Occupancy and Abundance of Stream Salamanders along a Specific Conductance Gradient

In the Central Appalachians (USA), mountaintop-removal mining accompanied by valley fills often leads to streams with elevated specific conductivity (SC). Thus, the ionic composition of freshwaters in this region is hypothesized to be a driver of the distribution and abundance of freshwater taxa, including stream salamanders. We examined the association between SC and stream salamander populations by conducting salamander counts in 30 southeastern Kentucky streams across a continuous gradient of SC that ranged from 30 to 1966 μS/cm. We counted 2319 salamanders across 5 species and, using a hierarchical Bayesian version of the N-mixture model, found a negative association between SC and salamander occupancy rates. This finding was consistent across adults and larvae of the 5 species we examined. Furthermore, we found that most salamander species and life stages showed reduced abundances given occupancy at greater SC levels. For example, estimated mean abundance given occupancy of larval Southern Two-lined Salamanders (Eurycea cirrigera) was 67.69 (95% credible interval 48.31–98.25) ind/10 m at 250 μS/cm and 2.30 (95% credible interval 1.46–3.93) ind/10 m at 2000 μS/cm. The consistent negative association across all species and life stages supports the hypothesis that salamander distributions and abundances are negatively associated with elevated SC of streams in southeastern Kentucky, even though physical and chemical environmental attributes, such as forest cover within stream catchments, were correlated with SC. Restoration of streams affected by mountaintop-removal mining should focus on restoring the ionic compositions that naturally occur in this region

Genomic Identification of the TOR Signaling Pathway as a Target of the Plant Alkaloid Antofine in the Phytopathogen Fusarium graminearum

Antofine, a phenanthroindolizidine alkaloid, is a bioactive natural product isolated from milkweeds that exhibits numerous biological activities, including anticancer, antimicrobial, antiviral, and anti-inflammatory properties. However, the direct targets and mode of action of antofine have not been determined. In this report, we show that antofine displays antifungal properties against the phytopathogen Fusarium graminearum, the cause of Fusarium head blight disease (FHB). FHB does devastating damage to agriculture, causing billions of dollars in economic losses annually. We therefore sought to understand the mode of action of antofine in F. graminearum using insights from yeast chemical genomic screens. We used haploinsufficiency profiling (HIP) to identify putative targets of antofine in yeast and identified three candidate targets, two of which had homologs in F. graminearum The Fusarium homologues of two targets, glutamate dehydrogenase (FgGDH) and resistance to rapamycin deletion 2 (FgRRD2), can bind antofine. Of the two genes, only the Fgrrd2 knockout displayed a los

Kinomic exploration of temozolomide and radiation resistance in Glioblastoma multiforme xenolines

Glioblastoma multiforme (GBM) represents the most common and deadly primary brain malignancy, particularly due to temozolomide (TMZ) and radiation (RT) resistance. To better understand resistance mechanisms, we examined global kinase activity (kinomic profiling) in both treatment sensitive and resistant human GBM patient-derived xenografts (PDX or “xenolines”)

Kinomic Profiling of Electromagnetic Navigational Bronchoscopy Specimens: A New Approach for Personalized Medicine

Purpose Researchers are currently seeking relevant lung cancer biomarkers in order to make informed decisions regarding therapeutic selection for patients in so-called “precision medicine.” However, there are challenges to obtaining adequate lung cancer tissue for molecular analyses. Furthermore, current molecular testing of tumors at the genomic or transcriptomic level are very indirect measures of biological response to a drug, particularly for small molecule inhibitors that target kinases. Kinase activity profiling is therefore theorized to be more reflective of in vivo biology than many current molecular analysis techniques. As a result, this study seeks to prove the feasibility of combining a novel minimally invasive biopsy technique that expands the number of lesions amenable for biopsy with subsequent ex vivo kinase activity analysis. Methods Eight patients with lung lesions of varying location and size were biopsied using the novel electromagnetic navigational bronchoscopy (ENB) technique. Basal kinase activity (kinomic) profiles and ex vivo interrogation of samples in combination with tyrosine kinase inhibitors erlotinib, crizotinib, and lapatinib were performed by PamStation 12 microarray analysis. Results Kinomic profiling qualitatively identified patient specific kinase activity profiles as well as patient and drug specific changes in kinase activity profiles following exposure to inhibitor. Thus, the study has verified the feasibility of ENB as a method for obtaining tissue in adequate quantities for kinomic analysis and has demonstrated the possible use of this tissue acquisition and analysis technique as a method for future study of lung cancer biomarkers. Conclusions We demonstrate the feasibility of using ENB-derived biopsies to perform kinase activity assessment in lung cancer patients

MARCKS regulates growth and radiation sensitivity and is a novel prognostic factor for glioma

Purpose This study assessed whether Myristoylated Alanine Rich C-Kinase Substrate (MARCKS) can regulate glioblastoma (GBM) growth, radiation sensitivity and clinical outcome. Experimental Design MARCKS protein levels were analyzed in five GBM explant cell lines and eight patient-derived xenograft tumors by immunoblot, and these levels were correlated to proliferation rates and intracranial growth rates, respectively. Manipulation of MARCKS protein levels was assessed by lentiviral-mediated shRNA knockdown in the U251 cell line and MARCKS over-expression in the U87 cell line. The effect of manipulation of MARCKS on proliferation, radiation sensitivity and senescence was assessed. MARCKS gene expression was correlated with survival outcomes in the Repository of Molecular Brain Neoplasia Data (REMBRANDT) Database and The Cancer Genome Atlas (TCGA). Results MARCKS protein expression was inversely correlated with GBM proliferation and intracranial xenograft growth rates. Genetic silencing of MARCKS promoted GBM proliferation and radiation resistance, while MARCKS overexpression greatly reduced GBM growth potential and induced senescence. We found MARCKS gene expression to be directly correlated with survival in both the REMBRANDT and TCGA databases. Specifically, patients with high MARCKS expressing tumors of the Proneural molecular subtype had significantly increased survival rates. This effect was most pronounced in tumors with unmethylated O6-methylguanine DNA methyltransferase (MGMT) promoters, a traditionally poor prognostic factor. Conclusions MARCKS levels impact GBM growth and radiation sensitivity. High MARCKS expressing GBM tumors are associated with improved survival, particularly with unmethylated MGMT promoters. These findings suggest the use of MARCKS as a novel target and biomarker for prognosis in the Proneural subtype of GBM

Targeting the effector domain of the myristoylated alanine rich C-kinase substrate enhances lung cancer radiation sensitivity

Lung cancer is the leading cause of cancer related deaths. Common molecular drivers of lung cancer are mutations in receptor tyrosine kinases (RTKs) leading to activation of the phosphatidylinositol 3-kinase (PI3K)/Akt pro-growth, pro-survival signaling pathways. Myristoylated alanine rich C-kinase substrate (MARCKS) is a protein that has the ability to mitigate this signaling cascade by sequestering the target of PI3K, phosphatidylinositol (4,5)-bisphosphate (PIP2). As such, MARCKS has been implicated as a tumor suppressor, though there is some evidence that MARCKS may be tumor promoting in certain cancer types. Since the MARCKS function depends on its phosphorylation status, which impacts its subcellular location, MARCKS role in cancer may depend highly on the signaling context. Currently, the importance of MARCKS in lung cancer biology is limited. Thus, we investigated MARCKS in both clinical specimens and cell culture models. Immunohistochemistry scoring of MARCKS protein expression in a diverse lung tumor tissue array revealed that the majority of squamous cell carcinomas stained positive for MARCKS while other histologies, such as adenocarcinomas, had lower levels. To study the importance of MARCKS in lung cancer biology, we used inducible overexpression of wild-type (WT) and non-phosphorylatable (NP)-MARCKS in A549 lung cancer cells that had a low level of endogenous MARCKS. We found that NP-MARCKS expression, but not WT-MARCKS, enhanced the radiosensitivity of A549 cells in part by inhibiting DNA repair as evidenced by prolonged radiation-induced DNA double strand breaks. We confirmed the importance of MARCKS phosphorylation status by treating several lung cancer cell lines with a peptide mimetic of the phosphorylation domain, the effector domain (ED), which effectively attenuated cell growth as measured by cell index. Thus, the MARCKS ED appears to be an important target for lung cancer therapeutic development

Interlaboratory Evaluation of in Vitro Cytotoxicity and Inflammatory Responses to Engineered Nanomaterials: The NIEHS Nano GO Consortium

Background: Differences in interlaboratory research protocols contribute to the conflicting data in the literature regarding engineered nanomaterial (ENM) bioactivity. Objectives: Grantees of a National Institute of Health Sciences (NIEHS)-funded consortium program performed two phases of in vitro testing with selected ENMs in an effort to identify and minimize sources of variability. Methods: Consortium program participants (CPPs) conducted ENM bioactivity evaluations on zinc oxide (ZnO), three forms of titanium dioxide (TiO2), and three forms of multiwalled carbon nanotubes (MWCNTs). In addition, CPPs performed bioassays using three mammalian cell lines (BEAS-2B, RLE-6TN, and THP-1) selected in order to cover two different species (rat and human), two different lung epithelial cells (alveolar type II and bronchial epithelial cells), and two different cell types (epithelial cells and macrophages). CPPs also measured cytotoxicity in all cell types while measuring inflammasome activation [interleukin-1β (IL-1β) release] using only THP-1 cells. Results: The overall in vitro toxicity profiles of ENM were as follows: ZnO was cytotoxic to all cell types at ≥ 50 μg/mL, but did not induce IL-1β. TiO2 was not cytotoxic except for the nanobelt form, which was cytotoxic and induced significant IL-1β production in THP-1 cells. MWCNTs did not produce cytotoxicity, but stimulated lower levels of IL-1β production in THP-1 cells, with the original MWCNT producing the most IL-1β. Conclusions: The results provide justification for the inclusion of mechanism-linked bioactivity assays along with traditional cytotoxicity assays for in vitro screening. In addition, the results suggest that conducting studies with multiple relevant cell types to avoid false-negative outcomes is critical for accurate evaluation of ENM bioactivity

'To live and die [for] Dixie': Irish civilians and the Confederate States of America

Around 20,000 Irishmen served in the Confederate army in the Civil War. As a result, they left behind, in various Southern towns and cities, large numbers of friends, family, and community leaders. As with native-born Confederates, Irish civilian support was crucial to Irish participation in the Confederate military effort. Also, Irish civilians served in various supporting roles: in factories and hospitals, on railroads and diplomatic missions, and as boosters for the cause. They also, however, suffered in bombardments, sieges, and the blockade. Usually poorer than their native neighbours, they could not afford to become 'refugees' and move away from the centres of conflict. This essay, based on research from manuscript collections, contemporary newspapers, British Consular records, and Federal military records, will examine the role of Irish civilians in the Confederacy, and assess the role this activity had on their integration into Southern communities. It will also look at Irish civilians in the defeat of the Confederacy, particularly when they came under Union occupation. Initial research shows that Irish civilians were not as upset as other whites in the South about Union victory. They welcomed a return to normalcy, and often 'collaborated' with Union authorities. Also, Irish desertion rates in the Confederate army were particularly high, and I will attempt to gauge whether Irish civilians played a role in this. All of the research in this paper will thus be put in the context of the Drew Gilpin Faust/Gary Gallagher debate on the influence of the Confederate homefront on military performance. By studying the Irish civilian experience one can assess how strong the Confederate national experiment was. Was it a nation without a nationalism

A qualitative study of cardiovascular disease risk communication in NHS Health Check using different risk calculators: protocol for the RIsk COmmunication in NHS Health Check (RICO) study. BMC family practice, 20(1), 11.

Background NHS Health Check is a national cardiovascular disease (CVD) risk assessment programme for 40–74 year olds in England, in which practitioners should assess and communicate CVD risk, supported by appropriate risk-management advice and goal-setting. This requires effective communication, to equip patients with knowledge and intention to act. Currently, the QRISK®2 10-year CVD risk score is most common way in which CVD risk is estimated. Newer tools, such as JBS3, allow manipulation of risk factors and can demonstrate the impact of positive actions. However, the use, and relative value, of these tools within CVD risk communication is unknown. We will explore practitioner and patient CVD risk perceptions when using QRISK®2 or JBS3, the associated advice or treatment offered by the practitioner, and patients’ responses. Methods RIsk COmmunication in NHS Health Check (RICO) is a qualitative study with quantitative process evaluation. Twelve general practices in the West Midlands of England will be randomised to one of two groups: usual practice, in which practitioners use QRISK®2 to assess and communicate CVD risk; intervention, in which practitioners use JBS3. Twenty Health Checks per practice will be video-recorded (n = 240, 120 per group), with patients stratified by age, gender and ethnicity. Post-Health Check, video-stimulated recall (VSR) interviews will be conducted with 48 patients (n = 24 per group) and all practitioners (n = 12–18), using video excerpts to enhance participant recall/reflection. Patient medical record reviews will detect health-protective actions in the first 12-weeks following a Health Check (e.g., lifestyle referrals, statin prescription). Risk communication, patient response and intentions for health-protective behaviours in each group will be explored through thematic analysis of video-recorded Health Checks (using Protection Motivation Theory as a framework) and VSR interviews. Process evaluation will include between-group comparisons of quantitatively coded Health Check content and post-Health Check patient outcomes. Finally, 10 patients with the most positive intentions or behaviours will be selected for case study analysis (using all data sources). Discussion This study will produce novel insights about the utility of QRISK®2 and JBS3 to promote patient and practitioner understanding and perception of CVD risk and associated implications for patient intentions with respect to health-protective behaviours (and underlying mechanisms). Recommendations for practice will be developed

Can auditors be independent? – Experimental evidence on the effects of client type

Recent regulatory initiatives stress that an independent oversight board, rather than the management board, should be the client of the auditor. In an experiment, we test whether the type of client affects auditors’ independence. Unique features of the German institutional setting enable us to realistically vary the type of auditors’ client as our treatment variable: we portray the client either as the management preferring aggressive accounting or the oversight board preferring conservative accounting. We measure auditors’ perceived client retention incentives and accountability pressure in a post-experiment questionnaire to capture potential threats to independence. We find that the type of auditors’ client affects auditors’ behaviour contingent on the degree of the perceived threats to independence. Our findings imply that both client retention incentives and accountability pressure represent distinctive threats to auditors’ independence and that the effectiveness of an oversight board in enhancing auditors’ independence depends on the underlying threat