Geometrical optics limit of stochastic electromagnetic fields

A method is described which elucidates propagation of an electromagnetic field generated by a stochastic, electromagnetic source within the short wavelength limit. The results can be used to determine statistical properties of fields using ray tracing methods

Calcium/calmodulin-dependent phosphorylation of vimentin in rat sertoli cells.

Ca2+-dependent protein phosphorylation and the role of calmodulin in this process was investigated in subcellular fractions of primary cultures of rat Sertoli cells. Significant Ca2+/calmodulin-dependent protein phosphorylation in Sertoli cells was restricted to the cytosol fraction. The calmodulin dependence of these effects was confirmed by using the calmodulin inhibitor trifluoperazine. One of the Ca2+/calmodulin-dependent phosphoproteins was identified as the intermediate filament protein vimentin, based on the following criteria: (i) migration pattern in two-dimensional polyacrylamide gels, (ii) Ca2+/calmodulin-dependent phosphorylation of a 58-kilodalton protein present in detergent-insoluble intermediate filament protein extract of Sertoli cells, and (iii) peptide mapping of the phosphoprotein. These data support a role for Ca2+/calmodulin-dependent protein phosphorylation in the modulation of Sertoli cell cytoskeletal components

Optical biopsy of lymph node morphology using optical coherence tomography

Optical diagnostic imaging techniques are increasingly being used in the clinical environment, allowing for improved screening and diagnosis while minimizing the number of invasive procedures. Diffuse optical tomography, for example, is capable of whole-breast imaging and is being developed as an alternative to traditional X-ray mammography. While this may eventually be a very effective screening method, other optical techniques are better suited for imaging on the cellular and molecular scale. Optical Coherence Tomography (OCT), for instance, is capable of high-resolution cross-sectional imaging of tissue morphology. In a manner analogous to ultrasound imaging except using optics, pulses of near-infrared light are sent into the tissue while coherence-gated reflections are measured interferometrically to form a cross-sectional image of tissue. In this paper we apply OCT techniques for the high-resolution three-dimensional visualization of lymph node morphology. We present the first reported OCT images showing detailed morphological structure and corresponding histological features of lymph nodes from a carcinogen-induced rat mammary tumor model, as well as from a human lymph node containing late stage metastatic disease. The results illustrate the potential for OCT to visualize detailed lymph node structures on the scale of micrometastases and the potential for the detection of metastatic nodal disease intraoperatively

Estrogenic activity assessment of environmental chemicals using in vitro assays: identification of two new estrogenic compounds.

Environmental chemicals with estrogenic activities have been suggested to be associated with deleterious effects in animals and humans. To characterize estrogenic chemicals and their mechanisms of action, we established in vitro and cell culture assays that detect human estrogen receptor [alpha] (hER[alpha])-mediated estrogenicity. First, we assayed chemicals to determine their ability to modulate direct interaction between the hER[alpha] and the steroid receptor coactivator-1 (SRC-1) and in a competition binding assay to displace 17ss-estradiol (E(2)). Second, we tested the chemicals for estrogen-associated transcriptional activity in the yeast estrogen screen and in the estrogen-responsive MCF-7 human breast cancer cell line. The chemicals investigated in this study were o,p'-DDT (racemic mixture and enantiomers), nonylphenol mixture (NPm), and two poorly analyzed compounds in the environment, namely, tris-4-(chlorophenyl)methane (Tris-H) and tris-4-(chlorophenyl)methanol (Tris-OH). In both yeast and MCF-7 cells, we determined estrogenic activity via the estrogen receptor (ER) for o,p'-DDT, NPm, and for the very first time, Tris-H and Tris-OH. However, unlike estrogens, none of these xenobiotics seemed to be able to induce ER/SRC-1 interactions, most likely because the conformation of the activated receptor would not allow direct contacts with this coactivator. However, these compounds were able to inhibit [(3)H]-E(2) binding to hER, which reveals a direct interaction with the receptor. In conclusion, the test compounds are estrogen mimics, but their molecular mechanism of action appears to be different from that of the natural hormone as revealed by the receptor/coactivator interaction analysis

SeekFusion - A Clinically Validated Fusion Transcript Detection Pipeline for PCR-Based Next-Generation Sequencing of RNA

Detecting gene fusions involving driver oncogenes is pivotal in clinical diagnosis and treatment of cancer patients. Recent developments in next-generation sequencing (NGS) technologies have enabled improved assays for bioinformatics-based gene fusions detection. In clinical applications, where a small number of fusions are clinically actionable, targeted polymerase chain reaction (PCR)-based NGS chemistries, such as the QIAseq RNAscan assay, aim to improve accuracy compared to standard RNA sequencing. Existing informatics methods for gene fusion detection in NGS-based RNA sequencing assays traditionally use a transcriptome-based spliced alignment approach or a de-novo assembly approach. Transcriptome-based spliced alignment methods face challenges with short read mapping yielding low quality alignments. De-novo assembly-based methods yield longer contigs from short reads that can be more sensitive for genomic rearrangements, but face performance and scalability challenges. Consequently, there exists a need for a method to efficiently and accurately detect fusions in targeted PCR-based NGS chemistries. We describe SeekFusion, a highly accurate and computationally efficient pipeline enabling identification of gene fusions from PCR-based NGS chemistries. Utilizing biological samples processed with the QIAseq RNAscan assay and in-silico simulated data we demonstrate that SeekFusion gene fusion detection accuracy outperforms popular existing methods such as STAR-Fusion, TOPHAT-Fusion and JAFFA-hybrid. We also present results from 4,484 patient samples tested for neurological tumors and sarcoma, encompassing details on some novel fusions identified

Real-time and non-invasive measurements of cell mechanical behaviour with optical coherence phase microscopy

There is an unmet need in tissue engineering for non-invasive, label-free monitoring of cell mechanical behaviour in their physiological environment. Here, we describe a novel optical coherence phase microscopy (OCPM) set-up which can map relative cell mechanical behaviour in monolayers and 3D systems non-invasively, and in real-time. 3T3 and MCF-7 cells were investigated, with MCF-7 demonstrating an increased response to hydrostatic stimulus indicating MCF-7 being softer than 3T3, demonstrating the ability to provide qualitative data on cell mechanical behaviour. Quantitative measurements of 6% agarose beads have been taken with commercial Cell Scale Microsquisher® system demonstrating that their mechanical properties are in the same order of magnitude of cells, indicating that this is an appropriate test sample for the novel method desctibed

Anticancer efficacy of the hypoxia-activated prodrug evofosfamide (TH-302) in osteolytic breast cancer murine models

Tumor hypoxia is a major cause of treatment failure for a variety of malignancies. However, hypoxia offers treatment opportunities, exemplified by the development of compounds that target hypoxic regions within tumors. Evofosfamide (TH-302) is a prodrug created by the conjugation of 2-nitroimidazole to bromo-isophosphoramide mustard (Br-IPM). When evofosfamide is delivered to hypoxic regions, the DNA cross-linking effector, Br-IPM, is released. This study assessed the cytotoxic activity of evofosfamide in vitro and its antitumor activity against osteolytic breast cancer either alone or in combination with paclitaxel in vivo. A panel of human breast cancer cell lines were treated with evofosfamide under hypoxia and assessed for cell viability. Osteolytic MDA-MB-231-TXSA cells were transplanted into the mammary fat pad, or into tibiae of mice, allowed to establish and treated with evofosfamide, paclitaxel, or both. Tumor burden was monitored using bioluminescence, and cancer-induced bone destruction was measured using micro-CT. In vitro, evofosfamide was selectively cytotoxic under hypoxic conditions. In vivo evofosfamide was tumor suppressive as a single agent and cooperated with paclitaxel to reduce mammary tumor growth. Breast cancer cells transplanted into the tibiae of mice developed osteolytic lesions. In contrast, treatment with evofosfamide or paclitaxel resulted in a significant delay in tumor growth and an overall reduction in tumor burden in bone, whereas combined treatment resulted in a significantly greater reduction in tumor burden in the tibia of mice. Evofosfamide cooperates with paclitaxel and exhibits potent tumor suppressive activity against breast cancer growth in the mammary gland and in bone.Vasilios Liapis, Irene Zinonos, Agatha Labrinidis, Shelley Hay, Vladimir Ponomarev, Vasilios Panagopoulos, Aneta Zysk, Mark DeNichilo, Wendy Ingman, Gerald J. Atkins, David M. Findlay, Andrew C. W. Zannettino, Andreas Evdokio

Characterization of a pneumococcal meningitis mouse model

<p>Abstract</p> <p>Background</p> <p><it>S. pneumoniae </it>is the most common causative agent of meningitis, and is associated with high morbidity and mortality. We aimed to develop an integrated and representative pneumococcal meningitis mouse model resembling the human situation.</p> <p>Methods</p> <p>Adult mice (C57BL/6) were inoculated in the cisterna magna with increasing doses of <it>S. pneumoniae </it>serotype 3 colony forming units (CFU; n = 24, 10⁴, 10⁵, 10⁶and 10⁷CFU) and survival studies were performed. Cerebrospinal fluid (CSF), brain, blood, spleen, and lungs were collected. Subsequently, mice were inoculated with 10⁴CFU <it>S. pneumoniae </it>serotype 3 and sacrificed at 6 (n = 6) and 30 hours (n = 6). Outcome parameters were bacterial outgrowth, clinical score, and cytokine and chemokine levels (using Luminex^®) in CSF, blood and brain. Meningeal inflammation, neutrophil infiltration, parenchymal and subarachnoidal hemorrhages, microglial activation and hippocampal apoptosis were assessed in histopathological studies.</p> <p>Results</p> <p>Lower doses of bacteria delayed onset of illness and time of death (median survival CFU 10⁴, 56 hrs; 10⁵, 38 hrs, 10⁶, 28 hrs. 10⁷, 24 hrs). Bacterial titers in brain and CSF were similar in all mice at the end-stage of disease independent of inoculation dose, though bacterial outgrowth in the systemic compartment was less at lower inoculation doses. At 30 hours after inoculation with 10⁴CFU of <it>S. pneumoniae</it>, blood levels of KC, IL6, MIP-2 and IFN- γ were elevated, as were brain homogenate levels of KC, MIP-2, IL-6, IL-1β and RANTES. Brain histology uniformly showed meningeal inflammation at 6 hours, and, neutrophil infiltration, microglial activation, and hippocampal apoptosis at 30 hours. Parenchymal and subarachnoidal and cortical hemorrhages were seen in 5 of 6 and 3 of 6 mice at 6 and 30 hours, respectively.</p> <p>Conclusion</p> <p>We have developed and validated a murine model of pneumococcal meningitis.</p

Visualizing Escherichia coli Sub-Cellular Structure Using Sparse Deconvolution Spatial Light Interference Tomography

Studying the 3D sub-cellular structure of living cells is essential to our understanding of biological function. However, tomographic imaging of live cells is challenging mainly because they are transparent, i.e., weakly scattering structures. Therefore, this type of imaging has been implemented largely using fluorescence techniques. While confocal fluorescence imaging is a common approach to achieve sectioning, it requires fluorescence probes that are often harmful to the living specimen. On the other hand, by using the intrinsic contrast of the structures it is possible to study living cells in a non-invasive manner. One method that provides high-resolution quantitative information about nanoscale structures is a broadband interferometric technique known as Spatial Light Interference Microscopy (SLIM). In addition to rendering quantitative phase information, when combined with a high numerical aperture objective, SLIM also provides excellent depth sectioning capabilities. However, like in all linear optical systems, SLIM's resolution is limited by diffraction. Here we present a novel 3D field deconvolution algorithm that exploits the sparsity of phase images and renders images with resolution beyond the diffraction limit. We employ this label-free method, called deconvolution Spatial Light Interference Tomography (dSLIT), to visualize coiled sub-cellular structures in E. coli cells which are most likely the cytoskeletal MreB protein and the division site regulating MinCDE proteins. Previously these structures have only been observed using specialized strains and plasmids and fluorescence techniques. Our results indicate that dSLIT can be employed to study such structures in a practical and non-invasive manner