First evidence of industrial fly-ash in an Antarctic ice core.

Spheroidal carbonaceous particles (SCPs) are a component of fly-ash, the particulate by-product of industrial high temperature combustion of fuel-oil and coal-series fuels. We provide the first evidence that these indelible markers of industrialisation have been deposited in Antarctic ice, thousands of kilometres from any potential source. The earliest observed particle was deposited in an ice layer from 1936 CE. While depositional fluxes are low, chemical analysis of individual SCPs indicates a coal combustion origin.

PDGFRß promotes oncogenic progression via STAT3/STAT5 hyperactivation in anaplastic large cell lymphoma.

BACKGROUND: Anaplastic large cell lymphoma (ALCL) is an aggressive non-Hodgkin T cell lymphoma commonly driven by NPM-ALK. AP-1 transcription factors, cJUN and JUNb, act as downstream effectors of NPM-ALK and transcriptionally regulate PDGFRß. Blocking PDGFRß kinase activity with imatinib effectively reduces tumor burden and prolongs survival, although the downstream molecular mechanisms remain elusive. METHODS AND RESULTS: In a transgenic mouse model that mimics PDGFRß-driven human ALCL in vivo, we identify PDGFRß as a driver of aggressive tumor growth. Mechanistically, PDGFRß induces the pro-survival factor Bcl-xL and the growth-enhancing cytokine IL-10 via STAT5 activation. CRISPR/Cas9 deletion of both STAT5 gene products, STAT5A and STAT5B, results in the significant impairment of cell viability compared to deletion of STAT5A, STAT5B or STAT3 alone. Moreover, combined blockade of STAT3/5 activity with a selective SH2 domain inhibitor, AC-4-130, effectively obstructs tumor development in vivo. CONCLUSIONS: We therefore propose PDGFRß as a novel biomarker and introduce PDGFRß-STAT3/5 signaling as an important axis in aggressive ALCL. Furthermore, we suggest that inhibition of PDGFRß or STAT3/5 improve existing therapies for both previously untreated and relapsed/refractory ALK+ ALCL patients.

Breast implant associated anaplastic large cell lymphoma: The UK experience. Recommendations on its management and implications for informed consent.

BACKGROUND: Breast implant-associated anaplastic large-cell lymphoma (BIA-ALCL) is a rare, Non-Hodgkin lymphoma arising in the capsule of breast implants. BIA-ALCL presents as a recurrent effusion and/or mass. Tumours exhibit CD30 expression and are negative for Anaplastic Lymphoma Kinase (ALK). We report the multi-disciplinary management of the UK series and how the stage of disease may be used to stratify treatment. METHODS: Between 2012 and 2016, 23 cases of BIA-ALCL were diagnosed in 15 regional centres throughout the UK. Data on breast implant surgeries, clinical features, treatment and follow-up were available for 18 patients. RESULTS: The mean lead-time from initial implant insertion to diagnosis was 10 years (range: 3-16). All cases were observed in patients with textured breast implants or expanders. Fifteen patients with breast implants presented with stage I disease (capsule confined), and were treated with implant removal and capsulectomy. One patient received adjuvant chest-wall radiotherapy. Three patients presented with extra-capsular masses (stage IIA). In addition to explantation, capsulectomy and excision of the mass, all patients received neo-/adjuvant chemotherapy with CHOP as first line. One patient progressed on CHOP but achieved pathological complete response (pCR) with Brentuximab Vedotin. After a mean follow-up of 23 months (range: 1-56) all patients reported here remain disease-free. DISCUSSION: BIA-ALCL is a rare neoplasm with a good prognosis. Our data support the recommendation that stage I disease be managed with surgery alone. Adjuvant chemotherapy may be required for more invasive disease and our experience has shown the efficacy of Brentuximab as a second line treatment.

Lysolecithin as feed additive enhances collagen expression and villus length in the jejunum of broiler chickens.

Adding lysolecithin to feed has reportedly improved the performance of broiler chickens. Lysolecithin is generated by phospholipase catalyzed hydrolysis of lecithin. The enzymatic reaction converts various phospholipids into the corresponding lysophospholipids, with lysophosphatidylcholine (LPC) one of the primary products. Here we compared supplementation with a commercial lysolecithin (Lysoforte®) with comparable levels of highly purified LPC for effects on broilers. Despite no differences in weight gain during the starter period, we discovered a significant increase in average villus length with lysolecithin and an increase in villus width with purified LPC. High-throughput gene expression microarray analyses revealed many more genes were regulated in the epithelium of the jejunum by lysolecithin compared to purified LPC. The most up-regulated genes and pathways were for collagen, extracellular matrix, and integrins. Staining sections of the jejunum with Picrosirius Red confirmed the increased deposition of collagen fibrils in the villi of broilers fed lysolecithin, but not purified LPC. Thus, lysolecithin elicits gene expression in the intestinal epithelium, leading to enhanced collagen deposition and villus length. Purified LPC alone as a supplement does not mimic these responses. Feed supplementation with lysolecithin triggers changes in the intestinal epithelium with the potential to improve overall gut health and performance.

Using public participation to sample trace metals in lake surface sediments: the OPAL Metals Survey.

Members of the public in England were invited in 2010 to take part in a national metals survey, by collecting samples of littoral sediment from a standing water body for geochemical analysis. To our knowledge, this is the first national sediment metals survey using public participation and reveals a snapshot of the extent of metals contamination in ponds and lakes across England. Hg, Ni, Cu, Zn and Pb concentrations exceeding sediment quality guidelines for the health of aquatic biota are ubiquitous in ponds and lakes, not just in areas with a legacy of industrial activity. To validate the public sampling approach, a calibration exercise was conducted at ten water bodies selected to represent a range of lakes found across England. Sediment concentrations of Hg, Ni, Cu, Zn and Pb were measured in samples of soil, stream and littoral and deep water sediment to assess inputs. Significant differences between littoral sediment metal concentrations occur due to local variability, but also organic content, especially in upland, peat soil catchments. Variability of metal concentrations between littoral samples is shown to be low in small (<20 ha) lowland lakes. Larger and upland lakes with more complex inputs and variation in organic content of littoral samples have a greater variability. Collection of littoral sediments in small lakes and ponds, with or without voluntary participation, can provide a reliable sampling technique for the preliminary assessment of metal contamination in standing waters. However, the heterogeneity of geology, soils and history/extent of metal contamination in the English landscape, combined with the random nature of sample collection, shows that systematic sampling for evaluating the full extent of metal contamination in lakes is still required.

Quality control in public participation assessments of water quality: the OPAL Water Survey.

BACKGROUND: Public participation in scientific data collection is a rapidly expanding field. In water quality surveys, the involvement of the public, usually as trained volunteers, generally includes the identification of aquatic invertebrates to a broad taxonomic level. However, quality assurance is often not addressed and remains a key concern for the acceptance of publicly-generated water quality data. The Open Air Laboratories (OPAL) Water Survey, launched in May 2010, aimed to encourage interest and participation in water science by developing a 'low-barrier-to-entry' water quality survey. During 2010, over 3000 participant-selected lakes and ponds were surveyed making this the largest public participation lake and pond survey undertaken to date in the UK. But the OPAL approach of using untrained volunteers and largely anonymous data submission exacerbates quality control concerns. A number of approaches were used in order to address data quality issues including: sensitivity analysis to determine differences due to operator, sampling effort and duration; direct comparisons of identification between participants and experienced scientists; the use of a self-assessment identification quiz; the use of multiple participant surveys to assess data variability at single sites over short periods of time; comparison of survey techniques with other measurement variables and with other metrics generally considered more accurate. These quality control approaches were then used to screen the OPAL Water Survey data to generate a more robust dataset. RESULTS: The OPAL Water Survey results provide a regional and national assessment of water quality as well as a first national picture of water clarity (as suspended solids concentrations). Less than 10 % of lakes and ponds surveyed were 'poor' quality while 26.8 % were in the highest water quality band. CONCLUSIONS: It is likely that there will always be a question mark over untrained volunteer generated data simply because quality assurance is uncertain, regardless of any post hoc data analyses. Quality control at all stages, from survey design, identification tests, data submission and interpretation can all increase confidence such that useful data can be generated by public participants.

Normal table of embryonic development in the four-toed salamander, Hemidactylium scutatum.

We present a complete staging table of normal development for the lungless salamander, Hemidactylium scutatum (Caudata: Plethodontidae). Terrestrial egg clutches from naturally ovipositing females were collected and maintained at 15 °C in the laboratory. Observations, photographs, and time-lapse movies of embryos were taken throughout the 45-day embryonic period. The complete normal table of development for H. scutatum is divided into 28 stages and extends previous analyses of H. scutatum embryonic development (Bishop, 1920; Humphrey, 1928). Early embryonic stage classifications through neurulation reflect criteria described for Xenopus laevis, Ambystoma maculatum and other salamanders. Later embryonic stage assignments are based on unique features of H. scutatum embryos. Additionally, we provide morphological analysis of gastrulation and neurulation, as well as details on external aspects of eye, gill, limb, pigmentation, and tail development to support future research related to phylogeny, comparative embryology, and molecular mechanisms of development.

Anaplastic large cell lymphoma-propagating cells are detectable by side population analysis and possess an expression profile reflective of a primitive origin.

Cancer stem cells or tumour-propagating cells (TPCs) have been identified for a number of cancers, but data pertaining to their existence in lymphoma so far remain elusive. We show for the first time that a small subset of cells purified from human anaplastic lymphoma kinase (ALK)-positive and -negative, anaplastic large cell lymphoma cell lines and primary patient tumours using the side population (SP) technique have serial tumour-propagating capacity both in vitro and in vivo; they give rise to both themselves and the bulk tumour population as well as supporting growth of the latter through the production of soluble factors. In vivo serial dilution assays utilising a variety of model systems inclusive of human cell lines, primary human tumours and nucleophosmin (NPM)-ALK-induced murine tumours demonstrate the TPC frequency to vary from as many as 1/54 to 1/1336 tumour cells. In addition, the SP cells express higher levels of pluripotency-associated transcription factors and are enriched for a gene expression profile consistent with early thymic progenitors. Finally, our data show that the SP cells express higher levels of the NPM-ALK oncogene and are sensitive to an ALK inhibitor.

Bcr-Abl-mediated redox regulation of the PI3K/AKT pathway.

Bcr-Abl causes chronic myelogenous leukemia, a myeloproliferative disorder characterized by clonal expansion of hematopoietic progenitor cells. In this study, inducible expression of Bcr-Abl in TonB.210 cells is associated with increased production of intracellular reactive oxygen species (ROS), which is thought to play a role in survival signaling when generated at specific levels. Elevated ROS in Bcr-Abl-expressing cells were found to activate PI3k/Akt pathway members such as Akt and GSK3beta as well as downstream targets beta-catenin and Mcl-1. The activation of these proteins was inhibited by the flavoprotein inhibitor diphenyleneiodonium, which is commonly used to inhibit NADPH oxidase (Nox). This indicated that increased ROS might be related to increased activity of one member of the Nox family. Knock-down experiments using siRNA suggest that Nox-4 is the main source of increased ROS following Bcr-Abl expression. We showed that Bcr-Abl-induced ROS could also increase survival pathway signaling through redox inhibition of PP1alpha, a serine threonine phosphatase that negatively regulates the PI3k/Akt pathway. Overall our results demonstrate that Bcr-Abl expression increases Nox-4-generated ROS, which in turn increases survival signaling through PI3k/Akt pathway by inhibition of PP1alpha, thus contributing to the high level of resistance to apoptosis seen in these Bcr-Abl-expressing cells.

Fusion tyrosine kinase mediated signalling pathways in the transformation of haematopoietic cells.

The fusion tyrosine kinases (FTKs) are generated by chromosomal translocations creating bipartite proteins in which the kinase is hyperactivated by an adjoining oligomerization domain. Autophosphorylation of the FTK generates a 'signalosome', an ensemble of signalling proteins that transduce signals to downstream pathways. At the earliest stages of oncogenesis, FTKs can mimic mitogenic cytokine signalling pathways involving the GAB-2 adaptor protein and signal transducers and activators of transcription (STAT) factors, generating replicative stress and thereby promoting a mutator phenotype. In parallel, FTKs couple to survival pathways that upregulate prosurvival proteins such as Bcl-xL, so preventing DNA-damage-induced apoptosis. Following transformation, FTKs induce resistance to genotoxic attack by upregulating DNA repair mechanisms such as STAT5-dependent RAD51 transcription. The phenomenon of 'oncogene addiction' reflects the continued requirement of an active FTK 'signalosome' to mediate survival and mitogenic signals involving the PI 3-kinase and mitogen-activated protein stress-activated protein kinase pathways, and the nuclear factor-kappa B, activator protein 1 and STAT transcription factors. The available data so far suggest that FTKs, with some possible exceptions, induce and maintain the transformed state using similar panoplies of signals, a finding with important therapeutic implications. The FTK signalling field has matured to an exciting phase in which rapid advances are facilitating rational drug design.

What have we learnt from mouse models of NPM-ALK-induced lymphomagenesis?

The nucleophosmin-anaplastic lymphoma kinase (NPM-ALK) is generated as a t(2;5) chromosomal breakpoint product, typically in CD30(+) anaplastic large cell lymphomas. Activation of the NPM-ALK tyrosine kinase by NPM dimerisation causes autophosphorylation at multiple tyrosine residues and the consequent recruitment of a 'signalosome' that couples the fusion protein to pathways regulating mitogenesis and apoptosis. This review focuses on recent advances in our understanding of the transforming signals induced by this fusion protein in mouse models.

The use of 3-[2-(N,N-diethyl-N-methylammonium)ethyl]-7-methoxy-4-methylcoumarin (AMMC) as a specific CYP2D6 probe in human liver microsomes.

Recently, a novel nonfluorescent probe 3-[2-(N,N-diethyl-N-methylammonium)-ethyl]-7-methoxy-4-methylcoumarin (AMMC), which produces a fluorescent metabolite AMHC (3-[2-(N,N-diethyl-N-methylammonium)ethyl]-7-hydroxy-4-methylcoumarin) was used with microsomes containing recombinant enzymes (rCYP) to monitor CYP2D6 inhibition in a microtiter plate assay. This article describes the studies that were performed in human liver microsomes (HLM) to establish the selectivity of AMMC toward CYP2D6. Metabolism studies in HLM showed that AMMC was converted to one metabolite identified by mass spectrometry as AMHC. Kinetic studies indicated an apparent K(m) of 3 microM with a V(max) of 20 pmol/min. mg of protein for the O-demethylation reaction. The O-demethylation of AMMC in HLM was inhibited significantly in the presence of a CYP2D6 inhibitory antibody. Using a panel of various HLM preparations (n = 12), a good correlation (r(2) = 0.95) was obtained between AMMC O-demethylation and bufuralol metabolism, a known CYP2D6 substrate, but not with probes for the other major xenobiotic metabolizing CYPs. Finally, only rCYP2D6 showed detectable metabolism in experiments conducted with rCYPs using AMMC at a concentration of 1.5 microM (near K(m)). However, at a concentration of 25 microM AMMC, rCYP1A also contributed significantly to the formation of AMHC. Knowing the experimental conditions under which AMMC was selective for CYP2D6, a microtiter assay was developed to study the inhibition of various compounds in HLM using the fluorescence of AMHC as an indication of CYP2D6 activity. The inhibition potential of various chemicals was found to be comparable to those determined using the standard CYP2D6 probe, bufuralol, which requires high-performance liquid chromatography separation for the analysis of its CYP2D6-mediated 1'-hydoxylated metabolite.

Essential role of neutrophils in germ cell-specific apoptosis following ischemia/reperfusion injury of the mouse testis.

This study investigates the role of neutrophils in ischemia-induced aspermatogenesis in the mouse. Previous studies in the rat have demonstrated that ischemia-inducing testicular torsion followed by torsion repair and reperfusion resulted in germ cell-specific apoptosis. This was correlated with an increase in neutrophil adhesion to subtunical venules, an increase in reactive oxygen species, and increased expression of several apoptosis-associated molecules. In the present investigation, wild-type C57BL/6 mice were subjected to various degrees and duration of testicular torsion. A torsion of 720 degrees for 2 h caused disruption of the seminiferous epithelium and significantly reduced testis weight and daily sperm production. An immunohistochemical method specific for apoptotic nuclei indicated that these effects were due to germ cell-specific apoptosis. An increase in myeloperoxidase (MPO) activity and an increase in the number of neutrophils adhering to testicular subtunical venules after torsion repair/reperfusion demonstrated an increase in neutrophil recruitment to the testis. In contrast, E-selectin knockout mice and wild-type mice rendered neutropenic showed a significant decrease in neutrophil recruitment as evidenced by MPO activity and microscopic examination of subtunical venules. Importantly, germ cell-specific apoptosis was also reduced. Thus, germ cell-specific apoptosis is observed after ischemia/reperfusion of the murine testis, and this apoptosis is directly linked to the recruitment of neutrophils to subtunical venules. Endothelial cell adhesion molecules, particularly E-selectin, play an important role in mediating this pathology.

Assays to predict the genotoxicity of the chromosomal mutagen etoposide -- focussing on the best assay.

The topoisomerase II inhibitor etoposide is used routinely to treat a variety of cancers in patients of all ages. As a result of its extensive use in the clinic and its association with secondary malignancies it has become a compound of great interest with regard to its genotoxic activity in vivo. This paper describes a series of assays that were employed to determine the in vivo genotoxicity of etoposide in a murine model system. The alkaline comet assay detected DNA damage in the bone marrow mononuclear compartment over the dose range of 10--100mg/kg and was associated with a large and dose dependent rise in the proportion of cells with severely damaged DNA. In contrast, the bone marrow micronucleus assay was found to be sensitive to genotoxic damage between the doses of 0.1--1mg/kg without any corresponding increases in cytotoxicity. An increase in the mutant frequency was undetectable at the Hprt locus at administered doses of 1 and 10mg/kg of etoposide, however, an increase in the mutant frequency was seen at the Aprt locus at these doses. We conclude that the BMMN assay is a good short-term predictor of the clastogenicity of etoposide at doses that do not result in cytotoxic activity, giving an indication of potential mutagenic effects. Moreover, the detection of mutants at the Aprt locus gives an indication of the potential of etoposide to cause chromosomal mutations that may lead to secondary malignancy.

The male rat carcinogens limonene and sodium saccharin are not mutagenic to male Big Blue rats.

Limonene and sodium saccharin are male rat specific carcinogens giving rise to renal and bladder tumours, respectively. Both compounds give negative results in genetic toxicity assays suggesting a non-genotoxic mode of action for their carcinogenicity. The alpha 2U-globulin accumulation theory has been invoked to explain the renal carcinogenicity of limonene: the accumulation of micro masses of calcium phosphate in the bladder, coupled with a high pH environment in the male rat bladder, has been suggested to be responsible for the bladder carcinogenicity of sodium saccharin. The implication of these proposed mechanisms is that limonene and sodium saccharin will not be mutagenic to the rat kidney and bladder, respectively. This proposal has been evaluated by assessing the mutagenic potential of the two chemicals to male lacI transgenic (Big Blue) rats. Male Big Blue rats were exposed for 10 consecutive days to either limonene in diet, at a dose level in excess of that used in the original National Toxicology Program gavage carcinogenicity bioassay, or to sodium saccharin in diet at the dose known to induce bladder tumours. The multi-site rat carcinogen 4-aminobiphenyl was used as a positive control for the experiment. Limonene failed to increase the mutant frequency in the liver or kidney of the rats, and sodium saccharin failed to increase the mutant frequency in the liver or bladder of the rats. 4-Aminobiphenyl was mutagenic to all three of these tissues. These results add further support to a non-genotoxic mechanism of carcinogenic action for both limonene and sodium saccharin.

Effect of topical fluoride on retention of pit and fissure sealants.

PURPOSE: The purpose of this study was to evaluate the clinical effect of topical fluoride on retention of light-cured (CLC) and self-cured (CSC) pit and fissure sealants. METHODS: CLC and CSC sealants were placed in vivo on opposite sides of the arch before and after fluoride treatment. A total of 122 sealants were placed on virgin permanent molars and premolars of 16 dental hygiene students enrolled in a two-year program. Sealant retention in both fluoridated and non-fluoridated teeth was evaluated at 6, 12, and 18 month intervals. RESULTS: Overall sealant retention for both fluoridated and non-fluoridated teeth at 6, 12 and 18 months was 68%, 48%, and 49%, respectively. There was a significant difference (p < 0.001) when fluoridated vs. non-fluoridated teeth were compared. Retention was greater on the fluoridated teeth, with respect to the sealant material (CLC-fluoride). Significant differences (p < 0.001) were found when CLC-fluoride and CLC-no fluoride treatment groups were compared. However, no significant differences were found in retention when CSC-fluoride and CSC-no fluoride groups were compared, or when CLC was compared to CSC irrespective of fluoridation. Significant differences (p < 0.0001) were found when sealant retention on molars was compared to premolars--retention of sealants was greater on premolars. CONCLUSION: This study suggests that sealant retention may not be adversely affected by a topical fluoride treatment applied immediately prior to placement.

Substrate-dependent modulation of CYP3A4 catalytic activity: analysis of 27 test compounds with four fluorometric substrates.

Inhibition of cytochrome P450 catalytic activity is a principal mechanism for pharmacokinetic drug-drug interactions. Rapid, in vitro testing for cytochrome P450 inhibition potential is part of the current paradigm for identifying drug candidates likely to give such interactions. We have explored the extent that qualitative and quantitative inhibition parameters are dependent on the cytochrome P450 (CYP) 3A4 probe substrate. Inhibition potential (e.g., IC(50) values from 8-point inhibition curves) or activation potential for most compounds varied dramatically depending on the fluorometric probe substrates for CYP3A4 [benzyloxyresorufin (BzRes), 7-benzyloxy-4-trifluoromethylcoumarin (BFC), 7-benzyloxyquinoline (BQ), and dibenzylfluorescein (DBF)]. For 21 compounds that were primarily inhibitors, the range of IC(50) values for the four substrates varied from 2.1- to 195-fold with an average of 29-fold. While the rank order of sensitivity among the fluorometric substrates varied among the individual inhibitors, on average, BFC dealkylation was the most sensitive to inhibition, while BQ dealkylation was least sensitive. Partial inhibition was observed with BzRes and BQ but not for BFC and DBF. BzRes was more prone to activation, whereas dramatic changes in IC(50) values were observed when the BQ concentration was below the S(50). Three different correlation analyses indicated that IC(50) values with BFC, BQ, and DBF correlated well with each other, whereas the response with BzRes correlated more weakly with the other substrates. One of these correlation analyses was extended to the percent inhibition of 10 microM inhibitor with the standard CYP3A4 probe substrates testosterone, midazolam, and nifedipine. In this analysis the responses with BQ, BFC and DBF correlated well with testosterone and midazolam but more poorly with nifedipine. In the aggregate, BFC and DBF appear more suitable as an initial screen for CYP3A4 inhibition. However, the substrate-dependent effects reported here and by others indicate that all CYP3A4 inhibition data should be interpreted with caution.

Molecular pathway of germ cell apoptosis following ischemia/reperfusion of the rat testis.

The present study investigates the molecular apoptotic pathway in germ cells following acute ischemia of the rat testis. Rats were subjected to ischemia-inducing torsion and testes were harvested after reperfusion. Apoptotic cells were identified with an antibody to single-stranded DNA. Seminiferous tubule RNA was examined by RNase protection assay or by reverse transcriptase-polymerase chain reaction (RT-PCR) for the presence and regulation of apoptotic molecules. Proteins from seminiferous tubules were used for Western blot analysis of cytochrome c. Germ cell apoptosis was maximal at 24 h after repair of torsion. Germ cells in stages II-III of the seminiferous epithelium cycle were the predominant early responders. The RNase protection assays revealed that Bcl-X(L) was the prominent mRNA species. Caspases 1, 2, 3, and Bax mRNA were consistently upregulated; however, the time of upregulation after torsion was variable. The Bcl-X(L) and Bcl-X(S) mRNAs were less consistently upregulated and there was no evidence for upregulation of Fas or Bcl-2. Fas ligand (FasL) was not detected by RNase protection assay, but RT-PCR revealed a significant increase in FasL expression 4 h after the repair of torsion. Western blot analysis for cytochrome c release demonstrated a significant increase 4 h after the repair of torsion. Results suggest that germ cell apoptosis following ischemia/reperfusion of the rat testis is initiated through the mitochondria-associated molecule Bax as well as Fas-FasL interactions.