32 research outputs found
sj-docx-1-tpx-10.1177_01926233221099273 – Supplemental material for Scientific and Regulatory Policy Committee Points to Consider: Primary Digital Histopathology Evaluation and Peer Review for Good Laboratory Practice (GLP) Nonclinical Toxicology Studies
No full textSupplemental material, sj-docx-1-tpx-10.1177_01926233221099273 for Scientific and Regulatory Policy Committee Points to Consider: Primary Digital Histopathology Evaluation and Peer Review for Good Laboratory Practice (GLP) Nonclinical Toxicology Studies by Thomas Forest, Famke Aeffner, Dinesh S. Bangari, Bhupinder Bawa, Jonathan Carter, James Fikes, Wanda High, Shim-mo Hayashi, Matthew Jacobsen, LuAnn McKinney, Daniel Rudmann, Thomas Steinbach, Vanessa Schumacher, Oliver Turner, Jerrold M. Ward and Cynthia J. Willson in Toxicologic Pathology</p
Fabrication and Characterization of PDMS Waveguides for Flexible Optrodes
No full textAbstract
With the growth of optogenetic research, the demand for optical probes tailored to specific applications is ever rising. Specifically, for applications like the coiled cochlea of the inner ear, where planar, stiff, and nonconformable probes can hardly be used, transitioning from commonly used stiff glass fibers to flexible probes is required, especially for long‐term use. Following this demand, polydimethylsiloxane (PDMS) with its lower Young's modulus compared to glass fibers can serve as material of choice. Hence, the long‐term usability of PDMS as a waveguide material with respect to variations in transmission and refractive index over time is investigated. Different manufacturing methods for PDMS‐based flexible waveguides are established and compared with the aim to minimize optical losses and thus maximize optical output power. Finally, the waveguides with lowest optical losses (−4.8 dB cm
−1
± 1.3 dB cm
−1
at 472 nm) are successfully inserted into the optogenetically modified cochlea of a Mongolian gerbil (
Meriones unguiculatus
), where optical stimuli delivered by the waveguides evoked robust neuronal responses in the auditory pathway.Deutsche Forschungsgemeinschaft https://doi.org/10.13039/50110000165
Using a Murine Erythroleukemia Cell Line to Evaluate the Potential Toxicity of Test Compounds
No full textiv, 36 p.The potential toxicologic effects of test compounds on hematopoiesis are
examined in preclinical animal studies by evaluating bone marrow and peripheral blood
hematology. If effects on bone marrow are observed, these can be further characterized
using in vitro cell culture systems. Murine erythroleukemia cells (MELC) are a well-characterized
cell line that can be induced to differentiate into erythroid progenitors, one
of the cell lineages present in bone marrow. The effects on MELC differentiation, cell
cycle, and Annexin V (AnnV) and propidium iodide (PI) staining were examined with
test compounds that had demonstrated effects on bone marrow in animals. DRUG 1,
DRUG 2, and chloramphenicol caused dose-dependent increases in double-positive
Ann V and PI MELC, indicating that the cells were in late apoptosis or necrosis. HMBA-induced
MELC were more sensitive to the compound-related increases in Ann V / PI
positive cells. The test compounds did not alter differentiation or cell cycle in HMBA-induced
cells. MELC may be prove to be a useful cell line for the study of test
compounds that demonstrate bone marrow toxicity in preclinical animal studies.Pathology Sciences and Services. Pharmacia. Kalamazoo, Michigan
A Comparison of the Toxic Effects of Two Oxazolidinones and Chloramphenicol on a Murine Erythroleukemia Cell Line
No full textiv, 25 p.As a result of the frequent use of penicillin and other common antibiotics,
numerous pathogens have developed mechanisms to evade these agents. Within the last
twenty years, however, a promising new class of drugs has been in development that
offers the potential to target bacteria that have become resistant to these antibiotics. This
class, the oxazolidinones, is a novel class of antimicrobial agents that are bacteriostatic
for Gram-positive bacteria, including those that are resistant to penicillin and other
commonly used antibiotics (Diekema and Jones 2000). Target organisms for the
oxazolidinones include methicillin-resistant Staphylococcus aureus, penicillin-resistant
Streptococcus pneumoniae, and vancomycin-resistant Enterococcus jaecium, among
others (Swaney et al. 1998). The oxazolidinones exhibit a unique mechanism of action
that enables their use for bacteria which are resistant to other antibiotics (Zurenko et ale
1996).
Early studies with the oxazolidinones demonstrated that they inhibit bacterial
protein synthesis, most likely at a step involving the binding of mRNA to the ribosome at
the initiation of translation (Diekema and Jones 2000). The drugs do not affect RNA and
DNA synthesis (Shinabarger et ale 1997), do not inhibit the formation of initiator tRNA,
and do not block the elongation or termination steps of prokaryotic translation
(Shinabarger et ale 1997). The binding of the oxazolidinones to the ribosomal site can be
inhibited by chloramphenicol and lincomycin; however, the oxazolidinones appear to act
through a mechanism that is distinct from the mechanisms of these drugs (Lin et al,
1997). Chloramphenicol and lincomycin act on the 50S subunit to inhibit the peptidyl
transferase reaction and the translation termination reaction. Oxazolidinones have not
been demonstrated to inhibit either peptidyl transferase or translation termination
(Diekema and Jones 1997). Instead, the oxazolidinones bind directly to the 50S
ribosomal subunit and prevent the formation of the 70S initiation complex in bacterial
translation systems (Lin et al. 1997, Swaney et a1. 1998, Burghardt et a1. 1998).
In early studies with the oxazolidinones, two compounds - eperezolid and
linezolid - emerged with good potential to progress to clinical trials because of their
significant in vitro activity against Gram-positive bacteria and their favorable toxicity
profiles (Diekema and Jones 2000). These two compounds also displayed significant
antibacterial activity in humans in vivo (Diekema and Jones 2000). Linezolid was
developed over eperezolid because it exhibited superior pharmacokinetics in phase I trials
in comparison with eperezolid (Diekema and Jones 2000). Recently, linezolid was
approved by the United States Food and Drug Administration for the treatment of Grampositive
skin and soft-tissue infection and community acquired pneumonia (Diekema and
Jones 2000).
Although linezolid has demonstrated an acceptable safety profile, dose- and timedependent,
reversible myelosuppression were observed in preclinical animal studies. The
myelosuppression was characterized by bone marrow hypocellularity, decreased
hematopoiesis, and decreased levels of circulating reticulocytes, erythrocytes, leukocytes,
and platelets. These effects occurred in both dogs and rats at exposure levels comparable
to those that would be used in humans. While the bone marrow effects affected multiple
lineages, the most principle effects seen in human clinical studies were decreased platelet
counts. (U.S. Food and Drug Administration, NDA 21-130,31,32 and Package Insert
2000). Based on this information, the current research focuses on investigating the
effects of two oxazolidinones on a murine erythroleukemia cell line (MEL C) as an in
vitro model of bone marrow erthyroid progenitors.Missing pp. 23-36.Preclinical Toxicology. Pharmacia, Inc. Kalamazoo, Michigan
Abstract 1710: Providing confidence around computational tissue analysis using heterogeneity assessments
No full textAbstract
Background: Though the techniques to interrogate the appearance of a biomarker in tissue sections have greatly advanced, there are limitations as to how representative an analysis of a tissue section is compared to the entire diseased tissue. Depending on the heterogeneous expression level of a biomarker, tissue sampling can result in different interpretations of the biomarker’s appearance, and hence could potentially lead to a false therapeutic intervention.
Hypothesis: Digital image analysis has demonstrated tremendous value in quantifying many features related to biomarker distribution and expression in biological tissues. The information can be collected for various indications and biomarkers and a phenotypic signature can be established that describes a biomarker representation across indications. Moreover, the assessment of new samples can be compared to the established phenotypic signature and a confidence score applied in support to the determined endpoint.
Approach: For a proof of concept, 6 prostate cancer samples were processed and a single section was collected after every 100microns. A total of 7 sections per sample were stained for the lymphocyte marker CD3, and the number of positive target cells were determined in the tumor and tumor microenvironment using tissue Image Analysis (tIA™). To assess how indicative the evaluation of a single tissue section would be for the entire tumor, the heterogeneity level was determined on the section level as well as by random grid analysis on each individual section. Both criteria were utilized to define an indication and biomarker specific confidence interval and heterogeneity score.
Conclusion: The combination of IHC and tIA is a powerful tool to convert complex data into meaningful interpretations. tIA is also a capable tool to catalogue valuable information about the biomarker’s expression pattern across different disease stages and hence could be used to evaluate how representative a single biomarker evaluation is in the grand scheme. Ultimately, we demonstrated a technique that can be applied to any biomarker and would assist in guiding therapeutic decisions.
Citation Format: Carsten Schnatwinkel, Daniel Rudmann, Famke Aeffner, Jasmeet Bajwa, Natalie Hutnick, Michael Sharp, Gerry Chu, JD Alvarez. Providing confidence around computational tissue analysis using heterogeneity assessments [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1710. doi:10.1158/1538-7445.AM2017-1710</jats:p
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Carried meaning in the Mahābhārata
No full textThe Mahābhārata describes itself as both a comprehensive and exhaustive text, incorporating a range of genres while presenting diverse perspectives through a matrix of interacting narratives. Its main story and subtales are the subject of productive contemporary studies that underscore the significance of the Sanskrit epic, though this scholarship is also famously criticized for overlooking literary inquiry. The following dissertation enacts a close reading of four subtales, Nala’s Tale, Rāma’s Tale, Sāvitrī’s Tale, and The Yakṣa’s Questions, in context with the larger work to uncover the implications of a literary study of the Mahābhārata. By conducting translations of passages from the epic, this dissertation builds sites of alliance among frame and subtale, literary and translation theory, critical analysis and contemporary scholarship, as well as the Mahābhārata and other works of literature in order to consider the ways in which meaning is generated throughout the text. Language, constituent parts, and operative principles are found to reverberate in the epic, eschewing didacticism and stasis for literary vitality. Themes of loss, love, disguise, and discovery veer throughout the subtales as sideshadows that at once collaborate and contradict to continuously redefine one another. The Mahābhārata’s self-conscious and reiterative reinterpretation of its own constructs presents critical insights on translation as dialogical correspondence, occurring within utterances as well as between languages. The act of translation, utilized by the poem itself to develop and proliferate significance, reveals difference and bears legibility within the epic.Asian Studie
Using gene-deleted mice to evaluate the role of tumor necrosis factor and gamma-interferon in disease
No full textTumor necrosis factor (TNF) and interferon gamma (IFN) are important proinflammatory cytokines and have multiple roles in immune system development and function. The purpose of the present study was to investigate the individual or combined roles of TNF and IFN in murine models of endotoxemia, asthma, and Pneumocystis carinii pneumonia using wild-type mice (WT) and mice deficient in TNF receptors 1 and 2 (TNFR1-2\sp{{-}/{-}}), IFN (IFN\gamma\sp{{-}/{-}}), and TNFR1, TNFR2, and IFN (TNFR1-2-INF\gamma\sp{{-}/{-}}). TNFR1-2-IFN\gamma\sp{{-}/{-}}mice were more resistant to intrapertioneally administered LPS and D-GalN/LPS as compared to WT, IFN\gamma\sp{{-}/{-}}, and TNFR1-2\sp{{-}/{-}} mice. The resistance to LPS in TNFR1-2-IFN\gamma\sp{{-}/{-}}mice was associated with lower serum concentrations of IL-6. In contrast, compared to WT mice, IFN\gamma\sp{{-}/{-}} and TNFR1-2\sp{{-}/{-}} were not protected from LPS alone, and this lack of protection was associated with elevated serum concentrations of IFN and IL-1 in TNFR1-2\sp{{-}/{-}} mice and higher serum concentrations of TNF in IFN\gamma\sp{{-}/{-}} mice. Thus, TNF and IFN are synergistic protagonists in LPS-mediated lethality and blocking both cytokines significantly improves outcome in murine IP endotoxemia. In a model of asthma, TNFR1-2\sp{{-}/{-}} mice and anti-murine TNF antibody treated WT mice did not have attenuated disease, but, instead had increased serum or lavage IgE levels. Additionally, compared to vehicle-only treated WT mice, WT mice treated chronically with mTNF had less severe pulmonary inflammation as characterized by lower numbers of bronchoalveolar eosinophils, decreased lung IL-4, and lower IL-5 concentrations in supernatants from stimulated lymph node cells. Taken together, TNF is not a critical protagonist in this murine model of asthma, and TNF may have important downregulatory effects in pulmonary inflammation. In a model of P. carinii infection, TNFR1-2-IFN\gamma\sp{{-}/{-}} mice had marked pneumonia at 4 weeks. In contrast, WT, TNFR1-2\sp{{-}/{-}} and IFN\gamma\sp{{-}/{-}} mice cleared infection. Thus, TNF and IFN are critical for a sufficient host response to P. carinii. The results from these three murine models demonstrate that TNF and IFN gave diverse and synergistic roles in the immune response to disease. Furthermore, changing the bioactivity of TNF and INF may be a rational approach for the treatment of a variety of diseases
Abstract 1674: Quantifying tumor-infiltrating leukocytes in hematoxylin stained NSCLC tissue samples using morphometric features
No full textAbstract
Quantification of tumor-infiltrating lymphocytes (TILs) in non-small cell lung cancers (NSCLC) is valuable for understanding patient prognosis and survival. TILs comprise a subset of tumor-infiltrating leukocytes that modulate immune evasion and response to therapy. Understanding the composition of TIL subsets, especially relative to the total tumor leukocyte population, may provide additional context for understanding NSCLC pathogenesis and patient response to treatment. However, availability of tissues and use of chromogenic assays can limit the number of TIL and leukocyte subset markers assayed in a tissue section. Therefore, this study evaluated the identification of total leukocyte component in NSCLC using morphometric parameters and routine TIL marker monoplex immunohistochemistry (IHC) assays to further identify the composition of TIL subsets. Computational Tissue Analysis (cTA™) tools were used to determine the morphometric parameters which could identify immune cells in the absence of biomarker stain. The morphometric features which characterized immune infiltrates were used to quantify the total immune cell population frequency in the tumor nests and surrounding stroma in hematoxylin-stained tissues. The leukocyte population identified with morphometric parameters was correlated with CD45+ cell frequencies identified by cTA based on biomarker staining in CD45-stained serial sections. This morphometric ruleset was then applied to CD3- and CD8-stained tissues to evaluate the frequency of CD3+ and CD8+ TILs in the context of total infiltrating leukocytes. The relative populations of CD3+ and CD8+ TILs were consistent with available literature demonstrating that the morphometric ruleset could be utilized to enable evaluation of TIL sub-types relative to total leukocyte population without the need for additional IHC stains. The approach could, therefore, provide an added dimension of analysis for tissues stained by IHC for identifying the total immune cell infiltrating component without requiring additional biomarker staining.
Citation Format: Elliott Ergon, Allison S. Harney, Nathan Martin, Will Paces, Famke Aeffner, Kristin Wilson, Janet Patterson-Kane, Karen Ryall, Daniel G. Rudmann, Brooke Hirsch, Joseph Krueger. Quantifying tumor-infiltrating leukocytes in hematoxylin stained NSCLC tissue samples using morphometric features [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1674. doi:10.1158/1538-7445.AM2017-1674</jats:p
