Knuckle Sandwich & Other Stories

Knuckle Sandwich & Other Stories is a collection of fiction written in the years between 2005 and 2008. The characters in these stories find themselves trying--and often failing--to cope with loss: loss of a romantic relationship, of a loved one, of youth, of innocence

Knuckle Sandwich & Other Stories

Knuckle Sandwich & Other Stories is a collection of fiction written in the years between 2005 and 2008. The characters in these stories find themselves trying--and often failing--to cope with loss: loss of a romantic relationship, of a loved one, of youth, of innocence

Pleiotropic and Novel Phenotypes in The \u3cem\u3eDrosophila\u3c/em\u3e Gut Caused by Mutation of \u3cem\u3eDrop-Dead\u3c/em\u3e

Normal gut function is vital for animal survival, and deviations from such function can contribute to malnutrition, inflammation, increased susceptibility to pathogens, diabetes, neurodegenerative diseases, and cancer. In the fruit fly Drosophila melanogaster, mutation of the gene drop-dead (drd) results in defective gut function, as measured by enlargement of the crop and reduced food movement through the gut, and drd mutation also causes the unrelated phenotypes of neurodegeneration, early adult lethality and female sterility. In the current work, adult drd mutant flies are also shown to lack the peritrophic matrix (PM), an extracellular barrier that lines the lumen of the midgut and is found in many insects including flies, mosquitos and termites. The use of a drd-gal4 construct to drive a GFP reporter in late pupae and adults revealed drd expression in the anterior cardia, which is the site of PM synthesis in Drosophila. Moreover, the ability of drd knockdown or rescue with several gal4 drivers to recapitulate or rescue the gut phenotypes (lack of a PM, reduced defecation, and reduced adult survival 10–40 days post-eclosion) was correlated to the level of expression of each driver in the anterior cardia. Surprisingly, however, knocking down drd expression only in adult flies, which has previously been shown not to affect survival, eliminated the PM without reducing defecation rate. These results demonstrate that drd mutant flies have a novel phenotype, the absence of a PM, which is functionally separable from the previously described gut dysfunction observed in these flies. As the first mutant Drosophila strain reported to lack a PM, drd mutants will be a useful tool for studying the synthesis of this structure

Electrical coupling between ventricular myocytes and myofibroblasts in the infarcted mouse heart

Aims: Recent studies have demonstrated electrotonic coupling between scar tissue and the surrounding myocardium in cryoinjured hearts. However, the electrical dynamics occurring at the myocyte-nonmyocyte interface in the fibrotic heart remain undefined. Here, we sought to develop an assay to interrogate the nonmyocyte cell type contributing to heterocellular coupling and to characterize, on a cellular scale, its voltage response in the infarct border zone of living hearts. Methods and results: We used two-photon laser scanning microscopy in conjunction with a voltage-sensitive dye to record transmembrane voltage changes simultaneously from cardiomyocytes and adjoined nonmyocytes in Langendorff-perfused mouse hearts with healing myocardial infarction. Transgenic mice with cardiomyocyte-restricted expression of a green fluorescent reporter protein underwent permanent coronary artery ligation and their hearts were subjected to voltage imaging 7-10 days later. Reporter-negative cells, i.e. nonmyocytes, in the infarct border zone exhibited depolarizing transients at a 1:1 coupling ratio with action potentials recorded simultaneously from adjacent, reporter-positive ventricular myocytes. The electrotonic responses in the nonmyocytes exhibited slower rates of de- and repolarization compared to the action potential waveform of juxtaposed myocytes. Voltage imaging in infarcted hearts expressing a fluorescent reporter specifically in myofibroblasts revealed that the latter were electrically coupled to border zone myocytes. Their voltage transient properties were indistinguishable from those of nonmyocytes in hearts with cardiomyocyte-restricted reporter expression. The density of connexin43 expression at myofibroblast-cardiomyocyte junctions was ∼5% of that in the intercalated disc regions of paired ventricular myocytes in the remote, uninjured myocardium, whereas the ratio of connexin45 to connexin43 expression levels at heterocellular contacts was ∼1%. Conclusion: Myofibroblasts contribute to the population of electrically coupled nonmyocytes in the infarct border zone. The slower kinetics of myofibroblast voltage responses may reflect low electrical conductivity across heterocellular junctions, in accordance with the paucity of connexin expression at myofibroblast-cardiomyocyte contacts

Aerobic Cometabolism of 1,4-Dioxane and cis-Dichloroethylene by Gellan Gum and Polyvinyl Alcohol/Sodium Alginate Hydrogel Beads Co-Encapsulated with Rhodococcus rhodochrous Strain ATCC 21198 and Slow-Release Growth Substrates: Batch, Column, and Toxicity Studies

1,4-Dioxane (dioxane) and cis-dichloroethylene (cDCE) are compounds commonly found in industrial cleaning and degreasing agents that are frequently present as groundwater contaminants. In an effort to develop a more effective treatment method for these compounds, hydrogel beads were fabricated with either gellan gum or a combination of polyvinyl alcohol (PVA) and sodium alginate (SA) co-encapsulated with Rhodococcus rhodochrous Strain ATCC 21198 (21198) and a slow-release growth substrate (SRS). The hydrogel beads were investigated in batch reactors and column systems to determine the long-term ability of these beads to successfully transform dioxane and cDCE through aerobic cometabolism. Tetra-butylorthosilicate (TBOS) and tetra-s-butylorthosilicate (T2BOS) were used to slowly release 1-butanol and 2-butanol, respectively, as growth substrates for 21198. The batch reactor studies conducted using gellan gum hydrogel beads co-encapsulated with 21198 and either TBOS or T2BOS demonstrated an ability to transform essentially all (>99%) of the added dioxane at a range of ~250-1000 µg/L and the added cDCE at ~250 µg/L throughout several additions. The beads containing TBOS showed significantly faster rates of transformation for both contaminants. An embryonic zebrafish toxicity study completed using the TBOS gellan gum beads showed no significant toxicity was generated after the complete transformation of cDCE. A column study was completed by packing a glass column with gellan gum beads co-encapsulated with 21198 and T2BOS. The influent media was composed of 10x diluted phosphate buffer mineral salt media (MSM) and added contaminants. Hydrogen peroxide (H2O2) was added to the influent media at concentrations of 25-100 mg/L to meet oxygen demands. The column had a hydraulic retention time (HRT) of 26 hours. This column initially demonstrated the ability to transform essentially all (>99%) influent cDCE (~300 µg/L) from 52 to 75 pore volumes (PV), while showing no evidence of transforming influent dioxane (~200 µg/L). A high-flow study was completed in which the influent flow rate was increased from 1 mL/hr to 90 mL/hr. This study provided evidence that cometabolism of cDCE was occurring due to the presence of cDCE epoxide in the effluent, but it also exposed the entire bead pack to the epoxide. This epoxide likely damaged the health of the column, and the column was unable to recover from this high-flow transient test and a subsequent stop-flow study. This was evidenced by a decrease in cDCE transformation. By the end of the column’s operation at 118 PV, a mass balance determined that a total of 71.5% of the influent cDCE was transformed. However, none of the influent dioxane was transformed. A column study was also completed by packing a glass column with side sampling ports with PVA/SA beads co-encapsulated with 21198 and TBOS. The influent media was made using a synthetic groundwater formulation and 200x diluted phosphate buffer MSM for additional nutrients. H2O2 was added at increasing concentrations (50-250 mg/L) over the operating period to meet oxygen demands. The influent pH was increased from 7.0 to 7.5 to 7.9 during the operating period in an effort to lower the rate of TBOS hydrolysis to 1-butanol. The column initially had an HRT of 28 hours from 0 to 7 PV. The flow rate was then doubled and the column had an HRT of 14 hours for the final 90 PV. The column demonstrated the ability to transform >95% of influent cDCE (~200 µg/L) when the influent media contained 250 mg/L H2O2 and was at a pH of 7.5. Side port sampling showed the presence of cDCE epoxide throughout the column, signifying the aerobic cometabolism of cDCE. There was evidence that nearly all of the transformation occurred directly at the influent end of the column. A visual orange tint, representing significant 21198 biostimulation in the bead, was evident mainly in that end of the column. By the end of the column’s operating period, samples from the lower side sampling port located near the influent also measured nearly equal in cDCE concentration to the column effluent, indicating that the majority of cometabolic transformation was occurring where visual biomass growth was observed. 1-butanol remained >50 mg/L in the effluent throughout the column’s operating period, indicating biostimulation did not occur throughout the column. Embryonic zebrafish toxicity studies demonstrated that the column effluent was significantly toxic when 1-butanol levels were ~120 mg/L and not significantly toxic when ~60 mg/L, suggesting that 1-butanol was the source of toxicity. More detailed studies are needed to establish factors causing the observed toxicity in the column effluent

Violent Extremism and Terrorism Online in 2023: The Year in Review

This report describes and discusses developments in the violent extremist and terrorist online scene(s) in the 12-month period from 1 January 2023 to 31 December 2023. It accomplishes this by surveying, synopsising, and integrating the findings of relevant articles and reports produced by academics, think-tanks, civil society, and governmental organisations; high quality press reports; and the first-hand online experience and primary research of the authors. The report is divided into four major parts: Part I focuses on the online aftermath of Hamas’ massive marauding terrorist attack in southern Israel on 7 October 2023; Part II on online Jihadism, particularly that linked to al-Qaeda and the so-called ‘Islamic State’ (hereafter IS); Part III focuses on the online activities of the extreme right, particularly its European and US variants; and Part IV details the responses of a range of legislators and Internet companies to online extremist and terrorist activity over the course of the last 12 months. We conclude by pointing to a five issues in the violent extremism and terrorism online realm(s) that bear watching in 2024

Image analysis as an adjunct to manual HER-2 immunohistochemical review: a diagnostic tool to standardize interpretation

Dobson L, Conway C, Hanley A, Johnson A, Costello S, O’Grady A, Connolly Y, Magee H, O’Shea D, Jeffers M & Kay E (2010) Histopathology57, 27–38 Image analysis as an adjunct to manual HER-2 immunohistochemical review: a diagnostic tool to standardize interpretatio

Intravital FRAP imaging using an E-cadherin-GFP mouse reveals disease- and drug-dependent dynamic regulation of cell-cell junctions in live tissue

E-cadherin-mediated cell-cell junctions play a prominent role in maintaining the epithelial architecture. The disruption or deregulation of these adhesions in cancer can lead to the collapse of tumor epithelia that precedes invasion and subsequent metastasis. Here we generated an E-cadherin-GFP mouse that enables intravital photobleaching and quantification of E-cadherin mobility in live tissue without affecting normal biology. We demonstrate the broad applications of this mouse by examining E-cadherin regulation in multiple tissues, including mammary, brain, liver, and kidney tissue, while specifically monitoring E-cadherin mobility during disease progression in the pancreas. We assess E-cadherin stability in native pancreatic tissue upon genetic manipulation involving Kras and p53 or in response to anti-invasive drug treatment and gain insights into the dynamic remodeling of E-cadherin during in situ cancer progression. FRAP in the E-cadherin-GFP mouse, therefore, promises to be a valuable tool to fundamentally expand our understanding of E-cadherin-mediated events in native microenvironments

Three-dimensional organotypic matrices from alternative collagen sources as pre-clinical models for cell biology.

Organotypic co-cultures bridge the gap between standard two-dimensional culture and mouse models. Such assays increase the fidelity of pre-clinical studies, to better inform lead compound development and address the increasing attrition rates of lead compounds within the pharmaceutical industry, which are often a result of screening in less faithful two-dimensional models. Using large-scale acid-extraction techniques, we demonstrate a step-by-step process to isolate collagen I from commercially available animal byproducts. Using the well-established rat tail tendon collagen as a benchmark, we apply our novel kangaroo tail tendon collagen as an alternative collagen source for our screening-ready three-dimensional organotypic co-culture platform. Both collagen sources showed equal applicability for invasive, proliferative or survival assessment of well-established cancer models and clinically relevant patient-derived cancer cell lines. Additional readouts were also demonstrated when comparing these alternative collagen sources for stromal contributions to stiffness, organization and ultrastructure via atomic force microscopy, second harmonic generation imaging and scanning electron microscopy, among other vital biological readouts, where only minor differences were found between the preparations. Organotypic co-cultures represent an easy, affordable and scalable model to investigate drug responses within a physiologically relevant 3D platform