manual creation of measurement points that can be placed in a volume image or drawn on a series of 2D slices to create a 3D object. This method is useful for single-click point measurements to measure a line distance between two objects or to create a polygon that encloses a region of interest, but it is difficult to apply to complex cellular network structures. Filament Tracer (Andor) allows automatic detection of the 3D neuronal filament-like however, this module has been developed to measure defined structures such as neurons, which are comprised of dendrites, axons and spines (tree-like structure). This module has been ingeniously utilized to make morphological measurements to non-neuronal cells, however, the output data provide information of an extended cellular network by using a software that depends on a defined cell shape rather than being an amorphous-shaped cellular model. To overcome the issue of analyzing amorphous-shaped cells and making the software more suitable to a biological application, Imaris developed Imaris Cell. This was a scientific project with the Eidgenössische Technische Hochschule, which has been developed to calculate the relationship between cells and organelles. While the software enables the detection of biological constraints, by forcing one nucleus per cell and using cell membranes to segment cells, it cannot be utilized to analyze fluorescence data that are not continuous because ideally it builds cell surface without void spaces. To our knowledge, at present no user-modifiable automated approach that provides morphometric information from 3D fluorescence images has been developed that achieves cellular spatial information of an undefined shape (Figure 1). We have developed an analytical platform using the Imaris core software module and Imaris XT interfaced to MATLAB (Mat Works, Inc.). These tools allow the 3D measurement of cells without a pre-defined shape and with inconsistent fluorescence network components. Furthermore, this method will allow researchers who have extended expertise in biological systems, but not familiarity to computer applications, to perform quantification of morphological changes in cell dynamics.

The point-spread function measure of resolution for the 3-D electrical resistivity experiment

NASA Astrophysics Data System (ADS)

Oldenborger, Greg A.; Routh, Partha S.

2009-02-01

The solution appraisal component of the inverse problem involves investigation of the relationship between our estimated model and the actual model. However, full appraisal is difficult for large 3-D problems such as electrical resistivity tomography (ERT). We tackle the appraisal problem for 3-D ERT via the point-spread functions (PSFs) of the linearized resolution matrix. The PSFs represent the impulse response of the inverse solution and quantify our parameter-specific resolving capability. We implement an iterative least-squares solution of the PSF for the ERT experiment, using on-the-fly calculation of the sensitivity via an adjoint integral equation with stored Green's functions and subgrid reduction. For a synthetic example, analysis of individual PSFs demonstrates the truly 3-D character of the resolution. The PSFs for the ERT experiment are Gaussian-like in shape, with directional asymmetry and significant off-diagonal features. Computation of attributes representative of the blurring and localization of the PSF reveal significant spatial dependence of the resolution with some correlation to the electrode infrastructure. Application to a time-lapse ground-water monitoring experiment demonstrates the utility of the PSF for assessing feature discrimination, predicting artefacts and identifying model dependence of resolution. For a judicious selection of model parameters, we analyse the PSFs and their attributes to quantify the case-specific localized resolving capability and its variability over regions of interest. We observe approximate interborehole resolving capability of less than 1-1.5m in the vertical direction and less than 1-2.5m in the horizontal direction. Resolving capability deteriorates significantly outside the electrode infrastructure.

Digital Single-Cell Analysis of Plant Organ Development Using 3DCellAtlas[OPEN

PubMed Central

Montenegro-Johnson, Thomas D.; Stamm, Petra; Strauss, Soeren; Topham, Alexander T.; Tsagris, Michail; Wood, Andrew T.A.; Smith, Richard S.; Bassel, George W.

2015-01-01

Diverse molecular networks underlying plant growth and development are rapidly being uncovered. Integrating these data into the spatial and temporal context of dynamic organ growth remains a technical challenge. We developed 3DCellAtlas, an integrative computational pipeline that semiautomatically identifies cell types and quantifies both 3D cellular anisotropy and reporter abundance at single-cell resolution across whole plant organs. Cell identification is no less than 97.8% accurate and does not require transgenic lineage markers or reference atlases. Cell positions within organs are defined using an internal indexing system generating cellular level organ atlases where data from multiple samples can be integrated. Using this approach, we quantified the organ-wide cell-type-specific 3D cellular anisotropy driving Arabidopsis thaliana hypocotyl elongation. The impact ethylene has on hypocotyl 3D cell anisotropy identified the preferential growth of endodermis in response to this hormone. The spatiotemporal dynamics of the endogenous DELLA protein RGA, expansin gene EXPA3, and cell expansion was quantified within distinct cell types of Arabidopsis roots. A significant regulatory relationship between RGA, EXPA3, and growth was present in the epidermis and endodermis. The use of single-cell analyses of plant development enables the dynamics of diverse regulatory networks to be integrated with 3D organ growth. PMID:25901089

Redundant function of DNA ligase 1 and 3 in alternative end-joining during immunoglobulin class switch recombination.

PubMed

Masani, Shahnaz; Han, Li; Meek, Katheryn; Yu, Kefei

2016-02-02

Nonhomologous end-joining (NHEJ) is the major DNA double-strand break (DSB) repair pathway in mammals and resolves the DSBs generated during both V(D)J recombination in developing lymphocytes and class switch recombination (CSR) in antigen-stimulated B cells. In contrast to the absolute requirement for NHEJ to resolve DSBs associated with V(D)J recombination, DSBs associated with CSR can be resolved in NHEJ-deficient cells (albeit at a reduced level) by a poorly defined alternative end-joining (A-EJ) pathway. Deletion of DNA ligase IV (Lig4), a core component of the NHEJ pathway, reduces CSR efficiency in a mouse B-cell line capable of robust cytokine-stimulated CSR in cell culture. Here, we report that CSR levels are not further reduced by deletion of either of the two remaining DNA ligases (Lig1 and nuclear Lig3) in Lig4(-/-) cells. We conclude that in the absence of Lig4, Lig1, and Lig3 function in a redundant manner in resolving switch region DSBs during CSR.

Evaluating Clouds in Long-Term Cloud-Resolving Model Simulations with Observational Data

NASA Technical Reports Server (NTRS)

Zeng, Xiping; Tao, Wei-Kuo; Zhang, Minghua; Peters-Lidard, Christa; Lang, Stephen; Simpson, Joanne; Kumar, Sujay; Xie, Shaocheng; Eastman, Joseph L.; Shie, Chung-Lin;

Phenotypic Characterization of Prostate Cancer LNCaP Cells Cultured within a Bioengineered Microenvironment

PubMed Central

Sieh, Shirly; Taubenberger, Anna V.; Rizzi, Simone C.; Sadowski, Martin; Lehman, Melanie L.; Rockstroh, Anja; An, Jiyuan; Clements, Judith A.; Nelson, Colleen C.; Hutmacher, Dietmar W.

2012-01-01

Biophysical and biochemical properties of the microenvironment regulate cellular responses such as growth, differentiation, morphogenesis and migration in normal and cancer cells. Since two-dimensional (2D) cultures lack the essential characteristics of the native cellular microenvironment, three-dimensional (3D) cultures have been developed to better mimic the natural extracellular matrix. To date, 3D culture systems have relied mostly on collagen and Matrigel™ hydrogels, allowing only limited control over matrix stiffness, proteolytic degradability, and ligand density. In contrast, bioengineered hydrogels allow us to independently tune and systematically investigate the influence of these parameters on cell growth and differentiation. In this study, polyethylene glycol (PEG) hydrogels, functionalized with the Arginine-glycine-aspartic acid (RGD) motifs, common cell-binding motifs in extracellular matrix proteins, and matrix metalloproteinase (MMP) cleavage sites, were characterized regarding their stiffness, diffusive properties, and ability to support growth of androgen-dependent LNCaP prostate cancer cells. We found that the mechanical properties modulated the growth kinetics of LNCaP cells in the PEG hydrogel. At culture periods of 28 days, LNCaP cells underwent morphogenic changes, forming tumor-like structures in 3D culture, with hypoxic and apoptotic cores. We further compared protein and gene expression levels between 3D and 2D cultures upon stimulation with the synthetic androgen R1881. Interestingly, the kinetics of R1881 stimulated androgen receptor (AR) nuclear translocation differed between 2D and 3D cultures when observed by immunofluorescent staining. Furthermore, microarray studies revealed that changes in expression levels of androgen responsive genes upon R1881 treatment differed greatly between 2D and 3D cultures. Taken together, culturing LNCaP cells in the tunable PEG hydrogels reveals differences in the cellular responses to androgen stimulation between the 2D and 3D environments. Therefore, we suggest that the presented 3D culture system represents a powerful tool for high throughput prostate cancer drug testing that recapitulates tumor microenvironment. PMID:22957009

Cellular uptake and retention measurements of alkylphosphocholines in the SK-BR-3 breast cancer and Molt-4 leukemia cell line using capillary gas chromatography.

PubMed

Brochez, V; Van Heuverswyn, D; Diniz, J A; De Potter, C R; Van den Eeckhout, E G

1999-05-01

The determination of cellular content of octadecylphosphocholine (D-19391) and hexadecylphosphocholine (HePC, D-18506), two anticancer agents of the alkylphosphocholine group, using capillary gas chromatography is described. The compounds' cytotoxicity was first determined by the MTT [3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyltetrazolium] assay, being indicative for the concentration used in the uptake and retention measurements. D-19391 was added to the SK-BR-3 breast cancer cell line and HePC to the Molt-4 leukemia cell line in concentrations of, respectively, 18.6 and 15.0 microM, during a 36-h incubation period at 37 degrees C, 5% CO2. HePC uptake in the leukemia cells was followed by a 24-h reversibility test in drug-free medium. Subsequently, sample clean-up was performed on a weak cation-exchange column. For the quantitative analysis, HePC was used as internal standard for the D-19391 measurements and vice versa. Derivatization of the samples with trimethylsilylbromide was followed by capillary gas chromatographic analysis. From these data we conclude that our uptake results are quite similar with those of a previous study of HePC cellular uptake in the more resistant Caco-2T colon cancer cell line. Without having investigated the mechanism that underlies the cellular uptake results obtained, our study points to no direct correlation between the compounds' cellular uptake and their cytotoxic effects.

Clinical performance of a free-breathing spatiotemporally accelerated 3-D time-resolved contrast-enhanced pediatric abdominal MR angiography.

PubMed

Zhang, Tao; Yousaf, Ufra; Hsiao, Albert; Cheng, Joseph Y; Alley, Marcus T; Lustig, Michael; Pauly, John M; Vasanawala, Shreyas S

2015-10-01

Pediatric contrast-enhanced MR angiography is often limited by respiration, other patient motion and compromised spatiotemporal resolution. To determine the reliability of a free-breathing spatiotemporally accelerated 3-D time-resolved contrast-enhanced MR angiography method for depicting abdominal arterial anatomy in young children. With IRB approval and informed consent, we retrospectively identified 27 consecutive children (16 males and 11 females; mean age: 3.8 years, range: 14 days to 8.4 years) referred for contrast-enhanced MR angiography at our institution, who had undergone free-breathing spatiotemporally accelerated time-resolved contrast-enhanced MR angiography studies. A radio-frequency-spoiled gradient echo sequence with Cartesian variable density k-space sampling and radial view ordering, intrinsic motion navigation and intermittent fat suppression was developed. Images were reconstructed with soft-gated parallel imaging locally low-rank method to achieve both motion correction and high spatiotemporal resolution. Quality of delineation of 13 abdominal arteries in the reconstructed images was assessed independently by two radiologists on a five-point scale. Ninety-five percent confidence intervals of the proportion of diagnostically adequate cases were calculated. Interobserver agreements were also analyzed. Eleven out of 13 arteries achieved acceptable image quality (mean score range: 3.9-5.0) for both readers. Fair to substantial interobserver agreement was reached on nine arteries. Free-breathing spatiotemporally accelerated 3-D time-resolved contrast-enhanced MR angiography frequently yields diagnostic image quality for most abdominal arteries in young children.

Investigation of temporal-resolved emission spectra of highly charged Al ions from laser-produced plasmas

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

Su, M. G., E-mail: sumg@nwnu.edu.cn; Sun, D. X.; Dong, C. Z.

2016-03-15

Temporal evolution of extreme ultraviolet emission from laser-produced aluminum (Al) plasma has been experimentally and theoretically investigated. Al plasmas have been measured by using the temporal-spatially resolved laser-produced plasma technique. The emission lines can be identified from 2p-3s, 3d, 4s, 4d, 5d transition lines from Al{sup 3+} to Al{sup 6+} ions. In order to quickly diagnose the plasma, the assumptions of a normalized Boltzmann distribution among the excited states and a steady-state collisional-radiative model are used to estimate the values of electron temperature and electron density in plasma. We succeeded in reproducing the simulated spectra related to the different timemore » delays, which are in good agreement with experiments. Temporal evolution behavior of highly charged Al ions in plasma has been analyzed, and the exponential decay about electron temperature and electron density has been obtained. The results indicate that the temporal-spatially resolved measurement is essential for accurate understanding of evolution behavior of highly charged ions in laser-produced plasmas.« less

In-situ straining and time-resolved electron tomography data acquisition in a transmission electron microscope.

PubMed

Hata, S; Miyazaki, S; Gondo, T; Kawamoto, K; Horii, N; Sato, K; Furukawa, H; Kudo, H; Miyazaki, H; Murayama, M

2017-04-01

This paper reports the preliminary results of a new in-situ three-dimensional (3D) imaging system for observing plastic deformation behavior in a transmission electron microscope (TEM) as a directly relevant development of the recently reported straining-and-tomography holder [Sato K et al. (2015) Development of a novel straining holder for transmission electron microscopy compatible with single tilt-axis electron tomography. Microsc. 64: 369-375]. We designed an integrated system using the holder and newly developed straining and image-acquisition software and then developed an experimental procedure for in-situ straining and time-resolved electron tomography (ET) data acquisition. The software for image acquisition and 3D visualization was developed based on the commercially available ET software TEMographyTM. We achieved time-resolved 3D visualization of nanometer-scale plastic deformation behavior in a Pb-Sn alloy sample, thus demonstrating the capability of this system for potential applications in materials science. © The Author 2016. Published by Oxford University Press on behalf of The Japanese Society of Microscopy. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Modeling Intrajunction Dispersion at a Well-Mixed Tidal River Junction

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

Wolfram, Phillip J.; Fringer, Oliver B.; Monsen, Nancy E.

In this paper, the relative importance of small-scale, intrajunction flow features such as shear layers, separation zones, and secondary flows on dispersion in a well-mixed tidal river junction is explored. A fully nonlinear, nonhydrostatic, and unstructured three-dimensional (3D) model is used to resolve supertidal dispersion via scalar transport at a well-mixed tidal river junction. Mass transport simulated in the junction is compared against predictions using a simple node-channel model to quantify the effects of small-scale, 3D intrajunction flow features on mixing and dispersion. The effects of three-dimensionality are demonstrated by quantifying the difference between two-dimensional (2D) and 3D model results.more » An intermediate 3D model that does not resolve the secondary circulation or the recirculating flow at the junction is also compared to the 3D model to quantify the relative sensitivity of mixing on intrajunction flow features. Resolution of complex flow features simulated by the full 3D model is not always necessary because mixing is primarily governed by bulk flow splitting due to the confluence–diffluence cycle. Finally, results in 3D are comparable to the 2D case for many flow pathways simulated, suggesting that 2D modeling may be reasonable for nonstratified and predominantly hydrostatic flows through relatively straight junctions, but not necessarily for the full junction network.« less

Modeling Intrajunction Dispersion at a Well-Mixed Tidal River Junction

DOE PAGES

Wolfram, Phillip J.; Fringer, Oliver B.; Monsen, Nancy E.; ...

2016-08-01

In this paper, the relative importance of small-scale, intrajunction flow features such as shear layers, separation zones, and secondary flows on dispersion in a well-mixed tidal river junction is explored. A fully nonlinear, nonhydrostatic, and unstructured three-dimensional (3D) model is used to resolve supertidal dispersion via scalar transport at a well-mixed tidal river junction. Mass transport simulated in the junction is compared against predictions using a simple node-channel model to quantify the effects of small-scale, 3D intrajunction flow features on mixing and dispersion. The effects of three-dimensionality are demonstrated by quantifying the difference between two-dimensional (2D) and 3D model results.more » An intermediate 3D model that does not resolve the secondary circulation or the recirculating flow at the junction is also compared to the 3D model to quantify the relative sensitivity of mixing on intrajunction flow features. Resolution of complex flow features simulated by the full 3D model is not always necessary because mixing is primarily governed by bulk flow splitting due to the confluence–diffluence cycle. Finally, results in 3D are comparable to the 2D case for many flow pathways simulated, suggesting that 2D modeling may be reasonable for nonstratified and predominantly hydrostatic flows through relatively straight junctions, but not necessarily for the full junction network.« less

The relative abundances of resolved l2CH2D2 and 13CH3D and mechanisms controlling isotopic bond ordering in abiotic and biotic methane gases

NASA Astrophysics Data System (ADS)

Young, E. D.; Kohl, I. E.; Lollar, B. Sherwood; Etiope, G.; Rumble, D.; Li, S.; Haghnegahdar, M. A.; Schauble, E. A.; McCain, K. A.; Foustoukos, D. I.; Sutclife, C.; Warr, O.; Ballentine, C. J.; Onstott, T. C.; Hosgormez, H.; Neubeck, A.; Marques, J. M.; Pérez-Rodríguez, I.; Rowe, A. R.; LaRowe, D. E.; Magnabosco, C.; Yeung, L. Y.; Ash, J. L.; Bryndzia, L. T.

2017-04-01

We report measurements of resolved 12CH2D2 and 13CH3D at natural abundances in a variety of methane gases produced naturally and in the laboratory. The ability to resolve 12CH2D2 from 13CH3D provides unprecedented insights into the origin and evolution of CH4. The results identify conditions under which either isotopic bond order disequilibrium or equilibrium are expected. Where equilibrium obtains, concordant Δ12CH2D2 and Δ13CH3D temperatures can be used reliably for thermometry. We find that concordant temperatures do not always match previous hypotheses based on indirect estimates of temperature of formation nor temperatures derived from CH4/H2 D/H exchange, underscoring the importance of reliable thermometry based on the CH4 molecules themselves. Where Δ12CH2D2 and Δ13CH3D values are inconsistent with thermodynamic equilibrium, temperatures of formation derived from these species are spurious. In such situations, while formation temperatures are unavailable, disequilibrium isotopologue ratios nonetheless provide novel information about the formation mechanism of the gas and the presence or absence of multiple sources or sinks. In particular, disequilibrium isotopologue ratios may provide the means for differentiating between methane produced by abiotic synthesis vs. biological processes. Deficits in 12CH2D2 compared with equilibrium values in CH4 gas made by surface-catalyzed abiotic reactions are so large as to point towards a quantum tunneling origin. Tunneling also accounts for the more moderate depletions in 13CH3D that accompany the low 12CH2D2 abundances produced by abiotic reactions. The tunneling signature may prove to be an important tracer of abiotic methane formation, especially where it is preserved by dissolution of gas in cool hydrothermal systems (e.g., Mars). Isotopologue signatures of abiotic methane production can be erased by infiltration of microbial communities, and Δ12CH2D2 values are a key tracer of microbial recycling.

The Relative Abundances of Resolved 12CH2D2 and 13CH3D and Mechanisms Controlling Isotopic Bond Ordering in Abiotic and Biotic Methane Gases

NASA Astrophysics Data System (ADS)

Young, E. D.; Kohl, I. E.; Sherwood Lollar, B.; Etiope, G.; Rumble, D.; Li, S.; Haghnegahdar, M. A.; Schauble, E. A.; McCain, K.; Foustoukos, D.; Sutcliffe, C. N.; Warr, O.; Ballentine, C. J.; Onstott, T. C.; Hosgormez, H.; Neubeck, A.; Marques, J. M.; Perez-Rodriguez, I. M.; Rowe, A. R.; LaRowe, D.; Magnabosco, C.; Bryndzia, T.

2016-12-01

We report measurements of resolved 12CH2D2 and 13CH3D at natural abundances in a variety of methane gases produced naturally and in the laboratory. The ability to resolve 12CH2D2 from 13CH3D provides unprecedented insights into the origin and evolution of CH4. The results identify conditions under which either isotopic bond order disequilibrium or equilibrium are expected. Where equilibrium obtains, concordant Δ12CH2D2 and Δ13CH3D temperatures can be used reliably for thermometry. We find that concordant temperatures do not always match previous hypotheses based on indirect estimates of temperature of formation nor temperatures derived from CH4/H2 D/H exchange, underscoring the importance of reliable thermometry based on the CH4 molecules themselves. Where Δ12CH2D2 and Δ13CH3D values are inconsistent with thermodynamic equilibrium, temperatures of formation derived from these species are spurious. In such situations, while formation temperatures are unavailable, disequilibrium isotopologue ratios nonetheless provide important information about the formation mechanism of the gas and the presence or absence of multiple sources or sinks. In particular, disequilibrium isotopologue ratios may provide the means for differentiating between methane produced by abiotic synthesis versus biological processes. Deficits in 12CH2D2 compared with equilibrium values in CH4 gas made by surface-catalyzed abiotic reactions are so large as to point towards a quantum tunneling origin. Tunneling also accounts for the more moderate depletions in 13CH3D that accompany the low 12CH2D2 abundances produced by abiotic reactions. The tunneling signature of abiotic CH4 formation may prove to be an important tracer of abiotic methane formation, especially where it is preserved by dissolution of gas in cool hydrothermal systems (e.g., Mars). Isotopologue signatures of abiotic methane production can be erased by infiltration of microbial communities, and Δ12CH2D2 values are a key tracer of microbial recycling.

Two-photon holographic optogenetics of neural circuits (Conference Presentation)

NASA Astrophysics Data System (ADS)

Yang, Weijian; Carrillo-Reid, Luis; Peterka, Darcy S.; Yuste, Rafael

2016-03-01

Optical manipulation of in vivo neural circuits with cellular resolution could be important for understanding cortical function. Despite recent progress, simultaneous optogenetic activation with cellular precision has either been limited to 2D planes, or a very small numbers of neurons over a limited volume. Here we demonstrate a novel paradigm for simultaneous 3D activation using a low repetition rate pulse-amplified fiber laser system and a spatial light modulator (SLM) to project 3D holographic excitation patterns on the cortex of mice in vivo for targeted volumetric 3D photoactivation. This method is compatible with two-photon imaging, and enables the simultaneous activation of multiple cells in 3D, using red-shifted opsins, such as C1V1 or ReaChR, while simultaneously imaging GFP-based sensors such as GCaMP6. This all-optical imaging and 3D manipulation approach achieves simultaneous reading and writing of cortical activity, and should be a powerful tool for the study of neuronal circuits.

Direct aqueous measurement of 25-hydroxyvitamin D levels in a cellular environment by LC-MS/MS using the novel chemical derivatization reagent MDBP.

PubMed

Müller, Miriam J; Bruns, Heiko; Volmer, Dietrich A

2017-04-01

Vitamin D measurements in biological fluids by mass spectrometry are challenging at very low concentration levels. As a result, chemical derivatization is often employed to enhance the ionization properties of low abundant vitamin D compounds. Cookson-type reagents such as 4-phenyl-1,2,4-triazoline-3,5-dione (PTAD) or similar derivatives work well but require careful, water-free experimental conditions, as traces of water inactivate the reagent and inhibit or stop the derivatization reactions, thus making quantitative measurements in aqueous samples impossible. We describe a novel electrospray liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay for determining 25-hydroxyvitamin D 3 (25(OH)D 3 ) directly in aqueous cellular systems using a new derivatization reagent, the ionic liquid 12-(maleimidyl)dodecyl-tri-n-butylphosphonium bromide (MDBP). The proof-of-concept for the MDBP assay was demonstrated by measuring the levels of 25(OH)D 3 in four different human cell types, namely T cells, helper T cells, B cells, and macrophages. In addition to the ability to determine the levels of 25(OH)D 3 directly in aqueous samples, the cellular integrity was maintained in our application. We show the time-dependent uptake of 25(OH)D 3 into the investigated cells to demonstrate the applicability of the new label. Furthermore, the MDBP derivatization technique may be equally useful in imaging mass spectrometry, where it could be used for response enhancements of spatially localized vitamin D metabolites on wet tissue surfaces, without destroying the integrity of the tissue surface. Graphical Abstract MDBP labelling of 25-hydroxyvitamin D in the extracellular space.

The Concerted Action of Type 2 and Type 3 Deiodinases Regulates the Cell Cycle and Survival of Basal Cell Carcinoma Cells.

PubMed

Miro, Caterina; Ambrosio, Raffaele; De Stefano, Maria Angela; Di Girolamo, Daniela; Di Cicco, Emery; Cicatiello, Annunziata Gaetana; Mancino, Giuseppina; Porcelli, Tommaso; Raia, Maddalena; Del Vecchio, Luigi; Salvatore, Domenico; Dentice, Monica

2017-04-01

Thyroid hormones (THs) mediate pleiotropic cellular processes involved in metabolism, cellular proliferation, and differentiation. The intracellular hormonal environment can be tailored by the type 1 and 2 deiodinase enzymes D2 and D3, which catalyze TH activation and inactivation respectively. In many cellular systems, THs exert well-documented stimulatory or inhibitory effects on cell proliferation; however, the molecular mechanisms by which they control rates of cell cycle progression have not yet been entirely clarified. We previously showed that D3 depletion or TH treatment influences the proliferation and survival of basal cell carcinoma (BCC) cells. Surprisingly, we also found that BCC cells express not only sustained levels of D3 but also robust levels of D2. The aim of the present study was to dissect the contribution of D2 to TH metabolism in the BCC context, and to identify the molecular changes associated with cell proliferation and survival induced by TH and mediated by D2 and D3. We used the CRISPR/Cas9 technology to genetically deplete D2 and D3 in BCC cells and studied the consequences of depletion on cell cycle progression and on cell death. Cell cycle progression was analyzed by fluorescence activated cell sorting analysis of synchronized cells, and the apoptosis rate by annexin V incorporation. Mechanistic investigations revealed that D2 inactivation accelerates cell cycle progression thereby enhancing the proportion of S-phase cells and cyclin D1 expression. Conversely, D3 mutagenesis drastically suppressed cell proliferation and enhanced apoptosis of BCC cells. Furthermore, the basal apoptotic rate was oppositely regulated in D2- and D3-depleted cells. Our results indicate that BCC cells constitute an example in which the TH signal is finely tuned by the concerted expression of opposite-acting deiodinases. The dual regulation of D2 and D3 expression plays a critical role in cell cycle progression and cell death by influencing cyclin D1-mediated entry into the G1-S phase. These findings reinforce the concept that TH is a potential therapeutic target in human BCC.

Three-Dimensional Reconstruction of Cloud-to-Ground Lightning Using High-Speed Video and VHF Broadband Interferometer

NASA Astrophysics Data System (ADS)

Li, Yun; Qiu, Shi; Shi, Lihua; Huang, Zhengyu; Wang, Tao; Duan, Yantao

2017-12-01

The time resolved three-dimensional (3-D) spatial reconstruction of lightning channels using high-speed video (HSV) images and VHF broadband interferometer (BITF) data is first presented in this paper. Because VHF and optical radiations in step formation process occur with time separation no more than 1 μs, the observation data of BITF and HSV at two different sites provide the possibility of reconstructing the time resolved 3-D channel of lightning. With the proposed procedures for 3-D reconstruction of leader channels, dart leaders as well as stepped leaders with complex multiple branches can be well reconstructed. The differences between 2-D speeds and 3-D speeds of leader channels are analyzed by comparing the development of leader channels in 2-D and 3-D space. Since return stroke (RS) usually follows the path of previous leader channels, the 3-D speeds of the return strokes are first estimated by combination with the 3-D structure of the preceding leaders and HSV image sequences. For the fourth RS, the ratios of the 3-D to 2-D RS speeds increase with height, and the largest ratio of the 3-D to 2-D return stroke speeds can reach 2.03, which is larger than the result of triggered lightning reported by Idone. Since BITF can detect lightning radiation in a 360° view, correlated BITF and HSV observations increase the 3-D detection probability than dual-station HSV observations, which is helpful to obtain more events and deeper understanding of the lightning process.

48 CFR 229.101 - Resolving tax problems.

Code of Federal Regulations, 2010 CFR

2010-10-01

... 48 Federal Acquisition Regulations System 3 2010-10-01 2010-10-01 false Resolving tax problems. 229.101 Section 229.101 Federal Acquisition Regulations System DEFENSE ACQUISITION REGULATIONS SYSTEM...) Within DoD, the agency-designated legal counsels are the defense agency General Counsels, the General...

Biomimetic Graphene-Based 3D Scaffold for Long-Term Cell Culture and Real-Time Electrochemical Monitoring.

PubMed

Hu, Xue-Bo; Liu, Yan-Ling; Wang, Wen-Jie; Zhang, Hai-Wei; Qin, Yu; Guo, Shan; Zhang, Xin-Wei; Fu, Lei; Huang, Wei-Hua

2018-01-16

Current achievements on electrochemical monitoring of cells are often gained on two-dimensional (2D) substrates, which fail in mimicking the cellular environments and accurately reproducing the cellular functions within a three-dimensional (3D) tissue. In this regard, 3D scaffold concurrently integrated with the function of cell culture and electrochemical sensing is conceivably a promising platform to monitor cells in real time under their in vivo-like 3D microenvironments. However, it is particularly challenging to construct such a multifunctional scaffold platform. Herein, we developed a 3-aminophenylboronic acid (APBA) functionalized graphene foam (GF) network, which combines the biomimetic property of APBA with the mechanical and electrochemical properties of GF. Hence, the GF network can serve as a 3D scaffold to culture cells for a long period with high viability and simultaneously as an electrode for highly sensitive electrochemical sensing. This allows monitoring of gaseous messengers H 2 S released from the cells cultured on the 3D scaffold in real time. This work represents considerable progress in fabricating 3D cell culture scaffold with electrochemical properties, thereby facilitating future studies of physiologically relevant processes.

TU-F-17A-04: Respiratory Phase-Resolved 3D MRI with Isotropic High Spatial Resolution: Determination of the Average Breathing Motion Pattern for Abdominal Radiotherapy Planning

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

Deng, Z; Pang, J; Yang, W

Purpose: To develop a retrospective 4D-MRI technique (respiratory phase-resolved 3D-MRI) for providing an accurate assessment of tumor motion secondary to respiration. Methods: A 3D projection reconstruction (PR) sequence with self-gating (SG) was developed for 4D-MRI on a 3.0T MRI scanner. The respiration-induced shift of the imaging target was recorded by SG signals acquired in the superior-inferior direction every 15 radial projections (i.e. temporal resolution 98 ms). A total of 73000 radial projections obtained in 8-min were retrospectively sorted into 10 time-domain evenly distributed respiratory phases based on the SG information. Ten 3D image sets were then reconstructed offline. The techniquemore » was validated on a motion phantom (gadolinium-doped water-filled box, frequency of 10 and 18 cycles/min) and humans (4 healthy and 2 patients with liver tumors). Imaging protocol included 8-min 4D-MRI followed by 1-min 2D-realtime (498 ms/frame) MRI as a reference. Results: The multiphase 3D image sets with isotropic high spatial resolution (1.56 mm) permits flexible image reformatting and visualization. No intra-phase motion-induced blurring was observed. Comparing to 2D-realtime, 4D-MRI yielded similar motion range (phantom: 10.46 vs. 11.27 mm; healthy subject: 25.20 vs. 17.9 mm; patient: 11.38 vs. 9.30 mm), reasonable displacement difference averaged over the 10 phases (0.74mm; 3.63mm; 1.65mm), and excellent cross-correlation (0.98; 0.96; 0.94) between the two displacement series. Conclusion: Our preliminary study has demonstrated that the 4D-MRI technique can provide high-quality respiratory phase-resolved 3D images that feature: a) isotropic high spatial resolution, b) a fixed scan time of 8 minutes, c) an accurate estimate of average motion pattern, and d) minimal intra-phase motion artifact. This approach has the potential to become a viable alternative solution to assess the impact of breathing on tumor motion and determine appropriate treatment margins. Comparison with 4D-CT in a clinical setting is warranted to assess the value of 4D-MRI in radiotherapy planning. This work supported in part by grant 1R03CA173273-01.« less

A Quantitative Three-Dimensional Image Analysis Tool for Maximal Acquisition of Spatial Heterogeneity Data.

PubMed

Allenby, Mark C; Misener, Ruth; Panoskaltsis, Nicki; Mantalaris, Athanasios

2017-02-01

Three-dimensional (3D) imaging techniques provide spatial insight into environmental and cellular interactions and are implemented in various fields, including tissue engineering, but have been restricted by limited quantification tools that misrepresent or underutilize the cellular phenomena captured. This study develops image postprocessing algorithms pairing complex Euclidean metrics with Monte Carlo simulations to quantitatively assess cell and microenvironment spatial distributions while utilizing, for the first time, the entire 3D image captured. Although current methods only analyze a central fraction of presented confocal microscopy images, the proposed algorithms can utilize 210% more cells to calculate 3D spatial distributions that can span a 23-fold longer distance. These algorithms seek to leverage the high sample cost of 3D tissue imaging techniques by extracting maximal quantitative data throughout the captured image.

Feasibility and Biocompatibility of 3D-Printed Photopolymerized and Laser Sintered Polymers for Neuronal, Myogenic, and Hepatic Cell Types.

PubMed

Rimington, Rowan P; Capel, Andrew J; Player, Darren J; Bibb, Richard J; Christie, Steven D R; Lewis, Mark P

2018-06-13

The integration of additive manufacturing (AM) technology within biological systems holds significant potential, specifically when refining the methods utilized for the creation of in vitro models. Therefore, examination of cellular interaction with the physical/physicochemical properties of 3D-printed polymers is critically important. In this work, skeletal muscle (C 2 C 12 ), neuronal (SH-SY5Y) and hepatic (HepG2) cell lines are utilized to ascertain critical evidence of cellular behavior in response to 3D-printed candidate polymers: Clear-FL (stereolithography, SL), PA-12 (laser sintering, LS), and VeroClear (PolyJet). This research outlines initial critical evidence for a framework of polymer/AM process selection when 3D printing biologically receptive scaffolds, derived from industry standard, commercially available AM instrumentation. C 2 C 12 , SH-SY5Y, and HepG2 cells favor LS polymer PA-12 for applications in which cellular adherence is necessitated. However, cell type specific responses are evident when cultured in the chemical leachate of photopolymers (Clear-FL and VeroClear). With the increasing prevalence of 3D-printed biointerfaces, the development of rigorous cell type specific biocompatibility data is imperative. Supplementing the currently limited database of functional 3D-printed biomaterials affords the opportunity for experiment-specific AM process and polymer selection, dependent on biological application and intricacy of design features required. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

1α,25-Dihydroxyvitamin D3-3β-bromoacetate, a potential cancer therapeutic agent: synthesis and molecular mechanism of action

PubMed Central

Ray, Rahul; Lambert, James R.

2011-01-01

Synthesis of 1α,25-dihydroxyvitamin D3-3β-bromoacetate (1,25(OH)2D3-3-BE), a potential anti-cancer agent is presented. We also report that mechanism of action of 1,25(OH)2D3-3-BE may involve reduction of its catabolism, as evidenced by the reduced and delayed expression of 1α,25-dihydroxyvitamin D3-24-hydroxylase (CYP24) gene in cellular assays. PMID:21392983

Adaptive-optics SLO imaging combined with widefield OCT and SLO enables precise 3D localization of fluorescent cells in the mouse retina.

PubMed

Zawadzki, Robert J; Zhang, Pengfei; Zam, Azhar; Miller, Eric B; Goswami, Mayank; Wang, Xinlei; Jonnal, Ravi S; Lee, Sang-Hyuck; Kim, Dae Yu; Flannery, John G; Werner, John S; Burns, Marie E; Pugh, Edward N

2015-06-01

Adaptive optics scanning laser ophthalmoscopy (AO-SLO) has recently been used to achieve exquisite subcellular resolution imaging of the mouse retina. Wavefront sensing-based AO typically restricts the field of view to a few degrees of visual angle. As a consequence the relationship between AO-SLO data and larger scale retinal structures and cellular patterns can be difficult to assess. The retinal vasculature affords a large-scale 3D map on which cells and structures can be located during in vivo imaging. Phase-variance OCT (pv-OCT) can efficiently image the vasculature with near-infrared light in a label-free manner, allowing 3D vascular reconstruction with high precision. We combined widefield pv-OCT and SLO imaging with AO-SLO reflection and fluorescence imaging to localize two types of fluorescent cells within the retinal layers: GFP-expressing microglia, the resident macrophages of the retina, and GFP-expressing cone photoreceptor cells. We describe in detail a reflective afocal AO-SLO retinal imaging system designed for high resolution retinal imaging in mice. The optical performance of this instrument is compared to other state-of-the-art AO-based mouse retinal imaging systems. The spatial and temporal resolution of the new AO instrumentation was characterized with angiography of retinal capillaries, including blood-flow velocity analysis. Depth-resolved AO-SLO fluorescent images of microglia and cone photoreceptors are visualized in parallel with 469 nm and 663 nm reflectance images of the microvasculature and other structures. Additional applications of the new instrumentation are discussed.

Freeform inkjet printing of cellular structures with bifurcations.

PubMed

Christensen, Kyle; Xu, Changxue; Chai, Wenxuan; Zhang, Zhengyi; Fu, Jianzhong; Huang, Yong

2015-05-01

Organ printing offers a great potential for the freeform layer-by-layer fabrication of three-dimensional (3D) living organs using cellular spheroids or bioinks as building blocks. Vascularization is often identified as a main technological barrier for building 3D organs. As such, the fabrication of 3D biological vascular trees is of great importance for the overall feasibility of the envisioned organ printing approach. In this study, vascular-like cellular structures are fabricated using a liquid support-based inkjet printing approach, which utilizes a calcium chloride solution as both a cross-linking agent and support material. This solution enables the freeform printing of spanning and overhang features by providing a buoyant force. A heuristic approach is implemented to compensate for the axially-varying deformation of horizontal tubular structures to achieve a uniform diameter along their axial directions. Vascular-like structures with both horizontal and vertical bifurcations have been successfully printed from sodium alginate only as well as mouse fibroblast-based alginate bioinks. The post-printing fibroblast cell viability of printed cellular tubes was found to be above 90% even after a 24 h incubation, considering the control effect. © 2014 Wiley Periodicals, Inc.

Time-resolved fluorescence microscopy to study biologically related applications using sol-gel derived and cellular media

NASA Astrophysics Data System (ADS)

Toury, Marion; Chandler, Lin; Allison, Archie; Campbell, David; McLoskey, David; Holmes-Smith, A. Sheila; Hungerford, Graham

2011-03-01

Fluorescence microscopy provides a non-invasive means for visualising dynamic protein interactions. As well as allowing the calculation of kinetic processes via the use of time-resolved fluorescence, localisation of the protein within cells or model systems can be monitored. These fluorescence lifetime images (FLIM) have become the preferred technique for elucidating protein dynamics due to the fact that the fluorescence lifetime is an absolute measure, in the main independent of fluorophore concentration and intensity fluctuations caused by factors such as photobleaching. In this work we demonstrate the use of a time-resolved fluorescence microscopy, employing a high repetition rate laser excitation source applied to study the influence of a metal surface on fluorescence tagged protein and to elucidate viscosity using the fluorescence lifetime probe DASPMI. These were studied in a cellular environment (yeast) and in a model system based on a sol-gel derived material, in which silver nanostructures were formed in situ using irradiation from a semiconductor laser in CW mode incorporated on a compact time-resolved fluorescence microscope (HORIBA Scientific DeltaDiode and DynaMyc).

Quantitative mapping of zinc fluxes in the mammalian egg reveals the origin of fertilization-induced zinc sparks

PubMed Central

Que, Emily L.; Bleher, Reiner; Duncan, Francesca E.; Kong, Betty Y.; Gleber, Sophie C.; Vogt, Stefan; Chen, Si; Garwin, Seth A.; Bayer, Amanda R.; Dravid, Vinayak; Woodruff, Teresa K.; O’Halloran, Thomas V.

2015-01-01

Fertilization of a mammalian egg induces a series of ‘zinc sparks’ that are necessary for inducing the egg-to-embryo transition. Despite the importance of these zinc efflux events little is known about their origin. To understand the molecular mechanism of the zinc spark we combined four physical approaches to resolve zinc distributions in single cells: a chemical probe for dynamic live-cell fluorescence imaging and a combination of scanning transmission electron microscopy with energy dispersive spectroscopy, X-ray fluorescence microscopy, and 3D elemental tomography for high resolution elemental mapping. We show that the zinc spark arises from a system of thousands of zinc-loaded vesicles, each of which contains, on average, 106 zinc atoms. These vesicles undergo dynamic movement during oocyte maturation and exocytosis at the time of fertilization. The discovery of these vesicles and the demonstration that zinc sparks originate from them provides a quantitative framework for understanding how zinc fluxes regulate cellular processes. PMID:25615666

An upwind method for the solution of the 3D Euler and Navier-Stokes equations on adaptively refined meshes

NASA Astrophysics Data System (ADS)

Aftosmis, Michael J.

1992-10-01

A new node based upwind scheme for the solution of the 3D Navier-Stokes equations on adaptively refined meshes is presented. The method uses a second-order upwind TVD scheme to integrate the convective terms, and discretizes the viscous terms with a new compact central difference technique. Grid adaptation is achieved through directional division of hexahedral cells in response to evolving features as the solution converges. The method is advanced in time with a multistage Runge-Kutta time stepping scheme. Two- and three-dimensional examples establish the accuracy of the inviscid and viscous discretization. These investigations highlight the ability of the method to produce crisp shocks, while accurately and economically resolving viscous layers. The representation of these and other structures is shown to be comparable to that obtained by structured methods. Further 3D examples demonstrate the ability of the adaptive algorithm to effectively locate and resolve multiple scale features in complex 3D flows with many interacting, viscous, and inviscid structures.

Hyperfine Structure Constants of Energetically High-lying Levels of Odd Parity of Atomic Vanadium

NASA Astrophysics Data System (ADS)

Güzelçimen, F.; Yapıcı, B.; Demir, G.; Er, A.; Öztürk, I. K.; Başar, Gö.; Kröger, S.; Tamanis, M.; Ferber, R.; Docenko, D.; Başar, Gü.

2014-09-01

High-resolution Fourier transform spectra of a vanadium-argon plasma have been recorded in the wavelength range of 365-670 nm (15,000-27,400 cm-1). Optical bandpass filters were used in the experimental setup to enhance the sensitivity of the Fourier transform spectrometer. In total, 138 atomic vanadium spectral lines showing resolved or partially resolved hyperfine structure have been analyzed to determine the magnetic dipole hyperfine structure constants A of the involved energy levels. One of the investigated lines has not been previously classified. As a result, the magnetic dipole hyperfine structure constants A for 90 energy levels are presented: 35 of them belong to the configuration 3d 34s4p and 55 to the configuration 3d 44p. Of these 90 constants, 67 have been determined for the first time, with 23 corresponding to the configuration 3d 34s4p and 44 to 3d 44p.

Engineering Three-Dimensional Collagen-IKVAV Matrix to Mimic Neural Microenvironment

PubMed Central

2013-01-01

Engineering the cellular microenvironment has great potential to create a platform technology toward engineering of tissue and organs. This study aims to engineer a neural microenvironment through fabrication of three-dimensional (3D) engineered collagen matrixes mimicking in-vivo-like conditions. Collagen was chemically modified with a pentapeptide epitope consisting of isoleucine-lysine-valine-alanine-valine (IKVAV) to mimic laminin structure supports of the neural extracellular matrix (ECM). Three-dimensional collagen matrixes with and without IKVAV peptide modification were fabricated by freeze-drying technology and chemical cross-linking with glutaraldehyde. Structural information of 3D collagen matrixes indicated interconnected pores structure with an average pore size of 180 μm. Our results indicated that culture of dorsal root ganglion (DRG) cells in 3D collagen matrix was greatly influenced by 3D culture method and significantly enhanced with engineered collagen matrix conjugated with IKVAV peptide. It may be concluded that an appropriate 3D culture of neurons enables DRG to positively improve the cellular fate toward further acceleration in tissue regeneration. PMID:23705903

Depth-resolved incoherent and coherent wide-field high-content imaging (Conference Presentation)

NASA Astrophysics Data System (ADS)

So, Peter T.

2016-03-01

Recent advances in depth-resolved wide-field imaging technique has enabled many high throughput applications in biology and medicine. Depth resolved imaging of incoherent signals can be readily accomplished with structured light illumination or nonlinear temporal focusing. The integration of these high throughput systems with novel spectroscopic resolving elements further enable high-content information extraction. We will introduce a novel near common-path interferometer and demonstrate its uses in toxicology and cancer biology applications. The extension of incoherent depth-resolved wide-field imaging to coherent modality is non-trivial. Here, we will cover recent advances in wide-field 3D resolved mapping of refractive index, absorbance, and vibronic components in biological specimens.

Four-dimensional volume-of-interest reconstruction for cone-beam computed tomography-guided radiation therapy.

PubMed

Ahmad, Moiz; Balter, Peter; Pan, Tinsu

2011-10-01

Data sufficiency are a major problem in four-dimensional cone-beam computed tomography (4D-CBCT) on linear accelerator-integrated scanners for image-guided radiotherapy. Scan times must be in the range of 4-6 min to avoid undersampling artifacts. Various image reconstruction algorithms have been proposed to accommodate undersampled data acquisitions, but these algorithms are computationally expensive, may require long reconstruction times, and may require algorithm parameters to be optimized. The authors present a novel reconstruction method, 4D volume-of-interest (4D-VOI) reconstruction which suppresses undersampling artifacts and resolves lung tumor motion for undersampled 1-min scans. The 4D-VOI reconstruction is much less computationally expensive than other 4D-CBCT algorithms. The 4D-VOI method uses respiration-correlated projection data to reconstruct a four-dimensional (4D) image inside a VOI containing the moving tumor, and uncorrelated projection data to reconstruct a three-dimensional (3D) image outside the VOI. Anatomical motion is resolved inside the VOI and blurred outside the VOI. The authors acquired a 1-min. scan of an anthropomorphic chest phantom containing a moving water-filled sphere. The authors also used previously acquired 1-min scans for two lung cancer patients who had received CBCT-guided radiation therapy. The same raw data were used to test and compare the 4D-VOI reconstruction with the standard 4D reconstruction and the McKinnon-Bates (MB) reconstruction algorithms. Both the 4D-VOI and the MB reconstructions suppress nearly all the streak artifacts compared with the standard 4D reconstruction, but the 4D-VOI has 3-8 times greater contrast-to-noise ratio than the MB reconstruction. In the dynamic chest phantom study, the 4D-VOI and the standard 4D reconstructions both resolved a moving sphere with an 18 mm displacement. The 4D-VOI reconstruction shows a motion blur of only 3 mm, whereas the MB reconstruction shows a motion blur of 13 mm. With graphics processing unit hardware used to accelerate computations, the 4D-VOI reconstruction required a 40-s reconstruction time. 4D-VOI reconstruction effectively reduces undersampling artifacts and resolves lung tumor motion in 4D-CBCT. The 4D-VOI reconstruction is computationally inexpensive compared with more sophisticated iterative algorithms. Compared with these algorithms, our 4D-VOI reconstruction is an attractive alternative in 4D-CBCT for reconstructing target motion without generating numerous streak artifacts.

Four-dimensional volume-of-interest reconstruction for cone-beam computed tomography-guided radiation therapy

PubMed Central

Ahmad, Moiz; Balter, Peter; Pan, Tinsu

2011-01-01

Purpose: Data sufficiency are a major problem in four-dimensional cone-beam computed tomography (4D-CBCT) on linear accelerator-integrated scanners for image-guided radiotherapy. Scan times must be in the range of 4–6 min to avoid undersampling artifacts. Various image reconstruction algorithms have been proposed to accommodate undersampled data acquisitions, but these algorithms are computationally expensive, may require long reconstruction times, and may require algorithm parameters to be optimized. The authors present a novel reconstruction method, 4D volume-of-interest (4D-VOI) reconstruction which suppresses undersampling artifacts and resolves lung tumor motion for undersampled 1-min scans. The 4D-VOI reconstruction is much less computationally expensive than other 4D-CBCT algorithms. Methods: The 4D-VOI method uses respiration-correlated projection data to reconstruct a four-dimensional (4D) image inside a VOI containing the moving tumor, and uncorrelated projection data to reconstruct a three-dimensional (3D) image outside the VOI. Anatomical motion is resolved inside the VOI and blurred outside the VOI. The authors acquired a 1-min. scan of an anthropomorphic chest phantom containing a moving water-filled sphere. The authors also used previously acquired 1-min scans for two lung cancer patients who had received CBCT-guided radiation therapy. The same raw data were used to test and compare the 4D-VOI reconstruction with the standard 4D reconstruction and the McKinnon-Bates (MB) reconstruction algorithms. Results: Both the 4D-VOI and the MB reconstructions suppress nearly all the streak artifacts compared with the standard 4D reconstruction, but the 4D-VOI has 3–8 times greater contrast-to-noise ratio than the MB reconstruction. In the dynamic chest phantom study, the 4D-VOI and the standard 4D reconstructions both resolved a moving sphere with an 18 mm displacement. The 4D-VOI reconstruction shows a motion blur of only 3 mm, whereas the MB reconstruction shows a motion blur of 13 mm. With graphics processing unit hardware used to accelerate computations, the 4D-VOI reconstruction required a 40-s reconstruction time. Conclusions: 4D-VOI reconstruction effectively reduces undersampling artifacts and resolves lung tumor motion in 4D-CBCT. The 4D-VOI reconstruction is computationally inexpensive compared with more sophisticated iterative algorithms. Compared with these algorithms, our 4D-VOI reconstruction is an attractive alternative in 4D-CBCT for reconstructing target motion without generating numerous streak artifacts. PMID:21992381

Measuring the Autocorrelation Function of Nanoscale Three-Dimensional Density Distribution in Individual Cells Using Scanning Transmission Electron Microscopy, Atomic Force Microscopy, and a New Deconvolution Algorithm.

PubMed

Li, Yue; Zhang, Di; Capoglu, Ilker; Hujsak, Karl A; Damania, Dhwanil; Cherkezyan, Lusik; Roth, Eric; Bleher, Reiner; Wu, Jinsong S; Subramanian, Hariharan; Dravid, Vinayak P; Backman, Vadim

2017-06-01

Essentially all biological processes are highly dependent on the nanoscale architecture of the cellular components where these processes take place. Statistical measures, such as the autocorrelation function (ACF) of the three-dimensional (3D) mass-density distribution, are widely used to characterize cellular nanostructure. However, conventional methods of reconstruction of the deterministic 3D mass-density distribution, from which these statistical measures can be calculated, have been inadequate for thick biological structures, such as whole cells, due to the conflict between the need for nanoscale resolution and its inverse relationship with thickness after conventional tomographic reconstruction. To tackle the problem, we have developed a robust method to calculate the ACF of the 3D mass-density distribution without tomography. Assuming the biological mass distribution is isotropic, our method allows for accurate statistical characterization of the 3D mass-density distribution by ACF with two data sets: a single projection image by scanning transmission electron microscopy and a thickness map by atomic force microscopy. Here we present validation of the ACF reconstruction algorithm, as well as its application to calculate the statistics of the 3D distribution of mass-density in a region containing the nucleus of an entire mammalian cell. This method may provide important insights into architectural changes that accompany cellular processes.

Measuring the Autocorrelation Function of Nanoscale Three-Dimensional Density Distribution in Individual Cells Using Scanning Transmission Electron Microscopy, Atomic Force Microscopy, and a New Deconvolution Algorithm

PubMed Central

Li, Yue; Zhang, Di; Capoglu, Ilker; Hujsak, Karl A.; Damania, Dhwanil; Cherkezyan, Lusik; Roth, Eric; Bleher, Reiner; Wu, Jinsong S.; Subramanian, Hariharan; Dravid, Vinayak P.; Backman, Vadim

2018-01-01

Essentially all biological processes are highly dependent on the nanoscale architecture of the cellular components where these processes take place. Statistical measures, such as the autocorrelation function (ACF) of the three-dimensional (3D) mass–density distribution, are widely used to characterize cellular nanostructure. However, conventional methods of reconstruction of the deterministic 3D mass–density distribution, from which these statistical measures can be calculated, have been inadequate for thick biological structures, such as whole cells, due to the conflict between the need for nanoscale resolution and its inverse relationship with thickness after conventional tomographic reconstruction. To tackle the problem, we have developed a robust method to calculate the ACF of the 3D mass–density distribution without tomography. Assuming the biological mass distribution is isotropic, our method allows for accurate statistical characterization of the 3D mass–density distribution by ACF with two data sets: a single projection image by scanning transmission electron microscopy and a thickness map by atomic force microscopy. Here we present validation of the ACF reconstruction algorithm, as well as its application to calculate the statistics of the 3D distribution of mass–density in a region containing the nucleus of an entire mammalian cell. This method may provide important insights into architectural changes that accompany cellular processes. PMID:28416035

Mechanisms of DNA Damage Response to Targeted Irradiation in Organotypic 3D Skin Cultures

PubMed Central

Acheva, Anna; Ghita, Mihaela; Patel, Gaurang; Prise, Kevin M.; Schettino, Giuseppe

2014-01-01

DNA damage (caused by direct cellular exposure and bystander signaling) and the complex pathways involved in its repair are critical events underpinning cellular and tissue response following radiation exposures. There are limited data addressing the dynamics of DNA damage induction and repair in the skin particularly in areas not directly exposed. Here we investigate the mechanisms regulating DNA damage, repair, intracellular signalling and their impact on premature differentiation and development of inflammatory-like response in the irradiated and surrounding areas of a 3D organotypic skin model. Following localized low-LET irradiation (225 kVp X-rays), low levels of 53BP1 foci were observed in the 3D model (3.8±0.28 foci/Gy/cell) with foci persisting and increasing in size up to 48 h post irradiation. In contrast, in cell monolayers 14.2±0.6 foci/Gy/cell and biphasic repair kinetics with repair completed before 24 h was observed. These differences are linked to differences in cellular status with variable level of p21 driving apoptotic signalling in 2D and accelerated differentiation in both the directly irradiated and bystander areas of the 3D model. The signalling pathways utilized by irradiated keratinocytes to induce DNA damage in non-exposed areas of the skin involved the NF-κB transcription factor and its downstream target COX-2. PMID:24505255

Fibronectin Deposition Participates in Extracellular Matrix Assembly and Vascular Morphogenesis

PubMed Central

Hielscher, Abigail; Ellis, Kim; Qiu, Connie; Porterfield, Josh; Gerecht, Sharon

2016-01-01

The extracellular matrix (ECM) has been demonstrated to facilitate angiogenesis. In particular, fibronectin has been documented to activate endothelial cells, resulting in their transition from a quiescent state to an active state in which the cells exhibit enhanced migration and proliferation. The goal of this study is to examine the role of polymerized fibronectin during vascular tubulogenesis using a 3 dimensional (3D) cell-derived de-cellularized matrix. A fibronectin-rich 3D de-cellularized ECM was used as a scaffold to study vascular morphogenesis of endothelial cells (ECs). Confocal analyses of several matrix proteins reveal high intra- and extra-cellular deposition of fibronectin in formed vascular structures. Using a small peptide inhibitor of fibronectin polymerization, we demonstrate that inhibition of fibronectin fibrillogenesis in ECs cultured atop de-cellularized ECM resulted in decreased vascular morphogenesis. Further, immunofluorescence and ultrastructural analyses reveal decreased expression of stromal matrix proteins in the absence of polymerized fibronectin with high co-localization of matrix proteins found in association with polymerized fibronectin. Evaluating vascular kinetics, live cell imaging showed that migration, migration velocity, and mean square displacement, are disrupted in structures grown in the absence of polymerized fibronectin. Additionally, vascular organization failed to occur in the absence of a polymerized fibronectin matrix. Consistent with these observations, we tested vascular morphogenesis following the disruption of EC adhesion to polymerized fibronectin, demonstrating that block of integrins α5β1 and αvβ3, abrogated vascular morphogenesis. Overall, fibronectin deposition in a 3D cell-derived de-cellularized ECM appears to be imperative for matrix assembly and vascular morphogenesis. PMID:26811931

Three-dimensional magnetotelluric axial anisotropic forward modeling and inversion

NASA Astrophysics Data System (ADS)

Cao, Hui; Wang, Kunpeng; Wang, Tao; Hua, Boguang

2018-06-01

Magnetotelluric (MT) data has been widely used to image underground electrical structural. However, when the significant axial resistivity anisotropy presents, how this influences three-dimensional MT data has not been resolved clearly yet. We here propose a scheme for three-dimensional modeling of MT data in presence of axial anisotropic resistivity, where the electromagnetic fields are decomposed into primary and secondary components. A 3D staggered-grid finite difference method is then used to resolve the resulting 3D governing equations. Numerical tests have completed to validate the correctness and accuracy of the present algorithm. A limited-memory Broyden-Fletcher-Goldfarb-Shanno method is then utilized to realize the 3D MT axial anisotropic inversion. The testing results show that, compared to the results of isotropic resistivity inversion, taking account the axial anisotropy can much improve the inverted results.

Development of a Time-Resolved Fluorescence Resonance Energy Transfer Ultrahigh-Throughput Screening Assay for Targeting the NSD3 and MYC Interaction. | Office of Cancer Genomics

Cancer.gov

Epigenetic modulators play critical roles in reprogramming of cellular functions, emerging as a new class of promising therapeutic targets. Nuclear receptor binding SET domain protein 3 (NSD3) is a member of the lysine methyltransferase family. Interestingly, the short isoform of NSD3 without the methyltransferase fragment, NSD3S, exhibits oncogenic activity in a wide range of cancers. We recently showed that NSD3S interacts with MYC, a central regulator of tumorigenesis, suggesting a mechanism by which NSD3S regulates cell proliferation through engaging MYC.

Clinical performance of a free-breathing spatiotemporally accelerated 3-D time-resolved contrast-enhanced pediatric abdominal MR angiography

PubMed Central

Yousaf, Ufra; Hsiao, Albert; Cheng, Joseph Y.; Alley, Marcus T.; Lustig, Michael; Pauly, John M.; Vasanawala, Shreyas S.

2015-01-01

Background Pediatric contrast-enhanced MR angiography is often limited by respiration, other patient motion and compromised spatiotemporal resolution. Objective To determine the reliability of a free-breathing spatiotemporally accelerated 3-D time-resolved contrast enhanced MR angiography method for depicting abdominal arterial anatomy in young children. Materials and methods With IRB approval and informed consent, we retrospectively identified 27 consecutive children (16 males and 11 females; mean age: 3.8 years, range: 14 days to 8.4 years) referred for contrast enhanced MR angiography at our institution, who had undergone free-breathing spatiotemporally accelerated time-resolved contrast enhanced MR angiography studies. An radio-frequency-spoiled gradient echo sequence with Cartesian variable density k-space sampling and radial view ordering, intrinsic motion navigation and intermittent fat suppression was developed. Images were reconstructed with soft-gated parallel imaging locally low-rank method to achieve both motion correction and high spatiotemporal resolution. Quality of delineation of 13 abdominal arteries in the reconstructed images was assessed independently by two radiologists on a five-point scale. Ninety-five percent confidence intervals of the proportion of diagnostically adequate cases were calculated. Interobserver agreements were also analyzed. Results Eleven out of 13 arteries achieved acceptable image quality (mean score range: 3.9–5.0) for both readers. Fair to substantial interobserver agreement was reached on nine arteries. Conclusion Free-breathing spatiotemporally accelerated 3-D time-resolved contrast enhanced MR angiography frequently yields diagnostic image quality for most abdominal arteries for pediatric contrast enhanced MR angiography. PMID:26040509

Usefulness of free-breathing readout-segmented echo-planar imaging (RESOLVE) for detection of malignant liver tumors: comparison with single-shot echo-planar imaging (SS-EPI).

PubMed

Tokoro, Hirokazu; Fujinaga, Yasunari; Ohya, Ayumi; Ueda, Kazuhiko; Shiobara, Aya; Kitou, Yoshihiro; Ueda, Hitoshi; Kadoya, Masumi

2014-10-01

We aimed to clarify the usefulness of free-breathing readout-segmented echo-planar imaging (RESOLVE), which is multi-shot echo-planar imaging based on a 2D-navigator-based reacquisition technique, for detecting malignant liver tumor. In 77 patients with malignant liver tumors, free-breathing RESOLVE and respiratory-triggered single-shot echo-planar imaging (SS-EPI) at 3-T MR unit were performed. We set a scan time up to approximately 5 min (300s) before examination, measured actual scan time and assessed (1) susceptibility and (2) motion artifacts in the right and left liver lobes (3, no artifact; 1, marked), and (3) detectability of malignant liver tumors (3, good; 1, poor) using a 3-point scale. The median actual scan time of RESOLVE/SS-EPI was 365/423s. The median scores of each factor in RESOLVE/SS-EPI were as following in this order: (1) 3/2 (right lobe); 3/3 (left lobe), (2) 2/3 (right lobe); 1/2 (left lobe), and (3) 3/3, respectively. Significant differences were noted between RESOLVE and SS-EPI in all evaluated factors (P<0.05) except for susceptibility of left lobe and detectability of the lesions. Despite the effect of motion artifacts, RESOLVE provides a comparable detectability of the lesion and the advantage of reducing scanning time compared with SS-EPI. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

High-strength cellular ceramic composites with 3D microarchitecture.

PubMed

Bauer, Jens; Hengsbach, Stefan; Tesari, Iwiza; Schwaiger, Ruth; Kraft, Oliver

2014-02-18

To enhance the strength-to-weight ratio of a material, one may try to either improve the strength or lower the density, or both. The lightest solid materials have a density in the range of 1,000 kg/m(3); only cellular materials, such as technical foams, can reach considerably lower values. However, compared with corresponding bulk materials, their specific strength generally is significantly lower. Cellular topologies may be divided into bending- and stretching-dominated ones. Technical foams are structured randomly and behave in a bending-dominated way, which is less weight efficient, with respect to strength, than stretching-dominated behavior, such as in regular braced frameworks. Cancellous bone and other natural cellular solids have an optimized architecture. Their basic material is structured hierarchically and consists of nanometer-size elements, providing a benefit from size effects in the material strength. Designing cellular materials with a specific microarchitecture would allow one to exploit the structural advantages of stretching-dominated constructions as well as size-dependent strengthening effects. In this paper, we demonstrate that such materials may be fabricated. Applying 3D laser lithography, we produced and characterized micro-truss and -shell structures made from alumina-polymer composite. Size-dependent strengthening of alumina shells has been observed, particularly when applied with a characteristic thickness below 100 nm. The presented artificial cellular materials reach compressive strengths up to 280 MPa with densities well below 1,000 kg/m(3).

Method to investigate temporal dynamics of ganglion and other retinal cells in the living human eye

NASA Astrophysics Data System (ADS)

Kurokawa, Kazuhiro; Liu, Zhuolin; Crowell, James; Zhang, Furu; Miller, Donald T.

2018-02-01

The inner retina is critical for visual processing, but much remains unknown about its neural circuitry and vulnerability to disease. A major bottleneck has been our inability to observe the structure and function of the cells composing these retinal layers in the living human eye. Here, we present a noninvasive method to observe both structural and functional information. Adaptive optics optical coherence tomography (AO-OCT) is used to resolve the inner retinal cells in all three dimensions and novel post processing algorithms are applied to extract structure and physiology down to the cellular level. AO-OCT captured the 3D mosaic of individual ganglion cell somas, retinal nerve fiber bundles of micron caliber, and microglial cells, all in exquisite detail. Time correlation analysis of the AO-OCT videos revealed notable temporal differences between the principal layers of the inner retina. The GC layer was more dynamic than the nerve fiber and inner plexiform layers. At the cellular level, we applied a customized correlation method to individual GCL somas, and found a mean time constant of activity of 0.57 s and spread of +/-0.1 s suggesting a range of physiological dynamics even in the same cell type. Extending our method to slower dynamics (from minutes to one year), time-lapse imaging and temporal speckle contrast revealed appendage and soma motion of resting microglial cells at the retinal surface.

Self-interference 3D super-resolution microscopy for deep tissue investigations.

PubMed

Bon, Pierre; Linarès-Loyez, Jeanne; Feyeux, Maxime; Alessandri, Kevin; Lounis, Brahim; Nassoy, Pierre; Cognet, Laurent

2018-06-01

Fluorescence localization microscopy has achieved near-molecular resolution capable of revealing ultra-structures, with a broad range of applications, especially in cellular biology. However, it remains challenging to attain such resolution in three dimensions and inside biological tissues beyond the first cell layer. Here we introduce SELFI, a framework for 3D single-molecule localization within multicellular specimens and tissues. The approach relies on self-interference generated within the microscope's point spread function (PSF) to simultaneously encode equiphase and intensity fluorescence signals, which together provide the 3D position of an emitter. We combined SELFI with conventional localization microscopy to visualize F-actin 3D filament networks and reveal the spatial distribution of the transcription factor OCT4 in human induced pluripotent stem cells at depths up to 50 µm inside uncleared tissue spheroids. SELFI paves the way to nanoscale investigations of native cellular processes in intact tissues.

Precipitation Processes Developed During ARM (1997), TOGA COARE (1992), GATE (1974), SCSMEX (1998), and KWAJEX (1999): Consistent 2D, Semi-3D and 3D Cloud Resolving Model Simulations

NASA Technical Reports Server (NTRS)

Tao, W-K.

2003-01-01

Real clouds and cloud systems are inherently three-dimensional (3D). Because of the limitations in computer resources, however, most cloud-resolving models (CRMs) today are still two-dimensional (2D). A few 3D CRMs have been used to study the response of clouds to large-scale forcing. In these 3D simulations, the model domain was small, and the integration time was 6 hours. Only recently have 3D experiments been performed for multi-day periods for tropical cloud systems with large horizontal domains at the National Center for Atmospheric Research (NACAR) and at NASA Goddard Space Flight Center . At Goddard, a 3D Goddard Cumulus Ensemble (GCE) model was used to simulate periods during TOGA COARE, SCSMEX and KWAJEX using 512 by 512 km domain (with 2 km resolution). The results indicate that surface precipitation and latent heating profiles are very similar between the 2D and 3D GCE model simulations. The reason for the strong similarity between the 2D and 3D CRM simulations is that the same observed large-scale advective tendencies of potential temperature, water vapor mixing ratio, and horizontal momentum were used as the main focusing in both the 2D and 3D models. Interestingly, the 2D and 3D versions of the CRM used at CSU showed significant differences in the rainfall and cloud statistics for three ARM cases. The major objectives of this paper are: (1) to assess the performance of the super-parameterization technique, (2) calculate and examine the surface energy (especially radiation) and water budgets, and (3) identify the differences and similarities in the organization and entrainment rates of convection between simulated 2D and 3D cloud systems.

Time-resolved 3D contrast-enhanced MRA on 3.0T: a non-invasive follow-up technique after stent-assisted coil embolization of the intracranial aneurysm.

PubMed

Choi, Jin Woo; Roh, Hong Gee; Moon, Won-Jin; Kim, Na Ra; Moon, Sung Gyu; Kang, Chung Hwan; Chun, Young Il; Kang, Hyun-Seung

2011-01-01

To evaluate the usefulness of time-resolved contrast enhanced magnetic resonance angiography (4D MRA) after stent-assisted coil embolization by comparing it with time of flight (TOF)-MRA. TOF-MRA and 4D MRA were obtained by 3T MRI in 26 patients treated with stent-assisted coil embolization (Enterprise:Neuroform = 7:19). The qualities of the MRA were rated on a graded scale of 0 to 4. We classified completeness of endovascular treatment into three categories. The degree of quality of visualization of the stented artery was compared between TOF and 4D MRA by the Wilcoxon signed rank test. We used the Mann-Whitney U test for comparing the quality of the visualization of the stented artery according to the stent type in each MRA method. The quality in terms of the visualization of the stented arteries in 4D MRA was significantly superior to that in 3D TOF-MRA, regardless of type of the stent (p < 0.001). The quality of the arteries which were stented with Neuroform was superior to that of the arteries stented with Enterprise in 3D TOF (p < 0.001) and 4D MRA (p = 0.008), respectively. 4D MRA provides a higher quality view of the stented parent arteries when compared with TOF.

High-resolution 3D simulations of NIF ignition targets performed on Sequoia with HYDRA

NASA Astrophysics Data System (ADS)

Marinak, M. M.; Clark, D. S.; Jones, O. S.; Kerbel, G. D.; Sepke, S.; Patel, M. V.; Koning, J. M.; Schroeder, C. R.

2015-11-01

Developments in the multiphysics ICF code HYDRA enable it to perform large-scale simulations on the Sequoia machine at LLNL. With an aggregate computing power of 20 Petaflops, Sequoia offers an unprecedented capability to resolve the physical processes in NIF ignition targets for a more complete, consistent treatment of the sources of asymmetry. We describe modifications to HYDRA that enable it to scale to over one million processes on Sequoia. These include new options for replicating parts of the mesh over a subset of the processes, to avoid strong scaling limits. We consider results from a 3D full ignition capsule-only simulation performed using over one billion zones run on 262,000 processors which resolves surface perturbations through modes l = 200. We also report progress towards a high-resolution 3D integrated hohlraum simulation performed using 262,000 processors which resolves surface perturbations on the ignition capsule through modes l = 70. These aim for the most complete calculations yet of the interactions and overall impact of the various sources of asymmetry for NIF ignition targets. This work was performed under the auspices of the Lawrence Livermore National Security, LLC, (LLNS) under Contract No. DE-AC52-07NA27344.

High-resolution, time-resolved MRA provides superior definition of lower-extremity arterial segments compared to 2D time-of-flight imaging.

PubMed

Thornton, F J; Du, J; Suleiman, S A; Dieter, R; Tefera, G; Pillai, K R; Korosec, F R; Mistretta, C A; Grist, T M

2006-08-01

To evaluate a novel time-resolved contrast-enhanced (CE) projection reconstruction (PR) magnetic resonance angiography (MRA) method for identifying potential bypass graft target vessels in patients with Class II-IV peripheral vascular disease. Twenty patients (M:F = 15:5, mean age = 58 years, range = 48-83 years), were recruited from routine MRA referrals. All imaging was performed on a 1.5 T MRI system with fast gradients (Signa LX; GE Healthcare, Waukesha, WI). Images were acquired with a novel technique that combined undersampled PR with a time-resolved acquisition to yield an MRA method with high temporal and spatial resolution. The method is called PR hyper time-resolved imaging of contrast kinetics (PR-hyperTRICKS). Quantitative and qualitative analyses were used to compare two-dimensional (2D) time-of-flight (TOF) and PR-hyperTRICKS in 13 arterial segments per lower extremity. Statistical analysis was performed with the Wilcoxon signed-rank test. Fifteen percent (77/517) of the vessels were scored as missing or nondiagnostic with 2D TOF, but were scored as diagnostic with PR-hyperTRICKS. Image quality was superior with PR-hyperTRICKS vs. 2D TOF (on a four-point scale, mean rank = 3.3 +/- 1.2 vs. 2.9 +/- 1.2, P < 0.0001). PR-hyperTRICKS produced images with high contrast-to-noise ratios (CNR) and high spatial and temporal resolution. 2D TOF images were of inferior quality due to moderate spatial resolution, inferior CNR, greater flow-related artifacts, and absence of temporal resolution. PR-hyperTRICKS provides superior preoperative assessment of lower limb ischemia compared to 2D TOF.

Predictability in cellular automata.

PubMed

Agapie, Alexandru; Andreica, Anca; Chira, Camelia; Giuclea, Marius

2014-01-01

Modelled as finite homogeneous Markov chains, probabilistic cellular automata with local transition probabilities in (0, 1) always posses a stationary distribution. This result alone is not very helpful when it comes to predicting the final configuration; one needs also a formula connecting the probabilities in the stationary distribution to some intrinsic feature of the lattice configuration. Previous results on the asynchronous cellular automata have showed that such feature really exists. It is the number of zero-one borders within the automaton's binary configuration. An exponential formula in the number of zero-one borders has been proved for the 1-D, 2-D and 3-D asynchronous automata with neighborhood three, five and seven, respectively. We perform computer experiments on a synchronous cellular automaton to check whether the empirical distribution obeys also that theoretical formula. The numerical results indicate a perfect fit for neighbourhood three and five, which opens the way for a rigorous proof of the formula in this new, synchronous case.

Associations of vitamin D with insulin resistance, obesity, type 2 diabetes, and metabolic syndrome.

PubMed

Wimalawansa, Sunil J

2018-01-01

The aim of this study is to determine the relationships of vitamin D with diabetes, insulin resistance obesity, and metabolic syndrome. Intra cellular vitamin D receptors and the 1-α hydroxylase enzyme are distributed ubiquitously in all tissues suggesting a multitude of functions of vitamin D. It plays an indirect but an important role in carbohydrate and lipid metabolism as reflected by its association with type 2 diabetes (T2D), metabolic syndrome, insulin secretion, insulin resistance, polycystic ovarian syndrome, and obesity. Peer-reviewed papers, related to the topic were extracted using key words, from PubMed, Medline, and other research databases. Correlations of vitamin D with diabetes, insulin resistance and metabolic syndrome were examined for this evidence-based review. In addition to the well-studied musculoskeletal effects, vitamin D decreases the insulin resistance, severity of T2D, prediabetes, metabolic syndrome, inflammation, and autoimmunity. Vitamin D exerts autocrine and paracrine effects such as direct intra-cellular effects via its receptors and the local production of 1,25(OH) 2 D 3 , especially in muscle and pancreatic β-cells. It also regulates calcium homeostasis and calcium flux through cell membranes, and activation of a cascade of key enzymes and cofactors associated with metabolic pathways. Cross-sectional, observational, and ecological studies reported inverse correlations between vitamin D status with hyperglycemia and glycemic control in patients with T2D, decrease the rate of conversion of prediabetes to diabetes, and obesity. However, no firm conclusions can be drawn from current studies, because (A) studies were underpowered; (B) few were designed for glycemic outcomes, (C) the minimum (or median) serum 25(OH) D levels achieved are not measured or reported; (D) most did not report the use of diabetes medications; (E) some trials used too little (F) others used too large, unphysiological and infrequent doses of vitamin D; and (G) relative paucity of rigorous clinical data on the effects of vitamin D sufficiency on non-calcium endpoints. Although a large number of observational studies support improving T2D, insulin resistance, obesity, and metabolic syndrome with vitamin D adequacy, there is a lack of conclusive evidence from randomized control clinical trials that, these disorders are prevented following optimization of serum levels of 25(OH)D. However, none of the currently conducted clinical studies would resolve these issues. Thus, specifically designed, new clinical studies are needed to be conducted in well-defined populations, following normalizing the serum vitamin D levels in vitamin D deficient prediabetes subjects, to test the hypothesis that hypovitaminosis D worsens these disorders and correction would alleviate it. Copyright © 2016 Elsevier Ltd. All rights reserved.

Tomo-PIV measurements of the flow field in the wake of a sphere

NASA Astrophysics Data System (ADS)

Eshbal, Lior; David, Tom; Rinsky, Vladislav; van Hout, Rene; Greenblatt, David

2017-11-01

A sphere can be considered as a prototypical 3D bluff body. In order to improve our understanding of its 3D wake flow, a combination of time-resolved planar particle image velocimetry (PIV) and tomographic PIV (tomo-PIV) was implemented. Experiments were performed in a closed-loop water channel facility and sphere Reynolds numbers ReD = UD/ ν = 400, 800, 1200 and 2000, where U is the free-stream velocity, ν the kinematic viscosity and D the sphere diameter. The measurement volume (Height x Length x Width, 5 x 5 x 1.5 D3) comprised the sphere and the downstream wake. Tomo-PIV snap-shots were correlated with the time-resolved PIV such that the 3D temporal evolution of the shed vortices became clear. At ReD = 400, this procedure revealed shed hairpin vortices having a vertical plane of symmetry in agreement with many dye visualization studies. However, the measurements also revealed weaker induced hairpins resulting from the interaction of the near-wake flow and the surrounding free stream. These induced vortices were not visible in previous dye and smoke visualizations and have only been observed in simulations. Data processing of the data at higher ReD is currently ongoing. Israel Science Foundation Grant No. 1596/14.

Automated 3D reconstruction of interiors with multiple scan views

NASA Astrophysics Data System (ADS)

Sequeira, Vitor; Ng, Kia C.; Wolfart, Erik; Goncalves, Joao G. M.; Hogg, David C.

1998-12-01

This paper presents two integrated solutions for realistic 3D model acquisition and reconstruction; an early prototype, in the form of a push trolley, and a later prototype in the form of an autonomous robot. The systems encompass all hardware and software required, from laser and video data acquisition, processing and output of texture-mapped 3D models in VRML format, to batteries for power supply and wireless network communications. The autonomous version is also equipped with a mobile platform and other sensors for the purpose of automatic navigation. The applications for such a system range from real estate and tourism (e.g., showing a 3D computer model of a property to a potential buyer or tenant) or as tool for content creation (e.g., creating 3D models of heritage buildings or producing broadcast quality virtual studios). The system can also be used in industrial environments as a reverse engineering tool to update the design of a plant, or as a 3D photo-archive for insurance purposes. The system is Internet compatible: the photo-realistic models can be accessed via the Internet and manipulated interactively in 3D using a common Web browser with a VRML plug-in. Further information and example reconstructed models are available on- line via the RESOLV web-page at http://www.scs.leeds.ac.uk/resolv/.

Structure of native oligomeric Sprouty2 by electron microscopy and its property of electroconductivity.

PubMed

Chen, Feng-Jung; Lee, Kuan-Wei; Lai, Chun-Chieh; Lee, Sue-Ping; Shen, Hsiao-Hsuian; Tsai, Shu-Ping; Liu, Bang-Hung; Wang, Ling-Mei; Liou, Gunn-Guang

2013-09-27

Receptor tyrosine kinases (RTKs) regulate many cellular processes, and Sprouty2 (Spry2) is known as an important regulator of RTK signaling pathways. Therefore, it is worth investigating the properties of Spry2 in more detail. In this study, we found that Spry2 is able to self-assemble into oligomers with a high-affinity KD value of approximately 16nM, as determined through BIAcore surface plasmon resonance analysis. The three-dimensional (3D) structure of Spry2 was resolved using an electron microscopy (EM) single-particle reconstruction approach, which revealed that Spry2 is donut-shaped with two lip-cover domains. Furthermore, the method of energy dispersive spectrum obtained through EM was analyzed to determine the elements carried by Spry2, and the results demonstrated that Spry2 is a silicon- and iron-containing protein. The silicon may contribute to the electroconductivity of Spry2, and this property exhibits a concentration-dependent feature. This study provides the first report of a silicon- and iron-containing protein, and its 3D structure may allow us (1) to study the potential mechanism through the signal transduction is controlled by switching the electronic transfer on or off and (2) to develop a new type of conductor or even semiconductor using biological or half-biological hybrid materials in the future. Copyright © 2013 Elsevier Inc. All rights reserved.

Evaluation of PBS Treatment and PEI Coating Effects on Surface Morphology and Cellular Response of 3D-Printed Alginate Scaffolds.

PubMed

Mendoza García, María A; Izadifar, Mohammad; Chen, Xiongbiao

2017-11-01

Three-dimensional (3D) printing is an emerging technology for the fabrication of scaffolds to repair/replace damaged tissue/organs in tissue engineering. This paper presents our study on 3D printed alginate scaffolds treated with phosphate buffered saline (PBS) and polyethyleneimine (PEI) coating and their impacts on the surface morphology and cellular response of the printed scaffolds. In our study, sterile alginate was prepared by means of the freeze-drying method and then, used to prepare the hydrogel for 3D printing into calcium chloride, forming 3D scaffolds. Scaffolds were treated with PBS for a time period of two days and seven days, respectively, and PEI coating; then they were seeded with Schwann cells (RSC96) for the examination of cellular response (proliferation and differentiation). In addition, swelling and stiffness (Young's modulus) of the treated scaffolds was evaluated, while their surface morphology was assessed using scanning electron microscopy (SEM). SEM images revealed significant changes in scaffold surface morphology due to degradation caused by the PBS treatment over time. Our cell proliferation assessment over seven days showed that a two-day PBS treatment could be more effective than seven-day PBS treatment for improving cell attachment and elongation. While PEI coating of alginate scaffolds seemed to contribute to cell growth, Schwann cells stayed round on the surface of alginate over the period of cell culture. In conclusion, PBS-treatment may offer the potential to induce surface physical cues due to degradation of alginate, which could improve cell attachment post cell-seeding of 3D-printed alginate scaffolds.

SUMO Modification Stabilizes Enterovirus 71 Polymerase 3D To Facilitate Viral Replication

PubMed Central

Liu, Yan; Shu, Bo; Meng, Jin; Zhang, Yuan; Zheng, Caishang; Ke, Xianliang; Gong, Peng; Hu, Qinxue; Wang, Hanzhong

2016-01-01

ABSTRACT Accumulating evidence suggests that viruses hijack cellular proteins to circumvent the host immune system. Ubiquitination and SUMOylation are extensively studied posttranslational modifications (PTMs) that play critical roles in diverse biological processes. Cross talk between ubiquitination and SUMOylation of both host and viral proteins has been reported to result in distinct functional consequences. Enterovirus 71 (EV71), an RNA virus belonging to the family Picornaviridae, is a common cause of hand, foot, and mouth disease. Little is known concerning how host PTM systems interact with enteroviruses. Here, we demonstrate that the 3D protein, an RNA-dependent RNA polymerase (RdRp) of EV71, is modified by small ubiquitin-like modifier 1 (SUMO-1) both during infection and in vitro. Residues K159 and L150/D151/L152 were responsible for 3D SUMOylation as determined by bioinformatics prediction combined with site-directed mutagenesis. Also, primer-dependent polymerase assays indicated that mutation of SUMOylation sites impaired 3D polymerase activity and virus replication. Moreover, 3D is ubiquitinated in a SUMO-dependent manner, and SUMOylation is crucial for 3D stability, which may be due to the interplay between the two PTMs. Importantly, increasing the level of SUMO-1 in EV71-infected cells augmented the SUMOylation and ubiquitination levels of 3D, leading to enhanced replication of EV71. These results together suggested that SUMO and ubiquitin cooperatively regulated EV71 infection, either by SUMO-ubiquitin hybrid chains or by ubiquitin conjugating to the exposed lysine residue through SUMOylation. Our study provides new insight into how a virus utilizes cellular pathways to facilitate its replication. IMPORTANCE Infection with enterovirus 71 (EV71) often causes neurological diseases in children, and EV71 is responsible for the majority of fatalities. Based on a better understanding of interplay between virus and host cell, antiviral drugs against enteroviruses may be developed. As a dynamic cellular process of posttranslational modification, SUMOylation regulates global cellular protein localization, interaction, stability, and enzymatic activity. However, little is known concerning how SUMOylation directly influences virus replication by targeting viral polymerase. Here, we found that EV71 polymerase 3D was SUMOylated during EV71 infection and in vitro. Moreover, the SUMOylation sites were determined, and in vitro polymerase assays indicated that mutations at SUMOylation sites could impair polymerase synthesis. Importantly, 3D is ubiquitinated in a SUMOylation-dependent manner that enhances the stability of the viral polymerase. Our findings indicate that the two modifications likely cooperatively enhance virus replication. Our study may offer a new therapeutic strategy against virus replication. PMID:27630238

Quantitative assessment of surface functionality effects on microglial uptake and retention of PAMAM dendrimers

NASA Astrophysics Data System (ADS)

Liaw, Kevin; Gök, Ozgul; DeRidder, Louis B.; Kannan, Sujatha; Kannan, Rangaramanujam M.

2018-04-01

Dendrimers are a promising class of polymeric nanoparticles for delivery of therapeutics and diagnostics. Polyamidoamine (PAMAM) dendrimers have shown significant efficacy in many animal models, with performance dependent on surface functionalities. Understanding the effects of end groups on biological interactions is critical for rational design of dendrimer-mediated therapies. In this study, we quantify the cellular trafficking kinetics (endocytosis and exocytosis) of generation 4 neutral (D4-OH), cationic (D4-NH2), anionic (D3.5-COOH), and generation 6 neutral (D6-OH) PAMAM dendrimers to investigate the nanoscale effects of surface functionality and size on cellular interactions. Resting and LPS-activated microglia were studied due to their central roles in dendrimer therapies for central nervous system disorders. D4-OH exhibits greater cellular uptake and lower retention than the larger D6-OH. D4-OH and D3.5-COOH exhibit similar trafficking kinetics, while D4-NH2 exhibits significant membrane interactions, resulting in faster cell association but lower internalization. Cationic charge may also enhance vesicular escape for greater cellular retention and preferential partitioning to nuclei. LPS activation further improves uptake of dendrimers, with smaller and cationic dendrimers experiencing the greatest increases in uptake compared to resting microglia. These studies have implications for the dependence of trafficking pathway on dendrimer properties and inform the design of dendrimer constructs tailored to specific therapeutic needs. Cationic dendrimers are ideal for delivering genetic materials to nuclei, but toxicity may be a limiting factor. Smaller, neutral dendrimers are best suited for delivering high levels of therapeutics in acute neuroinflammation, while larger or cationic dendrimers provide robust retention for sustained release of therapeutics in longer-term diseases.

Iao: The New Adaptive Optics Visible Imaging and Photometric System for AEOS

DTIC Science & Technology

2008-09-01

observations of binary stars, asteroids and planets such as Mercury and Mars [2,3,4]. The Visible Imager is also used to take time resolved photometry ...role it takes high spatial resolution imagery of resolved targets. These targets are primarily low Earth orbiting satellites acquired for the...albedo pattern: Comparing the AEOS and TES data sets [5] D.T. Hall et al. 2007, Journal of Spacecraft and Rockets, 44, 910-919, Time - Resolved I-Band

Motion robust high resolution 3D free-breathing pulmonary MRI using dynamic 3D image self-navigator.

PubMed

Jiang, Wenwen; Ong, Frank; Johnson, Kevin M; Nagle, Scott K; Hope, Thomas A; Lustig, Michael; Larson, Peder E Z

2018-06-01

To achieve motion robust high resolution 3D free-breathing pulmonary MRI utilizing a novel dynamic 3D image navigator derived directly from imaging data. Five-minute free-breathing scans were acquired with a 3D ultrashort echo time (UTE) sequence with 1.25 mm isotropic resolution. From this data, dynamic 3D self-navigating images were reconstructed under locally low rank (LLR) constraints and used for motion compensation with one of two methods: a soft-gating technique to penalize the respiratory motion induced data inconsistency, and a respiratory motion-resolved technique to provide images of all respiratory motion states. Respiratory motion estimation derived from the proposed dynamic 3D self-navigator of 7.5 mm isotropic reconstruction resolution and a temporal resolution of 300 ms was successful for estimating complex respiratory motion patterns. This estimation improved image quality compared to respiratory belt and DC-based navigators. Respiratory motion compensation with soft-gating and respiratory motion-resolved techniques provided good image quality from highly undersampled data in volunteers and clinical patients. An optimized 3D UTE sequence combined with the proposed reconstruction methods can provide high-resolution motion robust pulmonary MRI. Feasibility was shown in patients who had irregular breathing patterns in which our approach could depict clinically relevant pulmonary pathologies. Magn Reson Med 79:2954-2967, 2018. © 2017 International Society for Magnetic Resonance in Medicine. © 2017 International Society for Magnetic Resonance in Medicine.

3D critical layers in fully-developed turbulent flows

NASA Astrophysics Data System (ADS)

Saxton-Fox, Theresa; McKeon, Beverley

2016-11-01

Recent work has shown that 3D critical layers drive self-sustaining behavior of exact coherent solutions of the Navier-Stokes equations (Wang et al. 2007; Hall and Sherwin 2010; Park and Graham 2015). This study investigates the role of 3D critical layers in fully-developed turbulent flows. 3D critical layer effects are identified in instantaneous snapshots of turbulent boundary layers in both experimental and DNS data (Wu et al. 2014). Additionally, a 3D critical layer effect is demonstrated to appear using only a few resolvent response modes from the resolvent analysis of McKeon and Sharma 2010, with phase relationships appropriately chosen. Connections are sought to the thin shear layers observed in turbulent boundary layers (Klewicki and Hirschi 2004; Eisma et al. 2015) and to amplitude modulation observations (Mathis et al. 2009; Duvvuri and McKeon 2014). This research is made possible by the Department of Defense through the National Defense & Engineering Graduate Fellowship (NDSEG) Program and by the Air Force Office of Scientific Research Grant # FA9550-12-1-0060. The support of the Center for Turbulence Research (CTR) summer program at Stanford is gratefully acknowledged.

Polarization effects in the interaction between multi-level atoms and two optical fields

NASA Astrophysics Data System (ADS)

Colín-Rodríguez, R.; Flores-Mijangos, J.; Hernández-Gómez, S.; Jáuregui, R.; López-Hernández, O.; Mojica-Casique, C.; Ponciano-Ojeda, F.; Ramírez-Martínez, F.; Sahagún, D.; Volke-Sepúlveda, K.; Jiménez-Mier, J.

2015-06-01

Polarized velocity selective spectra for rubidium atoms in a room temperature cell are presented. The experiments were performed in the lambda configuration (D2 manifold) and in the 5s\\to 5{{p}3/2}\\to 5{{d}j} ladder configuration. For the lambda configuration the effect of the probe beam intensity in the absorption and polarization spectra are compared with results of a rate equation approximation. Good overall agreement between experiment and theory is found. The results indicate different saturation rates for each of the atomic transitions. Distinctive polarization signals with hyperfine-resolved components are found for the ladder 5{{d}3/2} and 5{{d}5/2} upper states. Fluorescence detection of the 420 nm that results from the second step in the cascade decay 5{{d}j}\\to 6{{p}{{j\\prime }}}\\to 5s was used in the ladder experiments. This fluorescence was also used for the detection of the 5{{p}3/2}\\to 6{{p}3/2} electric dipole forbidden transition in atomic rubidium that occurs at 911 nm. The 6{{p}3/2} hyperfine structure was resolved in this continuous wave, non-dipole excitation.

Multidimensional immunolabeling and 4D time-lapse imaging of vital ex vivo lung tissue

PubMed Central

Vierkotten, Sarah; Lindner, Michael; Königshoff, Melanie; Eickelberg, Oliver

2015-01-01

During the last decades, the study of cell behavior was largely accomplished in uncoated or extracellular matrix (ECM)-coated plastic dishes. To date, considerable cell biological efforts have tried to model in vitro the natural microenvironment found in vivo. For the lung, explants cultured ex vivo as lung tissue cultures (LTCs) provide a three-dimensional (3D) tissue model containing all cells in their natural microenvironment. Techniques for assessing the dynamic live interaction between ECM and cellular tissue components, however, are still missing. Here, we describe specific multidimensional immunolabeling of living 3D-LTCs, derived from healthy and fibrotic mouse lungs, as well as patient-derived 3D-LTCs, and concomitant real-time four-dimensional multichannel imaging thereof. This approach allowed the evaluation of dynamic interactions between mesenchymal cells and macrophages with their ECM. Furthermore, fibroblasts transiently expressing focal adhesions markers incorporated into the 3D-LTCs, paving new ways for studying the dynamic interaction between cellular adhesions and their natural-derived ECM. A novel protein transfer technology (FuseIt/Ibidi) shuttled fluorescently labeled α-smooth muscle actin antibodies into the native cells of living 3D-LTCs, enabling live monitoring of α-smooth muscle actin-positive stress fibers in native tissue myofibroblasts residing in fibrotic lesions of 3D-LTCs. Finally, this technique can be applied to healthy and diseased human lung tissue, as well as to adherent cells in conventional two-dimensional cell culture. This novel method will provide valuable new insights into the dynamics of ECM (patho)biology, studying in detail the interaction between ECM and cellular tissue components in their natural microenvironment. PMID:26092995

Compact plane illumination plugin device to enable light sheet fluorescence imaging of multi-cellular organisms on an inverted wide-field microscope

PubMed Central

Guan, Zeyi; Lee, Juhyun; Jiang, Hao; Dong, Siyan; Jen, Nelson; Hsiai, Tzung; Ho, Chih-Ming; Fei, Peng

2015-01-01

We developed a compact plane illumination plugin (PIP) device which enabled plane illumination and light sheet fluorescence imaging on a conventional inverted microscope. The PIP device allowed the integration of microscope with tunable laser sheet profile, fast image acquisition, and 3-D scanning. The device is both compact, measuring approximately 15 by 5 by 5 cm, and cost-effective, since we employed consumer electronics and an inexpensive device molding method. We demonstrated that PIP provided significant contrast and resolution enhancement to conventional microscopy through imaging different multi-cellular fluorescent structures, including 3-D branched cells in vitro and live zebrafish embryos. Imaging with the integration of PIP greatly reduced out-of-focus contamination and generated sharper contrast in acquired 2-D plane images when compared with the stand-alone inverted microscope. As a result, the dynamic fluid domain of the beating zebrafish heart was clearly segmented and the functional monitoring of the heart was achieved. Furthermore, the enhanced axial resolution established by thin plane illumination of PIP enabled the 3-D reconstruction of the branched cellular structures, which leads to the improvement on the functionality of the wide field microscopy. PMID:26819828

Compact plane illumination plugin device to enable light sheet fluorescence imaging of multi-cellular organisms on an inverted wide-field microscope.

PubMed

Guan, Zeyi; Lee, Juhyun; Jiang, Hao; Dong, Siyan; Jen, Nelson; Hsiai, Tzung; Ho, Chih-Ming; Fei, Peng

2016-01-01

We developed a compact plane illumination plugin (PIP) device which enabled plane illumination and light sheet fluorescence imaging on a conventional inverted microscope. The PIP device allowed the integration of microscope with tunable laser sheet profile, fast image acquisition, and 3-D scanning. The device is both compact, measuring approximately 15 by 5 by 5 cm, and cost-effective, since we employed consumer electronics and an inexpensive device molding method. We demonstrated that PIP provided significant contrast and resolution enhancement to conventional microscopy through imaging different multi-cellular fluorescent structures, including 3-D branched cells in vitro and live zebrafish embryos. Imaging with the integration of PIP greatly reduced out-of-focus contamination and generated sharper contrast in acquired 2-D plane images when compared with the stand-alone inverted microscope. As a result, the dynamic fluid domain of the beating zebrafish heart was clearly segmented and the functional monitoring of the heart was achieved. Furthermore, the enhanced axial resolution established by thin plane illumination of PIP enabled the 3-D reconstruction of the branched cellular structures, which leads to the improvement on the functionality of the wide field microscopy.

Extracting relevant information for cancer diagnosis from dynamic full field OCT through image processing and learning

NASA Astrophysics Data System (ADS)

Apelian, Clément; Gastaud, Clément; Boccara, A. Claude

2017-02-01

For a large number of cancer surgeries, the lack of reliable intraoperative diagnosis leads to reoperations or bad outcomes for the patients. To deliver better diagnosis, we developed Dynamic Full Field OCT (D-FFOCT) as a complement to FFOCT. FFOCT already presents interesting results for cancer diagnosis e.g. Mohs surgery and reaching 96% accuracy on prostate cancer. D-FFOCT accesses the dynamic processes of metabolism and gives new tools to diagnose the state of a tissue at the cellular level to complement FFOCT contrast. We developed a processing framework that intends to maximize the information provided by the FFOCT technology as well as D-FFOCT and synthetize this as a meaningful image. We use different time processing to generate metrics (standard deviation of time signals, decorrelation times and more) and spatial processing to sort out structures and the corresponding imaging modality, which is the most appropriate. Sorting was achieved through quadratic discriminant analysis in a N-dimension parametric space corresponding to our metrics. Combining the best imaging modalities for each structure leads to a rich morphology image. This image displaying the morphology is then colored to represent the dynamic behavior of these structures (slow or fast) and to be quickly analyzed by doctors. Therefore, we achieved a micron resolved image, rich of both FFOCT ability of imaging fixed and highly backscattering structures as well as D-FFOCT ability of imaging low level scattering cellular level details. We believe that this morphological contrast close to histology and the dynamic behavior contrast will push forward the limits of intraoperative diagnosis further on.

3D scanning and printing skeletal tissues for anatomy education.

PubMed

Thomas, Daniel B; Hiscox, Jessica D; Dixon, Blair J; Potgieter, Johan

2016-09-01

Detailed anatomical models can be produced with consumer-level 3D scanning and printing systems. 3D replication techniques are significant advances for anatomical education as they allow practitioners to more easily introduce diverse or numerous specimens into classrooms. Here we present a methodology for producing anatomical models in-house, with the chondrocranium cartilage from a spiny dogfish (Squalus acanthias) and the skeleton of a cane toad (Rhinella marina) as case studies. 3D digital replicas were produced using two consumer-level scanners and specimens were 3D-printed with selective laser sintering. The fidelity of the two case study models was determined with respect to key anatomical features. Larger-scale features of the dogfish chondrocranium and frog skeleton were all well-resolved and distinct in the 3D digital models, and many finer-scale features were also well-resolved, but some more subtle features were absent from the digital models (e.g. endolymphatic foramina in chondrocranium). All characters identified in the digital chondrocranium could be identified in the subsequent 3D print; however, three characters in the 3D-printed frog skeleton could not be clearly delimited (palatines, parasphenoid and pubis). Characters that were absent in the digital models or 3D prints had low-relief in the original scanned specimen and represent a minor loss of fidelity. Our method description and case studies show that minimal equipment and training is needed to produce durable skeletal specimens. These technologies support the tailored production of models for specific classes or research aims. © 2016 Anatomical Society.

The evolution of void-filled cosmological structures

NASA Technical Reports Server (NTRS)

Regos, Eniko; Geller, Margaret J.

1991-01-01

1D, 2D, and 3D simulations are used here to investigate the salient features in the evolution of void-filled cosmological structures in universes with arbitrary values of Omega. It is found that the growth of a void as a function of time decreases significantly at the time corresponding to Omega = 0.5. In models constructed in 2D and 3D, suitable initial conditions lead to cellular structure with faceted voids similar to those observed in redshift surveys. Matter compressed to planes flows more rapidly toward condensations at the intersections than would be expected for spherical infall. The peculiar streaming velocities for void diameters of 5000 km/s should be observable. The simulations provide a more physical basis and dynamics for the bubbly and Voronois tesselation models used to derive statistical properties of cellular large-scale structure.

Quantifying the added value of convection-permitting climate simulations in complex terrain: a systematic evaluation of WRF over the Himalayas

NASA Astrophysics Data System (ADS)

Karki, Ramchandra; Hasson, Shabeh ul; Gerlitz, Lars; Schickhoff, Udo; Scholten, Thomas; Böhner, Jürgen

2017-07-01

Mesoscale dynamical refinements of global climate models or atmospheric reanalysis have shown their potential to resolve intricate atmospheric processes, their land surface interactions, and subsequently, realistic distribution of climatic fields in complex terrains. Given that such potential is yet to be explored within the central Himalayan region of Nepal, we investigate the skill of the Weather Research and Forecasting (WRF) model with different spatial resolutions in reproducing the spatial, seasonal, and diurnal characteristics of the near-surface air temperature and precipitation as well as the spatial shifts in the diurnal monsoonal precipitation peak over the Khumbu (Everest), Rolwaling, and adjacent southern areas. Therefore, the ERA-Interim (0.75°) reanalysis has been dynamically refined to 25, 5, and 1 km (D1, D2, and D3) for one complete hydrological year (October 2014-September 2015), using the one-way nested WRF model run with mild nudging and parameterized convection for the outer but explicitly resolved convection for the inner domains. Our results suggest that D3 realistically reproduces the monsoonal precipitation, as compared to its underestimation by D1 but overestimation by D2. All three resolutions, however, overestimate precipitation from the westerly disturbances, owing to simulating anomalously higher intensity of few intermittent events. Temperatures are generally reproduced well by all resolutions; however, winter and pre-monsoon seasons feature a high cold bias for high elevations while lower elevations show a simultaneous warm bias. Unlike higher resolutions, D1 fails to realistically reproduce the regional-scale nocturnal monsoonal peak precipitation observed in the Himalayan foothills and its diurnal shift towards high elevations, whereas D2 resolves these characteristics but exhibits a limited skill in reproducing such a peak on the river valley scale due to the limited representation of the narrow valleys at 5 km resolution. Nonetheless, featuring a substantial skill over D1 and D2, D3 simulates almost realistic shapes of the seasonal and diurnal precipitation and the peak timings even on valley scales. These findings clearly suggest an added value of the convective-scale resolutions in realistically resolving the topoclimates over the central Himalayas, which in turn allows simulating their interactions with the synoptic-scale weather systems prevailing over high Asia.

[Evaluation of the resolving power of different angles in MPR images of 16DAS-MDCT].

PubMed

Kimura, Mikio; Usui, Junshi; Nozawa, Takeo

2007-03-20

In this study, we evaluated the resolving power of three-dimensional (3D) multiplanar reformation (MPR) images with various angles by using 16 data acquisition system multi detector row computed tomography (16DAS-MDCT) . We reconstructed the MPR images using data with a 0.75 mm slice thickness of the axial image in this examination. To evaluate resolving power, we used an original new phantom (RC phantom) that can be positioned at any slice angle in MPR images. We measured the modulation transfer function (MTF) by using the methods of measuring pre-sampling MTF, and used Fourier transform of image data of the square wave chart. The scan condition and image reconstruction condition that were adopted in this study correspond to the condition that we use for three-dimensional computed tomographic angiography (3D-CTA) examination of the head in our hospital. The MTF of MPR images showed minimum values at slice angles in parallel with the axial slice, and showed maximum values at the sagittal slice and coronal slice angles that are parallel to the Z-axis. With an oblique MPR image, MTF did not change with angle changes in the oblique sagittal slice plane, but in the oblique coronal slice plane, MTF increased as the tilt angle increased from the axial plane to the Z plane. As a result, we could evaluate the resolving power of a head 3D image by measuring the MTF of the axial image and sagittal image or the coronal image.

Precipitation processes developed during TOGA COARE (1992), GATE (1974), SCSMEX (1998), and KWAJEX (1999): 3D Cloud Resolving Model Simulation

NASA Technical Reports Server (NTRS)

Tao, W.-K.

2006-01-01

Real clouds and cloud systems are inherently three-dimensional (3D). Because of the limitations in computer resources, however, most cloud-resolving models (CRMs) today are still two-dimensional (2D). A few 3D CRMs have been used to study the response of clouds to large-scale forcing. In these 3D simulations, the model domain was small, and the integration time was 6 hours. Only recently have 3D experiments been performed for multi-day periods for tropical cloud systems with large horizontal domains at the National Center for Atmospheric Research (NCAR), NOAA GFDL, the U.K. Met. Office, Colorado State University and NASA Goddard Space Flight Center. An improved 3D Goddard Cumulus Ensemble (GCE) model was recently used to simulate periods during TOGA COARE (December 19-27, 1992), GATE (september 1-7, 1974), SCSMEX (May 18-26, June 2-11, 1998) and KWAJEX (August 7-13, August 18-21, and August 29-September 12, 1999) using a 512 by 512 km domain and 41 vertical layers. The major objectives of this paper are: (1) to identify the differences and similarities in the simulated precipitation processes and their associated surface and water energy budgets in TOGA COARE, GATE, KWAJEX, and SCSMEX, and (2) to asses the impact of microphysics, radiation budget and surface fluxes on the organization of convection in tropics.

Fluorescent BODIPY Rotor: Viscometer for Cellular Organelles and Membrane-Mimicking Vesicles

NASA Astrophysics Data System (ADS)

Kimball, J.; Raut, S.; Fudala, R.; Doan, H.; Maliwal, B.; Sabnis, N.; Lacko, A.; Gryczynski, I.; Dzyuba, S.; Gryczynski, Z.

2015-03-01

Many cellular processes, such as mass and signal transport, metabolism and protein-protein interactions are governed in part by diffusion, and thus affected by their local microviscosity. Changes in this microviscosity has also been linked to various diseases, including atherosclerosis, Alzheimer's disease and diabetes. Therefore, directly measuring the heterogeneous viscosity of cellular constitutes can lead to greater understanding of these processes. To this effect, a novel homodiemeric BODIPY dye was evaluated as a fluorescent rotor probe for this application. A linear dependence on viscosity in the range of typical cellular microviscosity was established for steady-state and time-resolved properties of the dye. It was then embedded in vitro to membrane-mimicking lipid vesicles (DPPC, POPC, and POPC plus cholesterol) and results indicated it to be a viable sensor for lifetime-based determination of microviscosity. The BODIPY dye was lastly endocytosed by SKOV3 cells and Fluorescence Lifetime Imaging Microscopy (FLIM) was performed, successfully mapping the viscosity of internal cell components. This work was supported by the NIH Grant R01EB12003, the NSF Grant CBET-1264608, and the INFOR Grant from TCU.

Metallic powder-bed based 3D printing of cellular scaffolds for orthopaedic implants: A state-of-the-art review on manufacturing, topological design, mechanical properties and biocompatibility.

PubMed

Tan, X P; Tan, Y J; Chow, C S L; Tor, S B; Yeong, W Y

2017-07-01

Metallic cellular scaffold is one of the best choices for orthopaedic implants as a replacement of human body parts, which could improve life quality and increase longevity for the people needed. Unlike conventional methods of making cellular scaffolds, three-dimensional (3D) printing or additive manufacturing opens up new possibilities to fabricate those customisable intricate designs with highly interconnected pores. In the past decade, metallic powder-bed based 3D printing methods emerged and the techniques are becoming increasingly mature recently, where selective laser melting (SLM) and selective electron beam melting (SEBM) are the two representatives. Due to the advantages of good dimensional accuracy, high build resolution, clean build environment, saving materials, high customisability, etc., SLM and SEBM show huge potential in direct customisable manufacturing of metallic cellular scaffolds for orthopaedic implants. Ti-6Al-4V to date is still considered to be the optimal materials for producing orthopaedic implants due to its best combination of biocompatibility, corrosion resistance and mechanical properties. This paper presents a state-of-the-art overview mainly on manufacturing, topological design, mechanical properties and biocompatibility of cellular Ti-6Al-4V scaffolds via SLM and SEBM methods. Current manufacturing limitations, topological shortcomings, uncertainty of biocompatible test were sufficiently discussed herein. Future perspectives and recommendations were given at the end. Copyright © 2017 Elsevier B.V. All rights reserved.

Three dimensional cultures: a tool to study normal acinar architecture vs. malignant transformation of breast cells.

PubMed

Pal, Anupama; Kleer, Celina G

2014-04-25

Invasive breast carcinomas are a group of malignant epithelial tumors characterized by the invasion of adjacent tissues and propensity to metastasize. The interplay of signals between cancer cells and their microenvironment exerts a powerful influence on breast cancer growth and biological behavior(1). However, most of these signals from the extracellular matrix are lost or their relevance is understudied when cells are grown in two dimensional culture (2D) as a monolayer. In recent years, three dimensional (3D) culture on a reconstituted basement membrane has emerged as a method of choice to recapitulate the tissue architecture of benign and malignant breast cells. Cells grown in 3D retain the important cues from the extracellular matrix and provide a physiologically relevant ex vivo system(2,3). Of note, there is growing evidence suggesting that cells behave differently when grown in 3D as compared to 2D(4). 3D culture can be effectively used as a means to differentiate the malignant phenotype from the benign breast phenotype and for underpinning the cellular and molecular signaling involved(3). One of the distinguishing characteristics of benign epithelial cells is that they are polarized so that the apical cytoplasm is towards the lumen and the basal cytoplasm rests on the basement membrane. This apico-basal polarity is lost in invasive breast carcinomas, which are characterized by cellular disorganization and formation of anastomosing and branching tubules that haphazardly infiltrates the surrounding stroma. These histopathological differences between benign gland and invasive carcinoma can be reproduced in 3D(6,7). Using the appropriate read-outs like the quantitation of single round acinar structures, or differential expression of validated molecular markers for cell proliferation, polarity and apoptosis in combination with other molecular and cell biology techniques, 3D culture can provide an important tool to better understand the cellular changes during malignant transformation and for delineating the responsible signaling.

Design and implementation of a novel mechanical testing system for cellular solids.

PubMed

Nazarian, Ara; Stauber, Martin; Müller, Ralph

2005-05-01

Cellular solids constitute an important class of engineering materials encompassing both man-made and natural constructs. Materials such as wood, cork, coral, and cancellous bone are examples of cellular solids. The structural analysis of cellular solid failure has been limited to 2D sections to illustrate global fracture patterns. Due to the inherent destructiveness of 2D methods, dynamic assessment of fracture progression has not been possible. Image-guided failure assessment (IGFA), a noninvasive technique to analyze 3D progressive bone failure, has been developed utilizing stepwise microcompression in combination with time-lapsed microcomputed tomographic imaging (microCT). This method allows for the assessment of fracture progression in the plastic region, where much of the structural deformation/energy absorption is encountered in a cellular solid. Therefore, the goal of this project was to design and fabricate a novel micromechanical testing system to validate the effectiveness of the stepwise IGFA technique compared to classical continuous mechanical testing, using a variety of engineered and natural cellular solids. In our analysis, we found stepwise compression to be a valid approach for IGFA with high precision and accuracy comparable to classical continuous testing. Therefore, this approach complements the conventional mechanical testing methods by providing visual insight into the failure propagation mechanisms of cellular solids. (c) 2005 Wiley Periodicals, Inc.

Measurement of replication structures at the nanometer scale using super-resolution light microscopy

PubMed Central

Baddeley, D.; Chagin, V. O.; Schermelleh, L.; Martin, S.; Pombo, A.; Carlton, P. M.; Gahl, A.; Domaing, P.; Birk, U.; Leonhardt, H.; Cremer, C.; Cardoso, M. C.

2010-01-01

DNA replication, similar to other cellular processes, occurs within dynamic macromolecular structures. Any comprehensive understanding ultimately requires quantitative data to establish and test models of genome duplication. We used two different super-resolution light microscopy techniques to directly measure and compare the size and numbers of replication foci in mammalian cells. This analysis showed that replication foci vary in size from 210 nm down to 40 nm. Remarkably, spatially modulated illumination (SMI) and 3D-structured illumination microscopy (3D-SIM) both showed an average size of 125 nm that was conserved throughout S-phase and independent of the labeling method, suggesting a basic unit of genome duplication. Interestingly, the improved optical 3D resolution identified 3- to 5-fold more distinct replication foci than previously reported. These results show that optical nanoscopy techniques enable accurate measurements of cellular structures at a level previously achieved only by electron microscopy and highlight the possibility of high-throughput, multispectral 3D analyses. PMID:19864256

Effects of the 3D bone-to-implant contact and bone stiffness on the initial stability of a dental implant: micro-CT and resonance frequency analyses.

PubMed

Hsu, J T; Huang, H L; Tsai, M T; Wu, A Y J; Tu, M G; Fuh, L J

2013-02-01

This study investigated the effects of bone stiffness (elastic modulus) and three-dimensional (3D) bone-to-implant contact ratio (BIC%) on the primary stabilities of dental implants using micro-computed tomography (micro-CT) and resonance frequency analyses. Artificial sawbone models with five values of elastic modulus (137, 123, 47.5, 22, and 12.4 MPa) comprising two types of trabecular structure (solid-rigid and cellular-rigid) were investigated for initial implant stability quotient (ISQ), measured using the wireless Osstell resonance frequency analyzer. Bone specimens were attached to 2 mm fibre-filled epoxy sheets mimicking the cortical shell. ISQ was measured after placing a dental implant into the bone specimen. Each bone specimen with an implant was subjected to micro-CT scanning to calculate the 3D BIC% values. The similarity of the cellular type of artificial bone to the trabecular structure might make it more appropriate for obtaining accurate values of primary implant stability than solid-bone blocks. For the cellular-rigid bone models, the ISQ increased with the elastic modulus of cancellous bone. The regression correlation coefficient was 0.96 for correlations of the ISQ with the elasticity of cancellous bone and with the 3D BIC%. The initial implant stability was moderately positively correlated with the elasticity of cancellous bone and with the 3D BIC%. Copyright © 2012 International Association of Oral and Maxillofacial Surgeons. Published by Elsevier Ltd. All rights reserved.

Classifying and Analyzing 3d Cell Motion in Jammed Microgels

NASA Astrophysics Data System (ADS)

Bhattacharjee, Tapomoy; Sawyer, W. Gregory; Angelini, Thomas

Soft granular polyelectrolyte microgels swell in liquid cell growth media to form a continuous elastic solid that can easily transition between solid to fluid state under a low shear stress. Such Liquid-like solids (LLS) have recently been used to create 3D cellular constructs as well as to support, culture and harvest cells in 3D. Current understanding of cell migration mechanics in 3D was established from experiments performed in natural and synthetic polymer networks. Spatial variation in network structure and the transience of degradable gels limit their usefulness in quantitative cell mechanics studies. By contrast, LLS growth media approximates a homogeneous continuum, enabling tractable cell mechanics measurements to be performed in 3D. Here, we introduce a process to understand and classify cytotoxic T cell motion in 3D by studying cellular motility in LLS media. General classification of T cell motion can be achieved with a very traditional statistical approach: the cell's mean squared displacement (MSD) as a function of delay time. We will also use Langevin approaches combined with the constitutive equations of the LLS medium to predict the statistics of T cell motion. National Science Foundation under Grant No. DMR-1352043.

Micro-/nano-engineered cellular responses for soft tissue engineering and biomedical applications.

PubMed

Tay, Chor Yong; Irvine, Scott Alexander; Boey, Freddy Y C; Tan, Lay Poh; Venkatraman, Subbu

2011-05-23

The development of biomedical devices and reconstruction of functional ex vivo tissues often requires the need to fabricate biomimetic surfaces with features of sub-micrometer precision. This can be achieved with the advancements in micro-/nano-engineering techniques, allowing researchers to manipulate a plethora of cellular behaviors at the cell-biomaterial interface. Systematic studies conducted on these 2D engineered surfaces have unraveled numerous novel findings that can potentially be integrated as part of the design consideration for future 2D and 3D biomaterials and will no doubt greatly benefit tissue engineering. In this review, recent developments detailing the use of micro-/nano-engineering techniques to direct cellular orientation and function pertinent to soft tissue engineering will be highlighted. Particularly, this article aims to provide valuable insights into distinctive cell interactions and reactions to controlled surfaces, which can be exploited to understand the mechanisms of cell growth on micro-/nano-engineered interfaces, and to harness this knowledge to optimize the performance of 3D artificial soft tissue grafts and biomedical applications. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Changing the picture of Earth's earliest fossils (3.5-1.9 Ga) with new approaches and new discoveries.

PubMed

Brasier, Martin D; Antcliffe, Jonathan; Saunders, Martin; Wacey, David

2015-04-21

New analytical approaches and discoveries are demanding fresh thinking about the early fossil record. The 1.88-Ga Gunflint chert provides an important benchmark for the analysis of early fossil preservation. High-resolution analysis of Gunflintia shows that microtaphonomy can help to resolve long-standing paleobiological questions. Novel 3D nanoscale reconstructions of the most ancient complex fossil Eosphaera reveal features hitherto unmatched in any crown-group microbe. While Eosphaera may preserve a symbiotic consortium, a stronger conclusion is that multicellular morphospace was differently occupied in the Paleoproterozoic. The 3.46-Ga Apex chert provides a test bed for claims of biogenicity of cell-like structures. Mapping plus focused ion beam milling combined with transmission electron microscopy data demonstrate that microfossil-like taxa, including species of Archaeoscillatoriopsis and Primaevifilum, are pseudofossils formed from vermiform phyllosilicate grains during hydrothermal alteration events. The 3.43-Ga Strelley Pool Formation shows that plausible early fossil candidates are turning up in unexpected environmental settings. Our data reveal how cellular clusters of unexpectedly large coccoids and tubular sheath-like envelopes were trapped between sand grains and entombed within coatings of dripstone beach-rock silica cement. These fossils come from Earth's earliest known intertidal to supratidal shoreline deposit, accumulated under aerated but oxygen poor conditions.

Changing the picture of Earth's earliest fossils (3.5–1.9 Ga) with new approaches and new discoveries

PubMed Central

Brasier, Martin D.; Antcliffe, Jonathan; Saunders, Martin; Wacey, David

2015-01-01

New analytical approaches and discoveries are demanding fresh thinking about the early fossil record. The 1.88-Ga Gunflint chert provides an important benchmark for the analysis of early fossil preservation. High-resolution analysis of Gunflintia shows that microtaphonomy can help to resolve long-standing paleobiological questions. Novel 3D nanoscale reconstructions of the most ancient complex fossil Eosphaera reveal features hitherto unmatched in any crown-group microbe. While Eosphaera may preserve a symbiotic consortium, a stronger conclusion is that multicellular morphospace was differently occupied in the Paleoproterozoic. The 3.46-Ga Apex chert provides a test bed for claims of biogenicity of cell-like structures. Mapping plus focused ion beam milling combined with transmission electron microscopy data demonstrate that microfossil-like taxa, including species of Archaeoscillatoriopsis and Primaevifilum, are pseudofossils formed from vermiform phyllosilicate grains during hydrothermal alteration events. The 3.43-Ga Strelley Pool Formation shows that plausible early fossil candidates are turning up in unexpected environmental settings. Our data reveal how cellular clusters of unexpectedly large coccoids and tubular sheath-like envelopes were trapped between sand grains and entombed within coatings of dripstone beach-rock silica cement. These fossils come from Earth’s earliest known intertidal to supratidal shoreline deposit, accumulated under aerated but oxygen poor conditions. PMID:25901305

Respiratory motion-resolved, self-gated 4D-MRI using Rotating Cartesian K-space (ROCK): Initial clinical experience on an MRI-guided radiotherapy system.

PubMed

Han, Fei; Zhou, Ziwu; Du, Dongsu; Gao, Yu; Rashid, Shams; Cao, Minsong; Shaverdian, Narek; Hegde, John V; Steinberg, Michael; Lee, Percy; Raldow, Ann; Low, Daniel A; Sheng, Ke; Yang, Yingli; Hu, Peng

2018-06-01

To optimize and evaluate the respiratory motion-resolved, self-gated 4D-MRI using Rotating Cartesian K-space (ROCK-4D-MRI) method in a 0.35 T MRI-guided radiotherapy (MRgRT) system. The study included seven patients with abdominal tumors treated on the MRgRT system. ROCK-4D-MRI and 2D-CINE, was performed immediately after one of the treatment fractions. Motion quantification based on 4D-MRI was compared with those based on 2D-CINE. The image quality of 4D-MRI was evaluated against 4D-CT. The gross tumor volumes (GTV) were defined based on individual respiratory phases of both 4D-MRI and 4D-CT and compared for their variability over the respiratory cycle. The motion measurements based on 4D-MRI matched well with 2D-CINE, with differences of 1.04 ± 0.52 mm in the superior-inferior and 0.54 ± 0.21 mm in the anterior-posterior directions. The image quality scores of 4D-MRI were significantly higher than 4D-CT, with better tumor contrast (3.29 ± 0.76 vs. 1.86 ± 0.90) and less motion artifacts (3.57 ± 0.53 vs. 2.29 ± 0.95). The GTVs were more consistent in 4D-MRI than in 4D-CT, with significantly smaller GTV variability (9.31 ± 4.58% vs. 34.27 ± 23.33%). Our study demonstrated the clinical feasibility of using the ROCK-4D-MRI to acquire high quality, respiratory motion-resolved 4D-MRI in a low-field MRgRT system. The 4D-MRI image could provide accurate dynamic information for radiotherapy treatment planning. Copyright © 2018 Elsevier B.V. All rights reserved.

Studying the loading effect of acidic type antioxidant on amorphous silica nanoparticle carriers

NASA Astrophysics Data System (ADS)

Ravinayagam, Vijaya; Rabindran Jermy, B.

2017-06-01

The study investigates the suitable nanosilica carriers to transport acidic type cargo molecules for potential targeted drug delivery application. Using phenolic acidic type antioxidant gallic acid (GA) as model compound, the present study investigates the loading effect of GA (0.3-15.9 mmol GA g-1 support) on textural characteristics of amorphous silica nanoparticles such as Q10 silica (1D), structured two-dimensional Si-MCM-41 (2D), and three-dimensional Si-SBA-16 (3D). The variation in the nature of textures after GA loading was analyzed using X-ray diffraction, N2 adsorption, FT-IR, scanning electron microscopy with energy dispersive X-ray spectroscopy, and high-resolution transmission electron microscopy. Among the nanocarriers, high adsorption of GA was found in the following order: Si-SBA-16 (3D)˜Si-KIT-6 (3D) > Si-MCM-41 (2D) > ultralarge pore FDU-12 (ULPFDU-12; 3D) > Q10 (1D)˜mesostructured cellular silica foam (MSU-F). 3D-type silicas Si-SBA-16 and KIT-6 were shown to maintain structural integrity at acidic condition (pH ˜3) and accommodate GA in non-crystalline form. In the case of ULPFDU-12 and MSU-F cellular foam, only crystalline deposition of GA occurs with a significant variation in the surface area and pore volume. [Figure not available: see fulltext.

Early detection of skin cancer via terahertz spectral profiling and 3D imaging.

PubMed

Rahman, Anis; Rahman, Aunik K; Rao, Babar

2016-08-15

Terahertz scanning reflectometry, terahertz 3D imaging and terahertz time-domain spectroscopy have been used to identify features in human skin biopsy samples diagnosed for basal cell carcinoma (BCC) and compared with healthy skin samples. It was found from the 3D images that the healthy skin samples exhibit regular cellular pattern while the BCC skin samples indicate lack of regular cell pattern. The skin is a highly layered structure organ; this is evident from the thickness profile via a scan through the thickness of the healthy skin samples, where, the reflected intensity of the terahertz beam exhibits fluctuations originating from different skin layers. Compared to the healthy skin samples, the BCC samples' profiles exhibit significantly diminished layer definition; thus indicating a lack of cellular order. In addition, terahertz time-domain spectroscopy reveals significant and quantifiable differences between the healthy and BCC skin samples. Thus, a combination of three different terahertz techniques constitutes a conclusive route for detecting the BCC condition on a cellular level compared to the healthy skin. Copyright © 2016 Elsevier B.V. All rights reserved.

Effect of pressure on infrared spectra of ice 7

NASA Technical Reports Server (NTRS)

Holzapfel, W. B.; Seiler, B.; Nicol, M.

1983-01-01

The effect of pressure on the infrared spectra of H2O and D2O ice VII was studied at room temperature and pressures between 2 and 15 GPa with a Fourier transform infrared spectrometer and a diamond anvil high pressure cell. Two librational modes, one bending mode, and various overtone bands are well resolved. The stretching modes, nu sub 1 and nu sub 3 are poorly resolved due to overlap with diamond window absorption. Differences between the spectra of H2O and D2O are discussed.

Paroxetine and Low-dose Risperidone Induce Serotonin 5-HT1A and Dopamine D2 Receptor Heteromerization in the Mouse Prefrontal Cortex.

PubMed

Kolasa, Magdalena; Solich, Joanna; Faron-Górecka, Agata; Żurawek, Dariusz; Pabian, Paulina; Łukasiewicz, Sylwia; Kuśmider, Maciej; Szafran-Pilch, Kinga; Szlachta, Marta; Dziedzicka-Wasylewska, Marta

2018-05-01

Recently, it has been shown that serotonin 5-HT 1A receptor interacts with dopamine D2 receptor in vitro. However, the existence of 5-HT 1A -D2 heteromers in native tissue remains unexplored. In the present study, we investigated 5-HT 1A -D2 receptor heteromerization in mice treated acutely or chronically with paroxetine (10 mg/kg) or risperidone (0.05 mg/kg). Receptor heteromerization was visualized and quantified in the mouse brain by in situ proximity ligation assay (PLA). Additionally, we aimed to determine the cellular localization of 5-HT 1A -D2 receptor heteromers in mouse adult primary neuronal cells by immunofluorescent staining with markers for astrocytes (GFAP) and neurons (NeuN and MAP2). The results from the current study demonstrated that 5-HT 1A and D2 receptor co-localization and heteromerization occurred in the mouse prefrontal cortex. Counterstaining after PLA confirmed neuronal (pyramidal and GABAergic) as well as astrocytal localization of 5-HT 1A -D2 receptor heteromers. Chronic administration of paroxetine or risperidone increased the level of 5-HT 1A -D2 receptor heteromers in the prefrontal cortex. These changes were not accompanied by any changes in the expression of mRNAs (measured by in situ hybridization) or densities of 5-HT 1A and D2 receptors (quantified by receptor autoradiography with [3H]8-OH-DPAT and [3H]domperidone, respectively), what all indicated that paroxetine and risperidone facilitated 5-HT 1A -D2 heteromer formation independently of the receptor expression. In vitro homogenous time-resolved FRET (HTRF) study confirmed the ability of tested drugs to influence the human 5-HT 1A -D2 heteromer formation. The obtained data indicate that the increase in 5-HT 1A -D2 receptor heteromerization is a common molecular characteristic of paroxetine and low-dose risperidone treatment. Copyright © 2018 IBRO. Published by Elsevier Ltd. All rights reserved.

Comparative Proteomic Analysis of Differentially Expressed Proteins Induced by Hydrogen Sulfide in Spinacia oleracea Leaves

PubMed Central

Chen, Juan; Liu, Ting-Wu; Hu, Wen-Jun; Simon, Martin; Wang, Wen-Hua; Chen, Juan; Liu, Xiang; Zheng, Hai-Lei

2014-01-01

Hydrogen sulfide (H2S), as a potential gaseous messenger molecule, has been suggested to play important roles in a wide range of physiological processes in plants. The aim of present study was to investigate which set of proteins is involved in H2S-regulated metabolism or signaling pathways. Spinacia oleracea seedlings were treated with 100 µM NaHS, a donor of H2S. Changes in protein expression profiles were analyzed by 2-D gel electrophoresis coupled with MALDI-TOF MS. Over 1000 protein spots were reproducibly resolved, of which the abundance of 92 spots was changed by at least 2-fold (sixty-five were up-regulated, whereas 27 were down-regulated). These proteins were functionally divided into 9 groups, including energy production and photosynthesis, cell rescue, development and cell defense, substance metabolism, protein synthesis and folding, cellular signal transduction. Further, we found that these proteins were mainly localized in cell wall, plasma membrane, chloroplast, mitochondria, nucleus, peroxisome and cytosol. Our results demonstrate that H2S is involved in various cellular and physiological activities and has a distinct influence on photosynthesis, cell defense and cellular signal transduction in S. oleracea leaves. These findings provide new insights into proteomic responses in plants under physiological levels of H2S. PMID:25181351

Serial analysis of 3D H-1 MRSI for patients with newly diagnosed GBM treated with combination therapy that includes bevacizumab.

PubMed

Nelson, Sarah J; Li, Yan; Lupo, Janine M; Olson, Marram; Crane, Jason C; Molinaro, Annette; Roy, Ritu; Clarke, Jennifer; Butowski, Nicholas; Prados, Michael; Cha, Soonmee; Chang, Susan M

2016-10-01

Interpretation of changes in the T1- and T2-weighted MR images from patients with newly diagnosed glioblastoma (GBM) treated with standard of care in conjunction with anti-angiogenic agents is complicated by pseudoprogression and pseudoresponse. The hypothesis being tested in this study was that 3D H-1 magnetic resonance spectroscopic imaging (MRSI) provides estimates of levels of choline, creatine, N-acetylaspartate (NAA), lactate and lipid that change in response to treatment and that metrics describing these characteristics are associated with survival. Thirty-one patients with newly diagnosed GBM and being treated with radiation therapy (RT), temozolomide, erlotinib and bevacizumab were recruited to receive serial MR scans that included 3-D lactate edited MRSI at baseline, mid-RT, post-RT and at specific follow-up time points. The data were processed to provide estimates of metrics representing changes in metabolite levels relative to normal appearing brain. Cox proportional hazards analysis was applied to examine the relationship of these parameters with progression free survival (PFS) and overall survival (OS). There were significant reductions in parameters that describe relative levels of choline to NAA and creatine, indicating that the treatment caused a decrease in tumor cellularity. Changes in the levels of lactate and lipid relative to the NAA from contralateral brain were consistent with vascular normalization. Metabolic parameters from the first serial follow-up scan were associated with PFS and OS, when accounting for age and extent of resection. Integrating metabolic parameters into the assessment of patients with newly diagnosed GBM receiving therapies that include anti-angiogenic agents may be helpful for tracking changes in tumor burden, resolving ambiguities in anatomic images caused by non-specific treatment effects and for predicting outcome.

Electronic structure of dense Pb overlayers on Si(111) investigated using angle-resolved photoemission

NASA Astrophysics Data System (ADS)

Choi, W. H.; Koh, H.; Rotenberg, E.; Yeom, H. W.

2007-02-01

Dense Pb overlayers on Si(111) are important as the wetting layer for anomalous Pb island growth as well as for their own complex “devil’s-staircase” phases. The electronic structures of dense Pb overlayers on Si(111) were investigated in detail by angle-resolved photoemission. Among the series of ordered phases found recently above one monolayer, the low-coverage 7×3 and the high-coverage 14×3 phases are studied; they are well ordered and form reproducibly in large areas. The band dispersions and Fermi surfaces of the two-dimensional (2D) electronic states of these overlayers are mapped out. A number of metallic surface-state bands are identified for both phases with complex Fermi contours. The basic features of the observed Fermi contours can be explained by overlapping 2D free-electron-like Fermi circles. This analysis reveals that the 2D electrons near the Fermi level of the 7×3 and 14×3 phases are mainly governed by strong 1×1 and 3×3 potentials, respectively. The origins of the 2D electronic states and their apparent Fermi surface shapes are discussed based on recent structure models.

Studies of Time-Resolved Fluorescence Spectroscopy and Resolved Absorption Spectra of Nucleic Acid Components.

NASA Astrophysics Data System (ADS)

Fu, Yingxian

1993-01-01

There is considerable uncertainty about dynamic aspects of the photophysics of the adenylyl chromophore, stemming from the discordant values reported for the room temperature fluorescence lifetimes (tau_1 = 5 ps, tau_2 = 330 ps for 9MeAde; tau_1 = 290 ps, tau_2 = 4.17 ns for ATP). Spectra reported in conjunction with these lifetimes create difficulties in assignment of emission. To clarify this situation I have investigated the fluorescence decay times and time -resolved emission spectra of adenylyl compounds under a variety of conditions (concentration, pH, solvent) using sub-ns laser excitation at 265 nm together with gated fast sampling (100 ps) detection and signal averaging. Multi -component decays and spectra are observed in aqueous solution. Major slow components (tau = 4.4 +/- 0.2 ns) with emission maxima at 380 nm are found for all components at pH 1.1 and for ATP at pH 4.4. At pH 7 a fast component (<100 ps) predominates. There is no marked evidence for a concentration dependence, the oscillator strengths are 10^ {-3}-10^{-5} and transitions must be classified as weakly forbidden. Single component emission is observed in acetonitrile and ethanol. The UV absorption spectra of biomolecules d(CG) and polyd(GC)cdotpolyd(GC) exhibit the different hypochromic effects due to different interactions between guanosine(G) and cytidine(C) in stacked form. The present work has been carried out to explain this quantitatively. To approach this problem the absorption spectra of G and C have been resolved into gaussian components using the PeakFit program. The absorption spectra (220-310 nm) of d(CG) and polyd(GC)cdotpolyd(GC) have been fitted with gaussian components of G and C (in the order of increasing energy, G1 and G2, and C1, C2 and C3, respectively), and the contribution to both spectra from individual gaussians is estimated in terms of oscillator strengths. The fitting results suggest that the small hypochromism in absorption spectrum of d(CG) may be attributed to the interactions between G1 and C1; the large hypochromism in absorption spectrum of polyd(GC)cdotpolyd(GC) probably originates from the interactions between G1, C1, C2 and C3. The present work has also resolved a series of absorption spectra of cytidyl chromophore in different pH aqueous solution and various solvents. Time-resolved emission spectra of GMP, dCMP and m^5 -dCMP in different pH aqueous solutions have been determined. The results show that pH affects the lifetimes and spectral characteristics of GMP significantly, but does not affect dCMP and m^5-dCMP.

Modeling of time dependent localized flow shear stress and its impact on cellular growth within additive manufactured titanium implants

PubMed Central

Zhang, Ziyu; Yuan, Lang; Lee, Peter D; Jones, Eric; Jones, Julian R

2014-01-01

Bone augmentation implants are porous to allow cellular growth, bone formation and fixation. However, the design of the pores is currently based on simple empirical rules, such as minimum pore and interconnects sizes. We present a three-dimensional (3D) transient model of cellular growth based on the Navier–Stokes equations that simulates the body fluid flow and stimulation of bone precursor cellular growth, attachment, and proliferation as a function of local flow shear stress. The model's effectiveness is demonstrated for two additive manufactured (AM) titanium scaffold architectures. The results demonstrate that there is a complex interaction of flow rate and strut architecture, resulting in partially randomized structures having a preferential impact on stimulating cell migration in 3D porous structures for higher flow rates. This novel result demonstrates the potential new insights that can be gained via the modeling tool developed, and how the model can be used to perform what-if simulations to design AM structures to specific functional requirements. PMID:24664988

Physically-Induced Cytoskeleton Remodeling of Cells in Three-Dimensional Culture

PubMed Central

Lee, Sheng-Lin; Nekouzadeh, Ali; Butler, Boyd; Pryse, Kenneth M.; McConnaughey, William B.; Nathan, Adam C.; Legant, Wesley R.; Schaefer, Pascal M.; Pless, Robert B.

2012-01-01

Characterizing how cells in three-dimensional (3D) environments or natural tissues respond to biophysical stimuli is a longstanding challenge in biology and tissue engineering. We demonstrate a strategy to monitor morphological and mechanical responses of contractile fibroblasts in a 3D environment. Cells responded to stretch through specific, cell-wide mechanisms involving staged retraction and reinforcement. Retraction responses occurred for all orientations of stress fibers and cellular protrusions relative to the stretch direction, while reinforcement responses, including extension of cellular processes and stress fiber formation, occurred predominantly in the stretch direction. A previously unreported role of F-actin clumps was observed, with clumps possibly acting as F-actin reservoirs for retraction and reinforcement responses during stretch. Responses were consistent with a model of cellular sensitivity to local physical cues. These findings suggest mechanisms for global actin cytoskeleton remodeling in non-muscle cells and provide insight into cellular responses important in pathologies such as fibrosis and hypertension. PMID:23300512

Advances in high-resolution imaging--techniques for three-dimensional imaging of cellular structures.

PubMed

Lidke, Diane S; Lidke, Keith A

2012-06-01

A fundamental goal in biology is to determine how cellular organization is coupled to function. To achieve this goal, a better understanding of organelle composition and structure is needed. Although visualization of cellular organelles using fluorescence or electron microscopy (EM) has become a common tool for the cell biologist, recent advances are providing a clearer picture of the cell than ever before. In particular, advanced light-microscopy techniques are achieving resolutions below the diffraction limit and EM tomography provides high-resolution three-dimensional (3D) images of cellular structures. The ability to perform both fluorescence and electron microscopy on the same sample (correlative light and electron microscopy, CLEM) makes it possible to identify where a fluorescently labeled protein is located with respect to organelle structures visualized by EM. Here, we review the current state of the art in 3D biological imaging techniques with a focus on recent advances in electron microscopy and fluorescence super-resolution techniques.

Cloning and characterization of the murine homolog of the sno proto-oncogene reveals a novel splice variant

NASA Technical Reports Server (NTRS)

Pelzer, T.; Lyons, G. E.; Kim, S.; Moreadith, R. W.; Blomqvist, C. G. (Principal Investigator)

1996-01-01

The cellular function(s) of the SNO protein remain undefined. To gain a better understanding of possible developmental roles of this cellular proto-oncogene, we have cloned two murine sno cDNAs and have investigated their expression patterns in embryonic and postnatal tissues. A single major transcript of 7.5 kb is detected in multiple tissues by Northern blot. However, reverse transcriptase polymerase chain reaction (RT-PCR) and RNAse protection assays revealed a novel splice variant in every tissue examined. Two isoforms, termed sno N and sno-dE3 (dE3, deletion within exon 3), were identified. The sno-dE3 isoform employs a novel 5' splice site located within the coding region of the third exon and deletes potential kinase recognition motifs. Transcripts of both sno isoforms accumulate ubiquitously but are most abundant in the developing central nervous system. The in situ hybridization patterns of sno expression during murine development suggest potential roles in tissues with a high degree of cellular proliferation. Expression in terminally differentiated tissues such as muscle and neurons indicates that SNO may have multiple functional activities.

Depth-varying density and organization of chondrocytes in immature and mature bovine articular cartilage assessed by 3d imaging and analysis.

PubMed

Jadin, Kyle D; Wong, Benjamin L; Bae, Won C; Li, Kelvin W; Williamson, Amanda K; Schumacher, Barbara L; Price, Jeffrey H; Sah, Robert L

2005-09-01

Articular cartilage is a heterogeneous tissue, with cell density and organization varying with depth from the surface. The objectives of the present study were to establish a method for localizing individual cells in three-dimensional (3D) images of cartilage and quantifying depth-associated variation in cellularity and cell organization at different stages of growth. Accuracy of nucleus localization was high, with 99% sensitivity relative to manual localization. Cellularity (million cells per cm3) decreased from 290, 310, and 150 near the articular surface in fetal, calf, and adult samples, respectively, to 120, 110, and 50 at a depth of 1.0 mm. The distance/angle to the nearest neighboring cell was 7.9 microm/31 degrees , 7.1 microm/31 degrees , and 9.1 microm/31 degrees for cells at the articular surface of fetal, calf, and adult samples, respectively, and increased/decreased to 11.6 microm/31 degrees , 12.0 microm/30 degrees , and 19.2 microm/25 degrees at a depth of 0.7 mm. The methodologies described here may be useful for analyzing the 3D cellular organization of cartilage during growth, maturation, aging, degeneration, and regeneration.

Depth-varying density and organization of chondrocytes in immature and mature bovine articular cartilage assessed by 3d imaging and analysis

NASA Technical Reports Server (NTRS)

Jadin, Kyle D.; Wong, Benjamin L.; Bae, Won C.; Li, Kelvin W.; Williamson, Amanda K.; Schumacher, Barbara L.; Price, Jeffrey H.; Sah, Robert L.

2005-01-01

Articular cartilage is a heterogeneous tissue, with cell density and organization varying with depth from the surface. The objectives of the present study were to establish a method for localizing individual cells in three-dimensional (3D) images of cartilage and quantifying depth-associated variation in cellularity and cell organization at different stages of growth. Accuracy of nucleus localization was high, with 99% sensitivity relative to manual localization. Cellularity (million cells per cm3) decreased from 290, 310, and 150 near the articular surface in fetal, calf, and adult samples, respectively, to 120, 110, and 50 at a depth of 1.0 mm. The distance/angle to the nearest neighboring cell was 7.9 microm/31 degrees , 7.1 microm/31 degrees , and 9.1 microm/31 degrees for cells at the articular surface of fetal, calf, and adult samples, respectively, and increased/decreased to 11.6 microm/31 degrees , 12.0 microm/30 degrees , and 19.2 microm/25 degrees at a depth of 0.7 mm. The methodologies described here may be useful for analyzing the 3D cellular organization of cartilage during growth, maturation, aging, degeneration, and regeneration.

In Situ 3D Coherent X-ray Diffraction Imaging of Shock Experiments: Possible?

NASA Astrophysics Data System (ADS)

Barber, John

2011-03-01

In traditional coherent X-ray diffraction imaging (CXDI), a 2D or quasi-2D object is illuminated by a beam of coherent X-rays to produce a diffraction pattern, which is then manipulated via a process known as iterative phase retrieval to reconstruct an image of the original 2D sample. Recently, there have been dramatic advances in methods for performing fully 3D CXDI of a sample from a single diffraction pattern [Raines et al, Nature 463 214-7 (2010)], and these methods have been used to image samples tens of microns in size using soft X-rays. In this work, I explore the theoretical possibility of applying 3D CXDI techniques to the in situ imaging of the interaction between a shock front and a polycrystal, a far more stringent problem. A delicate trade-off is required between photon energy, spot size, imaging resolution, and the dimensions of the experimental setup. In this talk, I will outline the experimental and computational requirements for performing such an experiment, and I will present images and movies from simulations of one such hypothetical experiment, including both the time-resolved X-ray diffraction patterns and the time-resolved sample imagery.

Rolled-up Functionalized Nanomembranes as Three-Dimensional Cavities for Single Cell Studies

PubMed Central

2014-01-01

We use micropatterning and strain engineering to encapsulate single living mammalian cells into transparent tubular architectures consisting of three-dimensional (3D) rolled-up nanomembranes. By using optical microscopy, we demonstrate that these structures are suitable for the scrutiny of cellular dynamics within confined 3D-microenvironments. We show that spatial confinement of mitotic mammalian cells inside tubular architectures can perturb metaphase plate formation, delay mitotic progression, and cause chromosomal instability in both a transformed and nontransformed human cell line. These findings could provide important clues into how spatial constraints dictate cellular behavior and function. PMID:24598026

Rainbow peacock spiders inspire miniature super-iridescent optics.

PubMed

Hsiung, Bor-Kai; Siddique, Radwanul Hasan; Stavenga, Doekele G; Otto, Jürgen C; Allen, Michael C; Liu, Ying; Lu, Yong-Feng; Deheyn, Dimitri D; Shawkey, Matthew D; Blackledge, Todd A

2017-12-22

Colour produced by wavelength-dependent light scattering is a key component of visual communication in nature and acts particularly strongly in visual signalling by structurally-coloured animals during courtship. Two miniature peacock spiders (Maratus robinsoni and M. chrysomelas) court females using tiny structured scales (~ 40 × 10 μm 2 ) that reflect the full visual spectrum. Using TEM and optical modelling, we show that the spiders' scales have 2D nanogratings on microscale 3D convex surfaces with at least twice the resolving power of a conventional 2D diffraction grating of the same period. Whereas the long optical path lengths required for light-dispersive components to resolve individual wavelengths constrain current spectrometers to bulky sizes, our nano-3D printed prototypes demonstrate that the design principle of the peacock spiders' scales could inspire novel, miniature light-dispersive components.

The partition dimension of cycle books graph

NASA Astrophysics Data System (ADS)

Santoso, Jaya; Darmaji

2018-03-01

Let G be a nontrivial and connected graph with vertex set V(G), edge set E(G) and S ⊆ V(G) with v ∈ V(G), the distance between v and S is d(v,S) = min{d(v,x)|x ∈ S}. For an ordered partition ∏ = {S 1, S 2, S 3,…, Sk } of V(G), the representation of v with respect to ∏ is defined by r(v|∏) = (d(v, S 1), d(v, S 2),…, d(v, Sk )). The partition ∏ is called a resolving partition of G if all representations of vertices are distinct. The partition dimension pd(G) is the smallest integer k such that G has a resolving partition set with k members. In this research, we will determine the partition dimension of Cycle Books {B}{Cr,m}. Cycle books graph {B}{Cr,m} is a graph consisting of m copies cycle Cr with the common path P 2. It is shown that the partition dimension of cycle books graph, pd({B}{C3,m}) is 3 for m = 2, 3, and m for m ≥ 4. pd({B}{C4,m}) is 3 + 2k for m = 3k + 2, 4 + 2(k ‑ 1) for m = 3k + 1, and 3 + 2(k ‑ 1) for m = 3k. pd({B}{C5,m}) is m + 1.

A linear 180 nm SOI CMOS antenna switch module using integrated passive device filters for cellular applications

NASA Astrophysics Data System (ADS)

Jie, Cui; Lei, Chen; Peng, Zhao; Xu, Niu; Yi, Liu

2014-06-01

A broadband monolithic linear single pole, eight throw (SP8T) switch has been fabricated in 180 nm thin film silicon-on-insulator (SOI) CMOS technology with a quad-band GSM harmonic filter in integrated passive devices (IPD) technology, which is developed for cellular applications. The antenna switch module (ASM) features 1.2 dB insertion loss with filter on 2G bands and 0.4 dB insertion loss in 3G bands, less than -45 dB isolation and maximum -103 dB intermodulation distortion for mobile front ends by applying distributed architecture and adaptive supply voltage generator.

Effect of an Antimicrobial Compound on Different Processes within the Oscillation of Min Proteins in E. coli Bacterial Cells

NASA Astrophysics Data System (ADS)

Giuliani, Maximiliano; Dutcher, John

2013-03-01

A key step in the life of a bacterium is its division into two daughter cells of equal size. This process is carefully controlled and regulated so that equal partitioning of the cellular machinery is obtained. In E. coli, this regulation is accomplished, in part, by the Min protein system. The Min proteins undergo an oscillation between the poles of rod-shaped E. coli bacteria. We use high magnification, time-resolved total internal reflection fluorescence microscopy to characterize the temporal distributions of different processes within the oscillation: the MinD-MinE interaction time, the residence time for membrane bound MinD, and the recruitment time for MinD to be observed at the opposite pole. We also characterize the change in each of these processes in the presence of the antimicrobial compound polymyxin B (PMB). We show that the times corresponding to the removal of MinD from one pole and the recruitment of MinD at the opposite pole are correlated. We explain this correlation through the existence of a concentration threshold. The effect of PMB on the concentration threshold is used to identify which process within the oscillation is most affected.

Nanoscale tissue engineering: spatial control over cell-materials interactions

PubMed Central

Wheeldon, Ian; Farhadi, Arash; Bick, Alexander G.; Jabbari, Esmaiel; Khademhosseini, Ali

2011-01-01

Cells interact with the surrounding environment by making tens to hundreds of thousands of nanoscale interactions with extracellular signals and features. The goal of nanoscale tissue engineering is to harness the interactions through nanoscale biomaterials engineering in order to study and direct cellular behaviors. Here, we review the nanoscale tissue engineering technologies for both two- and three-dimensional studies (2- and 3D), and provide a holistic overview of the field. Techniques that can control the average spacing and clustering of cell adhesion ligands are well established and have been highly successful in describing cell adhesion and migration in 2D. Extension of these engineering tools to 3D biomaterials has created many new hydrogel and nanofiber scaffolds technologies that are being used to design in vitro experiments with more physiologically relevant conditions. Researchers are beginning to study complex cell functions in 3D, however, there is a need for biomaterials systems that provide fine control over the nanoscale presentation of bioactive ligands in 3D. Additionally, there is a need for 2- and 3D techniques that can control the nanoscale presentation of multiple bioactive ligands and the temporal changes in cellular microenvironment. PMID:21451238

Biomimicry 3D gastrointestinal spheroid platform for the assessment of toxicity and inflammatory effects of zinc oxide nanoparticles.

PubMed

Chia, Sing Ling; Tay, Chor Yong; Setyawati, Magdiel I; Leong, David T

2015-02-11

Our current mechanistic understanding on the effects of engineered nanoparticles (NPs) on cellular physiology is derived mainly from 2D cell culture studies. However, conventional monolayer cell culture may not accurately model the mass transfer gradient that is expected in 3D tissue physiology and thus may lead to artifactual experimental conclusions. Herein, using a micropatterned agarose hydrogel platform, the effects of ZnO NPs (25 nm) on 3D colon cell spheroids of well-defined sizes are examined. The findings show that cell dimensionality plays a critical role in governing the spatiotemporal cellular outcomes like inflammatory response and cytotoxicity in response to ZnO NPs treatment. More importantly, ZnO NPs can induce different modes of cell death in 2D and 3D cell culture systems. Interestingly, the outer few layers of cells in 3D model could only protect the inner core of cells for a limited time and periodically slough off from the spheroids surface. These findings suggest that toxicological conclusions made from 2D cell models might overestimate the toxicity of ZnO NPs. This 3D cell spheroid model can serve as a reproducible platform to better reflect the actual cell response to NPs and to study a more realistic mechanism of nanoparticle-induced toxicity. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Localizing Protein in 3D Neural Stem Cell Culture: a Hybrid Visualization Methodology

PubMed Central

Fai, Stephen; Bennett, Steffany A.L.

2010-01-01

The importance of 3-dimensional (3D) topography in influencing neural stem and progenitor cell (NPC) phenotype is widely acknowledged yet challenging to study. When dissociated from embryonic or post-natal brain, single NPCs will proliferate in suspension to form neurospheres. Daughter cells within these cultures spontaneously adopt distinct developmental lineages (neurons, oligodendrocytes, and astrocytes) over the course of expansion despite being exposed to the same extracellular milieu. This progression recapitulates many of the stages observed over the course of neurogenesis and gliogenesis in post-natal brain and is often used to study basic NPC biology within a controlled environment. Assessing the full impact of 3D topography and cellular positioning within these cultures on NPC fate is, however, difficult. To localize target proteins and identify NPC lineages by immunocytochemistry, free-floating neurospheres must be plated on a substrate or serially sectioned. This processing is required to ensure equivalent cell permeabilization and antibody access throughout the sphere. As a result, 2D epifluorescent images of cryosections or confocal reconstructions of 3D Z-stacks can only provide spatial information about cell position within discrete physical or digital 3D slices and do not visualize cellular position in the intact sphere. Here, to reiterate the topography of the neurosphere culture and permit spatial analysis of protein expression throughout the entire culture, we present a protocol for isolation, expansion, and serial sectioning of post-natal hippocampal neurospheres suitable for epifluorescent or confocal immunodetection of target proteins. Connexin29 (Cx29) is analyzed as an example. Next, using a hybrid of graphic editing and 3D modelling softwares rigorously applied to maintain biological detail, we describe how to re-assemble the 3D structural positioning of these images and digitally map labelled cells within the complete neurosphere. This methodology enables visualization and analysis of the cellular position of target proteins and cells throughout the entire 3D culture topography and will facilitate a more detailed analysis of the spatial relationships between cells over the course of neurogenesis and gliogenesis in vitro. Both Imbeault and Valenzuela contributed equally and should be considered joint first authors. PMID:21258319

3D Structure Determination of Native Mammalian Cells using Cryo-FIB and Cryo-electron Tomography

PubMed Central

Wang, Ke; Strunk, Korrinn; Zhao, Gongpu; Gray, Jennifer L.; Zhang, Peijun

2012-01-01

Cryo-electron tomography (cryo-ET) has enabled high resolution three-dimensional (3D) structural analysis of virus and host cell interactions and many cell signaling events; these studies, however, have largely been limited to very thin, peripheral regions of eukaryotic cells or to small prokaryotic cells. Recent efforts to make thin, vitreous sections using cryo-ultramicrotomy have been successful, however, this method is technically very challenging and with many artifacts. Here, we report a simple and robust method for creating in situ, frozen-hydrated cell lamellas using a focused ion beam at cryogenic temperature (cryo-FIB), allowing access to any interior cellular regions of interest. We demonstrate the utility of cryo-FIB with high resolution 3D cellular structures from both bacterial cells and large mammalian cells. The method will not only facilitate high-throughput 3D structural analysis of biological specimens, but is also broadly applicable to sample preparation of thin films and surface materials without the need for FIB “lift-out”. PMID:22796867

3D imaging of intrinsic crystalline defects in zinc oxide by spectrally resolved two-photon fluorescence microscopy

NASA Astrophysics Data System (ADS)

Al-Tabich, A.; Inami, W.; Kawata, Y.; Jablonski, R.; Worasawat, S.; Mimura, H.

2017-05-01

We present a method for three-dimensional intrinsic defect imaging in zinc oxide (ZnO) by spectrally resolved two-photon fluorescence microscopy, based on the previously presented method of observing a photoluminescence distribution in wide-gap semiconductor crystals [Noor et al., Appl. Phys. Lett. 92(16), 161106 (2008)]. A tightly focused light beam radiated by a titanium-sapphire laser is used to obtain a two-photon excitation of selected area of the ZnO sample. Photoluminescence intensity of a specific spectral range is then selected by optical band pass filters and measured by a photomultiplier tube. Reconstruction of the specimen image is done by scanning the volume of interest by a piezoelectric positioning stage and measuring the spectrally resolved photoluminescence intensity at each point. The method has been proved to be effective at locating intrinsic defects of the ZnO crystalline structure in the volume of the crystal. The method was compared with other defect imaging and 3D imaging techniques like scanning tunneling microscopy and confocal microscopy. In both cases, our method shows superior penetration abilities and, as the only method, allows location of the defects of the chosen type in 3D. In this paper, we present the results of oxygen vacancies and zinc antisites imaging in ZnO nanorods.

Survey statistics of automated segmentations applied to optical imaging of mammalian cells.

PubMed

Bajcsy, Peter; Cardone, Antonio; Chalfoun, Joe; Halter, Michael; Juba, Derek; Kociolek, Marcin; Majurski, Michael; Peskin, Adele; Simon, Carl; Simon, Mylene; Vandecreme, Antoine; Brady, Mary

2015-10-15

The goal of this survey paper is to overview cellular measurements using optical microscopy imaging followed by automated image segmentation. The cellular measurements of primary interest are taken from mammalian cells and their components. They are denoted as two- or three-dimensional (2D or 3D) image objects of biological interest. In our applications, such cellular measurements are important for understanding cell phenomena, such as cell counts, cell-scaffold interactions, cell colony growth rates, or cell pluripotency stability, as well as for establishing quality metrics for stem cell therapies. In this context, this survey paper is focused on automated segmentation as a software-based measurement leading to quantitative cellular measurements. We define the scope of this survey and a classification schema first. Next, all found and manually filteredpublications are classified according to the main categories: (1) objects of interests (or objects to be segmented), (2) imaging modalities, (3) digital data axes, (4) segmentation algorithms, (5) segmentation evaluations, (6) computational hardware platforms used for segmentation acceleration, and (7) object (cellular) measurements. Finally, all classified papers are converted programmatically into a set of hyperlinked web pages with occurrence and co-occurrence statistics of assigned categories. The survey paper presents to a reader: (a) the state-of-the-art overview of published papers about automated segmentation applied to optical microscopy imaging of mammalian cells, (b) a classification of segmentation aspects in the context of cell optical imaging, (c) histogram and co-occurrence summary statistics about cellular measurements, segmentations, segmented objects, segmentation evaluations, and the use of computational platforms for accelerating segmentation execution, and (d) open research problems to pursue. The novel contributions of this survey paper are: (1) a new type of classification of cellular measurements and automated segmentation, (2) statistics about the published literature, and (3) a web hyperlinked interface to classification statistics of the surveyed papers at https://isg.nist.gov/deepzoomweb/resources/survey/index.html.

Vitamin D deficiency may be a risk factor for recurrent pregnancy losses by increasing cellular immunity and autoimmunity.

PubMed

Ota, Kuniaki; Dambaeva, Svetlana; Han, Ae-Ra; Beaman, Kenneth; Gilman-Sachs, Alice; Kwak-Kim, Joanne

2014-02-01

Do women with recurrent pregnancy losses (RPL) and low vitamin D have increased prevalence of auto- and cellular immune abnormalities when compared with women with RPL who have normal vitamin D, and does vitamin D have any effect on cellular immunity in vitro? A high proportion of women with RPL have vitamin D deficiency and the risk of auto- and cellular immune abnormalities is increased in women with RPL and vitamin D deficiency. Vitamin D deficiency in pregnant women is associated with increased risk of obstetrical complications such as pre-eclampsia, bacterial vaginosis associated preterm delivery, gestational diabetes mellitus and small-for-gestational age births. A retrospective cross-sectional study of 133 women with RPL who were enrolled in a 2-year period, together with laboratory experiments. Women with three or more consecutive spontaneous abortions prior to 20 weeks of gestation who were enrolled at the University clinic. Serum vitamin D level, cellular activity and autoimmune parameters in vivo and in vitro were measured. Sixty-three out of 133 women (47.4%) had low vitamin D (<30 ng/ml). The prevalence of antiphospholipid antibody (APA) was significantly higher in low vitamin D group (VDlow) (39.7%) than in the normal vitamin D group (VDnl) (22.9%) (P< 0.05) and the adjusted odds ratio (OR) for APA in VDlow was 2.22 with the 95% confidence interval (CI) of 1.0-4.7. The prevalence of antinuclear antigen antibody (VDlow versus VDnl; 23.8% versus 10.0%, OR 2.81, 95% CI 1.1-7.4), anti-ssDNA (19.0% versus 5.7%, OR 3.76, 95% CI 1.1-12.4) and thyroperoxidase antibody (33.3% versus 15.7%, OR 2.68, 95% CI 1.2-6.1) was significantly higher in VDlow than those of VDnl (P < 0.05 each). Peripheral blood CD19(+) B and CD56(+) NK cell levels and NK cytotoxicity at effector to target cell (E:T) ratio of 25:1 were significantly higher in VDlow when compared with those of VDnl (P < 0.05 each). Reduction (%) of NK cytotoxicity (at E:T ratio of 50:1 and 25:1) by IgG (12.5 mg/dl) was significantly lower in VDlow than those of VDnl (P < 0.05, P < 0.01, respectively). There were no differences in Th1/Th2 ratios between VDlow and VDnl. When vitamin D3 was added in NK cytotoxicity assay in vitro, NK cytotoxicity at E:T ratio of 50:1 was significantly suppressed with 10 nMol/L (nM) (11.9 ± 3.3%) and 100 nM (10.9 ± 3.7%) of vitamin D3 when compared with controls (15.3 ± 4.7%) (P < 0.01 each). TNF-α/IL-10 expressing CD3(+)/4(+) cell ratios were significantly decreased with 100 nM of vitamin D3 (31.3 ± 9.4, P < 0.05) when compared with controls (40.4 ± 11.3) in vitro. Additionally, INF-γ/IL-10 expressing CD3(+)/4(+) cell ratio was significantly decreased with 100 nM of vitamin D3 (12.1 ± 4.0, P < 0.05) when compared with controls (14.8 ± 4.6). IFN-γ and TNF-α secretion from NK cells were significantly decreased (P < 0.01 each), and IL-10, IL-1β, vascular endothelial growth factor and granulocyte colony stimulating factor levels were significantly increased (P < 0.01 each) with vitamin D3 100 nM when compared with those of controls. The prevalence of vitamin D deficiency in women with RPL in this study is open to a possible type I error since women with vitamin D supplementation were excluded from this study. Assessment of vitamin D level is recommended in women with RPL. Vitamin D supplementation should be explored further as a possible therapeutic option for RPL. This work was supported by the intramural funding from Department of Microbiology and Immunology, Chicago Medical School at Rosalind Franklin University of Medicine and Science. None of the authors has any conflict of interest to declare. N/A.

Radiosity diffusion model in 3D

NASA Astrophysics Data System (ADS)

Riley, Jason D.; Arridge, Simon R.; Chrysanthou, Yiorgos; Dehghani, Hamid; Hillman, Elizabeth M. C.; Schweiger, Martin

2001-11-01

We present the Radiosity-Diffusion model in three dimensions(3D), as an extension to previous work in 2D. It is a method for handling non-scattering spaces in optically participating media. We present the extension of the model to 3D including an extension to the model to cope with increased complexity of the 3D domain. We show that in 3D more careful consideration must be given to the issues of meshing and visibility to model the transport of light within reasonable computational bounds. We demonstrate the model to be comparable to Monte-Carlo simulations for selected geometries, and show preliminary results of comparisons to measured time-resolved data acquired on resin phantoms.

Developing multi-cellular tumor spheroid model (MCTS) in the chitosan/collagen/alginate (CCA) fibrous scaffold for anticancer drug screening.

PubMed

Wang, Jian-Zheng; Zhu, Yu-Xia; Ma, Hui-Chao; Chen, Si-Nan; Chao, Ji-Ye; Ruan, Wen-Ding; Wang, Duo; Du, Feng-guang; Meng, Yue-Zhong

2016-05-01

In this work, a 3D MCTS-CCA system was constructed by culturing multi-cellular tumor spheroid (MCTS) in the chitosan/collagen/alginate (CCA) fibrous scaffold for anticancer drug screening. The CCA scaffolds were fabricated by spray-spinning. The interactions between the components of the spray-spun fibers were evidenced by methods of Coomassie Blue stain, X-ray diffraction (XRD) and Fourier transform-infrared spectroscopy (FTIR). Co-culture indicated that MCF-7 cells showed a spatial growth pattern of multi-cellular tumor spheroid (MCTS) in the CCA fibrous scaffold with increased proliferation rate and drug-resistance to MMC, ADM and 5-Aza comparing with the 2D culture cells. Significant increases of total viable cells were found in 3D MCTS groups after drug administration by method of apoptotic analysis. Glucose-lactate analysis indicated that the metabolism of MCTS in CCA scaffold was closer to the tumor issue in vivo than the monolayer cells. In addition, MCTS showed the characteristic of epithelial mesenchymal transition (EMT) which is subverted by carcinoma cells to facilitate metastatic spread. These results demonstrated that MCTS in CCA scaffold possessed a more conservative phenotype of tumor than monolayer cells, and anticancer drug screening in 3D MCTS-CCA system might be superior to the 2D culture system. Copyright © 2016 Elsevier B.V. All rights reserved.

Imaging live cells at high spatiotemporal resolution for lab-on-a-chip applications.

PubMed

Chin, Lip Ket; Lee, Chau-Hwang; Chen, Bi-Chang

2016-05-24

Conventional optical imaging techniques are limited by the diffraction limit and difficult-to-image biomolecular and sub-cellular processes in living specimens. Novel optical imaging techniques are constantly evolving with the desire to innovate an imaging tool that is capable of seeing sub-cellular processes in a biological system, especially in three dimensions (3D) over time, i.e. 4D imaging. For fluorescence imaging on live cells, the trade-offs among imaging depth, spatial resolution, temporal resolution and photo-damage are constrained based on the limited photons of the emitters. The fundamental solution to solve this dilemma is to enlarge the photon bank such as the development of photostable and bright fluorophores, leading to the innovation in optical imaging techniques such as super-resolution microscopy and light sheet microscopy. With the synergy of microfluidic technology that is capable of manipulating biological cells and controlling their microenvironments to mimic in vivo physiological environments, studies of sub-cellular processes in various biological systems can be simplified and investigated systematically. In this review, we provide an overview of current state-of-the-art super-resolution and 3D live cell imaging techniques and their lab-on-a-chip applications, and finally discuss future research trends in new and breakthrough research areas of live specimen 4D imaging in controlled 3D microenvironments.

Predictability in Cellular Automata

PubMed Central

Agapie, Alexandru; Andreica, Anca; Chira, Camelia; Giuclea, Marius

2014-01-01

Modelled as finite homogeneous Markov chains, probabilistic cellular automata with local transition probabilities in (0, 1) always posses a stationary distribution. This result alone is not very helpful when it comes to predicting the final configuration; one needs also a formula connecting the probabilities in the stationary distribution to some intrinsic feature of the lattice configuration. Previous results on the asynchronous cellular automata have showed that such feature really exists. It is the number of zero-one borders within the automaton's binary configuration. An exponential formula in the number of zero-one borders has been proved for the 1-D, 2-D and 3-D asynchronous automata with neighborhood three, five and seven, respectively. We perform computer experiments on a synchronous cellular automaton to check whether the empirical distribution obeys also that theoretical formula. The numerical results indicate a perfect fit for neighbourhood three and five, which opens the way for a rigorous proof of the formula in this new, synchronous case. PMID:25271778

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

PubMed

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

2018-04-26

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

PARP13 and RNA regulation in immunity and cancer

PubMed Central

Todorova, Tanya; Bock, Florian; Chang, Paul

2015-01-01

Posttranscriptional regulation of RNA is an important mechanism for activating and resolving cellular stress responses. Poly(ADP-ribose) Polymerase-13 (PARP13), also known as ZC3HAV1 and Zinc-finger Antiviral Protein (ZAP), is an RNA-binding protein that regulates the stability, and translation of specific mRNAs, and modulates the miRNA silencing pathway to globally impact miRNA targets. These functions of PARP13 are important components of the cellular response to stress. In addition, the ability of PARP13 to restrict oncogenic viruses and to repress the pro-survival cytokine receptor TRAILR4 suggests that it can be protective against malignant transformation and cancer development. The relevance of PARP13 to human health and disease make it a promising therapeutic target. PMID:25851173

Revealing fine microstructural morphology in the living human retina using Optical Coherence Tomography with pancorrection

NASA Astrophysics Data System (ADS)

Torti, C.; Považay, B.; Hofer, B.; Unterhuber, A.; Hermann, B.; Drexler, W.

2008-09-01

Ultra-high speed optical coherence tomography employing an ultra-broadband light source has been combined with adaptive optics utilizing a single high stroke deformable mirror and chromatic aberration compensation. The reduction of motion artefacts, geometric and chromatic aberrations (pancorrection) permits to achieve an isotropic resolution of 2-3 μm in the human eye. The performance of this non-invasive imaging modality enables to resolve cellular structures including cone photoreceptors, nerve fibre bundles and collagenous plates of the lamina cribrosa, and retinal pigment epithelial (RPE) cells in the human retina in vivo with superior detail. Alterations of cellular morphology due to cone degeneration in a colour-blind subject are investigated in ultra-high resolution with selective depth sectioning for the first time.

Pilot Assessment of Brain Metabolism in Perinatally HIV-Infected Youths Using Accelerated 5D Echo Planar J-Resolved Spectroscopic Imaging.

PubMed

Iqbal, Zohaib; Wilson, Neil E; Keller, Margaret A; Michalik, David E; Church, Joseph A; Nielsen-Saines, Karin; Deville, Jaime; Souza, Raissa; Brecht, Mary-Lynn; Thomas, M Albert

2016-01-01

To measure cerebral metabolite levels in perinatally HIV-infected youths and healthy controls using the accelerated five dimensional (5D) echo planar J-resolved spectroscopic imaging (EP-JRESI) sequence, which is capable of obtaining two dimensional (2D) J-resolved spectra from three spatial dimensions (3D). After acquisition and reconstruction of the 5D EP-JRESI data, T1-weighted MRIs were used to classify brain regions of interest for HIV patients and healthy controls: right frontal white (FW), medial frontal gray (FG), right basal ganglia (BG), right occipital white (OW), and medial occipital gray (OG). From these locations, respective J-resolved and TE-averaged spectra were extracted and fit using two different quantitation methods. The J-resolved spectra were fit using prior knowledge fitting (ProFit) while the TE-averaged spectra were fit using the advanced method for accurate robust and efficient spectral fitting (AMARES). Quantitation of the 5D EP-JRESI data using the ProFit algorithm yielded significant metabolic differences in two spatial locations of the perinatally HIV-infected youths compared to controls: elevated NAA/(Cr+Ch) in the FW and elevated Asp/(Cr+Ch) in the BG. Using the TE-averaged data quantified by AMARES, an increase of Glu/(Cr+Ch) was shown in the FW region. A strong negative correlation (r < -0.6) was shown between tCh/(Cr+Ch) quantified using ProFit in the FW and CD4 counts. Also, strong positive correlations (r > 0.6) were shown between Asp/(Cr+Ch) and CD4 counts in the FG and BG. The complimentary results using ProFit fitting of J-resolved spectra and AMARES fitting of TE-averaged spectra, which are a subset of the 5D EP-JRESI acquisition, demonstrate an abnormal energy metabolism in the brains of perinatally HIV-infected youths. This may be a result of the HIV pathology and long-term combinational anti-retroviral therapy (cART). Further studies of larger perinatally HIV-infected cohorts are necessary to confirm these findings.

Angle-resolved studies of inner shell excitations for argon, krypton and xenon using third-generation synchrotron sources

NASA Astrophysics Data System (ADS)

Farhat, Ahmad H.

This dissertation, which is in the area of atomic physics, concentrates on the study of the interaction between VUV-soft X-ray radiation and atoms in the gas phase. The main area of interest is the study of Auger decay in atoms utilizing the process known as the resonance Auger effect, where an inner shell electron is excited to an unfilled orbital followed by the ejection of an Auger electron. The measurements in this thesis were performed by using the high resolution Atomic, Molecular and Optical Physics undulator beam line, which utilizes a spherical grating monochromator at the Advanced Light Source at Lawrence Berkeley National Laboratory. The research focused on three rare gases, argon, krypton and xenon. For argon, high resolution angular-resolved measurements of the 2p → 4s, 3d, 4d resonant Auger lines have been achieved. By measuring photoelectron spectra simultaneously at two different angles using efficient time of flight spectrometers, the angular distributions anisotropy parameters β have been measured, and relative intensities have been evaluated for each of the resolved final ionic states. For krypton, the resonant Auger decay of all the photoexcited Kr 3d3/2,5/2-1 nl (n = 5-9) states have been studied using an angle resolved two dimensional photoelectron spectroscopic technique, in which the electron yield was measured as a function of both photon energy and electron kinetic energy. Angular distributions, spectator and shake probabilities have been derived for the Kr 3d-1np/to4s- 14p-1mp + e/sp- (n = 5-9, m = 5-11) resonance Auger decay. The results show that the spectator-core coupling is strong at lower n (n = 5,6) but it lessens for higher n, with a shake up of m = n + 1 preferred. Finally for xenon, the autoionization resonances and angular distribution of the 4d → 6p decay spectrum were studied utilizing the Auger resonant Raman effect. Using this technique, β parameters of almost all 5p4/ (3P,/ 1D,/ 1S) 6p final ionic states were determined. These results contribute to our understanding of atomic structure and dynamics of inner shell processes and hopefully will stimulate further experimental and theoretical work.

Serum matrix metalloproteinase 3 in detecting remitting seronegative symmetrical synovitis with pitting edema syndrome: A case report.

PubMed

Kenzaka, Tsuneaki; Goda, Ken

2018-05-16

We report a case of remitting seronegative symmetrical synovitis with pitting edema (RS3PE) syndrome in a 71-year-old woman. She referred to our hospital with finger stiffness, edema of both hands and feet, pain of bilateral shoulder, wrist, metacarpophalangeal, proximal interphalangeal, and ankle joints. Rheumatoid factor was negative, human leukocyte antigen -B7 antigen was positive. Moreover, matrix metalloproteinase 3 (MMP-3) was high. She was diagnosed with RS3PE syndrome, and treatment with prednisolone (15 mg/d) was started. One week after prednisolone treatment initiation, CRP decreased to negative, and joint pain was almost completely resolved. However, hand stiffness persisted, and MMP-3 level was still high. Thus, prednisolone dose was increased to 20 mg/d, and the stiffness resolved. Twenty days after treatment initiation, MMP-3 was normalized. MMP-3 was more indicative of RS3PE syndrome symptoms than CRP. Thus, MMP-3 seems to be more sensitive to RS3PE syndrome symptoms.

Serum matrix metalloproteinase 3 in detecting remitting seronegative symmetrical synovitis with pitting edema syndrome: A case report

PubMed Central

Kenzaka, Tsuneaki; Goda, Ken

2018-01-01

We report a case of remitting seronegative symmetrical synovitis with pitting edema (RS3PE) syndrome in a 71-year-old woman. She referred to our hospital with finger stiffness, edema of both hands and feet, pain of bilateral shoulder, wrist, metacarpophalangeal, proximal interphalangeal, and ankle joints. Rheumatoid factor was negative, human leukocyte antigen -B7 antigen was positive. Moreover, matrix metalloproteinase 3 (MMP-3) was high. She was diagnosed with RS3PE syndrome, and treatment with prednisolone (15 mg/d) was started. One week after prednisolone treatment initiation, CRP decreased to negative, and joint pain was almost completely resolved. However, hand stiffness persisted, and MMP-3 level was still high. Thus, prednisolone dose was increased to 20 mg/d, and the stiffness resolved. Twenty days after treatment initiation, MMP-3 was normalized. MMP-3 was more indicative of RS3PE syndrome symptoms than CRP. Thus, MMP-3 seems to be more sensitive to RS3PE syndrome symptoms. PMID:29774220

Microfluidic engineering of neural stem cell niches for fate determination

PubMed Central

Ma, Jingyun; Li, Na; Wang, Liang; Shen, Liming; Sun, Yu; Wang, Yajun; Zhao, Jingyuan; Wei, Wenjuan; Ren, Yan; Liu, Jing

2017-01-01

Neural stem cell (NSC) transplantation has great therapeutic potential for neurodegenerative diseases and central nervous system injuries. Successful NSC replacement therapy requires precise control over the cellular behaviors. However, the regulation of NSC fate is largely unclear, which severely restricts the potential clinical applications. To develop an effective model, we designed an assembled microfluidic system to engineer NSC niches and assessed the effects of various culture conditions on NSC fate determination. Five types of NSC microenvironments, including two-dimensional (2D) cellular monolayer culture, 2D cellular monolayer culture on the extracellular matrix (ECM), dispersed cells in the ECM, three-dimensional (3D) spheroid aggregates, and 3D spheroids cultured in the ECM, were constructed within an integrated microfluidic chip simultaneously. In addition, we evaluated the influence of static and perfusion culture on NSCs. The efficiency of this approach was evaluated comprehensively by characterization of NSC viability, self-renewal, proliferation, and differentiation into neurons, astrocytes, or oligodendrocytes. Differences in the status and fate of NSCs governed by the culture modes and micro-niches were analyzed. NSCs in the microfluidic device demonstrated good viability, the 3D culture in the ECM facilitated NSC self-renewal and proliferation, and 2D culture in the static state and spheroid culture under perfusion conditions benefited NSC differentiation. Regulation of NSC self-renewal and differentiation on this microfluidic device could provide NSC-based medicinal products and references for distinct nerve disease therapy. PMID:28798841

Cellular internalization mechanism and intracellular trafficking of filamentous M13 phages displaying a cell-penetrating transbody and TAT peptide.

PubMed

Kim, Aeyung; Shin, Tae-Hwan; Shin, Seung-Min; Pham, Chuong D; Choi, Dong-Ki; Kwon, Myung-Hee; Kim, Yong-Sung

2012-01-01

Cellular internalization of bacteriophage by surface-displayed cell penetrating peptides has been reported, though the underlying mechanism remains elusive. Here we describe in detail the internalization mechanism and intracellular trafficking and stability of filamentous M13 phages, the cellular entry of which is mediated by surface-displayed cell-penetrating light chain variable domain 3D8 VL transbody (3D8 VL-M13) or TAT peptide (TAT-M13). Recombinant 3D8 VL-M13 and TAT-M13 phages were efficiently internalized into living mammalian cells via physiologically relevant, energy-dependent endocytosis and were recovered from the cells in their infective form with the yield of 3D8 VL-M13 being higher (0.005 ≈ 0.01%) than that of TAT-M13 (0.001 ≈ 0.005%). Biochemical and genetic studies revealed that 3D8 VL-M13 was internalized principally by caveolae-mediated endocytosis via interaction with heparan sulfate proteoglycans as cell surface receptors, whereas TAT-M13 was internalized by clathrin- and caveolae-mediated endocytosis utilizing chondroitin sulfate proteoglycans as cell surface receptors, suggesting that phage internalization occurs by physiological endocytotic mechanism through specific cell surface receptors rather than non-specific transcytotic pathways. Internalized 3D8 VL-M13 phages routed to the cytosol and remained stable for more than 18 h without further trafficking to other subcellular compartments, whereas TAT-M13 phages routed to several subcellular compartments before being degraded in lysosomes even after 2 h of internalization. Our results suggest that the internalizing mechanism and intracellular trafficking of filamentous M13 bacteriophages largely follow the attributes of the displayed cell-penetrating moiety. Efficient internalization and cytosolic localization of 3D8 VL transbody-displayed phages will provide a useful tool for intracellular delivery of polar macromolecules such as proteins, peptides, and siRNAs.

Cellular Internalization Mechanism and Intracellular Trafficking of Filamentous M13 Phages Displaying a Cell-Penetrating Transbody and TAT Peptide

PubMed Central

Shin, Seung-Min; Pham, Chuong D.; Choi, Dong-Ki; Kwon, Myung-Hee; Kim, Yong-Sung

2012-01-01

Cellular internalization of bacteriophage by surface-displayed cell penetrating peptides has been reported, though the underlying mechanism remains elusive. Here we describe in detail the internalization mechanism and intracellular trafficking and stability of filamentous M13 phages, the cellular entry of which is mediated by surface-displayed cell-penetrating light chain variable domain 3D8 VL transbody (3D8 VL-M13) or TAT peptide (TAT-M13). Recombinant 3D8 VL-M13 and TAT-M13 phages were efficiently internalized into living mammalian cells via physiologically relevant, energy-dependent endocytosis and were recovered from the cells in their infective form with the yield of 3D8 VL-M13 being higher (0.005∼0.01%) than that of TAT-M13 (0.001∼0.005%). Biochemical and genetic studies revealed that 3D8 VL-M13 was internalized principally by caveolae-mediated endocytosis via interaction with heparan sulfate proteoglycans as cell surface receptors, whereas TAT-M13 was internalized by clathrin- and caveolae-mediated endocytosis utilizing chondroitin sulfate proteoglycans as cell surface receptors, suggesting that phage internalization occurs by physiological endocytotic mechanism through specific cell surface receptors rather than non-specific transcytotic pathways. Internalized 3D8 VL-M13 phages routed to the cytosol and remained stable for more than 18 h without further trafficking to other subcellular compartments, whereas TAT-M13 phages routed to several subcellular compartments before being degraded in lysosomes even after 2 h of internalization. Our results suggest that the internalizing mechanism and intracellular trafficking of filamentous M13 bacteriophages largely follow the attributes of the displayed cell-penetrating moiety. Efficient internalization and cytosolic localization of 3D8 VL transbody-displayed phages will provide a useful tool for intracellular delivery of polar macromolecules such as proteins, peptides, and siRNAs. PMID:23251631

Revealing 3D Ultrastructure and Morphology of Stem Cell Spheroids by Electron Microscopy.

PubMed

Jaros, Josef; Petrov, Michal; Tesarova, Marketa; Hampl, Ales

2017-01-01

Cell culture methods have been developed in efforts to produce biologically relevant systems for developmental and disease modeling, and appropriate analytical tools are essential. Knowledge of ultrastructural characteristics represents the basis to reveal in situ the cellular morphology, cell-cell interactions, organelle distribution, niches in which cells reside, and many more. The traditional method for 3D visualization of ultrastructural components, serial sectioning using transmission electron microscopy (TEM), is very labor-intensive due to contentious TEM slice preparation and subsequent image processing of the whole collection. In this chapter, we present serial block-face scanning electron microscopy, together with complex methodology for spheroid formation, contrasting of cellular compartments, image processing, and 3D visualization. The described technique is effective for detailed morphological analysis of stem cell spheroids, organoids, as well as organotypic cell cultures.

3D-engineering of Cellularized Conduits for Peripheral Nerve Regeneration

NASA Astrophysics Data System (ADS)

Hu, Yu; Wu, Yao; Gou, Zhiyuan; Tao, Jie; Zhang, Jiumeng; Liu, Qianqi; Kang, Tianyi; Jiang, Shu; Huang, Siqing; He, Jiankang; Chen, Shaochen; Du, Yanan; Gou, Maling

2016-08-01

Tissue engineered conduits have great promise for bridging peripheral nerve defects by providing physical guiding and biological cues. A flexible method for integrating support cells into a conduit with desired architectures is wanted. Here, a 3D-printing technology is adopted to prepare a bio-conduit with designer structures for peripheral nerve regeneration. This bio-conduit is consisted of a cryopolymerized gelatin methacryloyl (cryoGelMA) gel cellularized with adipose-derived stem cells (ASCs). By modeling using 3D-printed “lock and key” moulds, the cryoGelMA gel is structured into conduits with different geometries, such as the designed multichannel or bifurcating and the personalized structures. The cryoGelMA conduit is degradable and could be completely degraded in 2-4 months in vivo. The cryoGelMA scaffold supports the attachment, proliferation and survival of the seeded ASCs, and up-regulates the expression of their neurotrophic factors mRNA in vitro. After implanted in a rat model, the bio-conduit is capable of supporting the re-innervation across a 10 mm sciatic nerve gap, with results close to that of the autografts in terms of functional and histological assessments. The study describes an indirect 3D-printing technology for fabricating cellularized designer conduits for peripheral nerve regeneration, and could lead to the development of future nerve bio-conduits for clinical use.

Modeling of flow-induced shear stress applied on 3D cellular scaffolds: Implications for vascular tissue engineering.

PubMed

Lesman, Ayelet; Blinder, Yaron; Levenberg, Shulamit

2010-02-15

Novel tissue-culture bioreactors employ flow-induced shear stress as a means of mechanical stimulation of cells. We developed a computational fluid dynamics model of the complex three-dimensional (3D) microstructure of a porous scaffold incubated in a direct perfusion bioreactor. Our model was designed to predict high shear-stress values within the physiological range of those naturally sensed by vascular cells (1-10 dyne/cm(2)), and will thereby provide suitable conditions for vascular tissue-engineering experiments. The model also accounts for cellular growth, which was designed as an added cell layer grown on all scaffold walls. Five model variants were designed, with geometric differences corresponding to cell-layer thicknesses of 0, 50, 75, 100, and 125 microm. Four inlet velocities (0.5, 1, 1.5, and 2 cm/s) were applied to each model. Wall shear-stress distribution and overall pressure drop calculations were then used to characterize the relation between flow rate, shear stress, cell-layer thickness, and pressure drop. The simulations showed that cellular growth within 3D scaffolds exposes cells to elevated shear stress, with considerably increasing average values in correlation to cell growth and inflow velocity. Our results provide in-depth analysis of the microdynamic environment of cells cultured within 3D environments, and thus provide advanced control over tissue development in vitro. 2009 Wiley Periodicals, Inc.

Ovarian mucinous tumors arising from mature cystic teratomas--a molecular genetic approach for understanding the cellular origin.

PubMed

Fujii, Kaho; Yamashita, Yoriko; Yamamoto, Toshimichi; Takahashi, Koji; Hashimoto, Katsunori; Miyata, Tomoko; Kawai, Kumi; Kikkawa, Fumitaka; Toyokuni, Shinya; Nagasaka, Tetsuro

2014-04-01

Mucinous tumors of the ovary are frequently associated with mature cystic teratomas, and it has been speculated that the mucinous tumors arise from teratoma components. The cellular origins of mature cystic teratomas are believed to be post-meiotic ovarian germ cells, and the analysis of microsatellite markers such as short tandem repeats is suitable for determining the cellular origin of tumors. In this study, we analyzed 3 ovarian mature cystic teratomas, all of which were associated with simultaneous ovarian mucinous tumors within the same ovary. Two of the 3 mucinous tumors were intestinal-type and the other was endocervical type. A laser capture microdissection technique was used to separate the epithelial component of the mucinous tumor, the components of the mature cystic teratoma, and control ovarian somatic tissue. Using short tandem repeat analysis based on 6 markers (D20S480, D6S2439, D6S1056, D9S1118, D4S2639, and D17S1290), we could distinguish the germ cell (homozygous) or somatic (heterozygous) origin of a given component in each sample. The epithelial components of the intestinal-type mucinous tumors in cases 1 and 2 were homozygous, and the epithelial component in case 3 (endocervical type) was heterozygous. All teratomatous components were homozygous, and the control components were heterozygous. In addition, we analyzed 3 mature cystic teratomas without mucinous tumors, and all 3 were homozygous in the tumor component. Our data suggest that the origin of mucinous tumors in the ovary may differ among histological subtypes, and intestinal-type mucinous tumors may arise from mature cystic teratomas, although endocervical-type mucinous tumors may not. Copyright © 2014 Elsevier Inc. All rights reserved.

Automated processing of label-free Raman microscope images of macrophage cells with standardized regression for high-throughput analysis.

PubMed

Milewski, Robert J; Kumagai, Yutaro; Fujita, Katsumasa; Standley, Daron M; Smith, Nicholas I

2010-11-19

Macrophages represent the front lines of our immune system; they recognize and engulf pathogens or foreign particles thus initiating the immune response. Imaging macrophages presents unique challenges, as most optical techniques require labeling or staining of the cellular compartments in order to resolve organelles, and such stains or labels have the potential to perturb the cell, particularly in cases where incomplete information exists regarding the precise cellular reaction under observation. Label-free imaging techniques such as Raman microscopy are thus valuable tools for studying the transformations that occur in immune cells upon activation, both on the molecular and organelle levels. Due to extremely low signal levels, however, Raman microscopy requires sophisticated image processing techniques for noise reduction and signal extraction. To date, efficient, automated algorithms for resolving sub-cellular features in noisy, multi-dimensional image sets have not been explored extensively. We show that hybrid z-score normalization and standard regression (Z-LSR) can highlight the spectral differences within the cell and provide image contrast dependent on spectral content. In contrast to typical Raman imaging processing methods using multivariate analysis, such as single value decomposition (SVD), our implementation of the Z-LSR method can operate nearly in real-time. In spite of its computational simplicity, Z-LSR can automatically remove background and bias in the signal, improve the resolution of spatially distributed spectral differences and enable sub-cellular features to be resolved in Raman microscopy images of mouse macrophage cells. Significantly, the Z-LSR processed images automatically exhibited subcellular architectures whereas SVD, in general, requires human assistance in selecting the components of interest. The computational efficiency of Z-LSR enables automated resolution of sub-cellular features in large Raman microscopy data sets without compromise in image quality or information loss in associated spectra. These results motivate further use of label free microscopy techniques in real-time imaging of live immune cells.

Long-lived and Well-resolved Mn2+ Ion Emissions in CuInS-ZnS Quantum Dots

PubMed Central

Cao, Sheng; Li, Chengming; Wang, Lin; Shang, Minghui; Wei, Guodong; Zheng, Jinju; Yang, Weiyou

2014-01-01

CuInS2 (CIS) quantum dots (QDs) have tunable photoluminescence (PL) behaviors in the visible and near infrared spectral range with markedly lower toxicity than the cadmium-based counterparts, making them very promising applications in light emitting and solar harvesting. However, there still remain material- and fabrication- related obstacles in realizing the high-performance CIS-based QDs with well-resolved Mn2+ d-d emission, long emission lifetimes as well as high efficiencies. Here, we demonstrate the growth of high-quality Mn2+-doped CuInS-ZnS (CIS-ZnS) QDs based on a multi-step hot-injection strategy. The resultant QDs exhibit a well-resolved Mn2+ d-d emission with a high PL quantum yield (QY) up to 66% and an extremely long excited state lifetime up to ~3.78 ms, which is nearly two times longer than the longest one of “green” QDs ever reported. It is promising that the synthesized Mn2+-doped CIS-ZnS QDs might open new doors for their practical applications in bioimaging and opto/electronic devices. PMID:25515207

Developing DIII-D To Prepare For ITER And The Path To Fusion Energy

NASA Astrophysics Data System (ADS)

Buttery, Richard; Hill, David; Solomon, Wayne; Guo, Houyang; DIII-D Team

2017-10-01

DIII-D pursues the advancement of fusion energy through scientific understanding and discovery of solutions. Research targets two key goals. First, to prepare for ITER we must resolve how to use its flexible control tools to rapidly reach Q =10, and develop the scientific basis to interpret results from ITER for fusion projection. Second, we must determine how to sustain a high performance fusion core in steady state conditions, with minimal actuators and a plasma exhaust solution. DIII-D will target these missions with: (i) increased electron heating and balanced torque neutral beams to simulate burning plasma conditions (ii) new 3D coil arrays to resolve control of transients (iii) off axis current drive to study physics in steady state regimes (iv) divertors configurations to promote detachment with low upstream density (v) a reactor relevant wall to qualify materials and resolve physics in reactor-like conditions. With new diagnostics and leading edge simulation, this will position the US for success in ITER and a unique knowledge to accelerate the approach to fusion energy. Supported by the US DOE under DE-FC02-04ER54698.

Characterization of Phenotypic and Transcriptional Differences in Human Pluripotent Stem Cells under 2D and 3D Culture Conditions.

PubMed

Kamei, Ken-Ichiro; Koyama, Yoshie; Tokunaga, Yumie; Mashimo, Yasumasa; Yoshioka, Momoko; Fockenberg, Christopher; Mosbergen, Rowland; Korn, Othmar; Wells, Christine; Chen, Yong

2016-11-01

Human pluripotent stem cells hold great promise for applications in drug discovery and regenerative medicine. Microfluidic technology is a promising approach for creating artificial microenvironments; however, although a proper 3D microenvironment is required to achieve robust control of cellular phenotypes, most current microfluidic devices provide only 2D cell culture and do not allow tuning of physical and chemical environmental cues simultaneously. Here, the authors report a 3D cellular microenvironment plate (3D-CEP), which consists of a microfluidic device filled with thermoresponsive poly(N-isopropylacrylamide)-β-poly(ethylene glycol) hydrogel (HG), which enables systematic tuning of both chemical and physical environmental cues as well as in situ cell monitoring. The authors show that H9 human embryonic stem cells (hESCs) and 253G1 human induced pluripotent stem cells in the HG/3D-CEP system maintain their pluripotent marker expression under HG/3D-CEP self-renewing conditions. Additionally, global gene expression analyses are used to elucidate small variations among different test environments. Interestingly, the authors find that treatment of H9 hESCs under HG/3D-CEP self-renewing conditions results in initiation of entry into the neural differentiation process by induction of PAX3 and OTX1 expression. The authors believe that this HG/3D-CEP system will serve as a versatile platform for developing targeted functional cell lines and facilitate advances in drug screening and regenerative medicine. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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

PubMed

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

2013-02-01

The in vitro generation of a three-dimensional (3-D) myocardial tissue-like construct employing cells, biomaterials, and biomolecules is a promising strategy in cardiac tissue regeneration, drug testing, and tissue engineering applications. Despite significant progress in this field, current cardiac tissue models are not yet able to stably maintain functional characteristics of cardiomyocytes for long-term culture and therapeutic purposes. The objective of this study was to fabricate bioactive 3-D chitosan nanofiber scaffolds using an electrospinning technique and exploring its potential for long-term cardiac function in the 3-D co-culture model. Chitosan is a natural polysaccharide biomaterial that is biocompatible, biodegradable, non-toxic, and cost effective. Electrospun chitosan was utilized to provide structural scaffolding characterized by scale and architectural resemblance to the extracellular matrix (ECM) in vivo. The chitosan fibers were coated with fibronectin via adsorption in order to enhance cellular adhesion to the fibers and migration into the interfibrous milieu. Ventricular cardiomyocytes were harvested from neonatal rats and studied in various culture conditions (i.e., mono- and co-cultures) for their viability and function. Cellular morphology and functionality were examined using immunofluorescent staining for alpha-sarcomeric actin (SM-actin) and gap junction protein, Connexin-43 (Cx43). Scanning electron microscopy (SEM) and light microscopy were used to investigate cellular morphology, spatial organization, and contractions. Calcium indicator was used to monitor calcium ion flux of beating cardiomyocytes. The results demonstrate that the chitosan nanofibers retained their cylindrical morphology in long-term cell cultures and exhibited good cellular attachment and spreading in the presence of adhesion molecule, fibronectin. Cardiomyocyte mono-cultures resulted in loss of cardiomyocyte polarity and islands of non-coherent contractions. However, the cardiomyocyte-fibroblast co-cultures resulted in polarized cardiomyocyte morphology and retained their morphology and function for long-term culture. The Cx43 expression in the fibroblast co-culture was higher than the cardiomyocytes mono-culture and endothelial cells co-culture. In addition, fibroblast co-cultures demonstrated synchronized contractions involving large tissue-like cellular networks. To our knowledge, this is the first attempt to test chitosan nanofiber scaffolds as a 3-D cardiac co-culture model. Our results demonstrate that chitosan nanofibers can serve as a potential scaffold that can retain cardiac structure and function. These studies will provide useful information to develop a strategy that allows us to generate engineered 3-D cardiac tissue constructs using biocompatible and biodegradable chitosan nanofiber scaffolds for many tissue engineering applications. Copyright © 2012 Wiley Periodicals, Inc.

Polymeric scaffolds for three-dimensional culture of nerve cells: a model of peripheral nerve regeneration

PubMed Central

Ayala-Caminero, Radamés; Pinzón-Herrera, Luis; Martinez, Carol A. Rivera; Almodovar, Jorge

2018-01-01

Understanding peripheral nerve repair requires the evaluation of 3D structures that serve as platforms for 3D cell culture. Multiple platforms for 3D cell culture have been developed, mimicking peripheral nerve growth and function, in order to study tissue repair or diseases. To recreate an appropriate 3D environment for peripheral nerve cells, key factors are to be considered including: selection of cells, polymeric biomaterials to be used, and fabrication techniques to shape and form the 3D scaffolds for cellular culture. This review focuses on polymeric 3D platforms used for the development of 3D peripheral nerve cell cultures. PMID:29515936

Echo-Planar Imaging Based J-Resolved Spectroscopic Imaging for Improved Metabolite Detection in Prostate Cancer

DTIC Science & Technology

2013-10-01

Scope: A major outcome is expected to be on improved detection ( specificity ) in differentiating malignant from benign prostate cancer using a novel...Digital Rectal Examination, prostate specific antigen , Four Dimensional (4D) Echo-Planar J-Resolved Spectroscopic Imaging (EP-JRESI); Citrate, Choline... prostate biopsy ranged from 3 to 8, while prostate - specific antigen varied from 2.8 to 20.6 ng/mL (mean of 6.84 ng/mL). A Siemens 3T MRI Scanner with

Engineering cancer microenvironments for in vitro 3-D tumor models

PubMed Central

Asghar, Waseem; El Assal, Rami; Shafiee, Hadi; Pitteri, Sharon; Paulmurugan, Ramasamy; Demirci, Utkan

2017-01-01

The natural microenvironment of tumors is composed of extracellular matrix (ECM), blood vasculature, and supporting stromal cells. The physical characteristics of ECM as well as the cellular components play a vital role in controlling cancer cell proliferation, apoptosis, metabolism, and differentiation. To mimic the tumor microenvironment outside the human body for drug testing, two-dimensional (2-D) and murine tumor models are routinely used. Although these conventional approaches are employed in preclinical studies, they still present challenges. For example, murine tumor models are expensive and difficult to adopt for routine drug screening. On the other hand, 2-D in vitro models are simple to perform, but they do not recapitulate natural tumor microenvironment, because they do not capture important three-dimensional (3-D) cell–cell, cell–matrix signaling pathways, and multi-cellular heterogeneous components of the tumor microenvironment such as stromal and immune cells. The three-dimensional (3-D) in vitro tumor models aim to closely mimic cancer microenvironments and have emerged as an alternative to routinely used methods for drug screening. Herein, we review recent advances in 3-D tumor model generation and highlight directions for future applications in drug testing. PMID:28458612

A Causal Relation between Bioluminescence and Oxygen to Quantify the Cell Niche

PubMed Central

Lambrechts, Dennis; Roeffaers, Maarten; Goossens, Karel; Hofkens, Johan; Van de Putte, Tom; Schrooten, Jan; Van Oosterwyck, Hans

2014-01-01

Bioluminescence imaging assays have become a widely integrated technique to quantify effectiveness of cell-based therapies by monitoring fate and survival of transplanted cells. To date these assays are still largely qualitative and often erroneous due to the complexity and dynamics of local micro-environments (niches) in which the cells reside. Here, we report, using a combined experimental and computational approach, on oxygen that besides being a critical niche component responsible for cellular energy metabolism and cell-fate commitment, also serves a primary role in regulating bioluminescent light kinetics. We demonstrate the potential of an oxygen dependent Michaelis-Menten relation in quantifying intrinsic bioluminescence intensities by resolving cell-associated oxygen gradients from bioluminescent light that is emitted from three-dimensional (3D) cell-seeded hydrogels. Furthermore, the experimental and computational data indicate a strong causal relation of oxygen concentration with emitted bioluminescence intensities. Altogether our approach demonstrates the importance of oxygen to evolve towards quantitative bioluminescence and holds great potential for future microscale measurement of oxygen tension in an easily accessible manner. PMID:24840204

Quantitative mapping of zinc fluxes in the mammalian egg reveals the origin of fertilization-induced zinc sparks

DOE PAGES

Que, Emily L.; Bleher, Reiner; Duncan, Francesca E.; ...

2014-12-15

Fertilization of a mammalian egg induces a series of ‘zinc sparks’ that are necessary for inducing the egg-to-embryo transition. Despite the importance of these zinc efflux events little is known about their origin. To understand the molecular mechanism of the zinc spark we combined four physical approaches to resolve zinc distributions in single cells: a chemical probe for dynamic live-cell fluorescence imaging and a combination of scanning transmission electron microscopy with energy dispersive spectroscopy, X-ray fluorescence microscopy, and 3D elemental tomography for high resolution elemental mapping. Here we show that the zinc spark arises from a system of thousands ofmore » zinc-loaded vesicles, each of which contains, on average, 106 zinc atoms. These vesicles undergo dynamic movement during oocyte maturation and exocytosis at the time of fertilization. We conclude that the discovery of these vesicles and the demonstration that zinc sparks originate from them provides a quantitative framework for understanding how zinc fluxes regulate cellular processes.« less

Shedding light on cell compartmentation in the candidate phylum Poribacteria by high resolution visualisation and transcriptional profiling

NASA Astrophysics Data System (ADS)

Jahn, Martin T.; Markert, Sebastian M.; Ryu, Taewoo; Ravasi, Timothy; Stigloher, Christian; Hentschel, Ute; Moitinho-Silva, Lucas

2016-10-01

Assigning functions to uncultivated environmental microorganisms continues to be a challenging endeavour. Here, we present a new microscopy protocol for fluorescence in situ hybridisation-correlative light and electron microscopy (FISH-CLEM) that enabled, to our knowledge for the first time, the identification of single cells within their complex microenvironment at electron microscopy resolution. Members of the candidate phylum Poribacteria, common and uncultivated symbionts of marine sponges, were used towards this goal. Cellular 3D reconstructions revealed bipolar, spherical granules of low electron density, which likely represent carbon reserves. Poribacterial activity profiles were retrieved from prokaryotic enriched sponge metatranscriptomes using simulation-based optimised mapping. We observed high transcriptional activity for proteins related to bacterial microcompartments (BMC) and we resolved their subcellular localisation by combining FISH-CLEM with immunohistochemistry (IHC) on ultra-thin sponge tissue sections. In terms of functional relevance, we propose that the BMC-A region may be involved in 1,2-propanediol degradation. The FISH-IHC-CLEM approach was proven an effective toolkit to combine -omics approaches with functional studies and it should be widely applicable in environmental microbiology.

A causal relation between bioluminescence and oxygen to quantify the cell niche.

PubMed

Lambrechts, Dennis; Roeffaers, Maarten; Goossens, Karel; Hofkens, Johan; Vande Velde, Greetje; Van de Putte, Tom; Schrooten, Jan; Van Oosterwyck, Hans

2014-01-01

Bioluminescence imaging assays have become a widely integrated technique to quantify effectiveness of cell-based therapies by monitoring fate and survival of transplanted cells. To date these assays are still largely qualitative and often erroneous due to the complexity and dynamics of local micro-environments (niches) in which the cells reside. Here, we report, using a combined experimental and computational approach, on oxygen that besides being a critical niche component responsible for cellular energy metabolism and cell-fate commitment, also serves a primary role in regulating bioluminescent light kinetics. We demonstrate the potential of an oxygen dependent Michaelis-Menten relation in quantifying intrinsic bioluminescence intensities by resolving cell-associated oxygen gradients from bioluminescent light that is emitted from three-dimensional (3D) cell-seeded hydrogels. Furthermore, the experimental and computational data indicate a strong causal relation of oxygen concentration with emitted bioluminescence intensities. Altogether our approach demonstrates the importance of oxygen to evolve towards quantitative bioluminescence and holds great potential for future microscale measurement of oxygen tension in an easily accessible manner.

On the Sensitivity of Atmospheric Ensembles to Cloud Microphysics in Long-Term Cloud-Resolving Model Simulations

NASA Technical Reports Server (NTRS)

Zeng, Xiping; Tao, Wei-Kuo; Lang, Stephen; Hou, Arthur Y.; Zhang, Minghua; Simpson, Joanne

2008-01-01

Month-long large-scale forcing data from two field campaigns are used to drive a cloud-resolving model (CRM) and produce ensemble simulations of clouds and precipitation. Observational data are then used to evaluate the model results. To improve the model results, a new parameterization of the Bergeron process is proposed that incorporates the number concentration of ice nuclei (IN). Numerical simulations reveal that atmospheric ensembles are sensitive to IN concentration and ice crystal multiplication. Two- (2D) and three-dimensional (3D) simulations are carried out to address the sensitivity of atmospheric ensembles to model dimensionality. It is found that the ensembles with high IN concentration are more sensitive to dimensionality than those with low IN concentration. Both the analytic solutions of linear dry models and the CRM output show that there are more convective cores with stronger updrafts in 3D simulations than in 2D, which explains the differing sensitivity of the ensembles to dimensionality at different IN concentrations.

Pathways of energy transfer in LHCII revealed by room-temperature 2D electronic spectroscopy.

PubMed

Wells, Kym L; Lambrev, Petar H; Zhang, Zhengyang; Garab, Gyözö; Tan, Howe-Siang

2014-06-21

We present here the first room-temperature 2D electronic spectroscopy study of energy transfer in the plant light-harvesting complex II, LHCII. Two-dimensional electronic spectroscopy has been used to study energy transfer dynamics in LHCII trimers from the chlorophyll b Qy band to the chlorophyll a Qy band. Observing cross-peak regions corresponding to couplings between different excitonic states reveals partially resolved fine structure at the exciton level that cannot be isolated by pump-probe or linear spectroscopy measurements alone. Global analysis of the data has been performed to identify the pathways and time constants of energy transfer. The measured waiting time (Tw) dependent 2D spectra are found to be composed of 2D decay-associated spectra with three timescales (0.3 ps, 2.3 ps and >20 ps). Direct and multistep cascading pathways from the high-energy chlorophyll b states to the lowest-energy chlorophyll a states have been resolved occurring on time scales of hundreds of femtoseconds to picoseconds.

Case report of asthma associated with 3D printing.

PubMed

House, R; Rajaram, N; Tarlo, S M

2017-12-02

Three-dimensional (3D) printing is being increasingly used in manufacturing and by small business entrepreneurs and home hobbyists. Exposure to airborne emissions during 3D printing raises the issue of whether there may be adverse health effects associated with these emissions. We present a case of a worker who developed asthma while using 3D printers, which illustrates that respiratory problems may be associated with 3D printer emissions. The patient was a 28-year-old self-employed businessman with a past history of asthma in childhood, which had resolved completely by the age of eight. He started using 10 fused deposition modelling 3D printers with acrylonitrile-butadiene-styrene filaments in a small work area of approximately 3000 cubic feet. Ten days later, he began to experience recurrent chest tightness, shortness of breath and coughing at work. After 3 months, his work environment was modified by reducing the number of printers, changing to polylactic acid filaments and using an air purifier with an high-efficiency particulate air filter and organic cartridge. His symptoms improved gradually, although he still needed periodic treatment with a salbutamol inhaler. While still symptomatic, a methacholine challenge indicated a provocation concentration causing a 20% fall in FEV1 (PC20) of 4 mg/ml, consistent with mild asthma. Eventually, his symptoms resolved completely and a second methacholine challenge after symptom resolution was normal (PC20 > 16 mg/ml). This case indicates that workers may develop respiratory problems, including asthma when using 3D printers. Further investigation of the specific airborne emissions and health problems from 3D printing is warranted. © The Author 2017. Published by Oxford University Press on behalf of the Society of Occupational Medicine. All rights reserved. For Permissions, please email: journals.permissions@oup.com

Fluorimetric Studies of a Trans-Membrane Protein and Its Interactions with Differently Functionalized Silver Nanoparticles.

PubMed

Gambucci, Marta; Tarpani, Luigi; Zampini, Giulia; Massaro, Giuseppina; Nocchetti, Morena; Sassi, Paola; Latterini, Loredana

2018-06-18

Trans-membrane proteins play important roles in the inter-cellular signaling to regulate the interactions among adjacent cells and influence cell fate. The study of the interactions between membrane proteins and nanomaterials is paramount for the design of nanomaterial-based therapies. In the present work, the fluorescence properties of the trans-membrane receptor Notch2 have been investigated. In particular, steady state and time resolved fluorescence methods have been used to characterize the emission of tryptophan residues of Notch2 and then this emission is used to monitor the impact of silver colloids on protein behavior. To this aim, silver colloids are prepared with two different methods to make sure they bear hydrophilic (citrate ions, C-AgNPs) or hydrophobic (dodecanethiol molecules D-AgNPs) capping agents; the preparation procedures are tightly controlled in order to obtain metal cores with similar size distributions (7.4 ± 2.5 and 5.0 ± 0.8 nm, respectively), thus making easier the comparison of the results. The occurrence of strong interactions between Notch2 and D-AgNPs is suggested by the efficient and statistically relevant quenching of the stationary protein emission already at low nanoparticle concentrations (ca. 12% quenching with [D-AgNPs] = 0.6nM). The quenching becomes even more pronounced (ca. 60%) when [D-AgNPs] is raised to 8.72nM. On the other hand, the addition of increasing concentrations of C-AgNPs to Notch2 does not affect the protein fluorescence (intensity variations below 5%) indicating that negligible interactions are taking place. The fluorescence data, recorded in the presence of increasing concentrations of silver nanoparticles, are then analyzed through the Stern-Volmer equation and the sphere of action model to discuss the nature of the interactions. The effect of D-AgNPs on the fluorescence decay times of Notch2 is also investigated and a decrease of the average decay time is observed (from 4.64 to 3.42 ns). The observed variations of the stationary and time-resolved fluorescence behavior of the protein are discussed in terms of static and collisional interactions. These results document that the capping shell is able to drive the protein-particle interactions, which have likely a hydrophobic nature.

Four-dimensional (4D) tracking of high-temperature microparticles

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

Wang, Zhehui, E-mail: zwang@lanl.gov; Liu, Q.; Waganaar, W.

High-speed tracking of hot and molten microparticles in motion provides rich information about burning plasmas in magnetic fusion. An exploding-wire apparatus is used to produce moving high-temperature metallic microparticles and to develop four-dimensional (4D) or time-resolved 3D particle tracking techniques. The pinhole camera model and algorithms developed for computer vision are used for scene calibration and 4D reconstructions. 3D positions and velocities are then derived for different microparticles. Velocity resolution approaches 0.1 m/s by using the local constant velocity approximation.

Four-dimensional (4D) tracking of high-temperature microparticles

NASA Astrophysics Data System (ADS)

Wang, Zhehui; Liu, Q.; Waganaar, W.; Fontanese, J.; James, D.; Munsat, T.

2016-11-01

High-speed tracking of hot and molten microparticles in motion provides rich information about burning plasmas in magnetic fusion. An exploding-wire apparatus is used to produce moving high-temperature metallic microparticles and to develop four-dimensional (4D) or time-resolved 3D particle tracking techniques. The pinhole camera model and algorithms developed for computer vision are used for scene calibration and 4D reconstructions. 3D positions and velocities are then derived for different microparticles. Velocity resolution approaches 0.1 m/s by using the local constant velocity approximation.

Four-dimensional (4D) tracking of high-temperature microparticles

DOE PAGES

Wang, Zhehui; Liu, Qiuguang; Waganaar, Bill; ...

2016-07-08

High-speed tracking of hot and molten microparticles in motion provides rich information about burning plasmas in magnetic fusion. An exploding-wire apparatus is used to produce moving high-temperature metallic microparticles and to develop four-dimensional (4D) or time-resolved 3D particle tracking techniques. The pinhole camera model and algorithms developed for computer vision are used for scene calibration and 4D reconstructions. 3D positions and velocities are then derived for different microparticles. As a result, velocity resolution approaches 0.1 m/s by using the local constant velocity approximation.

Four-dimensional (4D) tracking of high-temperature microparticles.

PubMed

Wang, Zhehui; Liu, Q; Waganaar, W; Fontanese, J; James, D; Munsat, T

2016-11-01

High-speed tracking of hot and molten microparticles in motion provides rich information about burning plasmas in magnetic fusion. An exploding-wire apparatus is used to produce moving high-temperature metallic microparticles and to develop four-dimensional (4D) or time-resolved 3D particle tracking techniques. The pinhole camera model and algorithms developed for computer vision are used for scene calibration and 4D reconstructions. 3D positions and velocities are then derived for different microparticles. Velocity resolution approaches 0.1 m/s by using the local constant velocity approximation.

Super-resolved Mirau digital holography by structured illumination

NASA Astrophysics Data System (ADS)

Ganjkhani, Yasaman; Charsooghi, Mohammad A.; Akhlaghi, Ehsan A.; Moradi, Ali-Reza

2017-12-01

In this paper, we apply structured illumination toward super-resolved 3D imaging in a common-path digital holography arrangement. Digital holographic microscopy (DHM) provides non-invasive 3D images of transparent samples as well as 3D profiles of reflective surfaces. A compact and vibration-immune arrangement for DHM may be obtained through the use of a Mirau microscope objective. However, high-magnification Mirau objectives have a low working distance and are expensive. Low-magnification ones, on the other hand, suffer from low lateral resolution. Structured illumination has been widely used for resolution improvement of intensity images, but the technique can also be readily applied to DHM. We apply structured illumination to Mirau DHM by implementing successive sinusoidal gratings with different orientations onto a spatial light modulator (SLM) and forming its image on the specimen. Moreover, we show that, instead of different orientations of 1D gratings, alternative single 2D gratings, e.g. checkerboard or hexagonal patterns, can provide resolution enhancement in multiple directions. Our results show a 35% improvement in the resolution power of the DHM. The presented arrangement has the potential to serve as a table-top device for high resolution holographic microscopy.

Genetic Algorithm Calibration of Probabilistic Cellular Automata for Modeling Mining Permit Activity

USGS Publications Warehouse

Louis, S.J.; Raines, G.L.

2003-01-01

We use a genetic algorithm to calibrate a spatially and temporally resolved cellular automata to model mining activity on public land in Idaho and western Montana. The genetic algorithm searches through a space of transition rule parameters of a two dimensional cellular automata model to find rule parameters that fit observed mining activity data. Previous work by one of the authors in calibrating the cellular automaton took weeks - the genetic algorithm takes a day and produces rules leading to about the same (or better) fit to observed data. These preliminary results indicate that genetic algorithms are a viable tool in calibrating cellular automata for this application. Experience gained during the calibration of this cellular automata suggests that mineral resource information is a critical factor in the quality of the results. With automated calibration, further refinements of how the mineral-resource information is provided to the cellular automaton will probably improve our model.

The 3D model control of image processing

NASA Technical Reports Server (NTRS)

Nguyen, An H.; Stark, Lawrence

1989-01-01

Telerobotics studies remote control of distant robots by a human operator using supervisory or direct control. Even if the robot manipulators has vision or other senses, problems arise involving control, communications, and delay. The communication delays that may be expected with telerobots working in space stations while being controlled from an Earth lab have led to a number of experiments attempting to circumvent the problem. This delay in communication is a main motivating factor in moving from well understood instantaneous hands-on manual control to less well understood supervisory control; the ultimate step would be the realization of a fully autonomous robot. The 3-D model control plays a crucial role in resolving many conflicting image processing problems that are inherent in resolving in the bottom-up approach of most current machine vision processes. The 3-D model control approach is also capable of providing the necessary visual feedback information for both the control algorithms and for the human operator.

Three-dimensional extracellular matrix scaffolds by microfluidic fabrication for long-term spontaneously contracted cardiomyocyte culture.

PubMed

Mei, Jeng-Chun; Wu, Aden Yuan Kun; Wu, Po-Chen; Cheng, Nai-Chen; Tsai, Wei-Bor; Yu, Jiashing

2014-11-01

To repair damaged cardiac tissue, the important principle of in vitro cell culture is to mimic the in vivo cell growth environment. Thus, micro-sized cells are more suitably cultured in three-dimensional (3D) than in two-dimensional (2D) microenvironments (ex: culture dish). With the matching dimensions of works produced by microfluidic technology, chemical engineering and biochemistry applications have used this technology extensively in cellular works. The 3D scaffolds produced in our investigation has essential properties, such has high mass transfer efficiency, and variable pore sizes, to adapt to various needs of different cell types. In addition to the malleability of these innovative scaffolds, fabrication procedure was effortless and fast. Primary neonatal mice cardiomyocytes were successfully harvested and cultured in 3D scaffolds made of gelatin and collagen. Gelatin and gelatin-collagen scaffold were produced by the formation of microbubbles through a microfluidic device, and the mechanical properties of gelatin scaffold and gelatin-collagen scaffold were measured. Cellular properties in the microbubbles were also monitored. Fluorescence staining results assured that cardiomyocytes could maintain in vivo morphology in 3D gelatin scaffold. In addition, it was found that 3D scaffold could prolong the contraction behavior of cardiomyocytes compared with a conventional 2D culture dish. Spontaneously contracted behavior was maintained for the longest (about 1 month) in the 3D gelatin scaffold, about 19 days in the 3D gelatin-collagen scaffold. To sum up, this 3D platform for cell culture has promising potential for myocardial tissue engineering.

Precipitation Processes developed during ARM (1997), TOGA COARE(1992), GATE(1 974), SCSMEX(1998) and KWAJEX(1999): Consistent 2D and 3D Cloud Resolving Model Simulations

NASA Technical Reports Server (NTRS)

Tao, W.-K.; Shie, C.-H.; Simpson, J.; Starr, D.; Johnson, D.; Sud, Y.

2003-01-01

Real clouds and clouds systems are inherently three dimensional (3D). Because of the limitations in computer resources, however, most cloud-resolving models (CRMs) today are still two-dimensional (2D). A few 3D CRMs have been used to study the response of clouds to large-scale forcing. In these 3D simulations, the model domain was small, and the integration time was 6 hours. Only recently have 3D experiments been performed for multi-day periods for tropical cloud system with large horizontal domains at the National Center for Atmospheric Research. The results indicate that surface precipitation and latent heating profiles are very similar between the 2D and 3D simulations of these same cases. The reason for the strong similarity between the 2D and 3D CRM simulations is that the observed large-scale advective tendencies of potential temperature, water vapor mixing ratio, and horizontal momentum were used as the main forcing in both the 2D and 3D models. Interestingly, the 2D and 3D versions of the CRM used in CSU and U.K. Met Office showed significant differences in the rainfall and cloud statistics for three ARM cases. The major objectives of this project are to calculate and axamine: (1)the surface energy and water budgets, (2) the precipitation processes in the convective and stratiform regions, (3) the cloud upward and downward mass fluxes in the convective and stratiform regions; (4) cloud characteristics such as size, updraft intensity and lifetime, and (5) the entrainment and detrainment rates associated with clouds and cloud systems that developed in TOGA COARE, GATE, SCSMEX, ARM and KWAJEX. Of special note is that the analyzed (model generated) data sets are all produced by the same current version of the GCE model, i.e. consistent model physics and configurations. Trajectory analyse and inert tracer calculation will be conducted to identify the differences and similarities in the organization of convection between simulated 2D and 3D cloud systems.

Rapid 3D Refractive‐Index Imaging of Live Cells in Suspension without Labeling Using Dielectrophoretic Cell Rotation

PubMed Central

Habaza, Mor; Kirschbaum, Michael; Guernth‐Marschner, Christian; Dardikman, Gili; Barnea, Itay; Korenstein, Rafi; Duschl, Claus

2016-01-01

A major challenge in the field of optical imaging of live cells is achieving rapid, 3D, and noninvasive imaging of isolated cells without labeling. If successful, many clinical procedures involving analysis and sorting of cells drawn from body fluids, including blood, can be significantly improved. A new label‐free tomographic interferometry approach is presented. This approach provides rapid capturing of the 3D refractive‐index distribution of single cells in suspension. The cells flow in a microfluidic channel, are trapped, and then rapidly rotated by dielectrophoretic forces in a noninvasive and precise manner. Interferometric projections of the rotated cell are acquired and processed into the cellular 3D refractive‐index map. Uniquely, this approach provides full (360°) coverage of the rotation angular range around any axis, and knowledge on the viewing angle. The experimental demonstrations presented include 3D, label‐free imaging of cancer cells and three types of white blood cells. This approach is expected to be useful for label‐free cell sorting, as well as for detection and monitoring of pathological conditions resulting in cellular morphology changes or occurrence of specific cell types in blood or other body fluids. PMID:28251046

Sulphur isotope fractionation during the reduction of elemental sulphur and thiosulphate by Dethiosulfovibrio spp.

PubMed

Surkov, Alexander V; Böttcher, Michael E; Kuever, Jan

2012-01-01

Stable sulphur isotope fractionation was investigated during reduction of thiosulphate and elemental sulphur at 28°C by growing batch cultures of the sulphur- and thiosulphate-reducing bacteria Dethiosulfovibrio marinus (type strain DSM 12537) and Dethiosulfovibrio russensis (type strain DSM 12538), using citrate as carbon and energy source. The cell-specific thiosulphate reduction rate in the growth phase was 7.4±3.9 fmol cell(-1) d(-1). The hydrogen sulphide produced was enriched in (32)S by 10.3±1 ‰ compared with total thiosulphate sulphur, close to previous experimental results observed for other sulphate- and non-sulphate-reducing bacteria. Elemental sulphur reduction yields sulphur isotope enrichment factors between-1.3 and-5.2 ‰ for D. russensis and-1.7 and-5.1 ‰ for D. marinus. The smaller fractionation effects are observed in the exponential growth phase (cellular rates between 5 and 70 fmol S° cell(-1) d(-1)) and enhanced discrimination under conditions of citrate depletion and cell lysis (cellular rates between 0.3 and 3 fmol S° cell(-1) d(-1)).

Absolute earthquake locations using 3-D versus 1-D velocity models below a local seismic network: example from the Pyrenees

NASA Astrophysics Data System (ADS)

Theunissen, T.; Chevrot, S.; Sylvander, M.; Monteiller, V.; Calvet, M.; Villaseñor, A.; Benahmed, S.; Pauchet, H.; Grimaud, F.

2018-03-01

Local seismic networks are usually designed so that earthquakes are located inside them (primary azimuthal gap <<180°) and close to the seismic stations (0-100 km). With these local or near-regional networks (0°-5°), many seismological observatories still routinely locate earthquakes using 1-D velocity models. Moving towards 3-D location algorithms requires robust 3-D velocity models. This work takes advantage of seismic monitoring spanning more than 30 yr in the Pyrenean region. We investigate the influence of a well-designed 3-D model with station corrections including basins structure and the geometry of the Mohorovicic discontinuity on earthquake locations. In the most favourable cases (GAP < 180° and distance to the first station lower than 15 km), results using 1-D velocity models are very similar to 3-D results. The horizontal accuracy in the 1-D case can be higher than in the 3-D case if lateral variations in the structure are not properly resolved. Depth is systematically better resolved in the 3-D model even on the boundaries of the seismic network (GAP > 180° and distance to the first station higher than 15 km). Errors on velocity models and accuracy of absolute earthquake locations are assessed based on a reference data set made of active seismic, quarry blasts and passive temporary experiments. Solutions and uncertainties are estimated using the probabilistic approach of the NonLinLoc (NLLoc) software based on Equal Differential Time. Some updates have been added to NLLoc to better focus on the final solution (outlier exclusion, multiscale grid search, S-phases weighting). Errors in the probabilistic approach are defined to take into account errors on velocity models and on arrival times. The seismicity in the final 3-D catalogue is located with a horizontal uncertainty of about 2.0 ± 1.9 km and a vertical uncertainty of about 3.0 ± 2.0 km.

3D Imaging Millimeter Wave Circular Synthetic Aperture Radar

PubMed Central

Zhang, Renyuan; Cao, Siyang

2017-01-01

In this paper, a new millimeter wave 3D imaging radar is proposed. The user just needs to move the radar along a circular track, and high resolution 3D imaging can be generated. The proposed radar uses the movement of itself to synthesize a large aperture in both the azimuth and elevation directions. It can utilize inverse Radon transform to resolve 3D imaging. To improve the sensing result, the compressed sensing approach is further investigated. The simulation and experimental result further illustrated the design. Because a single transceiver circuit is needed, a light, affordable and high resolution 3D mmWave imaging radar is illustrated in the paper. PMID:28629140

Biogenic iron mineralization at Iron Mountain, CA with implications for detection with the Mars Curiosity rover

USGS Publications Warehouse

Williams, Amy J.; Sumner, Dawn Y.; Alpers, Charles N.; Campbell, Kate M.; Nordstrom, D. Kirk

2014-01-01

(Introduction) Microbe-mineral interactions and biosignature preservation in oxidized sulfidic ore bodies (gossans) are prime candidates for astrobiological study. Such oxidized iron systems have been proposed as analogs for some Martian environments. Recent studies identified microbial fossils preserved as mineral-coated filaments. This study documents microbially-mediated mineral biosignatures in hydrous ferric oxide (HFO) and ferric oxyhydroxysulfates (FOHS) in three environments at Iron Mountain, CA. We investigated microbial community preservation via HFO and FOHS precipitation and the formation of filamentous mineral biosignatures. These environments included 1) actively precipitating (1000's yrs), naturally weathered HFO from in situ gossan, and 3) remobilized iron deposits, which contained lithified clastics and zones of HFO precipitate. We used published biogenicity criteria as guidelines to characterize the biogenicity of mineral filaments. These criteria included A) an actively precipitating environment where microbes are known to be coated in minerals, B) presence of extant microbial communities with carbon signatures, C) structures observable as a part of the host rock, and D) biological morphology, including cellular lumina, multiple member population, numerous taxa, variable and 3-D preservation, biological size ranges, uniform diameter, and evidence of flexibility. This study explores the relevance and detection of these biosignatures to possible Martian biosignatures. Similar filamentous biosignatures are resolvable by the Mars Hand Lens Imager (MAHLI) onboard the Mars Science Laboratory (MSL) rover, Curiosity, and may be identifiable as biogenic if present on Mars.

Rapid construction of mechanically- confined multi- cellular structures using dendrimeric intercellular linker.

PubMed

Mo, Xuejun; Li, Qiushi; Yi Lui, Lena Wai; Zheng, Baixue; Kang, Chiang Huen; Nugraha, Bramasta; Yue, Zhilian; Jia, Rui Rui; Fu, Hong Xia; Choudhury, Deepak; Arooz, Talha; Yan, Jie; Lim, Chwee Teck; Shen, Shali; Hong Tan, Choon; Yu, Hanry

2010-10-01

Tissue constructs that mimic the in vivo cell-cell and cell-matrix interactions are especially useful for applications involving the cell- dense and matrix- poor internal organs. Rapid and precise arrangement of cells into functional tissue constructs remains a challenge in tissue engineering. We demonstrate rapid assembly of C3A cells into multi- cell structures using a dendrimeric intercellular linker. The linker is composed of oleyl- polyethylene glycol (PEG) derivatives conjugated to a 16 arms- polypropylenimine hexadecaamine (DAB) dendrimer. The positively charged multivalent dendrimer concentrates the linker onto the negatively charged cell surface to facilitate efficient insertion of the hydrophobic oleyl groups into the cellular membrane. Bringing linker- treated cells into close proximity to each other via mechanical means such as centrifugation and micromanipulation enables their rapid assembly into multi- cellular structures within minutes. The cells exhibit high levels of viability, proliferation, three- dimensional (3D) cell morphology and other functions in the constructs. We constructed defined multi- cellular structures such as rings, sheets or branching rods that can serve as potential tissue building blocks to be further assembled into complex 3D tissue constructs for biomedical applications. 2010 Elsevier Ltd. All rights reserved.

Diplomatic Solutions to Additive Challenges

DTIC Science & Technology

Additive manufacturing (AM) technology, colloquially known as 3D printing , will bring significant benefits to society, but also poses great risks...regimes, are not sufficient to address the challenges presented by 3D printing technology. The DOS should evaluate and promote unconventional strategies...from printed weapons proliferation. Working with other nations to resolve the appropriate balance between development and security, and to promote norms

Hidden phase in parent Fe-pnictide superconductors

NASA Astrophysics Data System (ADS)

Ali, Khadiza; Adhikary, Ganesh; Thakur, Sangeeta; Patil, Swapnil; Mahatha, Sanjoy K.; Thamizhavel, A.; De Ninno, Giovanni; Moras, Paolo; Sheverdyaeva, Polina M.; Carbone, Carlo; Petaccia, Luca; Maiti, Kalobaran

2018-02-01

We investigate the origin of exoticity in Fe-based systems via studying the fermiology of CaFe2As2 employing angle-resolved photoemission spectroscopy. While the Fermi surfaces (FSs) at 200 K and 31 K are observed to exhibit two-dimensional and three-dimensional (3D) topology, respectively, the FSs at intermediate temperatures reveal the emergence of the 3D topology at a temperature much lower than the structural and magnetic phase transition temperature (170 K, for the sample under scrutiny). This leads to the conclusion that the evolution of FS topology is not directly driven by the structural transition. In addition, we discover the existence in ambient conditions of energy bands related to the cT phase. These bands are distinctly resolved in the high-photon energy spectra exhibiting strong Fe 3 d character. They gradually move to higher binding energies due to thermal compression with cooling, leading to the emergence of 3D topology in the Fermi surface. These results reveal the so-far hidden existence of a cT phase under ambient conditions, which is argued to lead to quantum fluctuations responsible for the exotic electronic properties in Fe-pnictide superconductors.

Ambiguity Resolution for Phase-Based 3-D Source Localization under Fixed Uniform Circular Array.

PubMed

Chen, Xin; Liu, Zhen; Wei, Xizhang

2017-05-11

Under fixed uniform circular array (UCA), 3-D parameter estimation of a source whose half-wavelength is smaller than the array aperture would suffer from a serious phase ambiguity problem, which also appears in a recently proposed phase-based algorithm. In this paper, by using the centro-symmetry of UCA with an even number of sensors, the source's angles and range can be decoupled and a novel algorithm named subarray grouping and ambiguity searching (SGAS) is addressed to resolve angle ambiguity. In the SGAS algorithm, each subarray formed by two couples of centro-symmetry sensors can obtain a batch of results under different ambiguities, and by searching the nearest value among subarrays, which is always corresponding to correct ambiguity, rough angle estimation with no ambiguity is realized. Then, the unambiguous angles are employed to resolve phase ambiguity in a phase-based 3-D parameter estimation algorithm, and the source's range, as well as more precise angles, can be achieved. Moreover, to improve the practical performance of SGAS, the optimal structure of subarrays and subarray selection criteria are further investigated. Simulation results demonstrate the satisfying performance of the proposed method in 3-D source localization.

Human/Computer Transaction Tasks: An Annotated Bibliography.

DTIC Science & Technology

1982-05-01

Operations (Manpower, Personnel, and Training, OP-01). The subproject was directed toward resolving fundamental human engineering design issues in...1978 and results were used at the Navy Personnel Research and Development Center in research to resolve fundamental human engineering design issues for...Dialogue Monitor and analysis of Sthe data obtained are briefly discussed. Alden, D. G., Daniels, P. 3., and Kanarick, A. F. Keyboard design and

Temperature dependence of laser-induced fluorescence of Tb3+Tb3+ in molten LiCl-KCl eutectic

NASA Astrophysics Data System (ADS)

C., E.; -E., Jung | S.; | W., Bae; Cha | I., A.; Bae | Y., J.; | K., Park; Song

2011-01-01

Fluorescence spectra and lifetimes originated from both 5D3 →7FJ and 5D4 →7FJ transitions of Tb3+ were measured using time-resolved laser fluorescence spectroscopy in order to investigate the excited state relaxation in a molten salt medium. A cross-relaxation energy transfer of 5D3 →5D4 resulted in rise and decay behaviors in fluorescence signal waveforms of 5D4 →7FJ transitions. The fluorescence intensity ratios of 5D4 →7F5 to 5D3 →7F4 decreased drastically when the temperature of molten salt increased. This result suggests that the cross-relaxation effect becomes weakened with increasing temperature. In addition, a strong increase of the 5D4 emission over the 5D3 emission was observed at high Tb3+ concentration.

Performance of high-temperature superconducting band-pass filters with high selectivity for base transceiver applications of digital cellular communication systems

NASA Astrophysics Data System (ADS)

Kwak, J. S.; Lee, J. H.; Kim, C. O.; Hong, J. P.; Han, S. K.; Char, K.

2002-07-01

Highly selective high-temperature superconducting band-pass filters based on spiral meander line structures have been developed for base transceiver station applications of digital cellular communication systems. The filter comprised 12-pole microstrip line resonators with a circuit size of 0.5 × 17 × 41 mm3. The filter was designed to have a bandwidth of 25 MHz at a centre frequency of 834 MHz. Particularly, the physical size of each resonator was chosen not only to reduce far-field radiation, but also to have reasonable tunability in the filter. Device characteristics exhibited a low insertion loss of 0.4 dB with a 0.2 dB ripple and a return loss better than 10 dB in the pass-band at 65 K. The out-of-band signals were attenuated better than 60 dB at about 3.5 MHz from the lower band edge, and 3.8 MHz from the higher band edge.

Linear Dichroism in Angle-Resolved Core-Level Photoemission Spectra Reflecting 4f Ground-State Symmetry of Strongly Correlated Cubic Pr Compounds

NASA Astrophysics Data System (ADS)

Hamamoto, Satoru; Fujioka, Shuhei; Kanai, Yuina; Yamagami, Kohei; Nakatani, Yasuhiro; Nakagawa, Koya; Fujiwara, Hidenori; Kiss, Takayuki; Higashiya, Atsushi; Yamasaki, Atsushi; Kadono, Toshiharu; Imada, Shin; Tanaka, Arata; Tamasaku, Kenji; Yabashi, Makina; Ishikawa, Tetsuya; Matsumoto, Keisuke T.; Onimaru, Takahiro; Takabatake, Toshiro; Sekiyama, Akira

2017-12-01

We report experimentally observed linear dichroism in angle-resolved core-level photoemission spectra of PrIr2Zn20 and PrB6 with cubic symmetry. The different anisotropic 4f charge distributions between the compounds due to the crystalline-electric-field splitting are responsible for the difference in the linear dichroism, which has been verified by spectral simulations with the full multiplet theory for a single-site Pr3+ ion with cubic symmetry. The observed linear dichroism and polarization-dependent spectra in two different photoelectron directions for PrIr2Zn20 are reproduced by theoretical analysis for the Γ3 ground state, whereas those of the Pr 3d and 4d core levels indicate the Γ5 ground state for PrB6.

Resolved Star Formation in Galaxies Using Slitless Spectroscopy

NASA Astrophysics Data System (ADS)

Pirzkal, Norbert; Finkelstein, Steven L.; Larson, Rebecca L.; Malhotra, Sangeeta; Rhoads, James E.; Ryan, Russell E.; Tilvi, Vithal; FIGS Team

2018-06-01

The ability to spatially resolve individual star-formation regions in distant galaxies and simultaneously extract their physical properties via emission lines is a critical step forward in studying the evolution of galaxies. While efficient, deep slitless spectroscopic observations offer a blurry view of the summed properties of galaxies. We present our studies of resolved star formation over a wide range of redshifts, including high redshift Ly-a sources. The unique capabilities of the WFC3 IR Grism and our two-dimensional emission line method (EM2D) allows us to accurately identify the specific spatial origin of emission lines in galaxies, thus creating a spatial map of star-formation sites in any given galaxy. This method requires the use of multiple position angles on the sky to accurately derive both the location and the observed wavelengths of these emission lines. This has the added benefit of producing better defined redshifts for these sources. Building on our success in applying the EM2D method towards galaxies with [OII]. [OIII], and Ha emission lines, we have also applied EM2D to high redshift (z>6) Ly-a emitting galaxies. We are also able to produce accurate 2D emission line maps (MAP2D) of the Ly-a emission in WFC3 IR grism observations, looking for evidence that a significant amount of resonant scattering is taking place in high redshift galaxies such as in a newly identified z=7.5 Faint Infrared Galaxy Survey (FIGS) Ly-a galaxy.

On the partition dimension of comb product of path and complete graph

NASA Astrophysics Data System (ADS)

Darmaji, Alfarisi, Ridho

2017-08-01

For a vertex v of a connected graph G(V, E) with vertex set V(G), edge set E(G) and S ⊆ V(G). Given an ordered partition Π = {S1, S2, S3, …, Sk} of the vertex set V of G, the representation of a vertex v ∈ V with respect to Π is the vector r(v|Π) = (d(v, S1), d(v, S2), …, d(v, Sk)), where d(v, Sk) represents the distance between the vertex v and the set Sk and d(v, Sk) = min{d(v, x)|x ∈ Sk}. A partition Π of V(G) is a resolving partition if different vertices of G have distinct representations, i.e., for every pair of vertices u, v ∈ V(G), r(u|Π) ≠ r(v|Π). The minimum k of Π resolving partition is a partition dimension of G, denoted by pd(G). Finding the partition dimension of G is classified to be a NP-Hard problem. In this paper, we will show that the partition dimension of comb product of path and complete graph. The results show that comb product of complete grapph Km and path Pn namely p d (Km⊳Pn)=m where m ≥ 3 and n ≥ 2 and p d (Pn⊳Km)=m where m ≥ 3, n ≥ 2 and m ≥ n.

Deterministic Integration of Biological and Soft Materials onto 3D Microscale Cellular Frameworks

PubMed Central

McCracken, Joselle M.; Xu, Sheng; Badea, Adina; Jang, Kyung-In; Yan, Zheng; Wetzel, David J.; Nan, Kewang; Lin, Qing; Han, Mengdi; Anderson, Mikayla A.; Lee, Jung Woo; Wei, Zijun; Pharr, Matt; Wang, Renhan; Su, Jessica; Rubakhin, Stanislav S.; Sweedler, Jonathan V.

2018-01-01

Complex 3D organizations of materials represent ubiquitous structural motifs found in the most sophisticated forms of matter, the most notable of which are in life-sustaining hierarchical structures found in biology, but where simpler examples also exist as dense multilayered constructs in high-performance electronics. Each class of system evinces specific enabling forms of assembly to establish their functional organization at length scales not dissimilar to tissue-level constructs. This study describes materials and means of assembly that extend and join these disparate systems—schemes for the functional integration of soft and biological materials with synthetic 3D microscale, open frameworks that can leverage the most advanced forms of multilayer electronic technologies, including device-grade semiconductors such as monocrystalline silicon. Cellular migration behaviors, temporal dependencies of their growth, and contact guidance cues provided by the nonplanarity of these frameworks illustrate design criteria useful for their functional integration with living matter (e.g., NIH 3T3 fibroblast and primary rat dorsal root ganglion cell cultures). PMID:29552634

Depth resolved hyperspectral imaging spectrometer based on structured light illumination and Fourier transform interferometry

PubMed Central

Choi, Heejin; Wadduwage, Dushan; Matsudaira, Paul T.; So, Peter T.C.

2014-01-01

A depth resolved hyperspectral imaging spectrometer can provide depth resolved imaging both in the spatial and the spectral domain. Images acquired through a standard imaging Fourier transform spectrometer do not have the depth-resolution. By post processing the spectral cubes (x, y, λ) obtained through a Sagnac interferometer under uniform illumination and structured illumination, spectrally resolved images with depth resolution can be recovered using structured light illumination algorithms such as the HiLo method. The proposed scheme is validated with in vitro specimens including fluorescent solution and fluorescent beads with known spectra. The system is further demonstrated in quantifying spectra from 3D resolved features in biological specimens. The system has demonstrated depth resolution of 1.8 μm and spectral resolution of 7 nm respectively. PMID:25360367

FRET Imaging in Three-dimensional Hydrogels

PubMed Central

Taboas, Juan M.

2016-01-01

Imaging of Förster resonance energy transfer (FRET) is a powerful tool for examining cell biology in real-time. Studies utilizing FRET commonly employ two-dimensional (2D) culture, which does not mimic the three-dimensional (3D) cellular microenvironment. A method to perform quenched emission FRET imaging using conventional widefield epifluorescence microscopy of cells within a 3D hydrogel environment is presented. Here an analysis method for ratiometric FRET probes that yields linear ratios over the probe activation range is described. Measurement of intracellular cyclic adenosine monophosphate (cAMP) levels is demonstrated in chondrocytes under forskolin stimulation using a probe for EPAC1 activation (ICUE1) and the ability to detect differences in cAMP signaling dependent on hydrogel material type, herein a photocrosslinking hydrogel (PC-gel, polyethylene glycol dimethacrylate) and a thermoresponsive hydrogel (TR-gel). Compared with 2D FRET methods, this method requires little additional work. Laboratories already utilizing FRET imaging in 2D can easily adopt this method to perform cellular studies in a 3D microenvironment. It can further be applied to high throughput drug screening in engineered 3D microtissues. Additionally, it is compatible with other forms of FRET imaging, such as anisotropy measurement and fluorescence lifetime imaging (FLIM), and with advanced microscopy platforms using confocal, pulsed, or modulated illumination. PMID:27500354

Roles of Diffusion Dynamics in Stem Cell Signaling and Three-Dimensional Tissue Development.

PubMed

McMurtrey, Richard J

2017-09-15

Recent advancements in the ability to construct three-dimensional (3D) tissues and organoids from stem cells and biomaterials have not only opened abundant new research avenues in disease modeling and regenerative medicine but also have ignited investigation into important aspects of molecular diffusion in 3D cellular architectures. This article describes fundamental mechanics of diffusion with equations for modeling these dynamic processes under a variety of scenarios in 3D cellular tissue constructs. The effects of these diffusion processes and resultant concentration gradients are described in the context of the major molecular signaling pathways in stem cells that both mediate and are influenced by gas and nutrient concentrations, including how diffusion phenomena can affect stem cell state, cell differentiation, and metabolic states of the cell. The application of these diffusion models and pathways is of vital importance for future studies of developmental processes, disease modeling, and tissue regeneration.

Biomimetic strategies for the glioblastoma microenvironment

NASA Astrophysics Data System (ADS)

Cha, Junghwa; Kim, Pilnam

2017-12-01

Glioblastoma multiforme (GBM) is a devastating type of tumor with high mortality, caused by extensive infiltration into adjacent tissue and rapid recurrence. Most therapies for GBM have focused on the cytotoxicity, and have not targeted GBM spread. However, there have been numerous attempts to improve therapy by addressing GBM invasion, through understanding and mimicking its behavior using three-dimensional (3D) experimental models. Compared with two-dimensional models and in vivo animal models, 3D GBM models can capture the invasive motility of glioma cells within a 3D environment comprising many cellular and non-cellular components. Based on tissue engineering techniques, GBM invasion has been investigated within a biologically relevant environment, from biophysical and biochemical perspectives, to clarify the pro-invasive factors of GBM. This review discusses the recent progress in techniques for modeling the microenvironments of GBM tissue and suggests future directions with respect to recreating the GBM microenvironment and preclinical applications.

High-speed 3D imaging of cellular activity in the brain using axially-extended beams and light sheets.

PubMed

Hillman, Elizabeth Mc; Voleti, Venkatakaushik; Patel, Kripa; Li, Wenze; Yu, Hang; Perez-Campos, Citlali; Benezra, Sam E; Bruno, Randy M; Galwaduge, Pubudu T

2018-06-01

As optical reporters and modulators of cellular activity have become increasingly sophisticated, the amount that can be learned about the brain via high-speed cellular imaging has increased dramatically. However, despite fervent innovation, point-scanning microscopy is facing a fundamental limit in achievable 3D imaging speeds and fields of view. A range of alternative approaches are emerging, some of which are moving away from point-scanning to use axially-extended beams or sheets of light, for example swept confocally aligned planar excitation (SCAPE) microscopy. These methods are proving effective for high-speed volumetric imaging of the nervous system of small organisms such as Drosophila (fruit fly) and D. Rerio (Zebrafish), and are showing promise for imaging activity in the living mammalian brain using both single and two-photon excitation. This article describes these approaches and presents a simple model that demonstrates key advantages of axially-extended illumination over point-scanning strategies for high-speed volumetric imaging, including longer integration times per voxel, improved photon efficiency and reduced photodamage. Copyright © 2018 Elsevier Ltd. All rights reserved.

Modeling of time dependent localized flow shear stress and its impact on cellular growth within additive manufactured titanium implants.

PubMed

Zhang, Ziyu; Yuan, Lang; Lee, Peter D; Jones, Eric; Jones, Julian R

2014-11-01

Bone augmentation implants are porous to allow cellular growth, bone formation and fixation. However, the design of the pores is currently based on simple empirical rules, such as minimum pore and interconnects sizes. We present a three-dimensional (3D) transient model of cellular growth based on the Navier-Stokes equations that simulates the body fluid flow and stimulation of bone precursor cellular growth, attachment, and proliferation as a function of local flow shear stress. The model's effectiveness is demonstrated for two additive manufactured (AM) titanium scaffold architectures. The results demonstrate that there is a complex interaction of flow rate and strut architecture, resulting in partially randomized structures having a preferential impact on stimulating cell migration in 3D porous structures for higher flow rates. This novel result demonstrates the potential new insights that can be gained via the modeling tool developed, and how the model can be used to perform what-if simulations to design AM structures to specific functional requirements. © 2014 Wiley Periodicals, Inc.

Chapter 6: cubic membranes the missing dimension of cell membrane organization.

PubMed

Almsherqi, Zakaria A; Landh, Tomas; Kohlwein, Sepp D; Deng, Yuru

2009-01-01

Biological membranes are among the most fascinating assemblies of biomolecules: a bilayer less than 10 nm thick, composed of rather small lipid molecules that are held together simply by noncovalent forces, defines the cell and discriminates between "inside" and "outside", survival, and death. Intracellular compartmentalization-governed by biomembranes as well-is a characteristic feature of eukaryotic cells, which allows them to fulfill multiple and highly specialized anabolic and catabolic functions in strictly controlled environments. Although cellular membranes are generally visualized as flat sheets or closely folded isolated objects, multiple observations also demonstrate that membranes may fold into "unusual", highly organized structures with 2D or 3D periodicity. The obvious correlation of highly convoluted membrane organizations with pathological cellular states, for example, as a consequence of viral infection, deserves close consideration. However, knowledge about formation and function of these highly organized 3D periodic membrane structures is scarce, primarily due to the lack of appropriate techniques for their analysis in vivo. Currently, the only direct way to characterize cellular membrane architecture is by transmission electron microscopy (TEM). However, deciphering the spatial architecture solely based on two-dimensionally projected TEM images is a challenging task and prone to artifacts. In this review, we will provide an update on the current progress in identifying and analyzing 3D membrane architectures in biological systems, with a special focus on membranes with cubic symmetry, and their potential role in physiological and pathophysiological conditions. Proteomics and lipidomics approaches in defined experimental cell systems may prove instrumental to understand formation and function of 3D membrane morphologies.

Modification of measurement methods for evaluation of tissue-engineered cartilage function and biochemical properties using nanosecond pulsed laser

NASA Astrophysics Data System (ADS)

Ishihara, Miya; Sato, Masato; Kutsuna, Toshiharu; Ishihara, Masayuki; Mochida, Joji; Kikuchi, Makoto

2008-02-01

There is a demand in the field of regenerative medicine for measurement technology that enables determination of functions and components of engineered tissue. To meet this demand, we developed a method for extracellular matrix characterization using time-resolved autofluorescence spectroscopy, which enabled simultaneous measurements with mechanical properties using relaxation of laser-induced stress wave. In this study, in addition to time-resolved fluorescent spectroscopy, hyperspectral sensor, which enables to capture both spectral and spatial information, was used for evaluation of biochemical characterization of tissue-engineered cartilage. Hyperspectral imaging system provides spectral resolution of 1.2 nm and image rate of 100 images/sec. The imaging system consisted of the hyperspectral sensor, a scanner for x-y plane imaging, magnifying optics and Xenon lamp for transmmissive lighting. Cellular imaging using the hyperspectral image system has been achieved by improvement in spatial resolution up to 9 micrometer. The spectroscopic cellular imaging could be observed using cultured chondrocytes as sample. At early stage of culture, the hyperspectral imaging offered information about cellular function associated with endogeneous fluorescent biomolecules.

Time-resolved 3D MR velocity mapping at 3T: improved navigator-gated assessment of vascular anatomy and blood flow.

PubMed

Markl, Michael; Harloff, Andreas; Bley, Thorsten A; Zaitsev, Maxim; Jung, Bernd; Weigang, Ernst; Langer, Mathias; Hennig, Jürgen; Frydrychowicz, Alex

2007-04-01

To evaluate an improved image acquisition and data-processing strategy for assessing aortic vascular geometry and 3D blood flow at 3T. In a study with five normal volunteers and seven patients with known aortic pathology, prospectively ECG-gated cine three-dimensional (3D) MR velocity mapping with improved navigator gating, real-time adaptive k-space ordering and dynamic adjustment of the navigator acceptance criteria was performed. In addition to morphological information and three-directional blood flow velocities, phase-contrast (PC)-MRA images were derived from the same data set, which permitted 3D isosurface rendering of vascular boundaries in combination with visualization of blood-flow patterns. Analysis of navigator performance and image quality revealed improved scan efficiencies of 63.6%+/-10.5% and temporal resolution (<50 msec) compared to previous implementations. Semiquantitative evaluation of image quality by three independent observers demonstrated excellent general image appearance with moderate blurring and minor ghosting artifacts. Results from volunteer and patient examinations illustrate the potential of the improved image acquisition and data-processing strategy for identifying normal and pathological blood-flow characteristics. Navigator-gated time-resolved 3D MR velocity mapping at 3T in combination with advanced data processing is a powerful tool for performing detailed assessments of global and local blood-flow characteristics in the aorta to describe or exclude vascular alterations. Copyright (c) 2007 Wiley-Liss, Inc.

From SHAPE Signatures to 3-D Structures | Center for Cancer Research

Cancer.gov

RNAs undergo extensive folding to form sophisticated based-paired secondary structures that are, in part, indicators of more complex three-dimensional structures.  These 3-D shapes are an integral part of the cellular gene-expression machinery. Deconstructing these structures is no small matter, yet it is critical to understanding their function.

Quantitative Large-Scale Three-Dimensional Imaging of Human Kidney Biopsies: A Bridge to Precision Medicine in Kidney Disease.

PubMed

Winfree, Seth; Dagher, Pierre C; Dunn, Kenneth W; Eadon, Michael T; Ferkowicz, Michael; Barwinska, Daria; Kelly, Katherine J; Sutton, Timothy A; El-Achkar, Tarek M

2018-06-05

Kidney biopsy remains the gold standard for uncovering the pathogenesis of acute and chronic kidney diseases. However, the ability to perform high resolution, quantitative, molecular and cellular interrogation of this precious tissue is still at a developing stage compared to other fields such as oncology. Here, we discuss recent advances in performing large-scale, three-dimensional (3D), multi-fluorescence imaging of kidney biopsies and quantitative analysis referred to as 3D tissue cytometry. This approach allows the accurate measurement of specific cell types and their spatial distribution in a thick section spanning the entire length of the biopsy. By uncovering specific disease signatures, including rare occurrences, and linking them to the biology in situ, this approach will enhance our understanding of disease pathogenesis. Furthermore, by providing accurate quantitation of cellular events, 3D cytometry may improve the accuracy of prognosticating the clinical course and response to therapy. Therefore, large-scale 3D imaging and cytometry of kidney biopsy is poised to become a bridge towards personalized medicine for patients with kidney disease. © 2018 S. Karger AG, Basel.

High-throughput 3D whole-brain quantitative histopathology in rodents

PubMed Central

Vandenberghe, Michel E.; Hérard, Anne-Sophie; Souedet, Nicolas; Sadouni, Elmahdi; Santin, Mathieu D.; Briet, Dominique; Carré, Denis; Schulz, Jocelyne; Hantraye, Philippe; Chabrier, Pierre-Etienne; Rooney, Thomas; Debeir, Thomas; Blanchard, Véronique; Pradier, Laurent; Dhenain, Marc; Delzescaux, Thierry

2016-01-01

Histology is the gold standard to unveil microscopic brain structures and pathological alterations in humans and animal models of disease. However, due to tedious manual interventions, quantification of histopathological markers is classically performed on a few tissue sections, thus restricting measurements to limited portions of the brain. Recently developed 3D microscopic imaging techniques have allowed in-depth study of neuroanatomy. However, quantitative methods are still lacking for whole-brain analysis of cellular and pathological markers. Here, we propose a ready-to-use, automated, and scalable method to thoroughly quantify histopathological markers in 3D in rodent whole brains. It relies on block-face photography, serial histology and 3D-HAPi (Three Dimensional Histology Analysis Pipeline), an open source image analysis software. We illustrate our method in studies involving mouse models of Alzheimer’s disease and show that it can be broadly applied to characterize animal models of brain diseases, to evaluate therapeutic interventions, to anatomically correlate cellular and pathological markers throughout the entire brain and to validate in vivo imaging techniques. PMID:26876372

Bilayered nanofibrous 3D hierarchy as skin rudiment by emulsion electrospinning for burn wound management.

PubMed

Pal, Pallabi; Dadhich, Prabhash; Srivas, Pavan Kumar; Das, Bodhisatwa; Maulik, Dhrubajyoti; Dhara, Santanu

2017-08-22

Mimicking skin extracellular matrix hierarchy, the present work aims to develop a bilayer skin graft comprising a porous cotton-wool-like 3D layer with membranous structure of PCL-chitosan nanofibers. Emulsion electrospinning with differential stirring periods of PCL-chitosan emulsion results in development of a bilayer 3D structure with varied morphology. The electrospun membrane has fiber diameter ∼274 nm and pore size ∼1.16 μm while fluffy 3D layer has fiber diameter ∼1.62 μm and pore size ∼62 μm. The 3D layer was further coated with collagen I isolated from Cirrhinus cirrhosus fish scales to improve biofunctionality. Surface coating with collagen I resulted in bundling the fibers together, thereby increasing their average diameter to 2.80 μm and decreasing pore size to ∼45 μm. The architecture and composition of the scaffold promotes efficient cellular activity where interconnected porosity with ECM resembling collagen I coating assists cellular adhesion, infiltration, and proliferation from initial days of fibroblast seeding, while keratinocytes migrate on the surface only without infiltrating in the membranous nanofiber layer. Anatomy of the scaffold arising due to variation in pore size distribution at different layers thereby facilitates compartmentalization and prevents initial cellular transmigration. The scaffold also assists in extracellular matrix protein synthesis and keratinocyte stratification in vitro. Further, the scaffold effectively integrates and attaches with third-degree burn wound margins created in rat models and accelerates healing in comparison to standard Tegaderm dressing™. The bilayer scaffold is thus a promising, readily available, cost-effective, off-the-shelf matrix as a skin substitute.

Topological Optimization of Artificial Microstructure Strategies

DTIC Science & Technology

2015-04-02

a 3D microstructural architecture structure made from bulk metallic glass , 3DMGS, exhibiting a combination of ceramic-like high strength (>1000 MPa...Research Triangle Park, NC 27709-2211 materials, cellular structures, metallic glass REPORT DOCUMENTATION PAGE 11. SPONSOR/MONITOR’S REPORT NUMBER(S...demonstrate a 3D microstructural architecture structure made from bulk metallic glass , 3DMGS, exhibiting a combination of ceramic-like high strength

Cyclin D2 in the basal process of neural progenitors is linked to non-equivalent cell fates

PubMed Central

Tsunekawa, Yuji; Britto, Joanne M; Takahashi, Masanori; Polleux, Franck; Tan, Seong-Seng; Osumi, Noriko

2012-01-01

Asymmetric cell division plays an indispensable role during corticogenesis for producing new neurons while maintaining a self-renewing pool of apical progenitors. The cellular and molecular determinants favouring asymmetric division are not completely understood. Here, we identify a novel mechanism for generating cellular asymmetry through the active transportation and local translation of Cyclin D2 mRNA in the basal process. This process is regulated by a unique cis-regulatory sequence found in the 3′ untranslated region (3′UTR) of the mRNA. Unequal inheritance of Cyclin D2 protein to the basally positioned daughter cell with the basal process confers renewal of the apical progenitor after asymmetric division. Conversely, depletion of Cyclin D2 in the apically positioned daughter cell results in terminal neuronal differentiation. We demonstrate that Cyclin D2 is also expressed in the developing human cortex within similar domains, thus indicating that its role as a fate determinant is ancient and conserved. PMID:22395070

Photo- and electropatterning of hydrogel-encapsulated living cell arrays.

PubMed

Albrecht, Dirk R; Tsang, Valerie Liu; Sah, Robert L; Bhatia, Sangeeta N

2005-01-01

Living cells have the potential to serve as sensors, naturally integrating the response to stimuli to generate predictions about cell fate (e.g., differentiation, migration, proliferation, apoptosis). Miniaturized arrays of living cells further offer the capability to interrogate many cells in parallel and thereby enable high-throughput and/or combinatorial assays. However, the interface between living cells and synthetic chip platforms is a critical one wherein the cellular phenotype must be preserved to generate useful signals. While some cell types retain tissue-specific features on a flat (2-D) surface, it has become increasingly apparent that a 3-D physical environment will be required for others. In this paper, we present two independent methods for creating living cell arrays that are encapsulated within a poly(ethylene glycol)-based hydrogel to create a local 3-D microenvironment. First, 'photopatterning' selectively crosslinks hydrogel microstructures containing living cells with approximately 100 microm feature size. Second, 'electropatterning' utilizes dielectrophoretic forces to position cells within a prepolymer solution prior to crosslinking, forming cell patterns with micron resolution. We further combine these methods to obtain hierarchical control of cell positioning over length scales ranging from microns to centimeters. This level of microenvironmental control should enable the fabrication of next-generation cellular microarrays in which robust 3-D cultures of cells are presented with appropriate physical and chemical cues and, consequently, report on cellular responses that resemble in vivo behavior.

Tradeoff Analysis for Combat Service Support Wireless Communications Alternatives

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

Burnette, John R.; Thibodeau, Christopher C.; Greitzer, Frank L.

2002-02-28

As the Army moves toward more mobile and agile forces and continued sustainment of numerous high-cost legacy logistics management systems, the requirement for wireless connectivity and a wireless network to supporting organizations has become ever more critical. There are currently several Army communications initiatives underway to resolve this wireless connectivity issue. However, to fully appreciate and understand the value of these initiatives, a Tradeoff Analysis is needed. The present study seeks to identify and assess solutions. The analysis identified issues that impede Interim Brigade Combat Team (IBCT) communication system integration and outlined core requirements for sharing of logistics data betweenmore » the field and Army battle command systems. Then, the analysis examined wireless communication alternatives as possible solutions for IBCT logistics communications problems. The current baseline system was compared with possible alternatives involving tactical radio systems, wireless/near term digital radio, cellular satellite, and third-generation (3G) wireless technologies. Cellular satellite and 3G wireless technologies offer clear advantages and should be considered for later IBCTs.« less

Phenolic derivatives from soy flour ethanol extract are potent in vitro quinone reductase (QR) inducing agents.

PubMed

Bolling, Bradley W; Parkin, Kirk L

2008-11-26

The fractionation of soy flour directed by a cellular bioassay for induction of phase 2 detoxification enzymes was used to identify quinone reductase (QR) inducing agents. A phospholipid-depleted, 80% methanol-partitioned isolate from a crude ethanol extract of soy flour was resolved using normal phase medium-pressure liquid chromatography (MPLC). Early eluting fractions were found to be the most potent QR inducing agents among the separated fractions. Fraction 2 was the most potent, doubling QR at <2 mug/mL. Further fractionation of this isolate led to the identification of several constituents. Fatty acids and sn-1 and sn-2 monoacylglycerols were identified, but were not highly potent QR inducers. Benzofuran-3-carbaldehyde, 4-hydroxybenzaldeyde, 4-ethoxybenzoic acid, 4-ethoxycinnamic acid, benzofuran-2-carboxylic ethyl ester, and ferulic acid ethyl ester (FAEE) were also identified as QR inducing constituents of this fraction. FAEE was the most potent of the identified constituents, doubling QR specific activity at 3.2 muM in the cellular bioassay.

Angle-resolved light scattering of individual rod-shaped bacteria based on Fourier transform light scattering

NASA Astrophysics Data System (ADS)

Jo, Youngju; Jung, Jaehwang; Lee, Jee Woong; Shin, Della; Park, Hyunjoo; Nam, Ki Tae; Park, Ji-Ho; Park, Yongkeun

2014-05-01

Two-dimensional angle-resolved light scattering maps of individual rod-shaped bacteria are measured at the single-cell level. Using quantitative phase imaging and Fourier transform light scattering techniques, the light scattering patterns of individual bacteria in four rod-shaped species (Bacillus subtilis, Lactobacillus casei, Synechococcus elongatus, and Escherichia coli) are measured with unprecedented sensitivity in a broad angular range from -70° to 70°. The measured light scattering patterns are analyzed along the two principal axes of rod-shaped bacteria in order to systematically investigate the species-specific characteristics of anisotropic light scattering. In addition, the cellular dry mass of individual bacteria is calculated and used to demonstrate that the cell-to-cell variations in light scattering within bacterial species is related to the cellular dry mass and growth.

Cardiac muscle organization revealed in 3-D by imaging whole-mount mouse hearts using two-photon fluorescence and confocal microscopy.

PubMed

Sivaguru, Mayandi; Fried, Glenn; Sivaguru, Barghav S; Sivaguru, Vignesh A; Lu, Xiaochen; Choi, Kyung Hwa; Saif, M Taher A; Lin, Brian; Sadayappan, Sakthivel

2015-11-01

The ability to image the entire adult mouse heart at high resolution in 3-D would provide enormous advantages in the study of heart disease. However, a technique for imaging nuclear/cellular detail as well as the overall structure of the entire heart in 3-D with minimal effort is lacking. To solve this problem, we modified the benzyl alcohol:benzyl benzoate (BABB) clearing technique by labeling mouse hearts with periodic acid Schiff (PAS) stain. We then imaged the hearts with a combination of two-photon fluorescence microscopy and automated tile-scan imaging/stitching. Utilizing the differential spectral properties of PAS, we could identify muscle and nuclear compartments in the heart. We were also able to visualize the differences between a 3-month-old normal mouse heart and a mouse heart that had undergone heart failure due to the expression of cardiac myosin binding protein-C (cMyBP-C) gene mutation (t/t). Using 2-D and 3-D morphometric analysis, we found that the t/t heart had anomalous ventricular shape, volume, and wall thickness, as well as a disrupted sarcomere pattern. We further validated our approach using decellularized hearts that had been cultured with 3T3 fibroblasts, which were tracked using a nuclear label. We were able to detect the 3T3 cells inside the decellularized intact heart tissue, achieving nuclear/cellular resolution in 3-D. The combination of labeling, clearing, and two-photon microscopy together with tiling eliminates laborious and time-consuming physical sectioning, alignment, and 3-D reconstruction.

Effects of 1,25(OH)2 D3 and 25(OH)D3 on C2C12 Myoblast Proliferation, Differentiation, and Myotube Hypertrophy.

PubMed

van der Meijden, K; Bravenboer, N; Dirks, N F; Heijboer, A C; den Heijer, M; de Wit, G M J; Offringa, C; Lips, P; Jaspers, R T

2016-11-01

An adequate vitamin D status is essential to optimize muscle strength. However, whether vitamin D directly reduces muscle fiber atrophy or stimulates muscle fiber hypertrophy remains subject of debate. A mechanism that may affect the role of vitamin D in the regulation of muscle fiber size is the local conversion of 25(OH)D to 1,25(OH)2 D by 1α-hydroxylase. Therefore, we investigated in a murine C2C12 myoblast culture whether both 1,25(OH)2 D3 and 25(OH)D3 affect myoblast proliferation, differentiation, and myotube size and whether these cells are able to metabolize 25(OH)D3 and 1,25(OH)2 D3 . We show that myoblasts not only responded to 1,25(OH)2 D3 , but also to the precursor 25(OH)D3 by increasing their VDR mRNA expression and reducing their proliferation. In differentiating myoblasts and myotubes 1,25(OH)2 D3 as well as 25(OH)D3 stimulated VDR mRNA expression and in myotubes 1,25(OH)2 D3 also stimulated MHC mRNA expression. However, this occurred without notable effects on myotube size. Moreover, no effects on the Akt/mTOR signaling pathway as well as MyoD and myogenin mRNA levels were observed. Interestingly, both myoblasts and myotubes expressed CYP27B1 and CYP24 mRNA which are required for vitamin D3 metabolism. Although 1α-hydroxylase activity could not be shown in myotubes, after treatment with 1,25(OH)2 D3 or 25(OH)D3 myotubes showed strongly elevated CYP24 mRNA levels compared to untreated cells. Moreover, myotubes were able to convert 25(OH)D3 to 24R,25(OH)2 D3 which may play a role in myoblast proliferation and differentiation. These data suggest that skeletal muscle is not only a direct target for vitamin D3 metabolites, but is also able to metabolize 25(OH)D3 and 1,25(OH)2 D3 . J. Cell. Physiol. 231: 2517-2528, 2016. © 2016 The Authors. Journal of Cellular Physiology Published by Wiley Periodicals, Inc. © 2016 The Authors. Journal of Cellular Physiology Published by Wiley Periodicals, Inc.

Applying 3D Full Waveform Inversion in resolving fracture damage zones around a modelled geological disposal facility in granite

NASA Astrophysics Data System (ADS)

Bentham, H. L. M.; Morgan, J. V.; Angus, D. A.

2016-12-01

The UK has a large volume of high level and intermediate level radioactive waste and government policy is to dispose of this waste in a Geological Disposal Facility (GDF). This will be a highly-engineered facility capable of isolating radioactive waste within multiple protective barriers, deep underground, to ensure that no harmful quantities of radioactivity ever reach the surface environment. Although no specific GDF site in the UK has been chosen, granite is one of the candidate host rocks due to its strength, in engineering terms, and because of its low permeability in consideration of groundwater movement. We design time-lapse seismic surveys to characterise geological models of naturally fractured granite with GDF-related tunnel damage zones at a potential disposal depth of 1000 m (the UK GDF might be shallower). Additionally, we use effective medium models to calculate the velocity change when the fracture density is increased in the damage zones, and find a reduction of 60 m/s in P-wave velocity when the fracture density is doubled. Next, we simulate seismic surveys and apply 3D Full Waveform Inversion (FWI) to see how well we can recover the low-velocity damage zones. Furthermore we evaluate the effectiveness of using a survey design consisting of surface and tunnel receivers (a combined array) to resolve the target. After applying FWI we find the velocity anomaly within the damage zone can be resolved to within 2 m/s (3%) and the shape of the damage zone is resolved to 12.5 m (within a single grid cell). Using the combined array we are able to resolve the anomaly strength and shape more completely. When we add further complexity to the model by including tunnel infrastructure, we conclude the combined array is essential in recovering the tunnel damage zone. Our findings show that it is beneficial to use 3D FWI and novel survey designs for characterising subtle variations as may be present in granite, information that could assist in the GDF site selection process and also with GDF design.

Transparent metal model study of the use of a cellular growth front to form aligned monotectic composite materials

NASA Technical Reports Server (NTRS)

Kaukler, William F.

1988-01-01

The purpose of this work was to resolve a scientific controversy in the understanding of how second phase particles become aligned during unidirectional growth of a monotectic alloy. A second aspect was to make the first systematic observations of the solidification behavior of a monotectic alloy during cellular growth in-situ. This research provides the first systematic transparent model study of cellular solidification. An interface stability diagram was developed for the planar to cellular transition of the succinonitrile glycerol (SNG) system. A method was developed utilizing Fourier Transform Infrared Spectroscopy which allows quantitative compositional analysis of directionally solidified SNG along the growth axis. To determine the influence of cellular growth front on alignment for directionally solidified monotectic alloys, the planar and cellular growth morphology was observed in-situ for SNG between 8 and 17 percent glycerol and for a range of over two orders of magnitude G/R.

Electron tomography of whole cultured cells using novel transmission electron imaging technique.

PubMed

Okumura, Taiga; Shoji, Minami; Hisada, Akiko; Ominami, Yusuke; Ito, Sukehiro; Ushiki, Tatsuo; Nakajima, Masato; Ohshima, Takashi

2018-01-01

Since a three-dimensional (3D) cellular ultrastructure is significant for biological functions, it has been investigated using various electron microscopic techniques. Although transmission electron microscopy (TEM)-based techniques are traditionally used, cells must be embedded in resin and sliced into ultrathin sections in sample preparation processes. Block-face observation using a scanning electron microscope (SEM) has also been recently applied to 3D observation of cellular components, but this is a destructive inspection and does not allow re-examination. Therefore, we developed electron tomography using a transmission electron imaging technique called Plate-TEM. With Plate-TEM, the cells cultured directly on a scintillator plate are inserted into a conventional SEM equipped with a Plate-TEM observation system, and their internal structures are observed by detecting scintillation light produced by electrons passing through the cells. This technology has the following four advantages. First, the cells cultured on the plate can be observed at electron-microscopic resolution since they remain on the plate. Second, both surface and internal information can be obtained simultaneously by using electron- and photo-detectors, respectively, because a Plate-TEM detector is installed in an SEM. Third, the cells on the scintillator plate can also be inspected using light microscopy because the plate has transparent features. Finally, correlative observation with other techniques, such as conventional TEM, is possible after Plate-TEM observation because Plate-TEM is a non-destructive analysis technique. We also designed a sample stage to tilt the samples for tomography with Plate-TEM, by which 3D organization of cellular structures can be visualized as a whole cell. In the present study, Mm2T cells were investigated using our tomography system, resulting in 3D visualization of cell organelles such as mitochondria, lipid droplets, and microvilli. Correlative observations with various imaging techniques were also conducted by successive observations with light microscopy, SEM, Plate-TEM, and conventional TEM. Consequently, the Plate-TEM tomography technique encourages understanding of cellular structures at high resolution, which can contribute to cellular biological research. Copyright © 2017 Elsevier Ltd. All rights reserved.

Virtual probing system for medical volume data

NASA Astrophysics Data System (ADS)

Xiao, Yongfei; Fu, Yili; Wang, Shuguo

2007-12-01

Because of the huge computation in 3D medical data visualization, looking into its inner data interactively is always a problem to be resolved. In this paper, we present a novel approach to explore 3D medical dataset in real time by utilizing a 3D widget to manipulate the scanning plane. With the help of the 3D texture property in modern graphics card, a virtual scanning probe is used to explore oblique clipping plane of medical volume data in real time. A 3D model of the medical dataset is also rendered to illustrate the relationship between the scanning-plane image and the other tissues in medical data. It will be a valuable tool in anatomy education and understanding of medical images in the medical research.

Control of cell growth on 3D-printed cell culture platforms for tissue engineering.

PubMed

Tan, Zhikai; Liu, Tong; Zhong, Juchang; Yang, Yikun; Tan, Weihong

2017-12-01

Biocompatible tissue growth has excellent prospects for tissue engineering. These tissues are built over scaffolds, which can influence aspects such as cell adhesion, proliferation rate, morphology, and differentiation. However, the ideal 3D biological structure has not been developed yet. Here, we applied the electro-hydrodynamic jet (E-jet) 3D printing technology using poly-(lactic-co-glycolic acid, PLGA) solution to print varied culture platforms for engineered tissue structures. The effects of different parameters (electrical voltage, plotting speed, and needle sizes) on the outcome were investigated. We compared the biological compatibility of the 3D printed culture platforms with that of random fibers. Finally, we used the 3D-printed PLGA platforms to culture fibroblasts, the main cellular components of loose connective tissue. The results show that the E-jet printed platforms could guide and improve cell growth. These highly aligned fibers were able to support cellular alignment and proliferation. Cell angle was consistent with the direction of the fibers, and cells cultured on these fibers showed a much faster migration, potentially enhancing wound healing performance. Thus, the potential of this technology for 3D biological printing is large. This process can be used to grow biological scaffolds for the engineering of tissues. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 3281-3292, 2017. © 2017 Wiley Periodicals, Inc.

3D culture of murine neural stem cells on decellularized mouse brain sections.

PubMed

De Waele, Jorrit; Reekmans, Kristien; Daans, Jasmijn; Goossens, Herman; Berneman, Zwi; Ponsaerts, Peter

2015-02-01

Transplantation of neural stem cells (NSC) in diseased or injured brain tissue is widely studied as a potential treatment for various neurological pathologies. However, effective cell replacement therapy relies on the intrinsic capacity of cellular grafts to overcome hypoxic and/or immunological barriers after transplantation. In this context, it is hypothesized that structural support for grafted NSC will be of utmost importance. With this study, we present a novel decellularization protocol for 1.5 mm thick mouse brain sections, resulting in the generation of acellular three-dimensional (3D) brain sections. Next, the obtained 3D brain sections were seeded with murine NSC expressing both the eGFP and luciferase reporter proteins (NSC-eGFP/Luc). Using real-time bioluminescence imaging, the survival and growth of seeded NSC-eGFP/Luc cells was longitudinally monitored for 1-7 weeks in culture, indicating the ability of the acellular brain sections to support sustained ex vivo growth of NSC. Next, the organization of a 3D maze-like cellular structure was examined using confocal microscopy. Moreover, under mitogenic stimuli (EGF and hFGF-2), most cells in this 3D culture retained their NSC phenotype. Concluding, we here present a novel protocol for decellularization of mouse brain sections, which subsequently support long-term 3D culture of undifferentiated NSC. Copyright © 2014 Elsevier Ltd. All rights reserved.

FishFace: interactive atlas of zebrafish craniofacial development at cellular resolution

PubMed Central

2013-01-01

Background The vertebrate craniofacial skeleton may exhibit anatomical complexity and diversity, but its genesis and evolution can be understood through careful dissection of developmental programs at cellular resolution. Resources are lacking that include introductory overviews of skeletal anatomy coupled with descriptions of craniofacial development at cellular resolution. In addition to providing analytical guidelines for other studies, such an atlas would suggest cellular mechanisms underlying development. Description We present the Fish Face Atlas, an online, 3D-interactive atlas of craniofacial development in the zebrafish Danio rerio. Alizarin red-stained skulls scanned by fluorescent optical projection tomography and segmented into individual elements provide a resource for understanding the 3D structure of the zebrafish craniofacial skeleton. These data provide the user an anatomical entry point to confocal images of Alizarin red-stained zebrafish with transgenically-labelled pharyngeal arch ectomesenchyme, chondrocytes, and osteoblasts, which illustrate the appearance, morphogenesis, and growth of the mandibular and hyoid cartilages and bones, as viewed in live, anesthetized zebrafish during embryonic and larval development. Confocal image stacks at high magnification during the same stages provide cellular detail and suggest developmental and evolutionary hypotheses. Conclusion The FishFace Atlas is a novel learning tool for understanding craniofacial skeletal development, and can serve as a reference for a variety of studies, including comparative and mutational analyses. PMID:23714426

Changes in cellular degradation activity in young and old worker honeybees (Apis mellifera).

PubMed

Hsu, Chin-Yuan; Chuang, Yu-Lung; Chan, Yu-Pei

2014-02-01

The trophocytes and fat cells of honeybees (Apis mellifera) have been used in cellular senescence studies, but the changes of cellular degradation activity with aging in workers are unknown. In this study, cellular degradation activity was evaluated in the trophocytes and fat cells of young and old workers reared in a field hive. The results showed the following: (1) 20S proteosome activity decreased with aging, whereas its expression increased with aging; (2) the expression of microtubule-associated protein 1 light chain 3-II (LC3-II) and the 70 kD heat shock cognate protein (Hsc70) decreased with aging; (3) the size and number of autophagic vacuoles decreased with aging; (4) p62/SQSTM1 and polyubiquitin aggregate expression decreased with aging; (5) lysosomal efficiency decreased with aging; and (6) molecular target of rapamycin (mTOR) expression increased with aging. These results indicate that young workers have higher levels of cellular degradation activity than old workers and that aging results in a decline in the cellular degradation activity in worker honeybees. Copyright © 2013 Elsevier Inc. All rights reserved.

The KMOS3D Survey: Design, First Results, and the Evolution of Galaxy Kinematics from 0.7 <= z <= 2.7

NASA Astrophysics Data System (ADS)

Wisnioski, E.; Förster Schreiber, N. M.; Wuyts, S.; Wuyts, E.; Bandara, K.; Wilman, D.; Genzel, R.; Bender, R.; Davies, R.; Fossati, M.; Lang, P.; Mendel, J. T.; Beifiori, A.; Brammer, G.; Chan, J.; Fabricius, M.; Fudamoto, Y.; Kulkarni, S.; Kurk, J.; Lutz, D.; Nelson, E. J.; Momcheva, I.; Rosario, D.; Saglia, R.; Seitz, S.; Tacconi, L. J.; van Dokkum, P. G.

2015-02-01

We present the KMOS3D survey, a new integral field survey of over 600 galaxies at 0.7 < z < 2.7 using KMOS at the Very Large Telescope. The KMOS3D survey utilizes synergies with multi-wavelength ground- and space-based surveys to trace the evolution of spatially resolved kinematics and star formation from a homogeneous sample over 5 Gyr of cosmic history. Targets, drawn from a mass-selected parent sample from the 3D-HST survey, cover the star formation-stellar mass (M *) and rest-frame (U - V) - M * planes uniformly. We describe the selection of targets, the observations, and the data reduction. In the first-year of data we detect Hα emission in 191 M * = 3 × 109-7 × 1011 M ⊙ galaxies at z = 0.7-1.1 and z = 1.9-2.7. In the current sample 83% of the resolved galaxies are rotation dominated, determined from a continuous velocity gradient and v rot/σ0 > 1, implying that the star-forming "main sequence" is primarily composed of rotating galaxies at both redshift regimes. When considering additional stricter criteria, the Hα kinematic maps indicate that at least ~70% of the resolved galaxies are disk-like systems. Our high-quality KMOS data confirm the elevated velocity dispersions reported in previous integral field spectroscopy studies at z >~ 0.7. For rotation-dominated disks, the average intrinsic velocity dispersion decreases by a factor of two from 50 km s-1at z ~ 2.3 to 25 km s-1at z ~ 0.9. Combined with existing results spanning z ~ 0-3, we show that disk velocity dispersions follow an evolution that is consistent with the dependence of velocity dispersion on gas fractions predicted by marginally stable disk theory. Based on observations obtained at the Very Large Telescope (VLT) of the European Southern Observatory (ESO), Paranal, Chile (ESO program IDS 092A-0091, 093.A-0079).

CELLULAR AND MOLECULAR MECHANISMS OF ACTION OF LINURON: AN ANTIANDROGENIC HERBICIDE THAT PRODUCES REPRODUCTIVE MALFORMATIONS IN MALE RATS

EPA Science Inventory

Title: CELLULAR AND MOLECULAR MECHANISMS OF ACTION OF LINURON: AN ANTIANDROGENIC HERBICIDE THAT PRODUCES REPRODUCTIVE MALFORMATIONS IN MALE RATS. C Lambright1, J Ostby, K Bobseine, V Wilson, AK Hotchkiss2, PC Mann3 and LE Gray Jr1.

Antiandrogenic chemicals alter sex d...

Matrix-Assisted Three-Dimensional Printing of Cellulose Nanofibers for Paper Microfluidics.

PubMed

Shin, Sungchul; Hyun, Jinho

2017-08-09

A cellulose nanofiber (CNF), one of the most attractive green bioresources, was adopted for construction of microfluidic devices using matrix-assisted three-dimensional (3D) printing. CNF hydrogels can support structures printed using CAD design in a 3D hydrogel environment with the appropriate combination of rheological properties between the CNF hydrogel and ink materials. Amazingly, the structure printed freely in the bulky CNF hydrogels was able to retain its highly resolved 3D features in an ultrathin two-dimensional (2D) paper using a simple drying process. The dimensional change in the CNF hydrogels from 3D to 2D resulted from simple dehydration of the CNFs and provided transparent, stackable paper-based 3D channel devices. As a proof of principle, the rheological properties of the CNF hydrogels, the 3D structure of the ink, the formation of channels by evacuation of the ink, and the highly localized selectivity of the devices are described.

A vacuum-actuated microtissue stretcher for long-term exposure to oscillatory strain within a 3D matrix.

PubMed

Walker, Matthew; Godin, Michel; Pelling, Andrew E

2018-05-28

Although our understanding of cellular behavior in response to extracellular biological and mechanical stimuli has greatly advanced using conventional 2D cell culture methods, these techniques lack physiological relevance. To a cell, the extracellular environment of a 2D plastic petri dish is artificially flat, extremely rigid, static and void of matrix protein. In contrast, we developed the microtissue vacuum-actuated stretcher (MVAS) to probe cellular behavior within a 3D multicellular environment composed of innate matrix protein, and in response to continuous uniaxial stretch. An array format, compatibility with live imaging and high-throughput fabrication techniques make the MVAS highly suited for biomedical research and pharmaceutical discovery. We validated our approach by characterizing the bulk microtissue strain, the microtissue strain field and single cell strain, and by assessing F-actin expression in response to chronic cyclic strain of 10%. The MVAS was shown to be capable of delivering reproducible dynamic bulk strain amplitudes up to 13%. The strain at the single cell level was found to be 10.4% less than the microtissue axial strain due to cellular rotation. Chronic cyclic strain produced a 35% increase in F-actin expression consistent with cytoskeletal reinforcement previously observed in 2D cell culture. The MVAS may further our understanding of the reciprocity shared between cells and their environment, which is critical to meaningful biomedical research and successful therapeutic approaches.

Decrease of reactive oxygen species-related biomarkers in the tissue-mimic 3D spheroid culture of human lung cells exposed to zinc oxide nanoparticles.

PubMed

Kim, Eunjoo; Jeon, Won Bae; Kim, Soonhyun; Lee, Soo-Keun

2014-05-01

Common 2-dimensional (2D) cell cultures do not adequately represent cell-cell and cell-matrix signaling and substantially different diffusion/transport pathways. To obtain tissue-mimic information on nanoparticle toxicity from in vitro cell tests, we used a 3-dimensional (3D) culture of human lung cells (A549) prepared with elastin-like peptides modified with an arginine-glycine-aspartate motif. The 3D cells showed different cellular phenotypes, gene expression profiles, and functionalities compared to the 2D cultured cells. In gene array analysis, 3D cells displayed the induced extracellular matrix (ECM)-related biological functions such as cell-to-cell signaling and interaction, cellular function and maintenance, connective tissue development and function, molecular transport, and tissue morphology. Additionally, the expression of ECM-related molecules, such as laminin, fibronectin, and insulin-like growth factor binding protein 3 (IGFBP3), was simultaneously induced at both mRNA and protein levels. When 0.08-50 microg/ml zinc oxide nanoparticles (ZnO-NPs) were administered to 2D and 3D cells, the cell proliferation was not significantly changed. The level of molecular markers for oxidative stress, such as superoxide dismutase (SOD), Bcl-2, ATP synthase, and Complex IV (cytochrome C oxidase), was significantly reduced in 2D culture when exposed to 10 microg/ml ZnO-NPs, but no significant decrease was detected in 3D culture when exposed to the same concentration of ZnO-NPs. In conclusion, the tissue-mimic phenotype and functionality of 3D cells could be achieved through the elevated expression of ECM components. The 3D cells were expected to help to better predict the nanotoxicity of ZnO-NPs at tissue-level by increased cell-cell and cell-ECM adhesion and signaling. The tissue-mimic morphology would also be useful to simulate the diffusion/transport of the nanoparticles in vitro.

Rapid, topology-based particle tracking for high-resolution measurements of large complex 3D motion fields.

PubMed

Patel, Mohak; Leggett, Susan E; Landauer, Alexander K; Wong, Ian Y; Franck, Christian

2018-04-03

Spatiotemporal tracking of tracer particles or objects of interest can reveal localized behaviors in biological and physical systems. However, existing tracking algorithms are most effective for relatively low numbers of particles that undergo displacements smaller than their typical interparticle separation distance. Here, we demonstrate a single particle tracking algorithm to reconstruct large complex motion fields with large particle numbers, orders of magnitude larger than previously tractably resolvable, thus opening the door for attaining very high Nyquist spatial frequency motion recovery in the images. Our key innovations are feature vectors that encode nearest neighbor positions, a rigorous outlier removal scheme, and an iterative deformation warping scheme. We test this technique for its accuracy and computational efficacy using synthetically and experimentally generated 3D particle images, including non-affine deformation fields in soft materials, complex fluid flows, and cell-generated deformations. We augment this algorithm with additional particle information (e.g., color, size, or shape) to further enhance tracking accuracy for high gradient and large displacement fields. These applications demonstrate that this versatile technique can rapidly track unprecedented numbers of particles to resolve large and complex motion fields in 2D and 3D images, particularly when spatial correlations exist.

Magnetic nanoparticles to recover cellular organelles and study the time resolved nanoparticle-cell interactome throughout uptake.

PubMed

Bertoli, Filippo; Davies, Gemma-Louise; Monopoli, Marco P; Moloney, Micheal; Gun'ko, Yurii K; Salvati, Anna; Dawson, Kenneth A

2014-08-27

Nanoparticles in contact with cells and living organisms generate quite novel interactions at the interface between the nanoparticle surface and the surrounding biological environment. However, a detailed time resolved molecular level description of the evolving interactions as nanoparticles are internalized and trafficked within the cellular environment is still missing and will certainly be required for the emerging arena of nanoparticle-cell interactions to mature. In this paper promising methodologies to map out the time resolved nanoparticle-cell interactome for nanoparticle uptake are discussed. Thus silica coated magnetite nanoparticles are presented to cells and their magnetic properties used to isolate, in a time resolved manner, the organelles containing the nanoparticles. Characterization of the recovered fractions shows that different cell compartments are isolated at different times, in agreement with imaging results on nanoparticle intracellular location. Subsequently the internalized nanoparticles can be further isolated from the recovered organelles, allowing the study of the most tightly nanoparticle-bound biomolecules, analogous to the 'hard corona' that so far has mostly been characterized in extracellular environments. Preliminary data on the recovered nanoparticles suggest that significant portion of the original corona (derived from the serum in which particles are presented to the cells) is preserved as nanoparticles are trafficked through the cells. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Nondestructive nanostraw intracellular sampling for longitudinal cell monitoring

PubMed Central

Cao, Yuhong; Chen, Haodong; Birey, Fikri; Leal-Ortiz, Sergio A.; Han, Crystal M.; Santiago, Juan G.; Paşca, Sergiu P.; Wu, Joseph C.; Melosh, Nicholas A.

2017-01-01

Here, we report a method for time-resolved, longitudinal extraction and quantitative measurement of intracellular proteins and mRNA from a variety of cell types. Cytosolic contents were repeatedly sampled from the same cell or population of cells for more than 5 d through a cell-culture substrate, incorporating hollow 150-nm-diameter nanostraws (NS) within a defined sampling region. Once extracted, the cellular contents were analyzed with conventional methods, including fluorescence, enzymatic assays (ELISA), and quantitative real-time PCR. This process was nondestructive with >95% cell viability after sampling, enabling long-term analysis. It is important to note that the measured quantities from the cell extract were found to constitute a statistically significant representation of the actual contents within the cells. Of 48 mRNA sequences analyzed from a population of cardiomyocytes derived from human induced pluripotent stem cells (hiPSC-CMs), 41 were accurately quantified. The NS platform samples from a select subpopulation of cells within a larger culture, allowing native cell-to-cell contact and communication even during vigorous activity such as cardiomyocyte beating. This platform was applied both to cell lines and to primary cells, including CHO cells, hiPSC-CMs, and human astrocytes derived in 3D cortical spheroids. By tracking the same cell or group of cells over time, this method offers an avenue to understand dynamic cell behavior, including processes such as induced pluripotency and differentiation. PMID:28223521

Mechanism of Facilitated Diffusion during a DNA Search in Crowded Environments.

PubMed

Krepel, Dana; Gomez, David; Klumpp, Stefan; Levy, Yaakov

2016-11-03

The key feature explaining the rapid recognition of a DNA target site by its protein lies in the combination of one- and three-dimensional (1D and 3D) diffusion, which allows efficient scanning of the many alternative sites. This facilitated diffusion mechanism is expected to be affected by cellular conditions, particularly crowding, given that up to 40% of the total cellular volume may by occupied by macromolecules. Using coarse-grained molecular dynamics and Monte Carlo simulations, we show that the crowding particles can enhance facilitated diffusion and accelerate search kinetics. This effect originates from a trade-off between 3D and 1D diffusion. The 3D diffusion coefficient is lower under crowded conditions, but it has little influence because the excluded volume effect of molecular crowding restricts its use. Largely prevented from using 3D diffusion, the searching protein dramatically increases its use of the hopping search mode, which results in a higher linear diffusion coefficient. The coefficient of linear diffusion also increases under crowded conditions as a result of increased collisions between the crowding particles and the searching protein. Overall, less 3D diffusion coupled with an increase in the use of the hopping and speed of 1D diffusion results in faster search kinetics under crowded conditions. Our study shows that the search kinetics and mechanism are modulated not only by the crowding occupancy but also by the properties of the crowding particles and the salt concentration.

Biodynamic imaging for phenotypic profiling of three-dimensional tissue culture

PubMed Central

Sun, Hao; Merrill, Daniel; An, Ran; Turek, John; Matei, Daniela; Nolte, David D.

2017-01-01

Abstract. Three-dimensional (3-D) tissue culture represents a more biologically relevant environment for testing new drugs compared to conventional two-dimensional cancer cell culture models. Biodynamic imaging is a high-content 3-D optical imaging technology based on low-coherence interferometry and digital holography that uses dynamic speckle as high-content image contrast to probe deep inside 3-D tissue. Speckle contrast is shown to be a scaling function of the acquisition time relative to the persistence time of intracellular transport and hence provides a measure of cellular activity. Cellular responses of 3-D multicellular spheroids to paclitaxel are compared among three different growth techniques: rotating bioreactor (BR), hanging-drop (HD), and nonadherent (U-bottom, UB) plate spheroids, compared with ex vivo living tissues. HD spheroids have the most homogeneous tissue, whereas BR spheroids display large sample-to-sample variability as well as spatial heterogeneity. The responses of BR-grown tumor spheroids to paclitaxel are more similar to those of ex vivo biopsies than the responses of spheroids grown using HD or plate methods. The rate of mitosis inhibition by application of taxol is measured through tissue dynamics spectroscopic imaging, demonstrating the ability to monitor antimitotic chemotherapy. These results illustrate the potential use of low-coherence digital holography for 3-D pharmaceutical screening applications. PMID:28301634

Biodynamic imaging for phenotypic profiling of three-dimensional tissue culture

NASA Astrophysics Data System (ADS)

Sun, Hao; Merrill, Daniel; An, Ran; Turek, John; Matei, Daniela; Nolte, David D.

2017-01-01

Three-dimensional (3-D) tissue culture represents a more biologically relevant environment for testing new drugs compared to conventional two-dimensional cancer cell culture models. Biodynamic imaging is a high-content 3-D optical imaging technology based on low-coherence interferometry and digital holography that uses dynamic speckle as high-content image contrast to probe deep inside 3-D tissue. Speckle contrast is shown to be a scaling function of the acquisition time relative to the persistence time of intracellular transport and hence provides a measure of cellular activity. Cellular responses of 3-D multicellular spheroids to paclitaxel are compared among three different growth techniques: rotating bioreactor (BR), hanging-drop (HD), and nonadherent (U-bottom, UB) plate spheroids, compared with ex vivo living tissues. HD spheroids have the most homogeneous tissue, whereas BR spheroids display large sample-to-sample variability as well as spatial heterogeneity. The responses of BR-grown tumor spheroids to paclitaxel are more similar to those of ex vivo biopsies than the responses of spheroids grown using HD or plate methods. The rate of mitosis inhibition by application of taxol is measured through tissue dynamics spectroscopic imaging, demonstrating the ability to monitor antimitotic chemotherapy. These results illustrate the potential use of low-coherence digital holography for 3-D pharmaceutical screening applications.

Chemistry of wood in 3D: new infrared imaging

Treesearch

Barbara L. Illman; Julia Sedlmair; Miriam Unger; Casey Crooks; Marli Oliveira; Carol Hirschmugl

2015-01-01

Chemical detection, mapping and imaging in three dimensions will help refine our understanding of wood properties and durability. We describe here a pioneering infrared method to create visual 3D images of the chemicals in wood, providing for the first time, spatial and architectural information at the cellular level without liquid extraction or prior fixation....

Emerging biomedical applications of time-resolved fluorescence spectroscopy

NASA Astrophysics Data System (ADS)

Lakowicz, Joseph R.; Szmacinski, Henryk; Koen, Peter A.

1994-07-01

Time-resolved fluorescence spectroscopy is presently regarded as a research tool in biochemistry, biophysics, and chemical physics. Advances in laser technology, the development of long-wavelength probes, and the use of lifetime-based methods are resulting in the rapid migration of time-resolved fluorescence to the clinical chemistry lab, to the patient's bedside, to flow cytometers, to the doctor's office, and even to home health care. Additionally, time-resolved imaging is now a reality in fluorescence microscopy, and will provide chemical imaging of a variety of intracellular analytes and/or cellular phenomena. In this overview paper we attempt to describe some of the opportunities available using chemical sensing based on fluorescence lifetimes, and to predict those applications of lifetime-based sensing which are most likely in the near future.

3D Printed Shock Mitigating Structures

NASA Astrophysics Data System (ADS)

Schrand, Amanda; Elston, Edwin; Dennis, Mitzi; Metroke, Tammy; Chen, Chenggang; Patton, Steven; Ganguli, Sabyasachi; Roy, Ajit

Here we explore the durability, and shock mitigating potential, of solid and cellular 3D printed polymers and conductive inks under high strain rate, compressive shock wave and high g acceleration conditions. Our initial designs include a simple circuit with 4 resistors embedded into circular discs and a complex cylindrical gyroid shape. A novel ink consisting of silver-coated carbon black nanoparticles in a thermoplastic polyurethane was used as the trace material. One version of the disc structural design has the advantage of allowing disassembly after testing for direct failure analysis. After increasing impacts, printed and traditionally potted circuits were examined for functionality. Additionally, in the open disc design, trace cracking and delamination of resistors were able to be observed. In a parallel study, we examined the shock mitigating behavior of 3D printed cellular gyroid structures on a Split Hopkinson Pressure Bar (SHPB). We explored alterations to the classic SHPB setup for testing the low impedance, cellular samples to most accurately reflect the stress state inside the sample (strain rates from 700 to 1750 s-1). We discovered that the gyroid can effectively absorb the impact of the test resulting in crushing the structure. Future studies aim to tailor the unit cell dimensions for certain frequencies, increase print accuracy and optimize material compositions for conductivity and adhesion to manufacture more durable devices.

Effects of Matrix Alignment and Mechanical Constraints on Cellular Behavior in 3D Engineered Microtissues

NASA Astrophysics Data System (ADS)

Bose, Prasenjit; Eyckmans, Jeroen; Chen, Christopher; Reich, Daniel

The adhesion of cells to the extracellular matrix (ECM) plays a crucial role in a variety of cellular functions. The main building blocks of the ECM are 3D networks of fibrous proteins whose structure and alignments varies with tissue type. However, the impact of ECM alignment on cellular behaviors such as cell adhesion, spreading, extension and mechanics remains poorly understood. We present results on the development of a microtissue-based system that enables control of the structure, orientation, and degree of fibrillar alignment in 3D fibroblast-populated collagen gels. The tissues self-assemble from cell-laden collagen gels placed in micro-fabricated wells containing sets of elastic pillars. The contractile action of the cells leads to controlled alignment of the fibrous collagen, depending on the number and location of the pillars in each well. The pillars are elastic, and are utilized to measure the contractile forces of the microtissues, and by incorporating magnetic material in selected pillars, time-varying forces can be applied to the tissues for dynamic stimulation and measurement of mechanical properties. Results on the effects of varying pillar shape, spacing, location, and stiffness on microtissue organization and contractility will be presented. This work is supported by NSF CMMI-1463011.

Effects of SiO2 and ZnO doping on mechanical and biological properties of 3D printed TCP scaffolds

PubMed Central

Fielding, Gary A.; Bandyopadhyay, Amit; Bose, Susmita

2011-01-01

Objectives To evaluate the effects of SiO2 (0.5 wt %) and ZnO (0.25 wt %) dopants on the mechanical and biological properties of tricalcium phosphate (TCP) scaffolds with three dimensionally (3D) interconnected pores. Methods Scaffolds were created with a commercial 3D printer. Post sintering phase analysis was determined by x-ray diffraction. Surface morphology of the scaffolds was examined by field emission electron microscopy. Mechanical strength was evaluated with a screw driven universal testing machine. MTT assay was used for cellular proliferation characteristics and cellular morphology was examined by field emission electron microscopy. Results Addition of dopants into TCP increased the average density of pure TCP from 90.8 ± 0.8% to 94.1 ± 1.6% and retarded the β to α phase transformation at high sintering temperatures, which resulted in up to 2.5 fold increase in compressive strength. In vitro cell-materials interaction studies, carried out using hFOB cells, confirmed that the addition of SiO2 and ZnO to the scaffolds facilitates faster cell proliferation when compared to pure TCP scaffolds. Significance Addition of SiO2 and ZnO dopants to the TCP scaffolds showed increased mechanical strength as well as increased cellular proliferation. PMID:22047943

The cellular lesion of humoral rejection: predominant recruitment of monocytes to peritubular and glomerular capillaries.

PubMed

Fahim, T; Böhmig, G A; Exner, M; Huttary, N; Kerschner, H; Kandutsch, S; Kerjaschki, D; Bramböck, A; Nagy-Bojarszky, K; Regele, H

2007-02-01

Accumulation of inflammatory cells within capillaries is a common morphologic feature of humoral renal allograft rejection and is most easily appreciated if it occurs in glomeruli. The aim of our study was to determine the amount and composition of immune cells within glomeruli and peritubular capillaries (PTC) in cellular and humoral allograft rejection. Immunofluorescent double-labeling for CD31 and CD3 or CD68 was used for phenotyping and enumerating immune cells within glomeruli and PTC. The major findings are: (1) accumulation of immune cells in PTC is far more common than it would be anticipated based on the assessment by conventional histology; (2) it is not the absolute number of immune cells accumulating within capillaries, but rather the composition of the intracapillary cell population that distinguishes humoral rejection from cellular rejection and (3) in C4d positive biopsies a predominantly monocytic cell population accumulates not only within glomeruli but also within PTC. The median value of monocyte/T-cell ratio within PTC was 2.3 in C4d positive biopsies but only 1 (p = 0.0008) in C4d negative biopsies. Given their prominent presence within capillaries and their extensive biological versatility monocytes might contribute to the capillary damage observed in acute and chronic allograft rejection.

Bringing macromolecular machinery to life using 3D animation.

PubMed

Iwasa, Janet H

2015-04-01

Over the past decade, there has been a rapid rise in the use of three-dimensional (3D) animation to depict molecular and cellular processes. Much of the growth in molecular animation has been in the educational arena, but increasingly, 3D animation software is finding its way into research laboratories. In this review, I will discuss a number of ways in which 3d animation software can play a valuable role in visualizing and communicating macromolecular structures and dynamics. I will also consider the challenges of using animation tools within the research sphere. Copyright © 2015. Published by Elsevier Ltd.

Electronic Structure at Oxide Interfaces

DTIC Science & Technology

2014-06-01

of materials with desired correlated electron properties such as ferromagnetism with a high Curie temperature, high transition temperature...approximation and therefore the canonical Mott picture is unable to account for the insulating behavior of these materials . We resolve this apparent...the two materials . LaTiO3 shows insulating behavior with a small excitation gap set by Ti d-d transitions and a wide energy separation between Ti d

Self-Expansion Construction of Ultralight Carbon Nanotube Aerogels with a 3D and Hierarchical Cellular Structure.

PubMed

Luo, Yufeng; Luo, Shu; Wu, Hengcai; Li, Mengya; Wang, Ke; Yan, Lingjia; Jiang, Kaili; Li, Qunqing; Fan, Shoushan; Wang, Jiaping

2017-07-01

A novel and simple strategy is developed to construct ultralight and 3D pure carbon nanotube (CNT) aerogels by the spontaneous expansion of superaligned CNT films soaked in a piranha (mixed H 2 SO 4 and H 2 O 2 ) solution, followed by cryodesiccation. The macroscopic CNT aerogels have an extremely low apparent density (0.12 mg cm -3 ), ultrahigh porosity (99.95%), high specific surface area (298 m 2 g -1 ), and a hierarchical cellular structure with giant and ultrathin CNT sheets as cell walls. The pure CNT aerogels show high adsorption abilities for various kinds of solvents, and have great potential in widespread applications such as energy storage, catalysis, and bioengineering. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Mechanism of the reaction, CH4+O(1D2)→CH3+OH, studied by ultrafast and state-resolved photolysis/probe spectroscopy of the CH4ṡO3 van der Waals complex

NASA Astrophysics Data System (ADS)

Miller, C. Cameron; van Zee, Roger D.; Stephenson, John C.

2001-01-01

The mechanism of the reaction CH4+O(1D2)→CH3+OH was investigated by ultrafast, time-resolved and state-resolved experiments. In the ultrafast experiments, short ultraviolet pulses photolyzed ozone in the CH4ṡO3 van der Waals complex to produce O(1D2). The ensuing reaction with CH4 was monitored by measuring the appearance rate of OH(v=0,1;J,Ω,Λ) by laser-induced fluorescence, through the OH A←X transition, using short probe pulses. These spectrally broad pulses, centered between 307 and 316 nm, probe many different OH rovibrational states simultaneously. At each probe wavelength, both a fast and a slow rise time were evident in the fluorescence signal, and the ratio of the fast-to-slow signal varied with probe wavelength. The distribution of OH(v,J,Ω,Λ) states, Pobs(v,J,Ω,Λ), was determined by laser-induced fluorescence using a high-resolution, tunable dye laser. The Pobs(v,J,Ω,Λ) data and the time-resolved data were analyzed under the assumption that different formation times represent different reaction mechanisms and that each mechanism produces a characteristic rovibrational distribution. The state-resolved and the time-resolved data can be fit independently using a two-mechanism model: Pobs(v,J,Ω,Λ) can be decomposed into two components, and the appearance of OH can be fit by two exponential rise times. However, these independent analyses are not mutually consistent. The time-resolved and state-resolved data can be consistently fit using a three-mechanism model. The OH appearance signals, at all probe wavelengths, were fit with times τfast≈0.2 ps, τinter≈0.5 ps and τslow≈5.4 ps. The slowest of these three is the rate for dissociation of a vibrationally excited methanol intermediate (CH3OH*) predicted by statistical theory after complete intramolecular energy redistribution following insertion of O(1D2) into CH4. The Pobs(v,J,Ω,Λ) was decomposed into three components, each with a linear surprisal, under the assumption that the mechanism producing OH at a statistical rate would be characterized by a statistical prior. Dissociation of a CH4O* intermediate before complete energy randomization was identified as producing OH at the intermediate rate and was associated with a population distribution with more rovibrational energy than the slow mechanism. The third mechanism produces OH promptly with a cold rovibrational distribution, indicative of a collinear abstraction mechanism. After these identifications were made, it was possible to predict the fraction of signal associated with each mechanism at different probe wavelengths in the ultrafast experiment, and the predictions proved consistent with measured appearance signals. This model also reconciles data from a variety of previous experiments. While this model is the simplest that is consistent with the data, it is not definitive for several reasons. First, the appearance signals measured in these experiments probe simultaneously many OH(v,J,Ω,Λ) states, which would tend to obfuscate differences in the appearance rate of specific rovibrational states. Second, only about half of the OH(v,J,Ω,Λ) states populated by this reaction could be probed by laser-induced fluorescence through the OH A←X band with our apparatus. Third, the cluster environment might influence the dynamics compared to the free bimolecular reaction.

Multifunctional Cellular Materials Based on 2D Nanomaterials: Prospects and Challenges.

PubMed

Qiu, Ling; He, Zijun; Li, Dan

2018-01-01

Recent advances in emerging 2D nanomaterial-based cellular materials (2D-CMs) open up new opportunities for the development of next generation cellular solids with exceptional properties. Herein, an overview of the current research status of 2D-CMs is provided and their future opportunities are highlighted. First, the unique features of 2D nanomaterials are introduced to illustrate why these nanoscale building blocks are promising for the development of novel cellular materials and what the new features of 2D nanoscale building blocks can offer when compared to their 0D and 1D counterparts. An in-depth discussion on the structure-property relationships of 2D-CMs is then provided, and the remarkable functions that can be achieved by engineering their cellular architecture are highlighted. Additionally, the use of 2D-CMs to tackle key challenges in different practical applications is demonstrated. In conclusion, a personal perspective on the challenges and future research directions of 2D-CMs is given. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Single-frequency 3D synthetic aperture imaging with dynamic metasurface antennas.

PubMed

Boyarsky, Michael; Sleasman, Timothy; Pulido-Mancera, Laura; Diebold, Aaron V; Imani, Mohammadreza F; Smith, David R

2018-05-20

Through aperture synthesis, an electrically small antenna can be used to form a high-resolution imaging system capable of reconstructing three-dimensional (3D) scenes. However, the large spectral bandwidth typically required in synthetic aperture radar systems to resolve objects in range often requires costly and complex RF components. We present here an alternative approach based on a hybrid imaging system that combines a dynamically reconfigurable aperture with synthetic aperture techniques, demonstrating the capability to resolve objects in three dimensions (3D), with measurements taken at a single frequency. At the core of our imaging system are two metasurface apertures, both of which consist of a linear array of metamaterial irises that couple to a common waveguide feed. Each metamaterial iris has integrated within it a diode that can be biased so as to switch the element on (radiating) or off (non-radiating), such that the metasurface antenna can produce distinct radiation profiles corresponding to different on/off patterns of the metamaterial element array. The electrically large size of the metasurface apertures enables resolution in range and one cross-range dimension, while aperture synthesis provides resolution in the other cross-range dimension. The demonstrated imaging capabilities of this system represent a step forward in the development of low-cost, high-performance 3D microwave imaging systems.

Three-dimensional locations of gold-labeled proteins in a whole mount eukaryotic cell obtained with 3nm precision using aberration-corrected scanning transmission electron microscopy.

PubMed

Dukes, Madeline J; Ramachandra, Ranjan; Baudoin, Jean-Pierre; Gray Jerome, W; de Jonge, Niels

2011-06-01

Three-dimensional (3D) maps of proteins within the context of whole cells are important for investigating cellular function. However, 3D reconstructions of whole cells are challenging to obtain using conventional transmission electron microscopy (TEM). We describe a methodology to determine the 3D locations of proteins labeled with gold nanoparticles on whole eukaryotic cells. The epidermal growth factor receptors on COS7 cells were labeled with gold nanoparticles, and critical-point dried whole-mount cell samples were prepared. 3D focal series were obtained with aberration-corrected scanning transmission electron microscopy (STEM), without tilting the specimen. The axial resolution was improved with deconvolution. The vertical locations of the nanoparticles in a whole-mount cell were determined with a precision of 3nm. From the analysis of the variation of the axial positions of the labels we concluded that the cellular surface was ruffled. To achieve sufficient stability of the sample under electron beam irradiation during the recording of the focal series, the sample was carbon coated. A quantitative method was developed to analyze the stability of the ultrastructure after electron beam irradiation using TEM. The results of this study demonstrate the feasibility of using aberration-corrected STEM to study the 3D nanoparticle distribution in whole cells. Copyright © 2011 Elsevier Inc. All rights reserved.

Three-dimensional locations of gold-labeled proteins in a whole mount eukaryotic cell obtained with 3 nm precision using aberration-corrected scanning transmission electron microscopy

PubMed Central

Dukes, Madeline J.; Ramachandra, Ranjan; Baudoin, Jean-Pierre; Jerome, W. Gray; de Jonge, Niels

2011-01-01

Three-dimensional (3D) maps of proteins within the context of whole cells are important for investigating cellular function. However, 3D reconstructions of whole cells are challenging to obtain using conventional transmission electron microscopy (TEM). We describe a methodology to determine the 3D locations of proteins labeled with gold nanoparticles on whole eukaryotic cells. The epidermal growth factor receptors on COS7 cells were labeled with gold nanoparticles, and critical-point dried whole-mount cell samples were prepared. 3D focal series were obtained with aberration-corrected scanning transmission electron microscopy (STEM), without tilting the specimen. The axial resolution was improved with deconvolution. The vertical locations of the nanoparticles in a whole-mount cell were determined with a precision of 3 nm. From the analysis of the variation of the axial positions of the labels we concluded that the cellular surface was ruffled. To achieve sufficient stability of the sample under the electron beam irradiation during the recording of the focal series, the sample was carbon coated. A quantitative method was developed to analyze the stability of the ultrastructure after electron beam irradiation using TEM. The results of this study demonstrate the feasibility of using aberration-corrected STEM to study the 3D nanoparticle distribution in whole cells. PMID:21440635

The effects of an RGD-PAMAM dendrimer conjugate in 3D spheroid culture on cell proliferation, expression and aggregation.

PubMed

Jiang, Li-Yang; Lv, Bing; Luo, Ying

2013-04-01

By presenting biomolecular ligands on the surface in high density, ligand-decorated dendrimers are capable of binding to membrane receptors and cells with specificity and avidity. Despite the various uses, fundamental investigations on ligand-dendrimer conjugates have mainly focused on their binding behavior with cells, whereas their potential bioactivity and applications in multicellular systems, especially in three-dimensional (3D) culture systems, remains untapped. In this study, a typical adhesive peptide ligand - RGD - was modified to generation 4 polyamidoamine (PAMAM), and the bioactivity of suspended RGD-PAMAM conjugates was investigated on cells cultured as multicellular spheroids. Our results demonstrate that the RGD-PAMAM conjugates, after being incorporated into the 3D spheroids, were able to promote cellular proliferation and aggregation, and affect the mRNA expression of extracellular factors by NIH 3T3 cells. These bioactive functions were multivalency-dependent, as none of similar effects was observed for monovalent RGD ligand. Our study suggests that multivalent ligand-dendrimer conjugates may act as a unique type of artificial factors to mediate the cellular microenvironment in 3D culture, a property attributable to the spatial organization of the ligands and possible "cell-gluing" function of multivalent conjugates. This new finding opens the door for further exploring multivalent ligand-dendrimer conjugates for applications in 3D cell culture and tissue engineering. Copyright © 2013 Elsevier Ltd. All rights reserved.

Two Distinct Families of Protein Kinases Are Required for Plant Growth under High External Mg2+ Concentrations in Arabidopsis1

PubMed Central

Mogami, Junro; Fujita, Yasunari; Yoshida, Takuya; Tsukiori, Yoshifumi; Nakagami, Hirofumi; Nomura, Yuko; Fujiwara, Toru; Nishida, Sho; Yanagisawa, Shuichi; Ishida, Tetsuya; Takahashi, Fuminori; Morimoto, Kyoko; Kidokoro, Satoshi; Mizoi, Junya; Shinozaki, Kazuo

2015-01-01

Protein phosphorylation events play key roles in maintaining cellular ion homeostasis in higher plants, and the regulatory roles of these events in Na+ and K+ transport have been studied extensively. However, the regulatory mechanisms governing Mg2+ transport and homeostasis in higher plants remain poorly understood, despite the vital roles of Mg2+ in cellular function. A member of subclass III sucrose nonfermenting-1-related protein kinase2 (SnRK2), SRK2D/SnRK2.2, functions as a key positive regulator of abscisic acid (ABA)-mediated signaling in response to water deficit stresses in Arabidopsis (Arabidopsis thaliana). Here, we used immunoprecipitation coupled with liquid chromatography-tandem mass spectrometry analyses to identify Calcineurin B-like-interacting protein kinase26 (CIPK26) as a novel protein that physically interacts with SRK2D. In addition to CIPK26, three additional CIPKs (CIPK3, CIPK9, and CIPK23) can physically interact with SRK2D in planta. The srk2d/e/i triple mutant lacking all three members of subclass III SnRK2 and the cipk26/3/9/23 quadruple mutant lacking CIPK26, CIPK3, CIPK9, and CIPK23 showed reduced shoot growth under high external Mg2+ concentrations. Similarly, several ABA biosynthesis-deficient mutants, including aba2-1, were susceptible to high external Mg2+ concentrations. Taken together, our findings provided genetic evidence that SRK2D/E/I and CIPK26/3/9/23 are required for plant growth under high external Mg2+ concentrations in Arabidopsis. Furthermore, we showed that ABA, a key molecule in water deficit stress signaling, also serves as a signaling molecule in plant growth under high external Mg2+ concentrations. These results suggested that SRK2D/E/I- and CIPK26/3/9/23-mediated phosphorylation signaling pathways maintain cellular Mg2+ homeostasis. PMID:25614064

Refraction statics and seismic imaging: 2-D versus 3-D solutions in the Western Desert of Egypt

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

El-Emam, A.; Nessim, M.

1994-12-31

Careful review of old geophysical and geological data from the Western Desert of Egypt led to the decision of shooting a 3-D seismic survey targeted to solve some of the encountered geophysical problems such as difficulty of tracing the very thin pay zone, identifying the stratigraphic plays and the main two problems of the seismic method in the Western Desert which are statics and poor imaging. In a case history form illustrated by examples, the result of the 3-D solutions will be shown. Furthermore, an analytical approach will be undertaken to clarify and highlight the sources of those geophysical problemsmore » and how the 3-D solution helped in resolving them.« less

Tempo-spatially resolved cellular dynamics of human immunodeficiency virus transacting activator of transcription (Tat) peptide-modified nanocargos in living cells

NASA Astrophysics Data System (ADS)

Wei, Lin; Yang, Qiaoyu; Xiao, Lehui

2014-08-01

Understanding the cellular uptake mechanism and intracellular fate of nanocarriers in living cells is of great importance for the rational design of efficient drug delivery cargos as well as the development of robust biomedical diagnostic probes. In present study, with a dual wavelength view darkfield microscope (DWVD), the tempo-spatially resolved dynamics of Tat peptide-functionalized gold nanoparticles (TGNPs, with size similar to viruses) in living HeLa cells were extensively explored. It was found that energy-dependent endocytosis (both clathrin- and caveolae-mediated processes were involved) was the prevailing pathway for the cellular uptake of TGNPs. The time-correlated dynamic spatial distribution information revealed that TGNPs could not actively target the cell nuclei, which is contrary to previous observations based on fixed cell results. More importantly, the inheritance of TGNPs to the daughter cells through mitosis was found to be the major route to metabolize TGNPs by HeLa cells. These understandings on the cellular uptake mechanism and intracellular fate of nanocargos in living cells would provide deep insight on how to improve and controllably manipulate their translocation efficiency for targeted drug delivery.Understanding the cellular uptake mechanism and intracellular fate of nanocarriers in living cells is of great importance for the rational design of efficient drug delivery cargos as well as the development of robust biomedical diagnostic probes. In present study, with a dual wavelength view darkfield microscope (DWVD), the tempo-spatially resolved dynamics of Tat peptide-functionalized gold nanoparticles (TGNPs, with size similar to viruses) in living HeLa cells were extensively explored. It was found that energy-dependent endocytosis (both clathrin- and caveolae-mediated processes were involved) was the prevailing pathway for the cellular uptake of TGNPs. The time-correlated dynamic spatial distribution information revealed that TGNPs could not actively target the cell nuclei, which is contrary to previous observations based on fixed cell results. More importantly, the inheritance of TGNPs to the daughter cells through mitosis was found to be the major route to metabolize TGNPs by HeLa cells. These understandings on the cellular uptake mechanism and intracellular fate of nanocargos in living cells would provide deep insight on how to improve and controllably manipulate their translocation efficiency for targeted drug delivery. Electronic supplementary information (ESI) available: Experimental section and additional supporting results as noted in the text. See DOI: 10.1039/c4nr02732a

Ambiguity resolving based on cosine property of phase differences for 3D source localization with uniform circular array

NASA Astrophysics Data System (ADS)

Chen, Xin; Wang, Shuhong; Liu, Zhen; Wei, Xizhang

2017-07-01

Localization of a source whose half-wavelength is smaller than the array aperture would suffer from serious phase ambiguity problem, which also appears in recently proposed phase-based algorithms. In this paper, by using the centro-symmetry of fixed uniform circular array (UCA) with even number of sensors, the source's angles and range can be decoupled and a novel ambiguity resolving approach is addressed for phase-based algorithms of source's 3-D localization (azimuth angle, elevation angle, and range). In the proposed method, by using the cosine property of unambiguous phase differences, ambiguity searching and actual-value matching are first employed to obtain actual phase differences and corresponding source's angles. Then, the unambiguous angles are utilized to estimate the source's range based on a one dimension multiple signal classification (1-D MUSIC) estimator. Finally, simulation experiments investigate the influence of step size in search and SNR on performance of ambiguity resolution and demonstrate the satisfactory estimation performance of the proposed method.

The partition dimension of subdivision of a graph

NASA Astrophysics Data System (ADS)

Amrullah, Baskoro, Edy Tri; Uttunggadewa, Saladin; Simanjuntak, Rinovia

2016-02-01

Let G = (V,E) be a connected graph, u,v ∈ V (G), e = uv ∈ E(G) and k be a positive integer. A k-subdivision of an edge e is a replacement of e = uv with a path u, x1, x2, x ..., xk, v. A graph G with a k-subdivided edge is denoted with S(G(e; k)). Let p be a positive integer and Π = {L1, L2, L3, …, Lp} be a p-partition of V (G). The representation of a vertex v with respect to Π, r(v|Π), is the vector (d(v, L1), d(v, L2), d(v, L3),…, d(v, Lp)) where d(v, Li) for i ∈ [1, p] is the minimum distance between v and the vertices of Li. The partition Π is called a resolving partition of G if r(w|Π) ≠ r(v|Π) for all w ≠ v ∈ V (G). The partition dimension, pd(G), of G is the smallest integer p such that G has a resolving p-partition. In this paper, we present sharp upper and lower bounds of the partition dimension of S(G(e; k)) for any graph G.

Inhibition of sirtuins 1 and 2 impairs cell survival and migration and modulates the expression of P-glycoprotein and MRP3 in hepatocellular carcinoma cell lines.

PubMed

Ceballos, María Paula; Decándido, Giulia; Quiroga, Ariel Darío; Comanzo, Carla Gabriela; Livore, Verónica Inés; Lorenzetti, Florencia; Lambertucci, Flavia; Chazarreta-Cifre, Lorena; Banchio, Claudia; Alvarez, María de Luján; Mottino, Aldo Domingo; Carrillo, María Cristina

2018-06-01

Sirtuins (SIRTs) 1 and 2 deacetylases are overexpressed in hepatocellular carcinoma (HCC) and are associated with tumoral progression and multidrug resistance (MDR). In this study we analyzed whether SIRTs 1 and 2 activities blockage was able to affect cellular survival and migration and to modulate p53 and FoxO1 acetylation in HepG2 and Huh7 cells. Moreover, we analyzed ABC transporters P-glycoprotein (P-gp) and multidrug resistance-associated protein 3 (MRP3) expression. We used cambinol and EX-527 as SIRTs inhibitors. Both drugs reduced cellular viability, number of colonies and cellular migration and augmented apoptosis. In 3D cultures, SIRTs inhibitors diminished spheroid growth and viability. 3D culture was less sensitive to drugs than 2D culture. The levels of acetylated p53 and FoxO1 increased after treatments. Drugs induced a decrease in ABC transporters mRNA and protein levels in HepG2 cells; however, only EX-527 was able to reduce MRP3 mRNA and protein levels in Huh7 cells. This is the first work demonstrating the regulation of MRP3 by SIRTs. In conclusion, both drugs decreased HCC cells survival and migration, suggesting SIRTs 1 and 2 activities blockage could be beneficial during HCC therapy. Downregulation of the expression of P-gp and MRP3 supports the potential application of SIRTs 1 and 2 inhibitions in combination with conventional chemotherapy. Copyright © 2018 Elsevier B.V. All rights reserved.

Time-resolved Analysis of Proteome Dynamics by Tandem Mass Tags and Stable Isotope Labeling in Cell Culture (TMT-SILAC) Hyperplexing*

PubMed Central

Welle, Kevin A.; Zhang, Tian; Hryhorenko, Jennifer R.; Shen, Shichen; Qu, Jun; Ghaemmaghami, Sina

2016-01-01

Recent advances in mass spectrometry have enabled system-wide analyses of protein turnover. By globally quantifying the kinetics of protein clearance and synthesis, these methodologies can provide important insights into the regulation of the proteome under varying cellular and environmental conditions. To facilitate such analyses, we have employed a methodology that combines metabolic isotopic labeling (Stable Isotope Labeling in Cell Culture - SILAC) with isobaric tagging (Tandem Mass Tags - TMT) for analysis of multiplexed samples. The fractional labeling of multiple time-points can be measured in a single mass spectrometry run, providing temporally resolved measurements of protein turnover kinetics. To demonstrate the feasibility of the approach, we simultaneously measured the kinetics of protein clearance and accumulation for more than 3000 proteins in dividing and quiescent human fibroblasts and verified the accuracy of the measurements by comparison to established non-multiplexed approaches. The results indicate that upon reaching quiescence, fibroblasts compensate for lack of cellular growth by globally downregulating protein synthesis and upregulating protein degradation. The described methodology significantly reduces the cost and complexity of temporally-resolved dynamic proteomic experiments and improves the precision of proteome-wide turnover data. PMID:27765818

Inter-Cellular Exchange of Cellular Components via VE-Cadherin-Dependent Trans-Endocytosis

PubMed Central

Sakurai, Takashi; Woolls, Melissa J.; Jin, Suk-Won

2014-01-01

Cell-cell communications typically involve receptor-mediated signaling initiated by soluble or cell-bound ligands. Here, we report a unique mode of endocytosis: proteins originating from cell-cell junctions and cytosolic cellular components from the neighboring cell are internalized, leading to direct exchange of cellular components between two adjacent endothelial cells. VE-cadherins form transcellular bridges between two endothelial cells that are the basis of adherence junctions. At such adherens junction sites, we observed the movement of the entire VE-cadherin molecule from one endothelial cell into the other with junctional and cytoplasmic components. This phenomenon, here termed trans-endocytosis, requires the establishment of a VE-cadherin homodimer in trans with internalization proceeding in a Rac1-, and actomyosin-dependent manner. Importantly, the trans-endocytosis is not dependent on any known endocytic pathway including clathrin-dependent endocytosis, macropinocytosis or phagocytosis. This novel form of cell-cell communications, leading to a direct exchange of cellular components, was observed in 2D and 3D-cultured endothelial cells as well as in the developing zebrafish vasculature. PMID:24603875

CellCognition: time-resolved phenotype annotation in high-throughput live cell imaging.

PubMed

Held, Michael; Schmitz, Michael H A; Fischer, Bernd; Walter, Thomas; Neumann, Beate; Olma, Michael H; Peter, Matthias; Ellenberg, Jan; Gerlich, Daniel W

2010-09-01

Fluorescence time-lapse imaging has become a powerful tool to investigate complex dynamic processes such as cell division or intracellular trafficking. Automated microscopes generate time-resolved imaging data at high throughput, yet tools for quantification of large-scale movie data are largely missing. Here we present CellCognition, a computational framework to annotate complex cellular dynamics. We developed a machine-learning method that combines state-of-the-art classification with hidden Markov modeling for annotation of the progression through morphologically distinct biological states. Incorporation of time information into the annotation scheme was essential to suppress classification noise at state transitions and confusion between different functional states with similar morphology. We demonstrate generic applicability in different assays and perturbation conditions, including a candidate-based RNA interference screen for regulators of mitotic exit in human cells. CellCognition is published as open source software, enabling live-cell imaging-based screening with assays that directly score cellular dynamics.

Calcium neuroimaging in behaving zebrafish larvae using a turn-key light field camera

NASA Astrophysics Data System (ADS)

Cruz Perez, Carlos; Lauri, Antonella; Symvoulidis, Panagiotis; Cappetta, Michele; Erdmann, Arne; Westmeyer, Gil Gregor

2015-09-01

Reconstructing a three-dimensional scene from multiple simultaneously acquired perspectives (the light field) is an elegant scanless imaging concept that can exceed the temporal resolution of currently available scanning-based imaging methods for capturing fast cellular processes. We tested the performance of commercially available light field cameras on a fluorescent microscopy setup for monitoring calcium activity in the brain of awake and behaving reporter zebrafish larvae. The plenoptic imaging system could volumetrically resolve diverse neuronal response profiles throughout the zebrafish brain upon stimulation with an aversive odorant. Behavioral responses of the reporter fish could be captured simultaneously together with depth-resolved neuronal activity. Overall, our assessment showed that with some optimizations for fluorescence microscopy applications, commercial light field cameras have the potential of becoming an attractive alternative to custom-built systems to accelerate molecular imaging research on cellular dynamics.

Calcium neuroimaging in behaving zebrafish larvae using a turn-key light field camera.

PubMed

Perez, Carlos Cruz; Lauri, Antonella; Symvoulidis, Panagiotis; Cappetta, Michele; Erdmann, Arne; Westmeyer, Gil Gregor

2015-09-01

Reconstructing a three-dimensional scene from multiple simultaneously acquired perspectives (the light field) is an elegant scanless imaging concept that can exceed the temporal resolution of currently available scanning-based imaging methods for capturing fast cellular processes. We tested the performance of commercially available light field cameras on a fluorescent microscopy setup for monitoring calcium activity in the brain of awake and behaving reporter zebrafish larvae. The plenoptic imaging system could volumetrically resolve diverse neuronal response profiles throughout the zebrafish brain upon stimulation with an aversive odorant. Behavioral responses of the reporter fish could be captured simultaneously together with depth-resolved neuronal activity. Overall, our assessment showed that with some optimizations for fluorescence microscopy applications, commercial light field cameras have the potential of becoming an attractive alternative to custom-built systems to accelerate molecular imaging research on cellular dynamics.

Isolating specific cell and tissue compartments from 3D images for quantitative regional distribution analysis using novel computer algorithms.

PubMed

Fenrich, Keith K; Zhao, Ethan Y; Wei, Yuan; Garg, Anirudh; Rose, P Ken

2014-04-15

Isolating specific cellular and tissue compartments from 3D image stacks for quantitative distribution analysis is crucial for understanding cellular and tissue physiology under normal and pathological conditions. Current approaches are limited because they are designed to map the distributions of synapses onto the dendrites of stained neurons and/or require specific proprietary software packages for their implementation. To overcome these obstacles, we developed algorithms to Grow and Shrink Volumes of Interest (GSVI) to isolate specific cellular and tissue compartments from 3D image stacks for quantitative analysis and incorporated these algorithms into a user-friendly computer program that is open source and downloadable at no cost. The GSVI algorithm was used to isolate perivascular regions in the cortex of live animals and cell membrane regions of stained spinal motoneurons in histological sections. We tracked the real-time, intravital biodistribution of injected fluorophores with sub-cellular resolution from the vascular lumen to the perivascular and parenchymal space following a vascular microlesion, and mapped the precise distributions of membrane-associated KCC2 and gephyrin immunolabeling in dendritic and somatic regions of spinal motoneurons. Compared to existing approaches, the GSVI approach is specifically designed for isolating perivascular regions and membrane-associated regions for quantitative analysis, is user-friendly, and free. The GSVI algorithm is useful to quantify regional differences of stained biomarkers (e.g., cell membrane-associated channels) in relation to cell functions, and the effects of therapeutic strategies on the redistributions of biomolecules, drugs, and cells in diseased or injured tissues. Copyright © 2014 Elsevier B.V. All rights reserved.

CORSEN, a new software dedicated to microscope-based 3D distance measurements: mRNA-mitochondria distance, from single-cell to population analyses.

PubMed

Jourdren, Laurent; Delaveau, Thierry; Marquenet, Emelie; Jacq, Claude; Garcia, Mathilde

2010-07-01

Recent improvements in microscopy technology allow detection of single molecules of RNA, but tools for large-scale automatic analyses of particle distributions are lacking. An increasing number of imaging studies emphasize the importance of mRNA localization in the definition of cell territory or the biogenesis of cell compartments. CORSEN is a new tool dedicated to three-dimensional (3D) distance measurements from imaging experiments especially developed to access the minimal distance between RNA molecules and cellular compartment markers. CORSEN includes a 3D segmentation algorithm allowing the extraction and the characterization of the cellular objects to be processed--surface determination, aggregate decomposition--for minimal distance calculations. CORSEN's main contribution lies in exploratory statistical analysis, cell population characterization, and high-throughput assays that are made possible by the implementation of a batch process analysis. We highlighted CORSEN's utility for the study of relative positions of mRNA molecules and mitochondria: CORSEN clearly discriminates mRNA localized to the vicinity of mitochondria from those that are translated on free cytoplasmic polysomes. Moreover, it quantifies the cell-to-cell variations of mRNA localization and emphasizes the necessity for statistical approaches. This method can be extended to assess the evolution of the distance between specific mRNAs and other cellular structures in different cellular contexts. CORSEN was designed for the biologist community with the concern to provide an easy-to-use and highly flexible tool that can be applied for diverse distance quantification issues.

Application of differential interference contrast with inverted microscopes to the in vitro perfused nephron.

PubMed

Horster, M; Gundlach, H

1979-12-01

The study of in vitro perfused individual nephron segments requires a microscope which provides: (1) easy access to the specimen for measurement of cellular solute flux and voltage; (2) an image with high resolution and contrast; (3) optical sectioning of the object at different levels; and (4) rapid recording of the morphological phenomena. This paper describes an example of commercially available apparatus meeting the above requirements, and illustrates its efficiency. The microscope is of the inverted type (Zeiss IM 35) equipped with differential-interference-contrast (DIC) with a long working distance, and an automatically controlled camera system. The microscopic image exhibits cellular and intercellular details in the unstained transporting mammalian nephron segments despite their tubular structure and great thickness and makes obvious function-structure correlations (e.g. cell volume changes); luminal and contraluminal cell borders are well resolved for controlled microelectrode impalement.

Fermi Surface of Metallic V_{2}O_{3} from Angle-Resolved Photoemission: Mid-level Filling of e_{g}^{π} Bands.

PubMed

Lo Vecchio, I; Denlinger, J D; Krupin, O; Kim, B J; Metcalf, P A; Lupi, S; Allen, J W; Lanzara, A

2016-10-14

Using angle resolved photoemission spectroscopy, we report the first band dispersions and distinct features of the bulk Fermi surface (FS) in the paramagnetic metallic phase of the prototypical metal-insulator transition material V_{2}O_{3}. Along the c axis we observe both an electron pocket and a triangular holelike FS topology, showing that both V 3d a_{1g} and e_{g}^{π} states contribute to the FS. These results challenge the existing correlation-enhanced crystal field splitting theoretical explanation for the transition mechanism and pave the way for the solution of this mystery.

Confocal reflectance quantitative phase microscope system for cellular membranes dynamics study (Conference Presentation)

NASA Astrophysics Data System (ADS)

Singh, Vijay Raj; Yaqoob, Zahid; So, Peter T. C.

2017-02-01

Quantitative phase microscopy (QPM) techniques developed so far primarily belongs to high speed transmitted light based systems that has enough sensitivity to resolve membrane fluctuations and dynamics, but has no depth resolution. Therefore, most biomechanics studies using QPM today is confined to simple cells, such as RBCs, without internal organelles. An important instrument that will greatly extend the biomedical applications of QPM is to develop next generation microscope with 3D capability and sufficient temporal resolution to study biomechanics of complex eukaryotic cells including the mechanics of their internal compartments. For eukaryotic cells, the depth sectioning capability is critical and should be sufficient to distinguish nucleic membrane fluctuations from plasma membrane fluctuations. Further, this microscope must provide high temporal resolution since typical eukaryotes membranes are substantially stiffer than RBCs. A confocal reflectance quantitative phase microscope is presented based on multi-pinhole scanning, with the capabilities of higher temporal resolution and sensitivity for nucleic and plasma membranes of eukaryotic cells. System hardware is developed based on an array of confocal pinhole generated by using the `ON' state of subset of micro-mirrors of digital micro-mirror device (DMD, from Texas Instruments) and high-speed raster scanning provides 14ms imaging speed in wide-field mode. A common path interferometer is integrated at the imaging arm for detection of specimens' quantitative phase information. Theoretical investigation of quantitative phase reconstructed from system is investigated and application of system is presented for dimensional fluctuations measurements of both cellular plasma and nucleic membranes of embryonic stem cells.

Fabrication and characterisation of a fully auxetic 3D lattice structure via selective electron beam melting

NASA Astrophysics Data System (ADS)

Warmuth, Franziska; Osmanlic, Fuad; Adler, Lucas; Lodes, Matthias A.; Körner, Carolin

2017-02-01

A three-dimensional fully auxetic cellular structure with negative Poisson’s ratio is presented. Samples are fabricated from Ti6Al4V powder via selective electron beam melting. The influence of the strut thickness and the amplitude of the strut on the mechanical properties and the deformation behaviour of cellular structures is studied.

Composite alginate gels for tunable cellular microenvironment mechanics

NASA Astrophysics Data System (ADS)

Khavari, Adele; Nydén, Magnus; Weitz, David A.; Ehrlicher, Allen J.

2016-08-01

The mechanics of the cellular microenvironment can be as critical as biochemistry in directing cell behavior. Many commonly utilized materials derived from extra-cellular-matrix create excellent scaffolds for cell growth, however, evaluating the relative mechanical and biochemical effects independently in 3D environments has been difficult in frequently used biopolymer matrices. Here we present 3D sodium alginate hydrogel microenvironments over a physiological range of stiffness (E = 1.85 to 5.29 kPa), with and without RGD binding sites or collagen fibers. We use confocal microscopy to measure the growth of multi-cellular aggregates (MCAs), of increasing metastatic potential in different elastic moduli of hydrogels, with and without binding factors. We find that the hydrogel stiffness regulates the growth and morphology of these cell clusters; MCAs grow larger and faster in the more rigid environments similar to cancerous breast tissue (E = 4-12 kPa) as compared to healthy tissue (E = 0.4-2 kpa). Adding binding factors from collagen and RGD peptides increases growth rates, and change maximum MCA sizes. These findings demonstrate the utility of these independently tunable mechanical/biochemistry gels, and that mechanical confinement in stiffer microenvironments may increase cell proliferation.

Identification and High-Resolution Imaging of α-Tocopherol from Human Cells to Whole Animals by TOF-SIMS Tandem Mass Spectrometry

NASA Astrophysics Data System (ADS)

Bruinen, Anne L.; Fisher, Gregory L.; Balez, Rachelle; van der Sar, Astrid M.; Ooi, Lezanne; Heeren, Ron M. A.

2018-06-01

A unique method for identification of biomolecular components in different biological specimens, while preserving the capability for high speed 2D and 3D molecular imaging, is employed to investigate cellular response to oxidative stress. The employed method enables observing the distribution of the antioxidant α-tocopherol and other molecules in cellular structures via time-of-flight secondary ion mass spectrometry (TOF-SIMS (MS1)) imaging in parallel with tandem mass spectrometry (MS2) imaging, collected simultaneously. The described method is employed to examine a network formed by neuronal cells differentiated from human induced pluripotent stem cells (iPSCs), a model for investigating human neurons in vitro. The antioxidant α-tocopherol is identified in situ within different cellular layers utilizing a 3D TOF-SIMS tandem MS imaging analysis. As oxidative stress also plays an important role in mediating inflammation, the study was expanded to whole body tissue sections of M. marinum-infected zebrafish, a model organism for tuberculosis. The TOF-SIMS tandem MS imaging results reveal an increased presence of α-tocopherol in response to the pathogen. [Figure not available: see fulltext.

Quantum Theory of Three-Dimensional Superresolution Using Rotating-PSF Imagery

NASA Astrophysics Data System (ADS)

Prasad, S.; Yu, Z.

The inverse of the quantum Fisher information (QFI) matrix (and extensions thereof) provides the ultimate lower bound on the variance of any unbiased estimation of a parameter from statistical data, whether of intrinsically quantum mechanical or classical character. We calculate the QFI for Poisson-shot-noise-limited imagery using the rotating PSF that can localize and resolve point sources fully in all three dimensions. We also propose an experimental approach based on the use of computer generated hologram and projective measurements to realize the QFI-limited variance for the problem of super-resolving a closely spaced pair of point sources at a highly reduced photon cost. The paper presents a preliminary analysis of quantum-limited three-dimensional (3D) pair optical super-resolution (OSR) problem with potential applications to astronomical imaging and 3D space-debris localization.

Differential regulation of cellular functions by the C-termini of transmembrane 4 L six family proteins in 2- or 3-dimensional environment.

PubMed

Cheong, Jin-Gyu; Song, Dae-Geun; Song, Haeng Eun; Berditchevski, Fedor; Nam, Seo Hee; Jung, Jae Woo; Kim, Hye-Jin; Kim, Ji Eon; Kim, Somi; Ryu, Jihye; Cho, Chang Yun; Lee, Kyung-Min; Lee, Jung Weon

2017-02-21

The transmembrane 4 L six family proteins TM4SF1, TM4SF4, and TM4SF5 share 40-50% overall sequence identity, but their C-terminus identity is limited. It may be likely that the C-termini of the members are important and unique for own regulatory functions. We thus examined how the TM4SF5 C-terminus affected cellular functions differentially from other family members. Using colon cancer cells expressing wildtype (WT), C-terminus-deleted, or chimeric mutants, diverse cellular functions were explored in 2-dimensional (2D) and 3-dimensional (3D) condition. The C-termini of the proteins were relatively comparable with respect to 2D cell proliferation, although each C-terminal-deletion mutant exhibited increased proliferation relative to the WT. Using chimeric constructs, we found that the TM4SF5 C-terminus was critical for regulating the diverse metastatic functions of TM4SF5, and could positively replace the C-termini of other family members. Replacement of the TM4SF1 or TM4SF4 C-terminus with that of TM4SF5 increased spheroids growth, transwell migration, and invasive dissemination from spheroids in 3D collagen gels. TM4SF5-mediated effects required its extracellular loop 2 linked to the C-terminus via the transmembrane domain 4, with causing c-Src activation. Altogether, the C-terminus of TM4SF5 appears to mediate pro-migratory roles, depending on a structural relay from the second extracellular loop to the C-terminus.

Resolving Orbital and Climate Keys of Earth and Extraterrestrial Environments with Dynamics (ROCKE-3D) 1.0: A General Circulation Model for Simulating the Climates of Rocky Planets

NASA Astrophysics Data System (ADS)

Way, M. J.; Aleinov, I.; Amundsen, David S.; Chandler, M. A.; Clune, T. L.; Del Genio, A. D.; Fujii, Y.; Kelley, M.; Kiang, N. Y.; Sohl, L.; Tsigaridis, K.

2017-07-01

Resolving Orbital and Climate Keys of Earth and Extraterrestrial Environments with Dynamics (ROCKE-3D) is a three-dimensional General Circulation Model (GCM) developed at the NASA Goddard Institute for Space Studies for the modeling of atmospheres of solar system and exoplanetary terrestrial planets. Its parent model, known as ModelE2, is used to simulate modern Earth and near-term paleo-Earth climates. ROCKE-3D is an ongoing effort to expand the capabilities of ModelE2 to handle a broader range of atmospheric conditions, including higher and lower atmospheric pressures, more diverse chemistries and compositions, larger and smaller planet radii and gravity, different rotation rates (from slower to more rapid than modern Earth’s, including synchronous rotation), diverse ocean and land distributions and topographies, and potential basic biosphere functions. The first aim of ROCKE-3D is to model planetary atmospheres on terrestrial worlds within the solar system such as paleo-Earth, modern and paleo-Mars, paleo-Venus, and Saturn’s moon Titan. By validating the model for a broad range of temperatures, pressures, and atmospheric constituents, we can then further expand its capabilities to those exoplanetary rocky worlds that have been discovered in the past, as well as those to be discovered in the future. We also discuss the current and near-future capabilities of ROCKE-3D as a community model for studying planetary and exoplanetary atmospheres.

Solving the inverse scattering problem in reflection-mode dynamic speckle-field phase microscopy (Conference Presentation)

NASA Astrophysics Data System (ADS)

Zhou, Renjie; So, Peter T. C.; Yaqoob, Zahid; Jin, Di; Hosseini, Poorya; Kuang, Cuifang; Singh, Vijay Raj; Kim, Yang-Hyo; Dasari, Ramachandra R.

2017-02-01

Most of the quantitative phase microscopy systems are unable to provide depth-resolved information for measuring complex biological structures. Optical diffraction tomography provides a non-trivial solution to it by 3D reconstructing the object with multiple measurements through different ways of realization. Previously, our lab developed a reflection-mode dynamic speckle-field phase microscopy (DSPM) technique, which can be used to perform depth resolved measurements in a single shot. Thus, this system is suitable for measuring dynamics in a layer of interest in the sample. DSPM can be also used for tomographic imaging, which promises to solve the long-existing "missing cone" problem in 3D imaging. However, the 3D imaging theory for this type of system has not been developed in the literature. Recently, we have developed an inverse scattering model to rigorously describe the imaging physics in DSPM. Our model is based on the diffraction tomography theory and the speckle statistics. Using our model, we first precisely calculated the defocus response and the depth resolution in our system. Then, we further calculated the 3D coherence transfer function to link the 3D object structural information with the axially scanned imaging data. From this transfer function, we found that in the reflection mode excellent sectioning effect exists in the low lateral spatial frequency region, thus allowing us to solve the "missing cone" problem. Currently, we are working on using this coherence transfer function to reconstruct layered structures and complex cells.

Resolving Orbital and Climate Keys of Earth and Extraterrestrial Environments with Dynamics (ROCKE-3D) 1.0: A General Circulation Model for Simulating the Climates of Rocky Planets

NASA Technical Reports Server (NTRS)

Way, M. J.; Aleinov, I.; Amundsen, David S.; Chandler, M. A.; Clune, T. L.; Del Genio, A.; Fujii, Y.; Kelley, M.; Kiang, N. Y.; Sohl, L.;

Imaging of oxygenation in 3D tissue models with multi-modal phosphorescent probes

NASA Astrophysics Data System (ADS)

Papkovsky, Dmitri B.; Dmitriev, Ruslan I.; Borisov, Sergei

2015-03-01

Cell-penetrating phosphorescence based probes allow real-time, high-resolution imaging of O2 concentration in respiring cells and 3D tissue models. We have developed a panel of such probes, small molecule and nanoparticle structures, which have different spectral characteristics, cell penetrating and tissue staining behavior. The probes are compatible with conventional live cell imaging platforms and can be used in different detection modalities, including ratiometric intensity and PLIM (Phosphorescence Lifetime IMaging) under one- or two-photon excitation. Analytical performance of these probes and utility of the O2 imaging method have been demonstrated with different types of samples: 2D cell cultures, multi-cellular spheroids from cancer cell lines and primary neurons, excised slices from mouse brain, colon and bladder tissue, and live animals. They are particularly useful for hypoxia research, ex-vivo studies of tissue physiology, cell metabolism, cancer, inflammation, and multiplexing with many conventional fluorophors and markers of cellular function.

Debio 0507 primarily forms diaminocyclohexane-Pt-d(GpG) and -d(ApG) DNA adducts in HCT116 cells

PubMed Central

King, C. L.; Ramachandran, S.; Collins, L.; Swenberg, J. A.; deKrafft, K. E.; Lin, W.; Cicurel, L.; Barbier, M.

2013-01-01

Purpose To characterize the cellular action mechanism of Debio 0507, we compared the major DNA adducts formed by Debio 0507- and oxaliplatin-treated HCT116 human colon carcinoma cells by a combination of inductively coupled plasma mass spectrometry (ICP-MS) and ultra-performance liquid chromatography mass spectrometry (UPLC-MS/MS). Methods HCT116 cells were treated with IC50 doses of Debio 0507 or oxaliplatin for 3 days. Total cellular Pt–DNA adducts were determined by ICP-MS. The DNA was digested, and the major Pt–DNA adducts formed by both drugs were characterized by UPLC/MS/MS essentially as described previously for cisplatin (Baskerville-Abraham et al. in Chem Res Toxicol 22:905–912, 2009). Results The Pt level/deoxynucleotide was 7.4/104 for DNA from Debio 0507-treated cells and 5.5/104 for oxaliplatin-treated cells following a 3-day treatment at the IC50 for each drug. UPLC-MS/MS in the positive ion mode confirmed the major Pt–DNA adducts formed by both drugs were dach-Pt-d(GpG) (904.2 m/z → 610 m/z and 904.2 m/z → 459 m/z) and dach-Pt-d(ApG) (888.2 m/z → 594 m/z and 888.2 m/z → 459 m/z). Conclusions These data show that the major DNA adducts formed by Debio 0507 are the dach-Pt-d(GpG) and dach-Pt-d(ApG) adducts and at equitoxic doses Debio 0507 and oxaliplatin form similar levels of dach-Pt-d(GpG) and dach-Pt-d(ApG) adducts. This suggests that the action mechanisms of Debio 0507 and oxaliplatin are similar at a cellular level. PMID:21968950

Understanding D-Ribose and Mitochondrial Function.

PubMed

Mahoney, Diane E; Hiebert, John B; Thimmesch, Amanda; Pierce, John T; Vacek, James L; Clancy, Richard L; Sauer, Andrew J; Pierce, Janet D

2018-01-01

Mitochondria are important organelles referred to as cellular powerhouses for their unique properties of cellular energy production. With many pathologic conditions and aging, mitochondrial function declines, and there is a reduction in the production of adenosine triphosphate. The energy carrying molecule generated by cellular respiration and by pentose phosphate pathway, an alternative pathway of glucose metabolism. D-ribose is a naturally occurring monosaccharide found in the cells and particularly in the mitochondria is essential in energy production. Without sufficient energy, cells cannot maintain integrity and function. Supplemental D-ribose has been shown to improve cellular processes when there is mitochondrial dysfunction. When individuals take supplemental D-ribose, it can bypass part of the pentose pathway to produce D-ribose-5-phosphate for the production of energy. In this article, we review how energy is produced by cellular respiration, the pentose pathway, and the use of supplemental D-ribose.

Organoids with cancer stem cell-like properties secrete exosomes and HSP90 in a 3D nanoenvironment.

PubMed

Eguchi, Takanori; Sogawa, Chiharu; Okusha, Yuka; Uchibe, Kenta; Iinuma, Ryosuke; Ono, Kisho; Nakano, Keisuke; Murakami, Jun; Itoh, Manabu; Arai, Kazuya; Fujiwara, Toshifumi; Namba, Yuri; Murata, Yoshiki; Ohyama, Kazumi; Shimomura, Manami; Okamura, Hirohiko; Takigawa, Masaharu; Nakatsura, Tetsuya; Kozaki, Ken-Ichi; Okamoto, Kuniaki; Calderwood, Stuart K

2018-01-01

Ability to form cellular aggregations such as tumorspheres and spheroids have been used as a morphological marker of malignant cancer cells and in particular cancer stem cells (CSC). However, the common definition of the types of cellular aggregation formed by cancer cells has not been available. We examined morphologies of 67 cell lines cultured on three dimensional morphology enhancing NanoCulture Plates (NCP) and classified the types of cellular aggregates that form. Among the 67 cell lines, 49 cell lines formed spheres or spheroids, 8 cell lines formed grape-like aggregation (GLA), 8 cell lines formed other types of aggregation, and 3 cell lines formed monolayer sheets. Seven GLA-forming cell lines were derived from adenocarcinoma among the 8 lines. A neuroendocrine adenocarcinoma cell line PC-3 formed asymmetric GLA with ductal structures on the NCPs and rapidly growing asymmetric tumors that metastasized to lymph nodes in immunocompromised mice. In contrast, another adenocarcinoma cell line DU-145 formed spheroids in vitro and spheroid-like tumors in vivo that did not metastasize to lymph nodes until day 50 after transplantation. Culture in the 3D nanoenvironment and in a defined stem cell medium enabled the neuroendocrine adenocarcinoma cells to form slowly growing large organoids that expressed multiple stem cell markers, neuroendocrine markers, intercellular adhesion molecules, and oncogenes in vitro. In contrast, the more commonly used 2D serum-contained environment reduced intercellular adhesion and induced mesenchymal transition and promoted rapid growth of the cells. In addition, the 3D stemness nanoenvironment promoted secretion of HSP90 and EpCAM-exosomes, a marker of CSC phenotype, from the neuroendocrine organoids. These findings indicate that the NCP-based 3D environment enables cells to form stem cell tumoroids with multipotency and model more accurately the in vivo tumor status at the levels of morphology and gene expression.

Organoids with cancer stem cell-like properties secrete exosomes and HSP90 in a 3D nanoenvironment

PubMed Central

Okusha, Yuka; Uchibe, Kenta; Iinuma, Ryosuke; Ono, Kisho; Nakano, Keisuke; Murakami, Jun; Itoh, Manabu; Arai, Kazuya; Fujiwara, Toshifumi; Namba, Yuri; Murata, Yoshiki; Ohyama, Kazumi; Shimomura, Manami; Okamura, Hirohiko; Takigawa, Masaharu; Nakatsura, Tetsuya; Kozaki, Ken-ichi; Okamoto, Kuniaki; Calderwood, Stuart K.

2018-01-01

Ability to form cellular aggregations such as tumorspheres and spheroids have been used as a morphological marker of malignant cancer cells and in particular cancer stem cells (CSC). However, the common definition of the types of cellular aggregation formed by cancer cells has not been available. We examined morphologies of 67 cell lines cultured on three dimensional morphology enhancing NanoCulture Plates (NCP) and classified the types of cellular aggregates that form. Among the 67 cell lines, 49 cell lines formed spheres or spheroids, 8 cell lines formed grape-like aggregation (GLA), 8 cell lines formed other types of aggregation, and 3 cell lines formed monolayer sheets. Seven GLA-forming cell lines were derived from adenocarcinoma among the 8 lines. A neuroendocrine adenocarcinoma cell line PC-3 formed asymmetric GLA with ductal structures on the NCPs and rapidly growing asymmetric tumors that metastasized to lymph nodes in immunocompromised mice. In contrast, another adenocarcinoma cell line DU-145 formed spheroids in vitro and spheroid-like tumors in vivo that did not metastasize to lymph nodes until day 50 after transplantation. Culture in the 3D nanoenvironment and in a defined stem cell medium enabled the neuroendocrine adenocarcinoma cells to form slowly growing large organoids that expressed multiple stem cell markers, neuroendocrine markers, intercellular adhesion molecules, and oncogenes in vitro. In contrast, the more commonly used 2D serum-contained environment reduced intercellular adhesion and induced mesenchymal transition and promoted rapid growth of the cells. In addition, the 3D stemness nanoenvironment promoted secretion of HSP90 and EpCAM-exosomes, a marker of CSC phenotype, from the neuroendocrine organoids. These findings indicate that the NCP-based 3D environment enables cells to form stem cell tumoroids with multipotency and model more accurately the in vivo tumor status at the levels of morphology and gene expression. PMID:29415026

Time-resolved wide-field optically sectioned fluorescence microscopy

NASA Astrophysics Data System (ADS)

Dupuis, Guillaume; Benabdallah, Nadia; Chopinaud, Aurélien; Mayet, Céline; Lévêque-Fort, Sandrine

2013-02-01

We present the implementation of a fast wide-field optical sectioning technique called HiLo microscopy on a fluorescence lifetime imaging microscope. HiLo microscopy is based on the fusion of two images, one with structured illumination and another with uniform illumination. Optically sectioned images are then digitally generated thanks to a fusion algorithm. HiLo images are comparable in quality with confocal images but they can be acquired faster over larger fields of view. We obtain 4D imaging by combining HiLo optical sectioning, time-gated detection, and z-displacement. We characterize the performances of this set-up in terms of 3D spatial resolution and time-resolved capabilities in both fixed- and live-cell imaging modes.

Agent-Based Computational Modeling of Cell Culture: Understanding Dosimetry In Vitro as Part of In Vitro to In Vivo Extrapolation

EPA Science Inventory

Quantitative characterization of cellular dose in vitro is needed for alignment of doses in vitro and in vivo. We used the agent-based software, CompuCell3D (CC3D), to provide a stochastic description of cell growth in culture. The model was configured so that isolated cells assu...

Tissue vascularization through 3D printing: Will technology bring us flow?

PubMed

Paulsen, S J; Miller, J S

2015-05-01

Though in vivo models provide the most physiologically relevant environment for studying tissue function, in vitro studies provide researchers with explicit control over experimental conditions and the potential to develop high throughput testing methods. In recent years, advancements in developmental biology research and imaging techniques have significantly improved our understanding of the processes involved in vascular development. However, the task of recreating the complex, multi-scale vasculature seen in in vivo systems remains elusive. 3D bioprinting offers a potential method to generate controlled vascular networks with hierarchical structure approaching that of in vivo networks. Bioprinting is an interdisciplinary field that relies on advances in 3D printing technology along with advances in imaging and computational modeling, which allow researchers to monitor cellular function and to better understand cellular environment within the printed tissue. As bioprinting technologies improve with regards to resolution, printing speed, available materials, and automation, 3D printing could be used to generate highly controlled vascularized tissues in a high throughput manner for use in regenerative medicine and the development of in vitro tissue models for research in developmental biology and vascular diseases. © 2015 Wiley Periodicals, Inc.

Fast Dynamic 3D MRSI with Compressed Sensing and Multiband Excitation Pulses for Hyperpolarized 13C Studies

PubMed Central

Larson, Peder E. Z.; Hu, Simon; Lustig, Michael; Kerr, Adam B.; Nelson, Sarah J.; Kurhanewicz, John; Pauly, John M.; Vigneron, Daniel B.

2010-01-01

Hyperpolarized 13C MRSI can detect not only the uptake of the pre-polarized molecule but also its metabolic products in vivo, thus providing a powerful new method to study cellular metabolism. Imaging the dynamic perfusion and conversion of these metabolites provides additional tissue information but requires methods for efficient hyperpolarization usage and rapid acquisitions. In this work, we have developed a time-resolved 3D MRSI method for acquiring hyperpolarized 13C data by combining compressed sensing methods for acceleration and multiband excitation pulses to efficiently use the magnetization. This method achieved a 2 sec temporal resolution with full volumetric coverage of a mouse, and metabolites were observed for up to 60 sec following injection of hyperpolarized [1-13C]-pyruvate. The compressed sensing acquisition used random phase encode gradient blips to create a novel random undersampling pattern tailored to dynamic MRSI with sampling incoherency in four (time, frequency and two spatial) dimensions. The reconstruction was also tailored to dynamic MRSI by applying a temporal wavelet sparsifying transform in order to exploit the inherent temporal sparsity. Customized multiband excitation pulses were designed with a lower flip angle for the [1-13C]-pyruvate substrate given its higher concentration than its metabolic products ([1-13C]-lactate and [1-13C]-alanine), thus using less hyperpolarization per excitation. This approach has enabled the monitoring of perfusion and uptake of the pyruvate, and the conversion dynamics to lactate and alanine throughout a volume with high spatial and temporal resolution. PMID:20939089

Human Milk Proresolving Mediators Stimulate Resolution of Acute Inflammation

PubMed Central

Dalli, Jesmond; Serhan, Charles N

2015-01-01

Human milk contains nutrients and bioactive products relevant to infant development and immunological protection. Here, we investigated the pro-resolving properties of milk using human milk lipid mediator isolates (HLMI) and determined their impact on resolution programs in vivo and with human macrophages. HLMI reduced maximum neutrophil numbers (14.6±1.2×106 to 11.0±1.0×106 cells/exudate) and shortened the resolution interval (Ri; 50% neutrophil reduction) 54% compared to peritonitis. Using rigorous liquid-chromatography tandem-mass spectrometry (LC-MS-MS)-based lipid mediator (LM) metabololipidomics, we demonstrated that human milk possesses a proresolving LM-SPM signature profile, containing specialized proresolving mediators (SPM; e.g. resolvins, protectins, maresins and lipoxins) at bioactive levels (pico-nanomolar concentrations) that enhanced human macrophage efferocytosis and bacterial containment. SPM identified in human milk included D-series resolvins, (e.g. Resolvin (Rv) D1, RvD2, RvD3, AT-RvD3 and RvD4), Protectin (PD)1, Maresin (MaR)1, E-series resolvins (e.g. RvE1, RvE2 and RvE3) and lipoxins (LXA4 and LXB4). Of the SPM identified in human milk, RvD2 and MaR1 (50 ng/mouse) individually shortened Ri ~75%. Milk from mastitis gave higher LTB4 and prostanoids and lower SPM levels. Taken together, these findings provide evidence that human milk has pro-resolving actions via comprehensive LM-SPM profiling, describing a potentially novel mechanism in maternal-infant biochemical imprinting. PMID:26462421

Controlled surface topography regulates collective 3D migration by epithelial-mesenchymal composite embryonic tissues.

PubMed

Song, Jiho; Shawky, Joseph H; Kim, YongTae; Hazar, Melis; LeDuc, Philip R; Sitti, Metin; Davidson, Lance A

2015-07-01

Cells in tissues encounter a range of physical cues as they migrate. Probing single cell and collective migratory responses to physically defined three-dimensional (3D) microenvironments and the factors that modulate those responses are critical to understanding how tissue migration is regulated during development, regeneration, and cancer. One key physical factor that regulates cell migration is topography. Most studies on surface topography and cell mechanics have been carried out with single migratory cells, yet little is known about the spreading and motility response of 3D complex multi-cellular tissues to topographical cues. Here, we examine the response to complex topographical cues of microsurgically isolated tissue explants composed of epithelial and mesenchymal cell layers from naturally 3D organized embryos of the aquatic frog Xenopus laevis. We control topography using fabricated micropost arrays (MPAs) and investigate the collective 3D migration of these multi-cellular systems in these MPAs. We find that the topography regulates both collective and individual cell migration and that dense MPAs reduce but do not eliminate tissue spreading. By modulating cell size through the cell cycle inhibitor Mitomycin C or the spacing of the MPAs we uncover how 3D topographical cues disrupt collective cell migration. We find surface topography can direct both single cell motility and tissue spreading, altering tissue-scale processes that enable efficient conversion of single cell motility into collective movement. Copyright © 2015 Elsevier Ltd. All rights reserved.

Polychromatic microdiffraction characterization of defect gradients in severely deformed materials.

PubMed

Barabash, Rozaliya I; Ice, Gene E; Liu, Wenjun; Barabash, Oleg M

2009-01-01

This paper analyzes local lattice rotations introduced in severely deformed polycrystalline titanium by friction stir welding. Nondestructive three-dimensional (3D) spatially resolved polychromatic X-ray microdiffraction, is used to resolve the local crystal structure of the restructured surface from neighboring local structures in the sample material. The measurements reveal strong gradients of strain and geometrically necessary dislocations near the surface and illustrate the potential of polychromatic microdiffraction for the study of deformation in complex materials systems.

Effects of vitamin E supplementation on cellular α-tocopherol concentrations of neutrophils in Holstein calves

PubMed Central

Higuchi, Hidetoshi; Ito, Erina; Iwano, Hidetoma; Oikawa, Shin; Nagahata, Hajime

2013-01-01

The effects of vitamin E supplementation on cellular α-tocopherol concentrations of neutrophils from Holstein calves and the mechanism of scavenger receptor class B type I (SR-BI)-mediated uptake of α-tocopherol were examined. Cellular α-tocopherol concentrations in vitamin E-treated calves increased from 3.5 ± 0.38 to 7.2 ± 0.84 μg/107 cells, respectively, within 14 d after vitamin E supplementation; these concentrations were significantly higher than those of control calves (P < 0.01). The expression indices of SR-BI [a major receptor that recognizes high-density lipoprotein (HDL)] mRNA in neutrophils were two to five times higher (P < 0.01) in neutrophils obtained from vitamin E-supplemented calves compared with those from control calves, and anti-SR-B1 antibody, ranging from 0.1 to 1.0 μg/mL, significantly (P < 0.01) decreased cellular α-tocopherol concentrations of neutrophils. Cytochalasin D and latrunculin B, major inhibitors of actin polymerization of neutrophils, significantly decreased cellular α-tocopherol concentrations of neutrophils (P < 0.01). Our results demonstrated that in vitamin E-supplemented calves: 1) α-tocopherol is mainly distributed with HDL, 2) α-tocopherol within HDL is recognized by SR-BI on the surface of neutrophils, and 3) rearrangement of the actin cytoskeleton is a crucial step for the uptake of α-tocopherol by neutrophils. PMID:24082403

Ion Dynamics of a BHT-600 Hall Thruster Measured with Time-Resolved Laser-Induced Fluorescence

DTIC Science & Technology

2016-01-14

LIF  Method V = 0 V V λ = λ0 λ > λ0 λ < λ0 Ion Velocity from  Doppler Shift: 0 5 10 15 20 25 30 0 0.1 0.2 0.3 0.4...0.5 0.6 0.7 0.8 0.9 1 Frequency (GHz) N o r m a l i z e d S i g n a l xneg28z6_AVG OG LIF OG Fit LIF Fit AFRL | BHT‐600 TRLIF | Jan 14, 2016 10...Distribution A: Approved for Public Release; Distribution Unlimited. PA# 16204 Time‐Resolved  LIF  Method Laser  Tapered Amp  D1  BS1  BS2  D2  O G C  

Development and characterization of a 3D multicell microtissue culture model of airway smooth muscle.

PubMed

West, Adrian R; Zaman, Nishat; Cole, Darren J; Walker, Matthew J; Legant, Wesley R; Boudou, Thomas; Chen, Christopher S; Favreau, John T; Gaudette, Glenn R; Cowley, Elizabeth A; Maksym, Geoffrey N

2013-01-01

Airway smooth muscle (ASM) cellular and molecular biology is typically studied with single-cell cultures grown on flat 2D substrates. However, cells in vivo exist as part of complex 3D structures, and it is well established in other cell types that altering substrate geometry exerts potent effects on phenotype and function. These factors may be especially relevant to asthma, a disease characterized by structural remodeling of the airway wall, and highlights a need for more physiologically relevant models of ASM function. We utilized a tissue engineering platform known as microfabricated tissue gauges to develop a 3D culture model of ASM featuring arrays of ∼0.4 mm long, ∼350 cell "microtissues" capable of simultaneous contractile force measurement and cell-level microscopy. ASM-only microtissues generated baseline tension, exhibited strong cellular organization, and developed actin stress fibers, but lost structural integrity and dissociated from the cantilevers within 3 days. Addition of 3T3-fibroblasts dramatically improved survival times without affecting tension development or morphology. ASM-3T3 microtissues contracted similarly to ex vivo ASM, exhibiting reproducible responses to a range of contractile and relaxant agents. Compared with 2D cultures, microtissues demonstrated identical responses to acetylcholine and KCl, but not histamine, forskolin, or cytochalasin D, suggesting that contractility is regulated by substrate geometry. Microtissues represent a novel model for studying ASM, incorporating a physiological 3D structure, realistic mechanical environment, coculture of multiple cells types, and comparable contractile properties to existing models. This new model allows for rapid screening of biochemical and mechanical factors to provide insight into ASM dysfunction in asthma.

Lectins interact differentially with purified human eosinophils, cultured cord blood-derived mast cells and the myeloid leukaemic cell line AML14.3D10: induction of interleukin-4 secretion is conserved among granulocytes, but is not proportional to agglutination or lectin-glycoprotein interaction.

PubMed

Hoffmann, H J; Dahl, C; Schiøtz, P O; Berglund, L; Dahl, R

2003-07-01

Atopy is closely associated with the cellular T helper type-2 (Th2) phenotype, that is dominated by the pleiotrophic cytokine IL-4. The cellular source of IL-4 has yet to be determined, although basophils have been proposed. Eosinophils and mast cells are likely contenders investigated here, and the eosinophil-like leukaemia line AML14.3D10 is compared to eosinophils as an in vitro culturable model for eosinophils. Lectins can cross-link-specific surface glycoproteins and are found in the ingested (processed foods) and inhaled (airborne pollen grains) human environment. Therefore it is of interest to determine whether lectins can elicit the release of IL-4 from Th2-associated granulocytes other than basophils. This study investigated the ability of eosinophils, AML14.3D10 and mast cells to secrete preformed IL-4 in response to stimulation with lectins, and explored molecular mechanisms underlying the interaction. Purified eosinophils and basophils, and cultured mast cells and AML14.3D10 cells were incubated with 1 micro m lectin. Agglutination was scored by microscopy. IL-4 secretion was measured by enzyme-linked immunosorbent assay. Biotinylated lectins were used to determine binding to cells by flow cytometry and in lectin blots of sodium dodecyl sulphate (SDS) gels. Purified human eosinophils, AML14.3D10 cells and cultured mast cells secrete IL-4 with a pattern similar to that found in basophils when stimulated with a panel of reactive and unreactive lectins. The lectin SNA induces IL-4 secretion from mast cells and basophils, but not from eosinophils or AML14.3D10. Eosinophils appear to secrete only pre-formed IL-4, whereas mast cells may synthesize IL-4 on ligation with the lectin LCA. Lectins that agglutinate the granulocytes investigated do not necessarily induce secretion of IL-4. Lectins that elicit secretion of IL-4 bind more to eosinophils than unreactive lectins as determined by flow cytometry and lectin blotting of SDS gels. As granulocytes with functions related to that of basophils, eosinophils, AML14.3D10 and cultured mast cells respond to stimulation with lectins similarly to basophils. This emphasizes the possibility that eosinophils and mast cells may be linked in their cellular heritage as the cellular partners, and lectins as ligands, may contribute to the maintenance of a Th2-favoured microenvironment that is thought to underlie the allergic march.

Time-resolved diffusion tomographic 2D and 3D imaging in highly scattering turbid media

NASA Technical Reports Server (NTRS)

Alfano, Robert R. (Inventor); Cai, Wei (Inventor); Liu, Feng (Inventor); Lax, Melvin (Inventor); Das, Bidyut B. (Inventor)

1999-01-01

A method for imaging objects in highly scattering turbid media. According to one embodiment of the invention, the method involves using a plurality of intersecting source/detectors sets and time-resolving equipment to generate a plurality of time-resolved intensity curves for the diffusive component of light emergent from the medium. For each of the curves, the intensities at a plurality of times are then inputted into the following inverse reconstruction algorithm to form an image of the medium: ##EQU1## wherein W is a matrix relating output at source and detector positions r.sub.s and r.sub.d, at time t, to position r, .LAMBDA. is a regularization matrix, chosen for convenience to be diagonal, but selected in a way related to the ratio of the noise, to fluctuations in the absorption (or diffusion) X.sub.j that we are trying to determine: .LAMBDA..sub.ij =.lambda..sub.j .delta..sub.ij with .lambda..sub.j =/<.DELTA.Xj.DELTA.Xj> Y is the data collected at the detectors, and X.sup.k is the kth iterate toward the desired absoption information. An algorithm, which combines a two dimensional (2D) matrix inversion with a one-dimensional (1D) Fourier transform inversion is used to obtain images of three dimensional hidden objects in turbid scattering media.

Time-resolved diffusion tomographic 2D and 3D imaging in highly scattering turbid media

NASA Technical Reports Server (NTRS)

Alfano, Robert R. (Inventor); Cai, Wei (Inventor); Gayen, Swapan K. (Inventor)

2000-01-01

A method for imaging objects in highly scattering turbid media. According to one embodiment of the invention, the method involves using a plurality of intersecting source/detectors sets and time-resolving equipment to generate a plurality of time-resolved intensity curves for the diffusive component of light emergent from the medium. For each of the curves, the intensities at a plurality of times are then inputted into the following inverse reconstruction algorithm to form an image of the medium: wherein W is a matrix relating output at source and detector positions r.sub.s and r.sub.d, at time t, to position r, .LAMBDA. is a regularization matrix, chosen for convenience to be diagonal, but selected in a way related to the ratio of the noise, to fluctuations in the absorption (or diffusion) X.sub.j that we are trying to determine: .LAMBDA..sub.ij =.lambda..sub.j .delta..sub.ij with .lambda..sub.j =/<.DELTA.Xj.DELTA.Xj> Y is the data collected at the detectors, and X.sup.k is the kth iterate toward the desired absorption information. An algorithm, which combines a two dimensional (2D) matrix inversion with a one-dimensional (1D) Fourier transform inversion is used to obtain images of three dimensional hidden objects in turbid scattering media.

Molecular counting of membrane receptor subunits with single-molecule localization microscopy

NASA Astrophysics Data System (ADS)

Krüger, Carmen; Fricke, Franziska; Karathanasis, Christos; Dietz, Marina S.; Malkusch, Sebastian; Hummer, Gerhard; Heilemann, Mike

2017-02-01

We report on quantitative single-molecule localization microscopy, a method that next to super-resolved images of cellular structures provides information on protein copy numbers in protein clusters. This approach is based on the analysis of blinking cycles of single fluorophores, and on a model-free description of the distribution of the number of blinking events. We describe the experimental and analytical procedures, present cellular data of plasma membrane proteins and discuss the applicability of this method.

Intravital FRET: Probing Cellular and Tissue Function in Vivo

PubMed Central

Radbruch, Helena; Bremer, Daniel; Mothes, Ronja; Günther, Robert; Rinnenthal, Jan Leo; Pohlan, Julian; Ulbricht, Carolin; Hauser, Anja E.; Niesner, Raluca

2015-01-01

The development of intravital Förster Resonance Energy Transfer (FRET) is required to probe cellular and tissue function in the natural context: the living organism. Only in this way can biomedicine truly comprehend pathogenesis and develop effective therapeutic strategies. Here we demonstrate and discuss the advantages and pitfalls of two strategies to quantify FRET in vivo—ratiometrically and time-resolved by fluorescence lifetime imaging—and show their concrete application in the context of neuroinflammation in adult mice. PMID:26006244

A liquid chromatography/tandem mass spectrometry assay for the analysis of atomoxetine in human plasma and in vitro cellular samples

PubMed Central

Appel, David I.; Brinda, Bryan; Markowitz, John S.; Newcorn, Jeffrey H.; Zhu, Hao-Jie

2012-01-01

A simple, rapid and sensitive method for quantification of atomoxetine by liquid chromatography- tandem mass spectrometry (LC-MS/MS) was developed. This assay represents the first LC-MS/MS quantification method for atomoxetine utilizing electrospray ionization. Deuterated atomoxetine (d3-atomoxetine) was adopted as the internal standard. Direct protein precipitation was utilized for sample preparation. This method was validated for both human plasma and in vitro cellular samples. The lower limit of quantification was 3 ng/ml and 10 nM for human plasma and cellular samples, respectively. The calibration curves were linear within the ranges of 3 ng/ml to 900 ng/ml and 10 nM to 10 μM for human plasma and cellular samples, respectively (r2 > 0.999). The intra- and inter-day assay accuracy and precision were evaluated using quality control samples at 3 different concentrations in both human plasma and cellular lysate. Sample run stability, assay selectivity, matrix effect, and recovery were also successfully demonstrated. The present assay is superior to previously published LC-MS and LC-MS/MS methods in terms of sensitivity or the simplicity of sample preparation. This assay is applicable to the analysis of atomoxetine in both human plasma and in vitro cellular samples. PMID:22275222

Phospholipase D Signaling Pathways and Phosphatidic Acid as Therapeutic Targets in Cancer

PubMed Central

Bruntz, Ronald C.; Lindsley, Craig W.

2014-01-01

Phospholipase D is a ubiquitous class of enzymes that generates phosphatidic acid as an intracellular signaling species. The phospholipase D superfamily plays a central role in a variety of functions in prokaryotes, viruses, yeast, fungi, plants, and eukaryotic species. In mammalian cells, the pathways modulating catalytic activity involve a variety of cellular signaling components, including G protein–coupled receptors, receptor tyrosine kinases, polyphosphatidylinositol lipids, Ras/Rho/ADP-ribosylation factor GTPases, and conventional isoforms of protein kinase C, among others. Recent findings have shown that phosphatidic acid generated by phospholipase D plays roles in numerous essential cellular functions, such as vesicular trafficking, exocytosis, autophagy, regulation of cellular metabolism, and tumorigenesis. Many of these cellular events are modulated by the actions of phosphatidic acid, and identification of two targets (mammalian target of rapamycin and Akt kinase) has especially highlighted a role for phospholipase D in the regulation of cellular metabolism. Phospholipase D is a regulator of intercellular signaling and metabolic pathways, particularly in cells that are under stress conditions. This review provides a comprehensive overview of the regulation of phospholipase D activity and its modulation of cellular signaling pathways and functions. PMID:25244928

Phospholipase D signaling pathways and phosphatidic acid as therapeutic targets in cancer.

PubMed

Bruntz, Ronald C; Lindsley, Craig W; Brown, H Alex

2014-10-01

Phospholipase D is a ubiquitous class of enzymes that generates phosphatidic acid as an intracellular signaling species. The phospholipase D superfamily plays a central role in a variety of functions in prokaryotes, viruses, yeast, fungi, plants, and eukaryotic species. In mammalian cells, the pathways modulating catalytic activity involve a variety of cellular signaling components, including G protein-coupled receptors, receptor tyrosine kinases, polyphosphatidylinositol lipids, Ras/Rho/ADP-ribosylation factor GTPases, and conventional isoforms of protein kinase C, among others. Recent findings have shown that phosphatidic acid generated by phospholipase D plays roles in numerous essential cellular functions, such as vesicular trafficking, exocytosis, autophagy, regulation of cellular metabolism, and tumorigenesis. Many of these cellular events are modulated by the actions of phosphatidic acid, and identification of two targets (mammalian target of rapamycin and Akt kinase) has especially highlighted a role for phospholipase D in the regulation of cellular metabolism. Phospholipase D is a regulator of intercellular signaling and metabolic pathways, particularly in cells that are under stress conditions. This review provides a comprehensive overview of the regulation of phospholipase D activity and its modulation of cellular signaling pathways and functions. Copyright © 2014 by The American Society for Pharmacology and Experimental Therapeutics.

Tropical Oceanic Precipitation Processes over Warm Pool: 2D and 3D Cloud Resolving Model Simulations

NASA Technical Reports Server (NTRS)

Tao, W.- K.; Johnson, D.

1998-01-01

Rainfall is a key link in the hydrologic cycle as well as the primary heat source for the atmosphere, The vertical distribution of convective latent-heat release modulates the large-scale circulations of the tropics, Furthermore, changes in the moisture distribution at middle and upper levels of the troposphere can affect cloud distributions and cloud liquid water and ice contents. How the incoming solar and outgoing longwave radiation respond to these changes in clouds is a major factor in assessing climate change. Present large-scale weather and climate models simulate cloud processes only crudely, reducing confidence in their predictions on both global and regional scales. One of the most promising methods to test physical parameterizations used in General Circulation Models (GCMS) and climate models is to use field observations together with Cloud Resolving Models (CRMs). The CRMs use more sophisticated and physically realistic parameterizations of cloud microphysical processes, and allow for their complex interactions with solar and infrared radiative transfer processes. The CRMs can reasonably well resolve the evolution, structure, and life cycles of individual clouds and cloud systems, The major objective of this paper is to investigate the latent heating, moisture and momenti,im budgets associated with several convective systems developed during the TOGA COARE IFA - westerly wind burst event (late December, 1992). The tool for this study is the Goddard Cumulus Ensemble (CCE) model which includes a 3-class ice-phase microphysical scheme, The model domain contains 256 x 256 grid points (using 2 km resolution) in the horizontal and 38 grid points (to a depth of 22 km depth) in the vertical, The 2D domain has 1024 grid points. The simulations are performed over a 7 day time period. We will examine (1) the precipitation processes (i.e., condensation/evaporation) and their interaction with warm pool; (2) the heating and moisture budgets in the convective and stratiform regions; (3) the cloud (upward-downward) mass fluxes in convective and stratiform regions; (4) characteristics of clouds (such as cloud size, updraft intensity and cloud lifetime) and the comparison of clouds with Radar observations. Differences and similarities in organization of convection between simulated 2D and 3D cloud systems. Preliminary results indicated that there is major differences between 2D and 3D simulated stratiform rainfall amount and convective updraft and downdraft mass fluxes.

Proteomics: a new approach to the study of disease.

PubMed

Chambers, G; Lawrie, L; Cash, P; Murray, G I

2000-11-01

The global analysis of cellular proteins has recently been termed proteomics and is a key area of research that is developing in the post-genome era. Proteomics uses a combination of sophisticated techniques including two-dimensional (2D) gel electrophoresis, image analysis, mass spectrometry, amino acid sequencing, and bio-informatics to resolve comprehensively, to quantify, and to characterize proteins. The application of proteomics provides major opportunities to elucidate disease mechanisms and to identify new diagnostic markers and therapeutic targets. This review aims to explain briefly the background to proteomics and then to outline proteomic techniques. Applications to the study of human disease conditions ranging from cancer to infectious diseases are reviewed. Finally, possible future advances are briefly considered, especially those which may lead to faster sample throughput and increased sensitivity for the detection of individual proteins. Copyright 2000 John Wiley & Sons, Ltd.

In vivo cellular imaging with microscopes enabled by MEMS scanners

NASA Astrophysics Data System (ADS)

Ra, Hyejun

High-resolution optical imaging plays an important role in medical diagnosis and biomedical research. Confocal microscopy is a widely used imaging method for obtaining cellular and sub-cellular images of biological tissue in reflectance and fluorescence modes. Its characteristic optical sectioning capability also enables three-dimensional (3-D) image reconstruction. However, its use has mostly been limited to excised tissues due to the requirement of high numerical aperture (NA) lenses for cellular resolution. Microscope miniaturization can enable in vivo imaging to make possible early cancer diagnosis and biological studies in the innate environment. In this dissertation, microscope miniaturization for in vivo cellular imaging is presented. The dual-axes confocal (DAC) architecture overcomes limitations of the conventional single-axis confocal (SAC) architecture to allow for miniaturization with high resolution. A microelectromechanical systems (MEMS) scanner is the central imaging component that is key in miniaturization of the DAC architecture. The design, fabrication, and characterization of the two-dimensional (2-D) MEMS scanner are presented. The gimbaled MEMS scanner is fabricated on a double silicon-on-insulator (SOI) wafer and is actuated by self-aligned vertical electrostatic combdrives. The imaging performance of the MEMS scanner in a DAC configuration is shown in a breadboard microscope setup, where reflectance and fluorescence imaging is demonstrated. Then, the MEMS scanner is integrated into a miniature DAC microscope. The whole imaging system is integrated into a portable unit for research in small animal models of human biology and disease. In vivo 3-D imaging is demonstrated on mouse skin models showing gene transfer and siRNA silencing. The siRNA silencing process is sequentially imaged in one mouse over time.

Looking inside the heart: a see-through view of the vascular tree

PubMed Central

Nehrhoff, Imke; Ripoll, Jorge; Samaniego, Rafael; Desco, Manuel; Gómez-Gaviro, Maria Victoria

2017-01-01

The ability to acquire 3D images of the heart and its vasculature at cellular resolution facilitates a more detailed study of many heart diseases. Here, we describe a novel technique to image in 3D the heart vasculature by combining the CUBIC clearing protocol combined with in vivo administration of fluorescent-labeled lectin. The use of these techniques in combination with Selective Plane Illumination Microscopy (SPIM) made it possible to obtain high resolution 3D images of the cardiac vascular tree. This methodological approach may enhance the visualization of 3D images of the cardiac vasculature remodeling associated with coronary disease. PMID:28663930

The development of a virtual 3D model of the renal corpuscle from serial histological sections for E-learning environments.

PubMed

Roth, Jeremy A; Wilson, Timothy D; Sandig, Martin

2015-01-01

Histology is a core subject in the anatomical sciences where learners are challenged to interpret two-dimensional (2D) information (gained from histological sections) to extrapolate and understand the three-dimensional (3D) morphology of cells, tissues, and organs. In gross anatomical education 3D models and learning tools have been associated with improved learning outcomes, but similar tools have not been created for histology education to visualize complex cellular structure-function relationships. This study outlines steps in creating a virtual 3D model of the renal corpuscle from serial, semi-thin, histological sections obtained from epoxy resin-embedded kidney tissue. The virtual renal corpuscle model was generated by digital segmentation to identify: Bowman's capsule, nuclei of epithelial cells in the parietal capsule, afferent arteriole, efferent arteriole, proximal convoluted tubule, distal convoluted tubule, glomerular capillaries, podocyte nuclei, nuclei of extraglomerular mesangial cells, nuclei of epithelial cells of the macula densa in the distal convoluted tubule. In addition to the imported images of the original sections the software generates, and allows for visualization of, images of virtual sections generated in any desired orientation, thus serving as a "virtual microtome". These sections can be viewed separately or with the 3D model in transparency. This approach allows for the development of interactive e-learning tools designed to enhance histology education of microscopic structures with complex cellular interrelationships. Future studies will focus on testing the efficacy of interactive virtual 3D models for histology education. © 2015 American Association of Anatomists.

Asymmetric cellular memory in bacteria exposed to antibiotics.

PubMed

Mathis, Roland; Ackermann, Martin

2017-03-09

The ability to form a cellular memory and use it for cellular decision-making could help bacteria to cope with recurrent stress conditions. We analyzed whether bacteria would form a cellular memory specifically if past events are predictive of future conditions. We worked with the asymmetrically dividing bacterium Caulobacter crescentus where past events are expected to only be informative for one of the two cells emerging from division, the sessile cell that remains in the same microenvironment and does not migrate. Time-resolved analysis of individual cells revealed that past exposure to low levels of antibiotics increases tolerance to future exposure for the sessile but not for the motile cell. Using computer simulations, we found that such an asymmetry in cellular memory could be an evolutionary response to situations where the two cells emerging from division will experience different future conditions. Our results raise the question whether bacteria can evolve the ability to form and use cellular memory conditionally in situations where it is beneficial.

Prospective Out-of-ecliptic White-light Imaging of Interplanetary Corotating Interaction Regions at Solar Maximum

NASA Astrophysics Data System (ADS)

Xiong, Ming; Davies, Jackie A.; Li, Bo; Yang, Liping; Liu, Ying D.; Xia, Lidong; Harrison, Richard A.; Keiji, Hayashi; Li, Huichao

2017-07-01

Interplanetary corotating interaction regions (CIRs) can be remotely imaged in white light (WL), as demonstrated by the Solar Mass Ejection Imager (SMEI) on board the Coriolis spacecraft and Heliospheric Imagers (HIs) on board the twin Solar TErrestrial RElations Observatory (STEREO) spacecraft. The interplanetary WL intensity, due to Thomson scattering of incident sunlight by free electrons, is jointly determined by the 3D distribution of electron number density and line-of-sight (LOS) weighting factors of the Thomson-scattering geometry. The 2D radiance patterns of CIRs in WL sky maps look very different from different 3D viewpoints. Because of the in-ecliptic locations of both the STEREO and Coriolis spacecraft, the longitudinal dimension of interplanetary CIRs has, up to now, always been integrated in WL imagery. To synthesize the WL radiance patterns of CIRs from an out-of-ecliptic (OOE) vantage point, we perform forward magnetohydrodynamic modeling of the 3D inner heliosphere during Carrington Rotation CR1967 at solar maximum. The mixing effects associated with viewing 3D CIRs are significantly minimized from an OOE viewpoint. Our forward modeling results demonstrate that OOE WL imaging from a latitude greater than 60° can (1) enable the garden-hose spiral morphology of CIRs to be readily resolved, (2) enable multiple coexisting CIRs to be differentiated, and (3) enable the continuous tracing of any interplanetary CIR back toward its coronal source. In particular, an OOE view in WL can reveal where nascent CIRs are formed in the extended corona and how these CIRs develop in interplanetary space. Therefore, a panoramic view from a suite of wide-field WL imagers in a solar polar orbit would be invaluable in unambiguously resolving the large-scale longitudinal structure of CIRs in the 3D inner heliosphere.

Meta!Blast computer game: a pipeline from science to 3D art to education

NASA Astrophysics Data System (ADS)

Schneller, William; Campbell, P. J.; Bassham, Diane; Wurtele, Eve Syrkin

2012-03-01

Meta!Blast (http://www.metablast.org) is designed to address the challenges students often encounter in understanding cell and metabolic biology. Developed by faculty and students in biology, biochemistry, computer science, game design, pedagogy, art and story, Meta!Blast is being created using Maya (http://usa.autodesk.com/maya/) and the Unity 3D (http://unity3d.com/) game engine, for Macs and PCs in classrooms; it has also been exhibited in an immersive environment. Here, we describe the pipeline from protein structural data and holographic information to art to the threedimensional (3D) environment to the game engine, by which we provide a publicly-available interactive 3D cellular world that mimics a photosynthetic plant cell.

Dual-color 3D superresolution microscopy by combined spectral-demixing and biplane imaging.

PubMed

Winterflood, Christian M; Platonova, Evgenia; Albrecht, David; Ewers, Helge

2015-07-07

Multicolor three-dimensional (3D) superresolution techniques allow important insight into the relative organization of cellular structures. While a number of innovative solutions have emerged, multicolor 3D techniques still face significant technical challenges. In this Letter we provide a straightforward approach to single-molecule localization microscopy imaging in three dimensions and two colors. We combine biplane imaging and spectral-demixing, which eliminates a number of problems, including color cross-talk, chromatic aberration effects, and problems with color registration. We present 3D dual-color images of nanoscopic structures in hippocampal neurons with a 3D compound resolution routinely achieved only in a single color. Copyright © 2015 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Attosecond sublevel beating and nonlinear dressing on the 3d-to-5p and 3p-to-5s core-transitions at 91.3 eV and 210.4 eV in krypton.

PubMed

Seres, Enikoe; Seres, Jozsef; Namba, Shinichi; Afa, John; Serrat, Carles

2017-12-11

Applying extreme ultraviolet (XUV) transient absorption spectroscopy, the dynamics of the two laser dressed transitions 3d 5/2 -to-5p 3/2 and 3p 3/2 -to-5s 1/2 at photon energies of 91.3 eV and 210.4 eV were examined with attosecond temporal resolution. The dressing process was modeled with density matrix equations which are found to describe very accurately both the experimentally observed transmission dynamics and the linear and nonlinear dressing oscillations at 0.75 PHz and 1.5 PHz frequencies. Furthermore, using Fourier transform XUV spectroscopy, quantum beats from the 3d 5/2 -3d 3/2 and 3p 3/2 -3p 1/2 sublevels at 0.3 PHz and 2.0 PHz were experimentally identified and resolved.

A single residue controls electron transfer gating in photosynthetic reaction centers

NASA Astrophysics Data System (ADS)

Shlyk, Oksana; Samish, Ilan; Matěnová, Martina; Dulebo, Alexander; Poláková, Helena; Kaftan, David; Scherz, Avigdor

2017-03-01

Interquinone QA- → QB electron-transfer (ET) in isolated photosystem II reaction centers (PSII-RC) is protein-gated. The temperature-dependent gating frequency “k” is described by the Eyring equation till levelling off at T ≥ 240 °K. Although central to photosynthesis, the gating mechanism has not been resolved and due to experimental limitations, could not be explored in vivo. Here we mimic the temperature dependency of “k” by enlarging VD1-208, the volume of a single residue at the crossing point of the D1 and D2 PSII-RC subunits in Synechocystis 6803 whole cells. By controlling the interactions of the D1/D2 subunits, VD1-208 (or 1/T) determines the frequency of attaining an ET-active conformation. Decelerated ET, impaired photosynthesis, D1 repair rate and overall cell physiology upon increasing VD1-208 to above 130 Å3, rationalize the >99% conservation of small residues at D1-208 and its homologous motif in non-oxygenic bacteria. The experimental means and resolved mechanism are relevant for numerous transmembrane protein-gated reactions.

Integrating Intracellular Dynamics Using CompuCell3D and Bionetsolver: Applications to Multiscale Modelling of Cancer Cell Growth and Invasion

PubMed Central

Andasari, Vivi; Roper, Ryan T.; Swat, Maciej H.; Chaplain, Mark A. J.

2012-01-01

In this paper we present a multiscale, individual-based simulation environment that integrates CompuCell3D for lattice-based modelling on the cellular level and Bionetsolver for intracellular modelling. CompuCell3D or CC3D provides an implementation of the lattice-based Cellular Potts Model or CPM (also known as the Glazier-Graner-Hogeweg or GGH model) and a Monte Carlo method based on the metropolis algorithm for system evolution. The integration of CC3D for cellular systems with Bionetsolver for subcellular systems enables us to develop a multiscale mathematical model and to study the evolution of cell behaviour due to the dynamics inside of the cells, capturing aspects of cell behaviour and interaction that is not possible using continuum approaches. We then apply this multiscale modelling technique to a model of cancer growth and invasion, based on a previously published model of Ramis-Conde et al. (2008) where individual cell behaviour is driven by a molecular network describing the dynamics of E-cadherin and -catenin. In this model, which we refer to as the centre-based model, an alternative individual-based modelling technique was used, namely, a lattice-free approach. In many respects, the GGH or CPM methodology and the approach of the centre-based model have the same overall goal, that is to mimic behaviours and interactions of biological cells. Although the mathematical foundations and computational implementations of the two approaches are very different, the results of the presented simulations are compatible with each other, suggesting that by using individual-based approaches we can formulate a natural way of describing complex multi-cell, multiscale models. The ability to easily reproduce results of one modelling approach using an alternative approach is also essential from a model cross-validation standpoint and also helps to identify any modelling artefacts specific to a given computational approach. PMID:22461894

Journey to the centre of the cell: Virtual reality immersion into scientific data.

PubMed

Johnston, Angus P R; Rae, James; Ariotti, Nicholas; Bailey, Benjamin; Lilja, Andrew; Webb, Robyn; Ferguson, Charles; Maher, Sheryl; Davis, Thomas P; Webb, Richard I; McGhee, John; Parton, Robert G

2018-02-01

Visualization of scientific data is crucial not only for scientific discovery but also to communicate science and medicine to both experts and a general audience. Until recently, we have been limited to visualizing the three-dimensional (3D) world of biology in 2 dimensions. Renderings of 3D cells are still traditionally displayed using two-dimensional (2D) media, such as on a computer screen or paper. However, the advent of consumer grade virtual reality (VR) headsets such as Oculus Rift and HTC Vive means it is now possible to visualize and interact with scientific data in a 3D virtual world. In addition, new microscopic methods provide an unprecedented opportunity to obtain new 3D data sets. In this perspective article, we highlight how we have used cutting edge imaging techniques to build a 3D virtual model of a cell from serial block-face scanning electron microscope (SBEM) imaging data. This model allows scientists, students and members of the public to explore and interact with a "real" cell. Early testing of this immersive environment indicates a significant improvement in students' understanding of cellular processes and points to a new future of learning and public engagement. In addition, we speculate that VR can become a new tool for researchers studying cellular architecture and processes by populating VR models with molecular data. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Investigating the interplay between substrate stiffness and ligand chemistry in directing mesenchymal stem cell differentiation within 3D macro-porous substrates.

PubMed

Haugh, Matthew G; Vaughan, Ted J; Madl, Christopher M; Raftery, Rosanne M; McNamara, Laoise M; O'Brien, Fergal J; Heilshorn, Sarah C

2018-07-01

Dimensionality can have a profound impact on stiffness-mediated differentiation of mesenchymal stem cells (MSCs). However, while we have begun to understand cellular response when encapsulated within 3D substrates, the behavior of cells within macro-porous substrates is relatively underexplored. The goal of this study was to determine the influence of macro-porous topographies on stiffness-mediated differentiation of MSCs. We developed macro-porous recombinant elastin-like protein (ELP) substrates that allow independent control of mechanical properties and ligand chemistry. We then used computational modeling to probe the impact of pore topography on the mechanical stimulus that cells are exposed to within these substrates, and finally we investigated stiffness induced biases towards adipogenic and osteogenic differentiation of MSCs within macro-porous substrates. Computational modeling revealed that there is significant heterogeneity in the mechanical stimuli that cells are exposed to within porous substrates and that this heterogeneity is predominantly due to the wide range of possible cellular orientations within the pores. Surprisingly, MSCs grown within 3D porous substrates respond to increasing substrate stiffness by up-regulating both osteogenesis and adipogenesis. These results demonstrate that within porous substrates the behavior of MSCs diverges from previously observed responses to substrate stiffness, emphasizing the importance of topography as a determinant of cellular behavior. Copyright © 2018 Elsevier Ltd. All rights reserved.

Effects of silica and zinc oxide doping on mechanical and biological properties of 3D printed tricalcium phosphate tissue engineering scaffolds.

PubMed

Fielding, Gary A; Bandyopadhyay, Amit; Bose, Susmita

2012-02-01

To evaluate the effects of silica (SiO(2)) (0.5 wt%) and zinc oxide (ZnO) (0.25 wt%) dopants on the mechanical and biological properties of tricalcium phosphate (TCP) scaffolds with three dimensionally (3D) interconnected pores. Scaffolds were created with a commercial 3D printer. Post sintering phase analysis was determined by X-ray diffraction. Surface morphology of the scaffolds was examined by field emission scanning electron microscopy (FESEM). Mechanical strength was evaluated with a screw driven universal testing machine. MTT assay was used for cellular proliferation characteristics and cellular morphology was examined by FESEM. Addition of dopants into TCP increased the average density of pure TCP from 90.8 ± 0.8% to 94.1 ± 1.6% and retarded the β to α phase transformation at high sintering temperatures, which resulted in up to 2.5 fold increase in compressive strength. In vitro cell-materials interaction studies, carried out using hFOB cells, confirmed that the addition of SiO(2) and ZnO to the scaffolds facilitated faster cell proliferation when compared to pure TCP scaffolds. Addition of SiO(2) and ZnO dopants to the TCP scaffolds showed increased mechanical strength as well as increased cellular proliferation. Copyright © 2011 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.

The carbocyanine dye DiD labels in vitro and in vivo neural stem cells of the subventricular zone as well as myelinated structures following in vivo injection in the lateral ventricle.

PubMed

Carradori, Dario; Barreau, Kristell; Eyer, Joël

2016-02-01

Carbocyanines are fluorescent lipophilic cationic dyes used since the early 1980s as neuronal tracers. Several applications of these compounds have been developed thanks to their low cell toxicity, lateral diffusion within the cellular membranes, and good photostability. 1,1'-Dioctadecyl-3,3,3',3'-tetramethylindodicarbocyanine 4-chlorobenzenesulfonate (DiD) is an interesting component of this family because, in addition to the classic carbocyanine properties, it has a longer wavelength compared with its analogues. That makes DiD an excellent carbocyanine for labeling cells and tissues with significant intrinsic fluorescence. Drug encapsulation, drug delivery, and cellular transplantation are also fields using DiD-based systems where having detailed knowledge about its behavior as a single entity is important. Recently, promising studies concerned neural stem cells from the subventricular zone of the lateral ventricle in the brain (their natural niche) and their potential therapeutic use. Here, we show that DiD is able to label these stem cells in vitro and present basilar information concerning its pharmacokinetics, concentrations, and microscope protocols. Moreover, when DiD is injected in vivo in the cerebrospinal fluid present in the lateral ventricle of rat, it also labels stem cells as well as myelinated structures of the caudoputamen. This analysis provides a database to consult when planning experiments concerning DiD and neural stem cells from the subventricular zone. © 2015 Wiley Periodicals, Inc.

Development and Optimization of Viable Human Platforms through 3D Printing

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

Parker, Paul R.; Moya, Monica L.; Wheeler, Elizabeth K.

2015-08-21

3D printing technology offers a unique method for creating cell cultures in a manner far more conducive to accurate representation of human tissues and systems. Here we print cellular structures capable of forming vascular networks and exhibiting qualities of natural tissues and human systems. This allows for cheaper and readily available sources for further study of biological and pharmaceutical agents.

Three-dimensional ghost imaging lidar via sparsity constraint

NASA Astrophysics Data System (ADS)

Gong, Wenlin; Zhao, Chengqiang; Yu, Hong; Chen, Mingliang; Xu, Wendong; Han, Shensheng

2016-05-01

Three-dimensional (3D) remote imaging attracts increasing attentions in capturing a target’s characteristics. Although great progress for 3D remote imaging has been made with methods such as scanning imaging lidar and pulsed floodlight-illumination imaging lidar, either the detection range or application mode are limited by present methods. Ghost imaging via sparsity constraint (GISC), enables the reconstruction of a two-dimensional N-pixel image from much fewer than N measurements. By GISC technique and the depth information of targets captured with time-resolved measurements, we report a 3D GISC lidar system and experimentally show that a 3D scene at about 1.0 km range can be stably reconstructed with global measurements even below the Nyquist limit. Compared with existing 3D optical imaging methods, 3D GISC has the capability of both high efficiency in information extraction and high sensitivity in detection. This approach can be generalized in nonvisible wavebands and applied to other 3D imaging areas.

Bioinspired Cellular Structures: Additive Manufacturing and Mechanical Properties

NASA Astrophysics Data System (ADS)

Stampfl, J.; Pettermann, H. E.; Liska, R.

Biological materials (e.g., wood, trabecular bone, marine skeletons) rely heavily on the use of cellular architecture, which provides several advantages. (1) The resulting structures can bear the variety of "real life" load spectra using a minimum of a given bulk material, featuring engineering lightweight design principles. (2) The inside of the structures is accessible to body fluids which deliver the required nutrients. (3) Furthermore, cellular architectures can grow organically by adding or removing individual struts or by changing the shape of the constituting elements. All these facts make the use of cellular architectures a reasonable choice for nature. Using additive manufacturing technologies (AMT), it is now possible to fabricate such structures for applications in engineering and biomedicine. In this chapter, we present methods that allow the 3D computational analysis of the mechanical properties of cellular structures with open porosity. Various different cellular architectures including disorder are studied. In order to quantify the influence of architecture, the apparent density is always kept constant. Furthermore, it is shown that how new advanced photopolymers can be used to tailor the mechanical and functional properties of the fabricated structures.

Elaboration and Characterization of Nano-Sized AlxMoyOz/Al Thermites

DTIC Science & Technology

2006-07-01

corresponding micrographs are not the experiments of time resolved shown here). In the Th. 1 material, the cinematography . micron-sized MoO 3...necessary to know cinelnalogra/)hy andspec’troscot)y- the effective power (P) delivered by the laser The time resolved cinematography (TRC) is source as...necessary to initiate the film paper ((D P 1.4 amm). The combustion rate 8 PU 614/2006 (Vr) is measured by ultra fast cinematography the lower are the

Multi-scale mechanics of granular solids from grain-resolved X-ray measurements

NASA Astrophysics Data System (ADS)

Hurley, R. C.; Hall, S. A.; Wright, J. P.

2017-11-01

This work discusses an experimental technique for studying the mechanics of three-dimensional (3D) granular solids. The approach combines 3D X-ray diffraction and X-ray computed tomography to measure grain-resolved strains, kinematics and contact fabric in the bulk of a granular solid, from which continuum strains, grain stresses, interparticle forces and coarse-grained elasto-plastic moduli can be determined. We demonstrate the experimental approach and analysis of selected results on a sample of 1099 stiff, frictional grains undergoing multiple uniaxial compression cycles. We investigate the inter-particle force network, elasto-plastic moduli and associated length scales, reversibility of mechanical responses during cyclic loading, the statistics of microscopic responses and microstructure-property relationships. This work serves to highlight both the fundamental insight into granular mechanics that is furnished by combined X-ray measurements and describes future directions in the field of granular materials that can be pursued with such approaches.

Three-dimensional turbulence-resolving modeling of the Venusian cloud layer and induced gravity waves

NASA Astrophysics Data System (ADS)

Lefèvre, Maxence; Spiga, Aymeric; Lebonnois, Sébastien

2017-01-01

The impact of the cloud convective layer of the atmosphere of Venus on the global circulation remains unclear. The recent observations of gravity waves at the top of the cloud by the Venus Express mission provided some answers. These waves are not resolved at the scale of global circulation models (GCM); therefore, we developed an unprecedented 3-D turbulence-resolving large-eddy simulations (LES) Venusian model using the Weather Research and Forecast terrestrial model. The forcing consists of three different heating rates: two radiative ones for solar and infrared and one associated with the adiabatic cooling/warming of the global circulation. The rates are extracted from the Laboratoire de Météorlogie Dynamique Venus GCM using two different cloud models. Thus, we are able to characterize the convection and associated gravity waves in function of latitude and local time. To assess the impact of the global circulation on the convective layer, we used rates from a 1-D radiative-convective model. The resolved layer, taking place between 1.0 × 105 and 3.8 × 104 Pa (48-53 km), is organized as polygonal closed cells of about 10 km wide with vertical wind of several meters per second. The convection emits gravity waves both above and below the convective layer leading to temperature perturbations of several tenths of kelvin with vertical wavelength between 1 and 3 km and horizontal wavelength from 1 to 10 km. The thickness of the convective layer and the amplitudes of waves are consistent with observations, though slightly underestimated. The global dynamics heating greatly modify the convective layer.

The ECM moves during primitive streak formation--computation of ECM versus cellular motion.

PubMed

Zamir, Evan A; Rongish, Brenda J; Little, Charles D

2008-10-14

Galileo described the concept of motion relativity--motion with respect to a reference frame--in 1632. He noted that a person below deck would be unable to discern whether the boat was moving. Embryologists, while recognizing that embryonic tissues undergo large-scale deformations, have failed to account for relative motion when analyzing cell motility data. A century of scientific articles has advanced the concept that embryonic cells move ("migrate") in an autonomous fashion such that, as time progresses, the cells and their progeny assemble an embryo. In sharp contrast, the motion of the surrounding extracellular matrix scaffold has been largely ignored/overlooked. We developed computational/optical methods that measure the extent embryonic cells move relative to the extracellular matrix. Our time-lapse data show that epiblastic cells largely move in concert with a sub-epiblastic extracellular matrix during stages 2 and 3 in primitive streak quail embryos. In other words, there is little cellular motion relative to the extracellular matrix scaffold--both components move together as a tissue. The extracellular matrix displacements exhibit bilateral vortical motion, convergence to the midline, and extension along the presumptive vertebral axis--all patterns previously attributed solely to cellular "migration." Our time-resolved data pose new challenges for understanding how extracellular chemical (morphogen) gradients, widely hypothesized to guide cellular trajectories at early gastrulation stages, are maintained in this dynamic extracellular environment. We conclude that models describing primitive streak cellular guidance mechanisms must be able to account for sub-epiblastic extracellular matrix displacements.

Engineering 3D Models of Tumors and Bone to Understand Tumor-Induced Bone Disease and Improve Treatments.

PubMed

Kwakwa, Kristin A; Vanderburgh, Joseph P; Guelcher, Scott A; Sterling, Julie A

2017-08-01

Bone is a structurally unique microenvironment that presents many challenges for the development of 3D models for studying bone physiology and diseases, including cancer. As researchers continue to investigate the interactions within the bone microenvironment, the development of 3D models of bone has become critical. 3D models have been developed that replicate some properties of bone, but have not fully reproduced the complex structural and cellular composition of the bone microenvironment. This review will discuss 3D models including polyurethane, silk, and collagen scaffolds that have been developed to study tumor-induced bone disease. In addition, we discuss 3D printing techniques used to better replicate the structure of bone. 3D models that better replicate the bone microenvironment will help researchers better understand the dynamic interactions between tumors and the bone microenvironment, ultimately leading to better models for testing therapeutics and predicting patient outcomes.

Detailed Characterization of a Nanosecond-Lived Excited State: X-ray and Theoretical Investigation of the Quintet State in Photoexcited [Fe(terpy) 2 ] 2+

DOE PAGES

Vanko, Gyorgy; Bordage, Amelie; Papai, Matyas; ...

2015-03-19

Theoretical predictions show that depending on the populations of the Fe 3d xy, 3d xz, and 3d yz orbitals two possible quintet states can exist for the high-spin state of the photoswitchable model system [Fe(terpy) 2] 2+. The differences in the structure and molecular properties of these 5B2 and 5E quintets are very small and pose a substantial challenge for experiments to resolve them. Yet for a better understanding of the physics of this system, which can lead to the design of novel molecules with enhanced photoswitching performance, it is vital to determine which high-spin state is reached in themore » transitions that follow the light excitation. The quintet state can be prepared with a short laser pulse and can be studied with cutting-edge time-resolved X-ray techniques. Here we report on the application of an extended set of X-ray spectroscopy and scattering techniques applied to investigate the quintet state of [Fe(terpy) 2] 2+ 80 ps after light excitation. High-quality X-ray absorption, nonresonant emission, and resonant emission spectra as well as X-ray diffuse scattering data clearly reflect the formation of the high-spin state of the [Fe(terpy) 2] 2+ molecule; moreover, extended X-ray absorption fine structure spectroscopy resolves the Fe–ligand bond-length variations with unprecedented bond-length accuracy in time-resolved experiments. With ab initio calculations we determine why, in contrast to most related systems, one configurational mode is insufficient for the description of the low-spin (LS)–high-spin (HS) transition. We identify the electronic structure origin of the differences between the two possible quintet modes, and finally, we unambiguously identify the formed quintet state as 5E, in agreement with our theoretical expectations.« less

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

Bailey, J.E.; Adams, R.; Carlson, A.L.

Stark-shift measurements using emission spectroscopy are a powerful tool for advancing understanding in many plasma physics experiments. The authors use simultaneous 2-D-spatial and time-resolved spectra to study the electric field evolution in the 20 TW Particle Beam Fusion Accelerator II ion diode acceleration gap. Fiber optic arrays transport light from the gap to remote streaked spectrographs operated in a multiplexed mode that enables recording time-resolved spectra from eight spatial locations on a single instrument. Design optimization and characterization measurements of the multiplexed spectrograph properties include the astigmatism, resolution, dispersion variation, and sensitivity. A semi-automated line-fitting procedure determines the Stark shiftmore » and the related uncertainties. Fields up to 10 MV/cm are measured with an accuracy {+-}2--4%. Detailed tests of the fitting procedure confirm that the wavelength shift uncertainties are accurate to better than {+-}20%. Development of an active spectroscopy probe technique that uses laser-induced fluorescence from an injected atomic beam to obtain 3-D space- and time-resolved measurements of the electric and magnetic fields is in progress.« less

Progress in the Development of a Global Quasi-3-D Multiscale Modeling Framework

NASA Astrophysics Data System (ADS)

Jung, J.; Konor, C. S.; Randall, D. A.

2017-12-01

The Quasi-3-D Multiscale Modeling Framework (Q3D MMF) is a second-generation MMF, which has following advances over the first-generation MMF: 1) The cloud-resolving models (CRMs) that replace conventional parameterizations are not confined to the large-scale dynamical-core grid cells, and are seamlessly connected to each other, 2) The CRMs sense the three-dimensional large- and cloud-scale environment, 3) Two perpendicular sets of CRM channels are used, and 4) The CRMs can resolve the steep surface topography along the channel direction. The basic design of the Q3D MMF has been developed and successfully tested in a limited-area modeling framework. Currently, global versions of the Q3D MMF are being developed for both weather and climate applications. The dynamical cores governing the large-scale circulation in the global Q3D MMF are selected from two cube-based global atmospheric models. The CRM used in the model is the 3-D nonhydrostatic anelastic Vector-Vorticity Model (VVM), which has been tested with the limited-area version for its suitability for this framework. As a first step of the development, the VVM has been reconstructed on the cubed-sphere grid so that it can be applied to global channel domains and also easily fitted to the large-scale dynamical cores. We have successfully tested the new VVM by advecting a bell-shaped passive tracer and simulating the evolutions of waves resulted from idealized barotropic and baroclinic instabilities. For improvement of the model, we also modified the tracer advection scheme to yield positive-definite results and plan to implement a new physics package that includes a double-moment microphysics and an aerosol physics. The interface for coupling the large-scale dynamical core and the VVM is under development. In this presentation, we shall describe the recent progress in the development and show some test results.

Three-Dimensional Terahertz Coded-Aperture Imaging Based on Matched Filtering and Convolutional Neural Network.

PubMed

Chen, Shuo; Luo, Chenggao; Wang, Hongqiang; Deng, Bin; Cheng, Yongqiang; Zhuang, Zhaowen

2018-04-26

As a promising radar imaging technique, terahertz coded-aperture imaging (TCAI) can achieve high-resolution, forward-looking, and staring imaging by producing spatiotemporal independent signals with coded apertures. However, there are still two problems in three-dimensional (3D) TCAI. Firstly, the large-scale reference-signal matrix based on meshing the 3D imaging area creates a heavy computational burden, thus leading to unsatisfactory efficiency. Secondly, it is difficult to resolve the target under low signal-to-noise ratio (SNR). In this paper, we propose a 3D imaging method based on matched filtering (MF) and convolutional neural network (CNN), which can reduce the computational burden and achieve high-resolution imaging for low SNR targets. In terms of the frequency-hopping (FH) signal, the original echo is processed with MF. By extracting the processed echo in different spike pulses separately, targets in different imaging planes are reconstructed simultaneously to decompose the global computational complexity, and then are synthesized together to reconstruct the 3D target. Based on the conventional TCAI model, we deduce and build a new TCAI model based on MF. Furthermore, the convolutional neural network (CNN) is designed to teach the MF-TCAI how to reconstruct the low SNR target better. The experimental results demonstrate that the MF-TCAI achieves impressive performance on imaging ability and efficiency under low SNR. Moreover, the MF-TCAI has learned to better resolve the low-SNR 3D target with the help of CNN. In summary, the proposed 3D TCAI can achieve: (1) low-SNR high-resolution imaging by using MF; (2) efficient 3D imaging by downsizing the large-scale reference-signal matrix; and (3) intelligent imaging with CNN. Therefore, the TCAI based on MF and CNN has great potential in applications such as security screening, nondestructive detection, medical diagnosis, etc.

Division of labor by dual feedback regulators controls JAK2/STAT5 signaling over broad ligand range.

PubMed

Bachmann, Julie; Raue, Andreas; Schilling, Marcel; Böhm, Martin E; Kreutz, Clemens; Kaschek, Daniel; Busch, Hauke; Gretz, Norbert; Lehmann, Wolf D; Timmer, Jens; Klingmüller, Ursula

2011-07-19

Cellular signal transduction is governed by multiple feedback mechanisms to elicit robust cellular decisions. The specific contributions of individual feedback regulators, however, remain unclear. Based on extensive time-resolved data sets in primary erythroid progenitor cells, we established a dynamic pathway model to dissect the roles of the two transcriptional negative feedback regulators of the suppressor of cytokine signaling (SOCS) family, CIS and SOCS3, in JAK2/STAT5 signaling. Facilitated by the model, we calculated the STAT5 response for experimentally unobservable Epo concentrations and provide a quantitative link between cell survival and the integrated response of STAT5 in the nucleus. Model predictions show that the two feedbacks CIS and SOCS3 are most effective at different ligand concentration ranges due to their distinct inhibitory mechanisms. This divided function of dual feedback regulation enables control of STAT5 responses for Epo concentrations that can vary 1000-fold in vivo. Our modeling approach reveals dose-dependent feedback control as key property to regulate STAT5-mediated survival decisions over a broad range of ligand concentrations.

Role of intercellular communications in breast cancer multicellular tumor spheroids after chemotherapy.

PubMed

Oktem, G; Bilir, A; Ayla, S; Yavasoglu, A; Goksel, G; Saydam, G; Uysal, A

2006-01-01

Tumor heterogeneity is an important feature that is especially involved in tumor aggressiveness. Multicellular tumor spheroids (MTS) may provide some benefits in different steps for investigation of the aggregation, organization, differentiation, and network formation of tumor cells in 3D space. This model offers a unique opportunity for improvements in the capability of a current strategy to detect the effect of an appropriate anticancer agent. The aim of this study was to investigate the cellular interactions and morphological changes following chemotherapy in a 3D breast cancer spheroid model. Distribution of the gap junction protein "connexin-43" and the tight junction protein "occludin" was investigated by immunohistochemistry. Cellular interactions were examined by using transmission and scanning electron microscopies as well as light microscopy with Giemsa staining after treating cells with doxorubicin, docetaxel, and doxorubicin/docetaxel combination. Statistical analyses showed significant changes and various alterations that were observed in all groups; however, the most prominent effect was detected in the doxorubicin/docetaxel combination group. Distinct composition as a vessel-like structure and a pseudoglandular pattern of control spheroids were detected in drug-administered groups. Immunohistochemical results were consistent with the ultrastructural changes. In conclusion, doxorubicin/docetaxel combination may be more effective than the single drug usage as shown in a 3D model. The MTS model has been found to be an appropriate and reliable method for the detection of the changes in the expression of cellular junction proteins as well as other cellular proteins occurring after chemotherapy. The MTS model can be used to validate the effects of various combinations or new chemotherapeutic agents as well as documentation of possible mechanisms of new drugs.

Excitation-dependent carrier lifetime and diffusion length in bulk CdTe determined by time-resolved optical pump-probe techniques

NASA Astrophysics Data System (ADS)

Ščajev, Patrik; Miasojedovas, Saulius; Mekys, Algirdas; Kuciauskas, Darius; Lynn, Kelvin G.; Swain, Santosh K.; JarašiÅ«nas, Kestutis

2018-01-01

We applied time-resolved pump-probe spectroscopy based on free carrier absorption and light diffraction on a transient grating for direct measurements of the carrier lifetime and diffusion coefficient D in high-resistivity single crystal CdTe (codoped with In and Er). The bulk carrier lifetime τ decreased from 670 ± 50 ns to 60 ± 10 ns with increase of excess carrier density N from 1016 to 5 × 1018 cm-3 due to the excitation-dependent radiative recombination rate. In this N range, the carrier diffusion length dropped from 14 μm to 6 μm due to lifetime decrease. Modeling of in-depth (axial) and in-plane (lateral) carrier diffusion provided the value of surface recombination velocity S = 6 × 105 cm/s for the untreated surface. At even higher excitations, in the 1019-3 × 1020 cm-3 density range, D increase from 5 to 20 cm2/s due to carrier degeneracy was observed.

Chronic administration of aripiprazole activates GSK3β-dependent signalling pathways, and up-regulates GABAA receptor expression and CREB1 activity in rats.

PubMed

Pan, Bo; Huang, Xu-Feng; Deng, Chao

2016-07-20

Aripiprazole is a D2-like receptor (D2R) partial agonist with a favourable clinical profile. Previous investigations indicated that acute and short-term administration of aripiprazole had effects on PKA activity, GSK3β-dependent pathways, GABAA receptors, NMDA receptor and CREB1 in the brain. Since antipsychotics are used chronically in clinics, the present study investigated the long-term effects of chronic oral aripiprazole treatment on these cellular signalling pathways, in comparison with haloperidol (a D2R antagonist) and bifeprunox (a potent D2R partial agonist). We found that the Akt-GSK3β pathway was activated by aripiprazole and bifeprunox in the prefrontal cortex; NMDA NR2A levels were reduced by aripiprazole and haloperidol. In the nucleus accumbens, all three drugs increased Akt-GSK3β signalling; in addition, both aripiprazole and haloperidol, but not bifeprunox, increased the expression of Dvl-3, β-catenin and GABAA receptors, NMDA receptor subunits, as well as CREB1 phosphorylation levels. The results suggest that chronic oral administration of aripiprazole affects schizophrenia-related cellular signalling pathways and markers (including Akt-GSK3β signalling, Dvl-GSK3β-β-catenin signalling, GABAA receptor, NMDA receptor and CREB1) in a brain-region-dependent manner; the selective effects of aripiprazole on these signalling pathways might be associated with its unique clinical effects.

VizieR Online Data Catalog: Investigating Tully-Fisher relation with KMOS3D (Ubler+,

NASA Astrophysics Data System (ADS)

Ubler, H.; Forster Schreiber, N. M.; Genzel, R.; Wisnioski, E.; Wuyts, S.; Lang, P.; Naab, T.; Burkert, A.; van Dokkum, P. G.; Tacconi, L. J.; Wilman, D. J.; Fossati, M.; Mendel, J. T.; Beifiori, A.; Belli, S.; Bender, R.; Brammer, G. B.; Chan, J.; Davies, R.; Fabricius, M.; Galametz, A.; Lutz, D.; Momcheva, I. G.; Nelson, E. J.; Saglia, R. P.; Seitz, S.; Tadaki, K.

2018-02-01

This work is based on the first 3yr of observations of KMOS3D multiyear near-infrared (near-IR) IFS survey of more than 600 mass-selected star-forming galaxies (SFGs) at 0.6<~z<~2.6 with the K-band Multi Object Spectrograph (KMOS; Sharples+ 2013Msngr.151...21S) on the Very Large Telescope. The KMOS3D survey and data reduction are described in detail by Wisnioski et al. 2015ApJ...799..209W The results presented in this paper build on the KMOS3D sample as of 2016 January, with 536 observed galaxies. Of these, 316 are detected in, and have spatially resolved, Hα emission free from skyline contamination from which two-dimensional velocity and dispersion maps are produced. (1 data file).

Spatially resolved D-T(2) correlation NMR of porous media.

PubMed

Zhang, Yan; Blümich, Bernhard

2014-05-01

Within the past decade, 2D Laplace nuclear magnetic resonance (NMR) has been developed to analyze pore geometry and diffusion of fluids in porous media on the micrometer scale. Many objects like rocks and concrete are heterogeneous on the macroscopic scale, and an integral analysis of microscopic properties provides volume-averaged information. Magnetic resonance imaging (MRI) resolves this spatial average on the contrast scale set by the particular MRI technique. Desirable contrast parameters for studies of fluid transport in porous media derive from the pore-size distribution and the pore connectivity. These microscopic parameters are accessed by 1D and 2D Laplace NMR techniques. It is therefore desirable to combine MRI and 2D Laplace NMR to image functional information on fluid transport in porous media. Because 2D Laplace resolved MRI demands excessive measuring time, this study investigates the possibility to restrict the 2D Laplace analysis to the sum signals from low-resolution pixels, which correspond to pixels of similar amplitude in high-resolution images. In this exploratory study spatially resolved D-T2 correlation maps from glass beads and mortar are analyzed. Regions of similar contrast are first identified in high-resolution images to locate corresponding pixels in low-resolution images generated with D-T2 resolved MRI for subsequent pixel summation to improve the signal-to-noise ratio of contrast-specific D-T2 maps. This method is expected to contribute valuable information on correlated sample heterogeneity from the macroscopic and the microscopic scales in various types of porous materials including building materials and rock. Copyright © 2014 Elsevier Inc. All rights reserved.

A Virtual Study of Grid Resolution on Experiments of a Highly-Resolved Turbulent Plume

NASA Astrophysics Data System (ADS)

Maisto, Pietro M. F.; Marshall, Andre W.; Gollner, Michael J.; Fire Protection Engineering Department Collaboration

2017-11-01

An accurate representation of sub-grid scale turbulent mixing is critical for modeling fire plumes and smoke transport. In this study, PLIF and PIV diagnostics are used with the saltwater modeling technique to provide highly-resolved instantaneous field measurements in unconfined turbulent plumes useful for statistical analysis, physical insight, and model validation. The effect of resolution was investigated employing a virtual interrogation window (of varying size) applied to the high-resolution field measurements. Motivated by LES low-pass filtering concepts, the high-resolution experimental data in this study can be analyzed within the interrogation windows (i.e. statistics at the sub-grid scale) and on interrogation windows (i.e. statistics at the resolved scale). A dimensionless resolution threshold (L/D*) criterion was determined to achieve converged statistics on the filtered measurements. Such a criterion was then used to establish the relative importance between large and small-scale turbulence phenomena while investigating specific scales for the turbulent flow. First order data sets start to collapse at a resolution of 0.3D*, while for second and higher order statistical moments the interrogation window size drops down to 0.2D*.

Selective self-assembly and light emission tuning of layered hybrid perovskites on patterned graphene.

PubMed

Guerra, Valentino L P; Kovaříček, Petr; Valeš, Václav; Drogowska, Karolina; Verhagen, Tim; Vejpravova, Jana; Horák, Lukáš; Listorti, Andrea; Colella, Silvia; Kalbáč, Martin

2018-02-15

The emission of light in two-dimensional (2-D) layered hybrid organic lead halide perovskites, namely (R-NH 3 ) 2 PbX 4 , can be effectively tuned using specific building blocks for the perovskite formation. Herein this behaviour is combined with a non-covalent graphene functionalization allowing excellent selectivity and spatial resolution of the perovskite film growth, promoting the formation of hybrid 2-D perovskite : graphene heterostructures with uniform coverage of up to centimeter scale graphene sheets and arbitrary shapes down to 5 μm. Using cryo-Raman microspectroscopy, highly resolved spectra of the perovskite phases were obtained and the Raman mapping served as a convenient spatially resolved technique for monitoring the distribution of the perovskite and graphene constituents on the substrate. In addition, the stability of the perovskite phase with respect to the thermal variation was inspected in situ by X-ray diffraction. Finally, time-resolved photoluminescence characterization demonstrated that the optical properties of the perovskite films grown on graphene are not hampered. Our study thus opens the door to smart fabrication routes for (opto)-electronic devices based on 2-D perovskites in contact with graphene with complex architectures.

4D Magnetic Resonance Velocimetry in a 3D printed brain aneurysm

NASA Astrophysics Data System (ADS)

Amili, Omid; Schiavazzi, Daniele; Coletti, Filippo

2016-11-01

Cerebral aneurysms are of great clinical importance. It is believed that hemodynamics play a critical role in the development, growth, and rupture of brain arteries with such condition. The flow structure in the aneurysm sac is complex, unsteady, and three-dimensional. Therefore the time-resolved measurement of the three-dimensional three-component velocity field is crucial to predict the clinical outcome. In this study magnetic resonance velocimetry is used to assess the fluid dynamics inside a 3D printed model of a giant intracranial aneurysm. We reach sub-millimeter resolution while resolving sixteen instances within the cardiac cycle. The physiological flow waveform is imposed using an in-house built pump in a flow circuit where the cardiovascular impedance is matched. The flow evolution over time is reconstructed in detail. The complex flow structure is characterized by vortical and helical motions that reside in the aneurysm for most part of the cycle. The 4D pressured distribution is also reconstructed from the velocity field. The present case study was used in a previous CFD challenge, therefore these results may provide useful experimental comparison for simulations performed by other research groups.

Three dimensional multi-cellular muscle-like tissue engineering in perfusion-based bioreactors.

PubMed

Cerino, Giulia; Gaudiello, Emanuele; Grussenmeyer, Thomas; Melly, Ludovic; Massai, Diana; Banfi, Andrea; Martin, Ivan; Eckstein, Friedrich; Grapow, Martin; Marsano, Anna

2016-01-01

Conventional tissue engineering strategies often rely on the use of a single progenitor cell source to engineer in vitro biological models; however, multi-cellular environments can better resemble the complexity of native tissues. Previous described co-culture models used skeletal myoblasts, as parenchymal cell source, and mesenchymal or endothelial cells, as stromal component. Here, we propose instead the use of adipose tissue-derived stromal vascular fraction cells, which include both mesenchymal and endothelial cells, to better resemble the native stroma. Percentage of serum supplementation is one of the crucial parameters to steer skeletal myoblasts toward either proliferation (20%) or differentiation (5%) in two-dimensional culture conditions. On the contrary, three-dimensional (3D) skeletal myoblast culture often simply adopts the serum content used in monolayer, without taking into account the new cell environment. When considering 3D cultures of mm-thick engineered tissues, homogeneous and sufficient oxygen supply is paramount to avoid formation of necrotic cores. Perfusion-based bioreactor culture can significantly improve the oxygen access to the cells, enhancing the viability and the contractility of the engineered tissues. In this study, we first investigated the influence of different serum supplementations on the skeletal myoblast ability to proliferate and differentiate during 3D perfusion-based culture. We tested percentages of serum promoting monolayer skeletal myoblast-proliferation (20%) and differentiation (5%) and suitable for stromal cell culture (10%) with a view to identify the most suitable condition for the subsequent co-culture. The 10% serum medium composition resulted in the highest number of mature myotubes and construct functionality. Co-culture with stromal vascular fraction cells at 10% serum also supported the skeletal myoblast differentiation and maturation, hence providing a functional engineered 3D muscle model that resembles the native multi-cellular environment. © 2015 Wiley Periodicals, Inc.

Differential regulation of cellular functions by the C-termini of transmembrane 4 L six family proteins in 2- or 3-dimensional environment

PubMed Central

Cheong, Jin-Gyu; Song, Dae-Geun; Song, Haeng Eun; Berditchevski, Fedor; Nam, Seo Hee; Jung, Jae Woo; Kim, Hye-Jin; Kim, Ji Eon; Kim, Somi; Ryu, Jihye; Cho, Chang Yun; Lee, Kyung-Min; Lee, Jung Weon

2017-01-01

The transmembrane 4 L six family proteins TM4SF1, TM4SF4, and TM4SF5 share 40-50% overall sequence identity, but their C-terminus identity is limited. It may be likely that the C-termini of the members are important and unique for own regulatory functions. We thus examined how the TM4SF5 C-terminus affected cellular functions differentially from other family members. Using colon cancer cells expressing wildtype (WT), C-terminus-deleted, or chimeric mutants, diverse cellular functions were explored in 2-dimensional (2D) and 3-dimensional (3D) condition. The C-termini of the proteins were relatively comparable with respect to 2D cell proliferation, although each C-terminal-deletion mutant exhibited increased proliferation relative to the WT. Using chimeric constructs, we found that the TM4SF5 C-terminus was critical for regulating the diverse metastatic functions of TM4SF5, and could positively replace the C-termini of other family members. Replacement of the TM4SF1 or TM4SF4 C-terminus with that of TM4SF5 increased spheroids growth, transwell migration, and invasive dissemination from spheroids in 3D collagen gels. TM4SF5-mediated effects required its extracellular loop 2 linked to the C-terminus via the transmembrane domain 4, with causing c-Src activation. Altogether, the C-terminus of TM4SF5 appears to mediate pro-migratory roles, depending on a structural relay from the second extracellular loop to the C-terminus. PMID:28129652

RTEMIS: Real-time Tumoroid and Environment Monitoring Using Impedance Spectroscopy and pH Sensing

NASA Astrophysics Data System (ADS)

Alexander, Frank A., Jr.

This research utilizes Electrical Impedance Spectroscopy, a technique classically used for electrochemical analysis and material characterization, as the basis for a non-destructive, label-free assay platform for three dimensional (3D) cellular spheroids. In this work, a linear array of microelectrodes is optimized to rapidly respond to changes located within a 3D multicellular model. In addition, this technique is coupled with an on chip micro-pH sensor for monitoring the environment around the cells. Finally, the responses of both impedance and pH are correlated with physical changes within the cellular model. The impedance analysis system realized through this work provides a foundation for the development of high-throughput drug screening systems that utilize multiple parallel sensing modalities including pH and impedance sensing in order to quickly assess the efficacy of specific drug candidates. The slow development of new drugs is mainly attributed to poor predictability of current chemosensitivity and resistivity assays, as well as genetic differences between the animal models used for tests and humans. In addition, monolayer cultures used in early experimentation are fundamentally different from the complex structure of organs in vivo. This requires the study of smaller 3D models (spheroids) that more efficiently replicate the conditions within the body. The main objective of this research was to develop a microfluidic system on a chip that is capable of deducing viability and morphology of 3D tumor spheroids by monitoring both the impedance of the cellular model and the pH of their local environment. This would provide a fast and reliable method for screening pharmaceutical compounds in a high-throughput system.

Resolving power of diffraction imaging with an objective: a numerical study.

PubMed

Wang, Wenjin; Liu, Jing; Lu, Jun Qing; Ding, Junhua; Hu, Xin-Hua

2017-05-01

Diffraction imaging in the far-field can detect 3D morphological features of an object for its coherent nature. We describe methods for accurate calculation and analysis of diffraction images of scatterers of single and double spheres by an imaging unit based on microscope objective at non-conjugate positions. A quantitative study of the calculated diffraction imaging in spectral domain has been performed to assess the resolving power of diffraction imaging. It has been shown numerically that with coherent illumination of 532 nm in wavelength the imaging unit can resolve single spheres of 2 μm or larger in diameters and double spheres separated by less than 300 nm between their centers.

Multiplexed 3D FRET imaging in deep tissue of live embryos

PubMed Central

Zhao, Ming; Wan, Xiaoyang; Li, Yu; Zhou, Weibin; Peng, Leilei

2015-01-01

Current deep tissue microscopy techniques are mostly restricted to intensity mapping of fluorophores, which significantly limit their applications in investigating biochemical processes in vivo. We present a deep tissue multiplexed functional imaging method that probes multiple Förster resonant energy transfer (FRET) sensors in live embryos with high spatial resolution. The method simultaneously images fluorescence lifetimes in 3D with multiple excitation lasers. Through quantitative analysis of triple-channel intensity and lifetime images, we demonstrated that Ca2+ and cAMP levels of live embryos expressing dual FRET sensors can be monitored simultaneously at microscopic resolution. The method is compatible with a broad range of FRET sensors currently available for probing various cellular biochemical functions. It opens the door to imaging complex cellular circuitries in whole live organisms. PMID:26387920

One-step fabrication of 3D silver paste electrodes into microfluidic devices for enhanced droplet-based cell sorting

NASA Astrophysics Data System (ADS)

Rao, Lang; Cai, Bo; Yu, Xiao-Lei; Guo, Shi-Shang; Liu, Wei; Zhao, Xing-Zhong

2015-05-01

3D microelectrodes are one-step fabricated into a microfluidic droplet separator by filling conductive silver paste into PDMS microchambers. The advantages of 3D silver paste electrodes in promoting droplet sorting accuracy are systematically demonstrated by theoretical calculation, numerical simulation and experimental validation. The employment of 3D electrodes also helps to decrease the droplet sorting voltage, guaranteeing that cells encapsulated in droplets undergo chip-based sorting processes are at better metabolic status for further potential cellular assays. At last, target droplet containing single cell are selectively sorted out from others by an appropriate electric pulse. This method provides a simple and inexpensive alternative to fabricate 3D electrodes, and it is expected our 3D electrode-integrated microfluidic droplet separator platform can be widely used in single cell operation and analysis.

Time-resolved inner-shell photoelectron spectroscopy: From a bound molecule to an isolated atom

NASA Astrophysics Data System (ADS)

Brauße, Felix; Goldsztejn, Gildas; Amini, Kasra; Boll, Rebecca; Bari, Sadia; Bomme, Cédric; Brouard, Mark; Burt, Michael; de Miranda, Barbara Cunha; Düsterer, Stefan; Erk, Benjamin; Géléoc, Marie; Geneaux, Romain; Gentleman, Alexander S.; Guillemin, Renaud; Ismail, Iyas; Johnsson, Per; Journel, Loïc; Kierspel, Thomas; Köckert, Hansjochen; Küpper, Jochen; Lablanquie, Pascal; Lahl, Jan; Lee, Jason W. L.; Mackenzie, Stuart R.; Maclot, Sylvain; Manschwetus, Bastian; Mereshchenko, Andrey S.; Mullins, Terence; Olshin, Pavel K.; Palaudoux, Jérôme; Patchkovskii, Serguei; Penent, Francis; Piancastelli, Maria Novella; Rompotis, Dimitrios; Ruchon, Thierry; Rudenko, Artem; Savelyev, Evgeny; Schirmel, Nora; Techert, Simone; Travnikova, Oksana; Trippel, Sebastian; Underwood, Jonathan G.; Vallance, Claire; Wiese, Joss; Simon, Marc; Holland, David M. P.; Marchenko, Tatiana; Rouzée, Arnaud; Rolles, Daniel

2018-04-01

Due to its element and site specificity, inner-shell photoelectron spectroscopy is a widely used technique to probe the chemical structure of matter. Here, we show that time-resolved inner-shell photoelectron spectroscopy can be employed to observe ultrafast chemical reactions and the electronic response to the nuclear motion with high sensitivity. The ultraviolet dissociation of iodomethane (CH3I ) is investigated by ionization above the iodine 4 d edge, using time-resolved inner-shell photoelectron and photoion spectroscopy. The dynamics observed in the photoelectron spectra appear earlier and are faster than those seen in the iodine fragments. The experimental results are interpreted using crystal-field and spin-orbit configuration interaction calculations, and demonstrate that time-resolved inner-shell photoelectron spectroscopy is a powerful tool to directly track ultrafast structural and electronic transformations in gas-phase molecules.

Resolved 12CH2D2 and 13CH3D in CH4 as Sensitive Indicators of Disequilibrium and Equilibrium during Microbial Methane Cycling

NASA Astrophysics Data System (ADS)

Ash, J. L.; Egger, M.; Slomp, C. P.; Kohl, I. E.; Treude, T.; Rumble, D.; Young, E. D.

2016-12-01

The ability to measure the relative concentrations of at least two doubly-substituted rare isotopologues of gases with biogeochemical relevance provides new constraints on sources and sinks of these gases. In particular, as shown recently for O2, the use of two independent, rare isotopologues allows for detection of thermodynamic intra-species equilibrium and disequilibrium. Here, we report the first measurements of fully resolved 13CH3D and 12CH2D2 from natural samples of microbial methane gas. A suite of sedimentary methane samples from the Bornholm Basin in the Baltic Sea was collected during IODP Exp. 347. Sample depths range from 2-20 meters below seafloor (mbsf). Methane concentrations decrease with depth, and mcrA (a marker for methanogenesis and methanotropy) is present throughout. See Figure. Both Δ13CH3D and Δ12CH2D2 increase with depth as methane concentrations decrease with the shallowest samples exhibiting disequilibrium by up to 2‰ in Δ13CH3D and 13‰ in Δ12CH2D2 while the deepest samples approach isotopic thermodynamic equilibrium (marked by grey bars in Figure). The Fe-mediated anaerobic oxidation of methane (Fe-AOM) has been inferred in these sediments by geochemical modeling . Slow methane cycling by methanogensis and methanotrophy is likely responsible for the approach to isotopic bond order equilibrium in CH4 with depth, consistent with Fe-AOM. While axenic culturing experiments generate methane with large deficits in 12CH2D2 (reported at this meeting), these data from the Baltic Sea demonstrate that isotopic equilibrium can be achieved during microbial recycling of methane. In the absence of Δ12CH2D2, the Δ13CH3D values alone could be misinterpreted as representing gradients in temperature due perhaps to exothermic organic matter degradation. The combination of both mass-18 rare isotopologues of methane provides the means to distinguish equilibrium from disequilibrium and probe microbial methane cycling even where Δ13CH3D suggests reasonable temperatures.

SPECT3D - A multi-dimensional collisional-radiative code for generating diagnostic signatures based on hydrodynamics and PIC simulation output

NASA Astrophysics Data System (ADS)

MacFarlane, J. J.; Golovkin, I. E.; Wang, P.; Woodruff, P. R.; Pereyra, N. A.

2007-05-01

SPECT3D is a multi-dimensional collisional-radiative code used to post-process the output from radiation-hydrodynamics (RH) and particle-in-cell (PIC) codes to generate diagnostic signatures (e.g. images, spectra) that can be compared directly with experimental measurements. This ability to post-process simulation code output plays a pivotal role in assessing the reliability of RH and PIC simulation codes and their physics models. SPECT3D has the capability to operate on plasmas in 1D, 2D, and 3D geometries. It computes a variety of diagnostic signatures that can be compared with experimental measurements, including: time-resolved and time-integrated spectra, space-resolved spectra and streaked spectra; filtered and monochromatic images; and X-ray diode signals. Simulated images and spectra can include the effects of backlighters, as well as the effects of instrumental broadening and time-gating. SPECT3D also includes a drilldown capability that shows where frequency-dependent radiation is emitted and absorbed as it propagates through the plasma towards the detector, thereby providing insights on where the radiation seen by a detector originates within the plasma. SPECT3D has the capability to model a variety of complex atomic and radiative processes that affect the radiation seen by imaging and spectral detectors in high energy density physics (HEDP) experiments. LTE (local thermodynamic equilibrium) or non-LTE atomic level populations can be computed for plasmas. Photoabsorption rates can be computed using either escape probability models or, for selected 1D and 2D geometries, multi-angle radiative transfer models. The effects of non-thermal (i.e. non-Maxwellian) electron distributions can also be included. To study the influence of energetic particles on spectra and images recorded in intense short-pulse laser experiments, the effects of both relativistic electrons and energetic proton beams can be simulated. SPECT3D is a user-friendly software package that runs on Windows, Linux, and Mac platforms. A parallel version of SPECT3D is supported for Linux clusters for large-scale calculations. We will discuss the major features of SPECT3D, and present example results from simulations and comparisons with experimental data.

A spatiotemporally controllable chemical gradient generator via acoustically oscillating sharp-edge structures.

PubMed

Huang, Po-Hsun; Chan, Chung Yu; Li, Peng; Nama, Nitesh; Xie, Yuliang; Wei, Cheng-Hsin; Chen, Yuchao; Ahmed, Daniel; Huang, Tony Jun

2015-11-07

The ability to generate stable, spatiotemporally controllable concentration gradients is critical for resolving the dynamics of cellular response to a chemical microenvironment. Here we demonstrate an acoustofluidic gradient generator based on acoustically oscillating sharp-edge structures, which facilitates in a step-wise fashion the rapid mixing of fluids to generate tunable, dynamic chemical gradients. By controlling the driving voltage of a piezoelectric transducer, we demonstrated that the chemical gradient profiles can be conveniently altered (spatially controllable). By adjusting the actuation time of the piezoelectric transducer, moreover, we generated pulsatile chemical gradients (temporally controllable). With these two characteristics combined, we have developed a spatiotemporally controllable gradient generator. The applicability and biocompatibility of our acoustofluidic gradient generator are validated by demonstrating the migration of human dermal microvascular endothelial cells (HMVEC-d) in response to a generated vascular endothelial growth factor (VEGF) gradient, and by preserving the viability of HMVEC-d cells after long-term exposure to an acoustic field. Our device features advantages such as simple fabrication and operation, compact and biocompatible device, and generation of spatiotemporally tunable gradients.

Additive Effects of Millimeter Waves and 2-Deoxyglucose Co-Exposure on the Human Keratinocyte Transcriptome.

PubMed

Soubere Mahamoud, Yonis; Aite, Meziane; Martin, Catherine; Zhadobov, Maxim; Sauleau, Ronan; Le Dréan, Yves; Habauzit, Denis

2016-01-01

Millimeter Waves (MMW) will be used in the next-generation of high-speed wireless technologies, especially in future Ultra-Broadband small cells in 5G cellular networks. Therefore, their biocompatibilities must be evaluated prior to their massive deployment. Using a microarray-based approach, we analyzed modifications to the whole genome of a human keratinocyte model that was exposed at 60.4 GHz-MMW at an incident power density (IPD) of 20 mW/cm2 for 3 hours in athermic conditions. No keratinocyte transcriptome modifications were observed. We tested the effects of MMWs on cell metabolism by co-treating MMW-exposed cells with a glycolysis inhibitor, 2-deoxyglucose (2dG, 20 mM for 3 hours), and whole genome expression was evaluated along with the ATP content. We found that the 2dG treatment decreased the cellular ATP content and induced a high modification in the transcriptome (632 coding genes). The affected genes were associated with transcriptional repression, cellular communication and endoplasmic reticulum homeostasis. The MMW/2dG co-treatment did not alter the keratinocyte ATP content, but it did slightly alter the transcriptome, which reflected the capacity of MMW to interfere with the bioenergetic stress response. The RT-PCR-based validation confirmed 6 MMW-sensitive genes (SOCS3, SPRY2, TRIB1, FAM46A, CSRNP1 and PPP1R15A) during the 2dG treatment. These 6 genes encoded transcription factors or inhibitors of cytokine pathways, which raised questions regarding the potential impact of long-term or chronic MMW exposure on metabolically stressed cells.

THE KMOS{sup 3D} SURVEY: DESIGN, FIRST RESULTS, AND THE EVOLUTION OF GALAXY KINEMATICS FROM 0.7 ≤ z ≤ 2.7

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

Wisnioski, E.; Förster Schreiber, N. M.; Wuyts, S.

2015-02-01

We present the KMOS{sup 3D} survey, a new integral field survey of over 600 galaxies at 0.7 < z < 2.7 using KMOS at the Very Large Telescope. The KMOS{sup 3D} survey utilizes synergies with multi-wavelength ground- and space-based surveys to trace the evolution of spatially resolved kinematics and star formation from a homogeneous sample over 5 Gyr of cosmic history. Targets, drawn from a mass-selected parent sample from the 3D-HST survey, cover the star formation-stellar mass (M {sub *}) and rest-frame (U – V) – M {sub *} planes uniformly. We describe the selection of targets, the observations, and themore » data reduction. In the first-year of data we detect Hα emission in 191 M {sub *} = 3 × 10{sup 9}-7 × 10{sup 11} M {sub ☉} galaxies at z = 0.7-1.1 and z = 1.9-2.7. In the current sample 83% of the resolved galaxies are rotation dominated, determined from a continuous velocity gradient and v {sub rot}/σ{sub 0} > 1, implying that the star-forming ''main sequence'' is primarily composed of rotating galaxies at both redshift regimes. When considering additional stricter criteria, the Hα kinematic maps indicate that at least ∼70% of the resolved galaxies are disk-like systems. Our high-quality KMOS data confirm the elevated velocity dispersions reported in previous integral field spectroscopy studies at z ≳ 0.7. For rotation-dominated disks, the average intrinsic velocity dispersion decreases by a factor of two from 50 km s{sup –1}at z ∼ 2.3 to 25 km s{sup –1}at z ∼ 0.9. Combined with existing results spanning z ∼ 0-3, we show that disk velocity dispersions follow an evolution that is consistent with the dependence of velocity dispersion on gas fractions predicted by marginally stable disk theory.« less

Key issues review: numerical studies of turbulence in stars

NASA Astrophysics Data System (ADS)

Arnett, W. David; Meakin, Casey

2016-10-01

Three major problems of single-star astrophysics are convection, magnetic fields and rotation. Numerical simulations of convection in stars now have sufficient resolution to be truly turbulent, with effective Reynolds numbers of \\text{Re}>{{10}4} , and some turbulent boundary layers have been resolved. Implications of these developments are discussed for stellar structure, evolution and explosion as supernovae. Methods for three-dimensional (3D) simulations of stars are compared and discussed for 3D atmospheres, solar rotation, core-collapse and stellar boundary layers. Reynolds-averaged Navier-Stokes (RANS) analysis of the numerical simulations has been shown to provide a novel and quantitative estimate of resolution errors. Present treatments of stellar boundaries require revision, even for early burning stages (e.g. for mixing regions during He-burning). As stellar core-collapse is approached, asymmetry and fluctuations grow, rendering spherically symmetric models of progenitors more unrealistic. Numerical resolution of several different types of three-dimensional (3D) stellar simulations are compared; it is suggested that core-collapse simulations may be under-resolved. The Rayleigh-Taylor instability in explosions has a deep connection to convection, for which the abundance structure in supernova remnants may provide evidence.

Accelerated echo-planar J-resolved spectroscopic imaging in the human brain using compressed sensing: a pilot validation in obstructive sleep apnea.

PubMed

Sarma, M K; Nagarajan, R; Macey, P M; Kumar, R; Villablanca, J P; Furuyama, J; Thomas, M A

2014-06-01

Echo-planar J-resolved spectroscopic imaging is a fast spectroscopic technique to record the biochemical information in multiple regions of the brain, but for clinical applications, time is still a constraint. Investigations of neural injury in obstructive sleep apnea have revealed structural changes in the brain, but determining the neurochemical changes requires more detailed measurements across multiple brain regions, demonstrating a need for faster echo-planar J-resolved spectroscopic imaging. Hence, we have extended the compressed sensing reconstruction of prospectively undersampled 4D echo-planar J-resolved spectroscopic imaging to investigate metabolic changes in multiple brain locations of patients with obstructive sleep apnea and healthy controls. Nonuniform undersampling was imposed along 1 spatial and 1 spectral dimension of 4D echo-planar J-resolved spectroscopic imaging, and test-retest reliability of the compressed sensing reconstruction of the nonuniform undersampling data was tested by using a brain phantom. In addition, 9 patients with obstructive sleep apnea and 11 healthy controls were investigated by using a 3T MR imaging/MR spectroscopy scanner. Significantly reduced metabolite differences were observed between patients with obstructive sleep apnea and healthy controls in multiple brain regions: NAA/Cr in the left hippocampus; total Cho/Cr and Glx/Cr in the right hippocampus; total NAA/Cr, taurine/Cr, scyllo-Inositol/Cr, phosphocholine/Cr, and total Cho/Cr in the occipital gray matter; total NAA/Cr and NAA/Cr in the medial frontal white matter; and taurine/Cr and total Cho/Cr in the left frontal white matter regions. The 4D echo-planar J-resolved spectroscopic imaging technique using the nonuniform undersampling-based acquisition and compressed sensing reconstruction in patients with obstructive sleep apnea and healthy brain is feasible in a clinically suitable time. In addition to brain metabolite changes previously reported by 1D MR spectroscopy, our results show changes of additional metabolites in patients with obstructive sleep apnea compared with healthy controls. © 2014 by American Journal of Neuroradiology.

Resolving Orbital and Climate Keys of Earth and Extraterrestrial Environments with Dynamics (ROCKE-3D) 1.0: A General Circulation Model for Simulating the Climates of Rocky Planets

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

Way, M. J.; Aleinov, I.; Amundsen, David S.

Resolving Orbital and Climate Keys of Earth and Extraterrestrial Environments with Dynamics (ROCKE-3D) is a three-dimensional General Circulation Model (GCM) developed at the NASA Goddard Institute for Space Studies for the modeling of atmospheres of solar system and exoplanetary terrestrial planets. Its parent model, known as ModelE2, is used to simulate modern Earth and near-term paleo-Earth climates. ROCKE-3D is an ongoing effort to expand the capabilities of ModelE2 to handle a broader range of atmospheric conditions, including higher and lower atmospheric pressures, more diverse chemistries and compositions, larger and smaller planet radii and gravity, different rotation rates (from slower tomore » more rapid than modern Earth’s, including synchronous rotation), diverse ocean and land distributions and topographies, and potential basic biosphere functions. The first aim of ROCKE-3D is to model planetary atmospheres on terrestrial worlds within the solar system such as paleo-Earth, modern and paleo-Mars, paleo-Venus, and Saturn’s moon Titan. By validating the model for a broad range of temperatures, pressures, and atmospheric constituents, we can then further expand its capabilities to those exoplanetary rocky worlds that have been discovered in the past, as well as those to be discovered in the future. We also discuss the current and near-future capabilities of ROCKE-3D as a community model for studying planetary and exoplanetary atmospheres.« less

Comment on "drug discovery: turning the titanic".

PubMed

Lesterhuis, W Joost; Bosco, Anthony; Lake, Richard A

2014-03-26

The pathobiology-based approach to research and development has been the dominant paradigm for successful drug discovery over the last decades. We propose that the molecular and cellular events that govern a resolving, rather than an evolving, disease may reveal new druggable pathways.

Time-resolved Sensing of Meso-scale Shock Compression with Multilayer Photonic Crystal Structures

NASA Astrophysics Data System (ADS)

Scripka, David; Lee, Gyuhyon; Summers, Christopher J.; Thadhani, Naresh

2017-06-01

Multilayer Photonic Crystal structures can provide spatially and temporally resolved data needed to validate theoretical and computational models relevant for understanding shock compression in heterogeneous materials. Two classes of 1-D photonic crystal multilayer structures were studied: optical microcavities (OMC) and distributed Bragg reflectors (DBR). These 0.5 to 5 micron thick structures were composed of SiO2, Al2O3, Ag, and PMMA layers fabricated primarily via e-beam evaporation. The multilayers have unique spectral signatures inherently linked to their time-resolved physical states. By observing shock-induced changes in these signatures, an optically-based pressure sensor was developed. Results to date indicate that both OMCs and DBRs exhibit nanosecond-resolved spectral shifts of several to 10s of nanometers under laser-driven shock compression loads of 0-10 GPa, with the magnitude of the shift strongly correlating to the shock load magnitude. Additionally, spatially and temporally resolved spectral shifts under heterogeneous laser-driven shock compression created by partial beam blocking have been successfully demonstrated. These results illustrate the potential for multilayer structures to serve as meso-scale sensors, capturing temporal and spatial pressure profile evolutions in shock-compressed heterogeneous materials, and revealing meso-scale pressure distributions across a shocked surface. Supported by DTRA Grant HDTRA1-12-1-005 and DoD, AFOSR, National Defense Science and Eng. Graduate Fellowship, 32 CFR 168a.

Imaging of cellular spread on a three-dimensional scaffold by means of a novel cell-labeling technique for high-resolution computed tomography.

PubMed

Thimm, Benjamin W; Hofmann, Sandra; Schneider, Philipp; Carretta, Roberto; Müller, Ralph

2012-03-01

Computed tomography (CT) represents a truly three-dimensional (3D) imaging technique that can provide high-resolution images on the cellular level. Thus, one approach to detect single cells is X-ray absorption-based CT, where cells are labeled with a dense, opaque material providing the required contrast for CT imaging. Within the present work, a novel cell-labeling method has been developed showing the feasibility of labeling fixed cells with iron oxide (FeO) particles for subsequent CT imaging and quantitative morphometry. A biotin-streptavidin detection system was exploited to bind FeO particles to its target endothelial cells. The binding of the particles was predominantly close to the cell centers on 2D surfaces as shown by light microscopy, scanning electron microscopy, and CT. When cells were cultured on porous, 3D polyurethane surfaces, significantly more FeO particles were detected compared with surfaces without cells and FeO particle labeling using CT. Here, we report on the implementation and evaluation of a novel cell detection method based on high-resolution CT. This system has potential in cell tracking for 3D in vitro imaging in the future.

The chemosensory brain requires a distributed cellular mechanism to harness information and resolve conflicts - is consciousness the forum?

PubMed

Lathe, Richard

2016-01-01

The central nervous system (CNS) evolved from a chemosensory epithelium, but a simple epithelium has limited means to resolve conflicts between early drives (e.g., approach vs. avoid). Understanding the role of "consciousness" as a resolution device, with specific focus on chemosensation and the olfactory system, is of appeal. I argue that consciousness is not the adjudicator, but is instead the forum that brings conflicting (conscious) inputs into a form that allows them to be (unconsciously) compared/contrasted, guiding rational action.

MDC9, a widely expressed cellular disintegrin containing cytoplasmic SH3 ligand domains

PubMed Central

1996-01-01

Cellular disintegrins are a family of proteins that are related to snake venom integrin ligands and metalloproteases. We have cloned and sequenced the mouse and human homologue of a widely expressed cellular disintegrin, which we have termed MDC9 (for metalloprotease/disintegrin/cysteine-rich protein 9). The deduced mouse and human protein sequences are 82% identical. MDC9 contains several distinct protein domains: a signal sequence is followed by a prodomain and a domain with sequence similarity to snake venom metalloproteases, a disintegrin domain, a cysteine-rich region, an EGF repeat, a membrane anchor, and a cytoplasmic tail. The cytoplasmic tail of MDC9 has two proline-rich sequences which can bind the SH3 domain of Src, and may therefore function as SH3 ligand domains. Western blot analysis shows that MDC9 is an approximately 84-kD glycoprotein in all mouse tissues examined, and in NIH 3T3 fibroblast and C2C12 myoblast mouse cell lines. MDC9 can be both cell surface biotinylated and 125I-labeled in NIH 3T3 mouse fibroblasts, indicating that the protein is present on the plasma membrane. Expression of MDC9 in COS-7 cells yields an 84-kD protein, and immunofluorescence analysis of COS-7 cells expressing MDC9 shows a staining pattern that is consistent with a plasma membrane localization. The apparent molecular mass of 84 kD suggests that MDC9 contains a membrane-anchored metalloprotease and disintegrin domain. We propose that MDC9 might function as a membrane-anchored integrin ligand or metalloprotease, or that MDC9 may combine both activities in one protein. PMID:8647900

3D and 4D Bioprinting of the Myocardium: Current Approaches, Challenges, and Future Prospects

PubMed Central

Ong, Chin Siang; Nam, Lucy; Ong, Kingsfield; Krishnan, Aravind; Huang, Chen Yu; Fukunishi, Takuma

2018-01-01

3D and 4D bioprinting of the heart are exciting notions in the modern era. However, myocardial bioprinting has proven to be challenging. This review outlines the methods, materials, cell types, issues, challenges, and future prospects in myocardial bioprinting. Advances in 3D bioprinting technology have significantly improved the manufacturing process. While scaffolds have traditionally been utilized, 3D bioprinters, which do not require scaffolds, are increasingly being employed. Improved understanding of the cardiac cellular composition and multiple strategies to tackle the issues of vascularization and viability had led to progress in this field. In vivo studies utilizing small animal models have been promising. 4D bioprinting is a new concept that has potential to advance the field of 3D bioprinting further by incorporating the fourth dimension of time. Clinical translation will require multidisciplinary collaboration to tackle the pertinent issues facing this field. PMID:29850546

Phenotypic feature quantification of patient derived 3D cancer spheroids in fluorescence microscopy image

NASA Astrophysics Data System (ADS)

Kang, Mi-Sun; Rhee, Seon-Min; Seo, Ji-Hyun; Kim, Myoung-Hee

2017-03-01

Patients' responses to a drug differ at the cellular level. Here, we present an image-based cell phenotypic feature quantification method for predicting the responses of patient-derived glioblastoma cells to a particular drug. We used high-content imaging to understand the features of patient-derived cancer cells. A 3D spheroid culture formation resembles the in vivo environment more closely than 2D adherent cultures do, and it allows for the observation of cellular aggregate characteristics. However, cell analysis at the individual level is more challenging. In this paper, we demonstrate image-based phenotypic screening of the nuclei of patient-derived cancer cells. We first stitched the images of each well of the 384-well plate with the same state. We then used intensity information to detect the colonies. The nuclear intensity and morphological characteristics were used for the segmentation of individual nuclei. Next, we calculated the position of each nucleus that is appeal of the spatial pattern of cells in the well environment. Finally, we compared the results obtained using 3D spheroid culture cells with those obtained using 2D adherent culture cells from the same patient being treated with the same drugs. This technique could be applied for image-based phenotypic screening of cells to determine the patient's response to the drug.

Clinical confocal microlaparoscope for real-time in vivo optical biopsies

NASA Astrophysics Data System (ADS)

Tanbakuchi, Anthony A.; Rouse, Andrew R.; Udovich, Joshua A.; Hatch, Kenneth D.; Gmitro, Arthur F.

2009-07-01

Successful treatment of cancer is highly dependent on the stage at which it is diagnosed. Early diagnosis, when the disease is still localized at its origin, results in very high cure rates-even for cancers that typically have poor prognosis. Biopsies are often used for diagnosis of disease. However, because biopsies are destructive, only a limited number can be taken. This leads to reduced sensitivity for detection due to sampling error. A real-time fluorescence confocal microlaparoscope has been developed that provides instant in vivo cellular images, comparable to those provided by histology, through a nondestructive procedure. The device includes an integrated contrast agent delivery mechanism and a computerized depth scan system. The instrument uses a fiber bundle to relay the image plane of a slit-scan confocal microlaparoscope into tissue. It has a 3-μm lateral resolution and a 25-μm axial resolution. Initial in vivo clinical testing using the device to image human ovaries has been done in 21 patients. Results indicate that the device can successfully image organs in vivo without complications. Results with excised tissue demonstrate that the instrument can resolve sufficient cellular detail to visualize the cellular changes associated with the onset of cancer.

C4d-the witness of humoral rejection.

PubMed

de Gouveia, R H; Vitorino, E; Ramos, S; Rebocho, M J; Queirós E Melo, J; Martins, A P; Moura, M L C

2009-04-01

Acute antibody-mediated (humoral) rejection is a major cause of morbidity, graft loss, and mortality among heart transplant patients. Herein we have presented our experience using C4d to characterize humoral rejection. All nonformalin-fixed cardiac graft biopsies (protocol or emergency) received between May 2007 and May 2008 were examined by immunofluorescence for C4d. One hundred twelve endomyocardial biopsies from 25 transplanted patients included 20 males and 5 females of ages ranging from 3 to 71 years. The number of biopsies per subject varied from 1 to 11; the timespan between transplantation and the diagnostic biopsies ranged from days to 8 years. Thirteen biopsies showed acute humoral rejection (intramyocardial capillaries positive for C4d); 31, acute cellular rejection (grades 1R, 2R); 7, both humoral and cellular rejection; and 1, acute humoral rejection and allograft vasculopathy. Some of the positive biopsies belonged to the same person, and some to transplanted individuals with signs and symptoms suggestive of rejection, while others did not. The persistence of humoral rejection, despite the disappearance of a cellular component, correlated with slower clinicoechocardiographic improvement. C4d positivity is a morphologic sign of humoral rejection. It may hasten the appearance and/or worsening of allograft vasculopathy independent of patient age or posttransplantation time.

Dynamic positional fate map of the primary heart-forming region.

PubMed

Cui, Cheng; Cheuvront, Tracey J; Lansford, Rusty D; Moreno-Rodriguez, Ricardo A; Schultheiss, Thomas M; Rongish, Brenda J

2009-08-15

Here we show the temporal-spatial orchestration of early heart morphogenesis at cellular level resolution, in vivo, and reconcile conflicting positional fate mapping data regarding the primary heart-forming field(s). We determined the positional fates of precardiac cells using a precision electroporation approach in combination with wide-field time-lapse microscopy in the quail embryo, a warm-blooded vertebrate (HH Stages 4 through 10). Contrary to previous studies, the results demonstrate the existence of a "continuous" circle-shaped heart field that spans the midline, appearing at HH Stage 4, which then expands to form a wide arc of progenitors at HH Stages 5-7. Our time-resolved image data show that a subset of these cardiac progenitor cells do not overlap with the expression of common cardiogenic factors, Nkx-2.5 and Bmp-2, until HH Stage 10, when a tubular heart has formed, calling into question when cardiac fate is specified and by which key factors. Sub-groups and anatomical bands (cohorts) of heart precursor cells dramatically change their relative positions in a process largely driven by endodermal folding and other large-scale tissue deformations. Thus, our novel dynamic positional fate maps resolve the origin of cardiac progenitor cells in amniotes. The data also establish the concept that tissue motion contributes significantly to cellular position fate - i.e., much of the cellular displacement that occurs during assembly of a midline heart tube (HH Stage 9) is NOT due to "migration" (autonomous motility), a commonly held belief. Computational analysis of our time-resolved data lays the foundation for more precise analyses of how cardiac gene regulatory networks correlate with early heart tissue morphogenesis in birds and mammals.

Spatially resolved bimodal spectroscopy for classification/evaluation of mouse skin inflammatory and pre-cancerous stages

NASA Astrophysics Data System (ADS)

Díaz-Ayil, Gilberto; Amouroux, Marine; Clanché, Fabien; Granjon, Yves; Blondel, Walter C. P. M.

2009-07-01

Spatially-resolved bimodal spectroscopy (multiple AutoFluorescence AF excitation and Diffuse Reflectance DR), was used in vivo to discriminate various healthy and precancerous skin stages in a pre-clinical model (UV-irradiated mouse): Compensatory Hyperplasia CH, Atypical Hyperplasia AH and Dysplasia D. A specific data preprocessing scheme was applied to intensity spectra (filtering, spectral correction and intensity normalization), and several sets of spectral characteristics were automatically extracted and selected based on their discrimination power, statistically tested for every pair-wise comparison of histological classes. Data reduction with Principal Components Analysis (PCA) was performed and 3 classification methods were implemented (k-NN, LDA and SVM), in order to compare diagnostic performance of each method. Diagnostic performance was studied and assessed in terms of Sensibility (Se) and Specificity (Sp) as a function of the selected features, of the combinations of 3 different inter-fibres distances and of the numbers of principal components, such that: Se and Sp ~ 100% when discriminating CH vs. others; Sp ~ 100% and Se > 95% when discriminating Healthy vs. AH or D; Sp ~ 74% and Se ~ 63% for AH vs. D.

Component-resolved diagnosis of plant food allergy by SPT.

PubMed

Asero, R; Jimeno, L; Barber, D

2008-12-01

Fruits and vegetables may contain both labile and stable allergens. The former induce only OAS, whereas stable allergens may induce systemic reactions. Component-resolved diagnosis (CRD) of allergy to plant foods is therefore essential for the clinical management of allergic patients. 80 adults allergic to plant foods underwent SPT with purified natural date palm profilin (Pho d 2), purified Mal d 1, a peach extract containing uniquely LTP, and with a kiwi extract containing uniquely stable allergens. 58 (72%) patients were monosensitized: 24 to Mal d 1, 24 to profilin, 7 to LTP, and 3 to kiwi. 22 patients were multi-sensitised: 14 to Mal d 1 and profilin, 2 to Mal d 1 and kiwi, 1 to LTP and profilin, 3 to LTP and Mal d 1, and 2 to LTP, Mal d 1 and profilin. Mal d 1 and LTP sensitisation were associated with apple and peach allergy, respectively, whereas profilin sensitisation was associated with allergy to melon, watermelon, banana, tomato and citrus fruits. 18/21 kiwi-allergic patients were sensitised to one of the cross-reacting allergens, but 2/18 reacted to kiwi-specific allergens as well. In patients with allergy to plant-derived foods CRD can be performed by SPT with purified allergen proteins. In the future, the availability of a larger number of purified natural or recombinant allergens for SPT will represent a simple means to classify food-allergic patients properly on the first visit.

Resolution of Site-Specific Conformational Heterogeneity in Proline-Rich Molecular Recognition by Src Homology 3 Domains.

PubMed

Horness, Rachel E; Basom, Edward J; Mayer, John P; Thielges, Megan C

2016-02-03

Conformational heterogeneity and dynamics are increasingly evoked in models of protein molecular recognition but are challenging to experimentally characterize. Here we combine the inherent temporal resolution of infrared (IR) spectroscopy with the spatial resolution afforded by selective incorporation of carbon-deuterium (C-D) bonds, which provide frequency-resolved absorptions within a protein IR spectrum, to characterize the molecular recognition of the Src homology 3 (SH3) domain of the yeast protein Sho1 with its cognate proline-rich (PR) sequence of Pbs2. The IR absorptions of C-D bonds introduced at residues along a peptide of the Pbs2 PR sequence report on the changes in the local environments upon binding to the SH3 domain. Interestingly, upon forming the complex the IR spectra of the peptides labeled with C-D bonds at either of the two conserved prolines of the PXXP consensus recognition sequence show more absorptions than there are C-D bonds, providing evidence for the population of multiple states. In contrast, the NMR spectra of the peptides labeled with (13)C at the same residues show only single resonances, indicating rapid interconversion on the NMR time scale. Thus, the data suggest that the SH3 domain recognizes its cognate peptide with a component of induced fit molecular recognition involving the adoption of multiples states, which have previously gone undetected due to interconversion between the populated states that is too fast to resolve using conventional methods.

A comparison of 4D time-resolved MRA with keyhole and 3D time-of-flight MRA at 3.0 T for the evaluation of cerebral aneurysms

PubMed Central

2012-01-01

Background A subarachnoid hemorrhage (SAH) due to the rupture of a cerebral aneurysm (CA) is a devastating event associated with high rates of mortality. Magnetic resonance angiography (MRA), as a noninvasive technique, is typically used initially. The object of our study is to evaluate the feasibility of 4D time-resolved MRA with keyhole (4D-TRAK) for the diagnostic accuracy and reliability of the detection and characterization of cerebral aneurysms (CAs), with a comparison of 3D time-of-flight MRA (3D-TOF-MRA) by using DSA as a reference. Methods 3D-TOF-MRA, 4D-TRAK and 3D-DSA were performed sequentially in 52 patients with suspected CAs. 4D-TRAK was acquired using a combination of sensitivity encoding (SENSE) and CE timing robust angiography (CENTRA) k-space sampling techniques at a contrast dose of 10 ml at 3 T. Accuracy, sensitivity, specificity of 4D-TRAK and 3D-TOF-MRA were calculated and compared for the detection of CAs on patient-based and aneurysm-based evaluation using 3D-DSA as a reference. Results The overall image quality of 4D-TRAK with a contrast dose of 10 ml was in the diagnostic range but still cannot be compared with that of 3D-TOF-MRA. In 52 patients with suspected CAs, fifty-eight CAs were confirmed on 3D-DSA finally. Fifty-one (with 2 false-positives and 9 false-negatives) and 58 (with 1 false-positive and 1 false-negative) CAs were visualized on 4D-TRAK and 3D-TOF-MRA, respectively. Accuracy, sensitivity and specificity on patient-based evaluation of 4D-TRAK and 3D-TOF-MRA were 92.31%, 93.33%, 85.71% and 98.08%, 100%, 85.71%, respectively, and 74.07%, 75.00%, 66.67% and 96.30%, 95.83%, 100% on aneurysm-based evaluation in patients with multiple CAs, respectively. Subgroup analysis revealed that for 19 very small CAs (maximal diameter <3 mm, measured on 3D-DSA), 9 were missed on 4D-TRAK and 1 on 3D-TOF-MRA (P = 0.008). However, for 39 CAs with maximal diameter ≥ 3 mm, the diagnostic accuracy is equally (39 on 4D-TRAK vs. 39 on 3D-TOF-MRA) (P = 1). In four larger CAs with maximal diameter ≥ 10 mm, 4D-TRAK provided a better characterization of morphology than 3D-TOF-MRA. Conclusion 4D-TRAK at a lower contrast dose of 10 ml with a combination of SENSE and CENTRA at 3 T could provide similar diagnostic accuracy rate for CAs with maximal diameter ≥ 3 mm, and a better characterization of morphology for larger CAs with maximal diameter ≥ 10 mm compared to 3D-TOF-MRA. However, further study is still needed to improve the “vascular edge” artifact and the compromise in spatial resolution in depiction of CAs with maximal diameter<3 mm. PMID:22784396

A comparison of 4D time-resolved MRA with keyhole and 3D time-of-flight MRA at 3.0 T for the evaluation of cerebral aneurysms.

PubMed

Wu, Qian; Li, Ming-Hua

2012-07-06

A subarachnoid hemorrhage (SAH) due to the rupture of a cerebral aneurysm (CA) is a devastating event associated with high rates of mortality. Magnetic resonance angiography (MRA), as a noninvasive technique, is typically used initially. The object of our study is to evaluate the feasibility of 4D time-resolved MRA with keyhole (4D-TRAK) for the diagnostic accuracy and reliability of the detection and characterization of cerebral aneurysms (CAs), with a comparison of 3D time-of-flight MRA (3D-TOF-MRA) by using DSA as a reference. 3D-TOF-MRA, 4D-TRAK and 3D-DSA were performed sequentially in 52 patients with suspected CAs. 4D-TRAK was acquired using a combination of sensitivity encoding (SENSE) and CE timing robust angiography (CENTRA) k-space sampling techniques at a contrast dose of 10 ml at 3 T. Accuracy, sensitivity, specificity of 4D-TRAK and 3D-TOF-MRA were calculated and compared for the detection of CAs on patient-based and aneurysm-based evaluation using 3D-DSA as a reference. The overall image quality of 4D-TRAK with a contrast dose of 10 ml was in the diagnostic range but still cannot be compared with that of 3D-TOF-MRA. In 52 patients with suspected CAs, fifty-eight CAs were confirmed on 3D-DSA finally. Fifty-one (with 2 false-positives and 9 false-negatives) and 58 (with 1 false-positive and 1 false-negative) CAs were visualized on 4D-TRAK and 3D-TOF-MRA, respectively. Accuracy, sensitivity and specificity on patient-based evaluation of 4D-TRAK and 3D-TOF-MRA were 92.31%, 93.33%, 85.71% and 98.08%, 100%, 85.71%, respectively, and 74.07%, 75.00%, 66.67% and 96.30%, 95.83%, 100% on aneurysm-based evaluation in patients with multiple CAs, respectively. Subgroup analysis revealed that for 19 very small CAs (maximal diameter <3 mm, measured on 3D-DSA), 9 were missed on 4D-TRAK and 1 on 3D-TOF-MRA (P = 0.008). However, for 39 CAs with maximal diameter ≥ 3 mm, the diagnostic accuracy is equally (39 on 4D-TRAK vs. 39 on 3D-TOF-MRA) (P = 1). In four larger CAs with maximal diameter ≥ 10 mm, 4D-TRAK provided a better characterization of morphology than 3D-TOF-MRA. 4D-TRAK at a lower contrast dose of 10 ml with a combination of SENSE and CENTRA at 3 T could provide similar diagnostic accuracy rate for CAs with maximal diameter ≥ 3 mm, and a better characterization of morphology for larger CAs with maximal diameter ≥ 10 mm compared to 3D-TOF-MRA. However, further study is still needed to improve the "vascular edge" artifact and the compromise in spatial resolution in depiction of CAs with maximal diameter<3 mm.

SpectraPLOT, Visualization Package with a User-Friendly Graphical Interface

NASA Astrophysics Data System (ADS)

Sebald, James; Macfarlane, Joseph; Golovkin, Igor

2017-10-01

SPECT3D is a collisional-radiative spectral analysis package designed to compute detailed emission, absorption, or x-ray scattering spectra, filtered images, XRD signals, and other synthetic diagnostics. The spectra and images are computed for virtual detectors by post-processing the results of hydrodynamics simulations in 1D, 2D, and 3D geometries. SPECT3D can account for a variety of instrumental response effects so that direct comparisons between simulations and experimental measurements can be made. SpectraPLOT is a user-friendly graphical interface for viewing a wide variety of results from SPECT3D simulations, and applying various instrumental effects to the simulated images and spectra. We will present SpectraPLOT's ability to display a variety of data, including spectra, images, light curves, streaked spectra, space-resolved spectra, and drilldown plasma property plots, for an argon-doped capsule implosion experiment example. Future SpectraPLOT features and enhancements will also be discussed.

Vasomodulatory effects of the angiotensin II type 1 receptor antagonist losartan on experimentally induced cerebral vasospasm after subarachnoid haemorrhage.

PubMed

Wanderer, Stefan; Mrosek, Jan; Gessler, Florian; Seifert, Volker; Konczalla, Juergen

2018-02-01

Cerebral vasospasm following subarachnoid haemorrhage (SAH) remains one of the major factors contributing to poor overall patient outcome. Prostaglandin F2-alpha (PGF2a) induces vasoconstriction. After SAH, PGF2a leads to cerebral inflammation and enhanced vasoconstriction, resulting in cerebral vasospasm. Losartan is already known to have beneficial effects in stroke models and also on several cerebral inflammatory processes. Therefore, the aim of the study was to analyse the effect of losartan on PGF2a-enhanced vasoconstriction after SAH. To investigate the effect of losartan on PGF2a-enhanced vasoconstriction after SAH, cerebral vasospasm was induced by a double-haemorrhage model. Rats were killed on day 3 and 5 after SAH followed by measurement of the isometric force of basilar artery ring segments in an organ bath. PGF2a induced a dose-dependent contraction. After pre-incubation with losartan, the maximum contraction (E max ) for sham-operated animals was significantly lowered [E max 6% in losartan 3 × 10 -4  molar (M) vs. 56% without losartan]. Also, after induced SAH, PGF2a induced no vasoconstriction in pre-incubated vessels with losartan 3 × 10 -4  M on day 3 (d3) as well as on day 5 (d5). For the vasorelaxative investigations, vessel segments were pre-incubated with PFG2a. Cumulative application of losartan completely resolved the pre-contraction in sham-operated animals (non SAH: 95% relaxation). After SAH, losartan not only resolved the pre-contraction (d5: 103%), but also exceeded the pre-contraction (d3: 119%). Therefore, a statistically significantly increased and earlier relaxation was calculated for all losartan concentrations [E max (d3/d5) and pD 2 (d3/d5)] compared with the solvent control group. In a physiological and pathophysiological setup, losartan reduces a PGF2-induced vasoconstriction and reverses a PGF2a-precontraction completely. This fact can be integrated in pushing forward further concepts trying to antagonise/prevent cerebral vasospasm after SAH.

Muscarinic receptor stimulation of D-aspartate uptake into human SH-SY5Y neuroblastoma cells is attenuated by hypoosmolarity.

PubMed

Foster, Daniel J; Heacock, Anne M; Fisher, Stephen K

2010-04-01

In addition to its function as an excitatory neurotransmitter, glutamate plays a major role as an osmolyte within the central nervous system (CNS). Accordingly, mechanisms that regulate glutamate release and uptake are of physiological importance not only during conditions in which cell volume remains constant but also when cells are subjected to hypoosmotic stress. In the present study, the ability of muscarinic cholinergic receptors (mAChRs) to regulate the uptake of glutamate (monitored as D-aspartate) into human SH-SY5Y neuroblastoma cells under isotonic or hypotonic conditions has been examined. In isotonic media, agonist activation of mAChRs resulted in a significant increase (250-300% of control) in the uptake of D-aspartate and, concurrently, a cellular redistribution of the excitatory amino acid transporter 3 (EAAT3) to the plasma membrane. mAChR-mediated increases in d-aspartate uptake were potently blocked by the EAAT3 inhibitor l-beta-threo-benzyl-aspartate. In hypotonic media, the ability of mAChR activation to facilitate D-aspartate uptake was significantly attenuated (40-50%), and the cellular distribution of EAAT3 was disrupted. Reduction of mAChR-stimulated D-aspartate uptake under hypoosmotic conditions could be fully reversed upon re-exposure of the cells to isotonic media. Under both isotonic and hypotonic conditions, mAChR-mediated increases in D-aspartate uptake depended on cytoskeletal integrity, protein kinase C and phosphatidylinositol 3-kinase activities, and the availability of intracellular Ca2+. In contrast, dependence on extracellular Ca2+ was observed only under isotonic conditions. The results suggest that, although the uptake of D-aspartate into SH-SY5Y cells is enhanced after mAChR activation, this process is markedly attenuated by hypoosmolarity.

Evaluation of two novel methods for assessing intracellular oxygen

NASA Astrophysics Data System (ADS)

Williams, Catrin F.; Kombrabail, M.; Vijayalakshmi, K.; White, Nick; Krishnamoorthy, G.; Lloyd, David

2012-08-01

The ability to resolve the spatio-temporal complexity of intracellular O2 distribution is the ‘Holy Grail’ of cellular physiology. In an effort to obtain a non-invasive approach of mapping intracellular O2 tensions, two methods of phosphorescent lifetime imaging microscopy were examined in the current study. These were picosecond time-resolved epiphosphorescence microscopy (single 0.5 µm focused spot) and two-photon confocal laser scanning microscopy with pinhole shifting. Both methods utilized nanoparticle-embedded Ru complex (45 nm diameter) as the phosphorescent probe, excited using pulsed outputs of Ti-sapphire Tsunami lasers (710-1050 nm). The former method used a 1 ps pulse width excitation beam with vertical polarization via a dichroic mirror (610 nm, XF43) and a 20× objective (NA 0.55, Nikon). Transmitted luminescence (1-2 × 104 counts s-1) was collected and time-correlated single photon counted decay times measured. Alternatively, an unmodified Zeiss LSM510 Confocal NLO microscope with 40× objective (NA 1.3) used successively shifted pinhole positions to collect image data from the lagging trail of the raster scan. Images obtained from two-photon excitation of a yeast (Schizosaccharomyces pombe) and a flagellate fish parasite (Spironucleus vortens), electroporated with Ru complex, indicated the intracellular location and magnitude of O2 gradients, thus confirming the feasibility of optical mapping under different external O2 concentrations. Both methods gave similar lifetimes for Ru complex phosphorescence under aerobic and anaerobic gas phases. Estimation of O2 tensions within individual fibroblasts (human dermal fibroblast (HDF)) and mammary adenocarcinoma (MCF-7) cells was possible using epiphosphorescence microscopy. MCF-7 cells showed lower intracellular O2 concentrations than HDF cells, possibly due to higher metabolic rates in the former. Future work should involve construction of higher resolution 3D maps of Ru coordinate complex lifetime distribution in cultured cell lines.

Extracting 3D Information from 1D and 2D Diagnostic Systems on the DIII-D Tokamak

NASA Astrophysics Data System (ADS)

Brookman, Michael

2017-10-01

The interpretation of tokamak data often hinges on assumptions of axisymetry and flux surface equilibria, neglecting 3D effects. This work discusses examples on the DIII-D tokamak where this assumption is an insufficient approximation, and explores the diagnostic information available to resolve 3D effects while preserving 1D profiles. Methods for extracting 3D data from the electron cyclotron emission radiometers, density profile reflectometer, and Thomson scattering system are discussed. Coordinating diagnostics around the tokamak shows the significance of 3D features, such as sawteeth[1] and resonant magnetic perturbations. A consequence of imposed 3D perturbations is a shift in major radius of measured profiles between diagnostics at different toroidal locations. Integrating different diagnostics requires a database containing information about their toroidal, poloidal, and radial locations. Through the data analysis framework OMFIT, it is possible to measure the magnitude of the apparent shifts from 3D effects and enforce consistency between diagnostics. Using the existing 1D and 2D diagnostic systems on DIII-D, this process allows the effects of the 3D perturbations on 1D profiles to be addressed. Supported by US DOE contracts DE-FC02-04ER54698, DE-FG03-97ER54415.

Nodule Detection in a Lung Region that's Segmented with Using Genetic Cellular Neural Networks and 3D Template Matching with Fuzzy Rule Based Thresholding

PubMed Central

Osman, Onur; Ucan, Osman N.

2008-01-01

Objective The purpose of this study was to develop a new method for automated lung nodule detection in serial section CT images with using the characteristics of the 3D appearance of the nodules that distinguish themselves from the vessels. Materials and Methods Lung nodules were detected in four steps. First, to reduce the number of region of interests (ROIs) and the computation time, the lung regions of the CTs were segmented using Genetic Cellular Neural Networks (G-CNN). Then, for each lung region, ROIs were specified with using the 8 directional search; +1 or -1 values were assigned to each voxel. The 3D ROI image was obtained by combining all the 2-Dimensional (2D) ROI images. A 3D template was created to find the nodule-like structures on the 3D ROI image. Convolution of the 3D ROI image with the proposed template strengthens the shapes that are similar to those of the template and it weakens the other ones. Finally, fuzzy rule based thresholding was applied and the ROI's were found. To test the system's efficiency, we used 16 cases with a total of 425 slices, which were taken from the Lung Image Database Consortium (LIDC) dataset. Results The computer aided diagnosis (CAD) system achieved 100% sensitivity with 13.375 FPs per case when the nodule thickness was greater than or equal to 5.625 mm. Conclusion Our results indicate that the detection performance of our algorithm is satisfactory, and this may well improve the performance of computer-aided detection of lung nodules. PMID:18253070

Synchrotron radiation microtomography of Taylor bubbles in capillary two-phase flow

NASA Astrophysics Data System (ADS)

Boden, Stephan; dos Santos Rolo, Tomy; Baumbach, Tilo; Hampel, Uwe

2014-07-01

We report on a study to measure the three-dimensional shape of Taylor bubbles in capillaries using synchrotron radiation in conjunction with ultrafast radiographic imaging. Moving Taylor bubbles in 2-mm round and square capillaries were radiographically scanned with an ultrahigh frame rate of up to 36,000 fps and 5.6-µm pixel separation. Consecutive images were properly processed to yield 2D transmission radiographs of high contrast-to-noise ratio. Application of 3D tomographic image reconstruction disclosed the 3D bubble shape. The results provide a reference data base for development of sophisticated interface resolving CFD computations.

The Use of 3D Telomere FISH for the Characterization of the Nuclear Architecture in EBV-Positive Hodgkin's Lymphoma.

PubMed

Knecht, Hans; Mai, Sabine

2017-01-01

The 3D nuclear architecture is closely related to cellular functions and chromosomes are organized in distinct territories. Quantitative 3D telomere FISH analysis (3D Q-FISH) and 3D super-resolution imaging (3D-SIM) at a resolution up to 80 nm as well as the recently developed combined quantitative 3D TRF2-telomere immune FISH technique (3D TRF2/Telo-Q-FISH) have substantially contributed to elucidate molecular pathogenic mechanisms of hematological diseases. Here we report the methods we applied to uncover major molecular steps involved in the pathogenesis of EBV-associated Hodgkin's lymphoma. These methods allowed us to identify the EBV-encoded oncoprotein LMP1 as a key element in the formation of Hodgkin (H-cell) and multinucleated Reed-Sternberg cells (RS-cell), the diagnostic tumor cell of classical Hodgkin's lymphoma (cHL). LMP1 mediates multinuclearity through downregulation of shelterin proteins, in particular telomere repeat binding factor 2 (TRF2).

Microscale screening systems for 3D cellular microenvironments: platforms, advances, and challenges

PubMed Central

Montanez-Sauri, Sara I.; Beebe, David J.; Sung, Kyung Eun

2015-01-01

The increasing interest in studying cells using more in vivo-like three-dimensional (3D) microenvironments has created a need for advanced 3D screening platforms with enhanced functionalities and increased throughput. 3D screening platforms that better mimic in vivo microenvironments with enhanced throughput would provide more in-depth understanding of the complexity and heterogeneity of microenvironments. The platforms would also better predict the toxicity and efficacy of potential drugs in physiologically relevant conditions. Traditional 3D culture models (e.g. spinner flasks, gyratory rotation devices, non-adhesive surfaces, polymers) were developed to create 3D multicellular structures. However, these traditional systems require large volumes of reagents and cells, and are not compatible with high throughput screening (HTS) systems. Microscale technology offers the miniaturization of 3D cultures and allows efficient screening of various conditions. This review will discuss the development, most influential works, and current advantages and challenges of microscale culture systems for screening cells in 3D microenvironments. PMID:25274061

Electronic structure and relaxation dynamics in a superconducting topological material

DOE PAGES

Neupane, Madhab; Ishida, Yukiaki; Sankar, Raman; ...

2016-03-03

Topological superconductors host new states of quantum matter which show a pairing gap in the bulk and gapless surface states providing a platform to realize Majorana fermions. Recently, alkaline-earth metal Sr intercalated Bi2Se3 has been reported to show superconductivity with a Tc~3K and a large shielding fraction. Here we report systematic normal state electronic structure studies of Sr0.06Bi2Se3 (Tc~2.5K) by performing photoemission spectroscopy. Using angle-resolved photoemission spectroscopy (ARPES), we observe a quantum well confined two-dimensional (2D) state coexisting with a topological surface state in Sr0.06Bi2Se3. Furthermore, our time-resolved ARPES reveals the relaxation dynamics showing different decay mechanism between the excitedmore » topological surface states and the two-dimensional states. Our experimental observation is understood by considering the intra-band scattering for topological surface states and an additional electron phonon scattering for the 2D states, which is responsible for the superconductivity. Our first-principles calculations agree with the more effective scattering and a shorter lifetime of the 2D states. In conclusion, our results will be helpful in understanding low temperature superconducting states of these topological materials.« less

Full-field dual-color 100-nm super-resolution imaging reveals organization and dynamics of mitochondrial and ER networks.

PubMed

Brunstein, Maia; Wicker, Kai; Hérault, Karine; Heintzmann, Rainer; Oheim, Martin

2013-11-04

Most structured illumination microscopes use a physical or synthetic grating that is projected into the sample plane to generate a periodic illumination pattern. Albeit simple and cost-effective, this arrangement hampers fast or multi-color acquisition, which is a critical requirement for time-lapse imaging of cellular and sub-cellular dynamics. In this study, we designed and implemented an interferometric approach allowing large-field, fast, dual-color imaging at an isotropic 100-nm resolution based on a sub-diffraction fringe pattern generated by the interference of two colliding evanescent waves. Our all-mirror-based system generates illumination pat-terns of arbitrary orientation and period, limited only by the illumination aperture (NA = 1.45), the response time of a fast, piezo-driven tip-tilt mirror (10 ms) and the available fluorescence signal. At low µW laser powers suitable for long-period observation of life cells and with a camera exposure time of 20 ms, our system permits the acquisition of super-resolved 50 µm by 50 µm images at 3.3 Hz. The possibility it offers for rapidly adjusting the pattern between images is particularly advantageous for experiments that require multi-scale and multi-color information. We demonstrate the performance of our instrument by imaging mitochondrial dynamics in cultured cortical astrocytes. As an illustration of dual-color excitation dual-color detection, we also resolve interaction sites between near-membrane mitochondria and the endoplasmic reticulum. Our TIRF-SIM microscope provides a versatile, compact and cost-effective arrangement for super-resolution imaging, allowing the investigation of co-localization and dynamic interactions between organelles--important questions in both cell biology and neurophysiology.

Self-organisation in Cellular Automata with Coalescent Particles: Qualitative and Quantitative Approaches

NASA Astrophysics Data System (ADS)

Hellouin de Menibus, Benjamin; Sablik, Mathieu

2017-06-01

This article introduces new tools to study self-organisation in a family of simple cellular automata which contain some particle-like objects with good collision properties (coalescence) in their time evolution. We draw an initial configuration at random according to some initial shift-ergodic measure, and use the limit measure to describe the asymptotic behaviour of the automata. We first take a qualitative approach, i.e. we obtain information on the limit measure(s). We prove that only particles moving in one particular direction can persist asymptotically. This provides some previously unknown information on the limit measures of various deterministic and probabilistic cellular automata: 3 and 4-cyclic cellular automata [introduced by Fisch (J Theor Probab 3(2):311-338, 1990; Phys D 45(1-3):19-25, 1990)], one-sided captive cellular automata [introduced by Theyssier (Captive Cellular Automata, 2004)], the majority-traffic cellular automaton, a self stabilisation process towards a discrete line [introduced by Regnault and Rémila (in: Mathematical Foundations of Computer Science 2015—40th International Symposium, MFCS 2015, Milan, Italy, Proceedings, Part I, 2015)]. In a second time we restrict our study to a subclass, the gliders cellular automata. For this class we show quantitative results, consisting in the asymptotic law of some parameters: the entry times [generalising K ůrka et al. (in: Proceedings of AUTOMATA, 2011)], the density of particles and the rate of convergence to the limit measure.

Vitamin D Combined with Aminolevulinate (ALA)-Mediated Photodynamic Therapy (PDT) for Human Psoriasis: A Proof-of-Principle Study

PubMed Central

Maytin, Edward V.; Honari, Golara; Khachemoune, Amor; Taylor, Charles R.; Ortel, Bernhard; Pogue, Brian W.; Sznycer-Taub, Nathaniel; Hasan, Tayyaba

2012-01-01

We previously showed that select agents (methotrexate or Vitamin D), when administered as a preconditioning regimen, are capable of promoting cellular differentiation of epithelial cancer cells while simultaneously enhancing the efficacy of 5-aminolevulinic acid (ALA)-mediated photodynamic therapy (PDT). In solid tumors, pretreatment with Vitamin D simultaneously promotes cellular differentiation and leads to selective accumulation of target porphyrins (mainly protoporphyrin IX, PpIX) within diseased tissue. However, questions of whether or not the effects upon cellular differentiation are inexorably linked to PpIX accumulation, and whether these effects might occur in hyperproliferative noncancerous tissues, have remained unanswered. In this paper, we reasoned that psoriasis, a human skin disease in which abnormal cellular proliferation and differentiation plays a major role, could serve as a useful model to test the effects of pro-differentiating agents upon PpIX levels in a non-neoplastic setting. In particular, Vitamin D, a treatment for psoriasis that restores (increases) differentiation, might increase PpIX levels in psoriatic lesions and facilitate their responsiveness to ALA-PDT. This concept was tested in a pilot study of 7 patients with bilaterally-matched psoriatic plaques. A regimen in which calcipotriol 0.005% ointment was applied for 3 days prior to ALA-PDT with blue light, led to preferential increases in PpIX (~130%), and reductions in thickness, redness, scaling, and itching in the pretreated plaques. The results suggest that a larger clinical trial is warranted to confirm a role for combination treatments with Vitamin D and ALA-PDT for psoriasis. PMID:23264699

Complexity growth in minimal massive 3D gravity

NASA Astrophysics Data System (ADS)

Qaemmaqami, Mohammad M.

2018-01-01

We study the complexity growth by using "complexity =action " (CA) proposal in the minimal massive 3D gravity (MMG) model which is proposed for resolving the bulk-boundary clash problem of topologically massive gravity (TMG). We observe that the rate of the complexity growth for Banados-Teitelboim-Zanelli (BTZ) black hole saturates the proposed bound by physical mass of the BTZ black hole in the MMG model, when the angular momentum parameter and the inner horizon of black hole goes to zero.

Drosophila melanogaster cellular repressor of E1A-stimulated genes is a lysosomal protein essential for fly development

PubMed Central

Kowalewski-Nimmerfall, Elisabeth; Schähs, Philipp; Maresch, Daniel; Rendic, Dubravko; Krämer, Helmut; Mach, Lukas

2014-01-01

Mammalian cellular repressor of E1A-stimulated genes is a lysosomal glycoprotein implicated in cellular growth and differentiation. The genome of the fruit fly Drosophila melanogaster encodes a putative orthologue (dCREG), suggesting evolutionarily conserved physiological functions of this protein. In D. melanogaster S2 cells, dCREG was found to localize in lysosomes. Further studies revealed that intracellular dCREG is subject of proteolytic maturation. Processing and turnover could be substantially reduced by RNAi-mediated silencing of cathepsin L. In contrast to mammalian cells, lysosomal delivery of dCREG does not depend on its carbohydrate moiety. Furthermore, depletion of the putative D. melanogaster lysosomal sorting receptor lysosomal enzyme receptor protein did not compromise cellular retention of dCREG. We also investigated the developmental consequences of dCREG ablation in whole D. melanogaster flies. Ubiquitous depletion of dCREG proved lethal at the late pupal stage once a knock-down efficiency of > 95% was achieved. These results demonstrate that dCREG is essential for proper completion of fly development. PMID:25173815

Drosophila melanogaster cellular repressor of E1A-stimulated genes is a lysosomal protein essential for fly development.

PubMed

Kowalewski-Nimmerfall, Elisabeth; Schähs, Philipp; Maresch, Daniel; Rendic, Dubravko; Krämer, Helmut; Mach, Lukas

2014-12-01

Mammalian cellular repressor of E1A-stimulated genes is a lysosomal glycoprotein implicated in cellular growth and differentiation. The genome of the fruit fly Drosophila melanogaster encodes a putative orthologue (dCREG), suggesting evolutionarily conserved physiological functions of this protein. In D. melanogaster S2 cells, dCREG was found to localize in lysosomes. Further studies revealed that intracellular dCREG is subject of proteolytic maturation. Processing and turnover could be substantially reduced by RNAi-mediated silencing of cathepsin L. In contrast to mammalian cells, lysosomal delivery of dCREG does not depend on its carbohydrate moiety. Furthermore, depletion of the putative D. melanogaster lysosomal sorting receptor lysosomal enzyme receptor protein did not compromise cellular retention of dCREG. We also investigated the developmental consequences of dCREG ablation in whole D. melanogaster flies. Ubiquitous depletion of dCREG proved lethal at the late pupal stage once a knock-down efficiency of >95% was achieved. These results demonstrate that dCREG is essential for proper completion of fly development. Copyright © 2014. Published by Elsevier B.V.

Assessing the risk of CMV reactivation and reconstitution of antiviral immune response post bone marrow transplantation by the QuantiFERON-CMV-assay and real time PCR.

PubMed

Krawczyk, Adalbert; Ackermann, Jessica; Goitowski, Birgit; Trenschel, Rudolf; Ditschkowski, Markus; Timm, Jörg; Ottinger, Hellmut; Beelen, Dietrich W; Grüner, Nico; Fiedler, Melanie

CMV reactivation is a major cause of severe complications in allogeneic hematopoietic stem cell transplant (HSCT) recipients. The risk of CMV reactivation depends on the serostatus (+/-) of the donor (D) and recipient (R). The reconstitution of CMV-specific T-cell responses after transplantation is crucial for the control of CMV reactivation. The study aimed to determine the cellular immune status correlating with protection from high-level CMV viremia (>5000 copies/ml) and disease. We monitored CMV-specific cellular immune responses in 9 high-risk (D-/R+), 14 intermediate risk (D+/R+) and 3 low risk individuals (D+/R-), and 8 CMV negative controls (D-/R-). Interferon- γ (IFN-γ) levels as a marker for the CD8+ T-cell response were determined by the QuantiFERON-CMV-assay and compared to viral loads determined by PCR. Early CMV reactivation was detected in all high-risk and 13/14 intermediate risk individuals. High-level viremia was detected in 5/7 high and 7/14 intermediate risk patients. Reconstitution of the CMV-specific cellular immune response started from 3 months after transplantation and resulted in protection against CMV reactivation. Re-establishing of CMV-specific T-cell immune responses with IFN- γ levels >8.9 IU/ml is crucial for protection from high-level CMV viremia. Monitoring of HSCT-recipients with the QuantiFERON-CMV-assay might be of great benefit to optimize antiviral treatment. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

3D image processing architecture for camera phones

NASA Astrophysics Data System (ADS)

Atanassov, Kalin; Ramachandra, Vikas; Goma, Sergio R.; Aleksic, Milivoje

2011-03-01

Putting high quality and easy-to-use 3D technology into the hands of regular consumers has become a recent challenge as interest in 3D technology has grown. Making 3D technology appealing to the average user requires that it be made fully automatic and foolproof. Designing a fully automatic 3D capture and display system requires: 1) identifying critical 3D technology issues like camera positioning, disparity control rationale, and screen geometry dependency, 2) designing methodology to automatically control them. Implementing 3D capture functionality on phone cameras necessitates designing algorithms to fit within the processing capabilities of the device. Various constraints like sensor position tolerances, sensor 3A tolerances, post-processing, 3D video resolution and frame rate should be carefully considered for their influence on 3D experience. Issues with migrating functions such as zoom and pan from the 2D usage model (both during capture and display) to 3D needs to be resolved to insure the highest level of user experience. It is also very important that the 3D usage scenario (including interactions between the user and the capture/display device) is carefully considered. Finally, both the processing power of the device and the practicality of the scheme needs to be taken into account while designing the calibration and processing methodology.

Three-dimensional drift kinetic response of high- β plasmas in the DIII-D tokamak

DOE PAGES

Wang, Zhirui R.; Lanctot, Matthew J.; Liu, Y. Q.; ...

2015-04-07

A quantitative interpretation of the experimentally measured high pressure plasma response to externally applied three-dimensional (3D) magnetic field perturbations, across the no-wall Troyon limit, is achieved. The key to success is the self-consistent inclusion of the drift kinetic resonance effects in numerical modeling using the MARS-K code. This resolves an outstanding issue of ideal magneto-hydrodynamic model, which signi cantly over-predicts the plasma induced field ampli fication near the no-wall limit, as compared to experiments. The self-consistent drift kinetic model leads to quantitative agreement not only for the measured 3D field amplitude and toroidal phase, but also for the measured internalmore » 3D displacement of the plasma.« less

Quantitative real-time analysis of collective cancer invasion and dissemination

NASA Astrophysics Data System (ADS)

Ewald, Andrew J.

2015-05-01

A grand challenge in biology is to understand the cellular and molecular basis of tissue and organ level function in mammals. The ultimate goals of such efforts are to explain how organs arise in development from the coordinated actions of their constituent cells and to determine how molecularly regulated changes in cell behavior alter the structure and function of organs during disease processes. Two major barriers stand in the way of achieving these goals: the relative inaccessibility of cellular processes in mammals and the daunting complexity of the signaling environment inside an intact organ in vivo. To overcome these barriers, we have developed a suite of tissue isolation, three dimensional (3D) culture, genetic manipulation, nanobiomaterials, imaging, and molecular analysis techniques to enable the real-time study of cell biology within intact tissues in physiologically relevant 3D environments. This manuscript introduces the rationale for 3D culture, reviews challenges to optical imaging in these cultures, and identifies current limitations in the analysis of complex experimental designs that could be overcome with improved imaging, imaging analysis, and automated classification of the results of experimental interventions.

Three-Dimensional Human iPSC-Derived Artificial Skeletal Muscles Model Muscular Dystrophies and Enable Multilineage Tissue Engineering.

PubMed

Maffioletti, Sara Martina; Sarcar, Shilpita; Henderson, Alexander B H; Mannhardt, Ingra; Pinton, Luca; Moyle, Louise Anne; Steele-Stallard, Heather; Cappellari, Ornella; Wells, Kim E; Ferrari, Giulia; Mitchell, Jamie S; Tyzack, Giulia E; Kotiadis, Vassilios N; Khedr, Moustafa; Ragazzi, Martina; Wang, Weixin; Duchen, Michael R; Patani, Rickie; Zammit, Peter S; Wells, Dominic J; Eschenhagen, Thomas; Tedesco, Francesco Saverio

2018-04-17

Generating human skeletal muscle models is instrumental for investigating muscle pathology and therapy. Here, we report the generation of three-dimensional (3D) artificial skeletal muscle tissue from human pluripotent stem cells, including induced pluripotent stem cells (iPSCs) from patients with Duchenne, limb-girdle, and congenital muscular dystrophies. 3D skeletal myogenic differentiation of pluripotent cells was induced within hydrogels under tension to provide myofiber alignment. Artificial muscles recapitulated characteristics of human skeletal muscle tissue and could be implanted into immunodeficient mice. Pathological cellular hallmarks of incurable forms of severe muscular dystrophy could be modeled with high fidelity using this 3D platform. Finally, we show generation of fully human iPSC-derived, complex, multilineage muscle models containing key isogenic cellular constituents of skeletal muscle, including vascular endothelial cells, pericytes, and motor neurons. These results lay the foundation for a human skeletal muscle organoid-like platform for disease modeling, regenerative medicine, and therapy development. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

An in vitro 3D model using collagen coated gelatin nanofibers for studying breast cancer metastasis.

PubMed

Janani, G; Pillai, Mamatha M; Selvakumar, R; Bhattacharyya, Amitava; Sabarinath, C

2017-02-07

The study of breast cancer metastasis is limited due to poor knowledge of molecular progression of breast tumor and varied heterogeneity. For a better understanding of tumor metastasis, a reliable 3D in vitro model bridging the gap between 2D cultures and in vivo animal model studies is essential. Our study is focused on two key points: (i) designing a 3D microenvironment for studying metastasis and (ii) simulating the metastasis milieu by inducing epithelial to mesenchymal transition (EMT) and mesenchymal to epithelial transition (MET). An electrospun gelatin nanofiber matrix (EGNF) was fabricated using electrospinning and further dip coated with different concentrations of collagen to obtain surface complexity and mechanical properties, similar to connective tissues. Nanofiber matrices were physically characterized by Fourier transform infrared spectroscopy (FTIR), atomic force microscopy (AFM), and field-emission scanning electron microscopy (FESEM). The FTIR, AFM, and FESEM results indicated the crosslinking and confirmed the presence of pores in the nanofiber matrices. Comparative studies on biocompatibility, cell attachment, and the proliferation of MCF-7 cells on EGNF and collagen coated gelatin nanofibrous matrix (CCGM) revealed higher cellular attachment and proliferation in CCGM. CCGM with human metastatic breast cancer cell line (MCF-7) was taken to study breast cancer metastasis using estrogen (induces EMT) and progesterone (induces MET) hormones for 24 h. Quantitative real-time PCR was used for quantifying the expression of metastasis related genes, and fluorescence microscopy for verifying the invasion of cells to the matrices. The expression of E-cadherin and matrix metalloproteinase 2 (MMP 2) confirmed the occurrence of EMT and MET. Live cell imaging and cellular attachment showed significant increase of cellular invasion in crosslinked 0.15% CCGM that serves as a suitable non-toxic, biocompatible, and affordable scaffold for studying breast cancer metastasis. Our findings suggested that CCGM can be used as a tissue-like 3D model for studying breast cancer metastatic events in vitro.