Influence of key factors on ammonia and nitrous oxide emission factors for excreta deposited by livestock and land-applied manure.

Ammonia (NH3) and nitrous oxide (N2O) emissions from livestock manure management have a significant impact on air quality and climate change. There is an increasing urgency to improve our understanding of drivers influencing these emissions. We analysed the DATAMAN ("DATAbase for MANaging greenhouse gas and ammonia emissions factors") database to identify key factors influencing (i) NH3 emission factors (EFs) for cattle and swine manure applied to land and (ii) N2O EFs for cattle and swine manure applied to land, and (iii) cattle urine, dung and sheep urine deposited during grazing. Slurry dry matter (DM) content, total ammoniacal nitrogen (TAN) concentration and method of application were significant drivers of NH3 EFs from cattle and swine slurry. Mixed effect models explained 14-59 % of the variance in NH3 EFs. Apart from the method of application, the significant influence of manure DM, manure TAN concentration or pH on NH3 EFs suggests mitigation strategies should focus on these. Identifying key factors influencing N2O EFs from manures and livestock grazing was more challenging, likely because of the complexities associated with microbial processes and soil physical properties impacting N2O production and emissions. Generally, significant factors were soil-related e.g. soil water content, pH, clay content, suggesting mitigations may need to consider the conditions of the receiving environment for manure spreading and grazing deposition. Total variability explained by terms in mixed effect model was on average 66 %, with the random effect 'experiment identification number' explaining, on average, 41 % of the total variability in the models. We suspect this term captured the effect of non-measured manure, soil and climate factors and any biases in application and measurement technique effects associated with individual experiments. This analysis has helped to improve our understanding of key factors of NH3 and N2O EFs for inclusion within models. With more studies over time, insights into the underlying processes influencing emissions will be further improved.

Effect of Time and Temperature of Air Jet on the Mechanical and Biological Behavior of a Universal Adhesive System.

OBJECTIVES: To evaluate the influence of heat application on the degree of conversion (DC) of the 3M Single Bond Universal Adhesive System, as well as its transdentinal cytotoxicity and microtensile bond strength to dentin. METHODS: Experimental groups were established according to the time and temperature of the air jet: G1: 5 seconds-25°C; G2: 10 seconds-25°C; G3: 20 seconds-25°C; G4: 5 seconds-50°C; G5: 10 seconds-50°C; G6: 20 seconds-50°C. In control group (G7), no treatment was performed. The DC was assessed using the Fourier transform infrared spectroscopy-attenuated total reflectance (FTIR-ATR) technique. For the transdentinal cytotoxicity test, dentin discs fitted in artificial pulp chambers (APC) received the application of the adhesive system and the air jets. For the microtensile bond strength, healthy molars were restored and submitted to the microtensile test after 24 hours and 6 months, respectively. RESULTS: Significant reduction in viability of Mouse Dental Papilla Cell-23 (MDPC-23), which exhibited morphological changes, was observed in all experimental groups compared to control (p<0.05). Although all tested protocols resulted in transdentinal diffusion of 2-hydroxyethyl methacrylate (HEMA), the group G6 presented the highest degree of monomeric conversion and the lowest cytotoxic effect, with higher dentin bond strength values in comparison to group G1 (p<0.05). CONCLUSIONS: Applying an air blast at 50°C for 20 seconds increases the DC and microtensile bond strength of the 3M Single Bond Universal Adhesive System to dentin, as well as reduces the transdentinal cytotoxicity of the material to pulp cells.

Meta-analysis of New Zealand's nitrous oxide emission factors for ruminant excreta supports disaggregation based on excreta form, livestock type and slope class.

Globally, animal excreta (dung and urine) deposition onto grazed pastures represents more than half of anthropogenic nitrous oxide (N2O) emissions. To account for these emissions, New Zealand currently employs urine and dung emission factor (EF3) values of 1.0% and 0.25%, respectively, for all livestock. These values are primarily based on field studies conducted on fertile, flatland pastures predominantly used for dairy cattle production but do not consider emissions from hill land pastures primarily used for sheep, deer and non-dairy cattle. The objective of this study was to determine the most suitable urine and dung EF3 values for dairy cattle, non-dairy cattle, and sheep grazing pastures on different slopes based on a meta-analysis of New Zealand EF3 studies. As none of the studies included deer excreta, deer EF3 values were estimated from cattle and sheep values. The analysis revealed that a single dung EF3 value should be maintained, although the value should be reduced from 0.25% to 0.12%. Furthermore, urine EF3 should be disaggregated by livestock type (cattle > sheep) and topography (flatland and low sloping hill country > medium and steep sloping hill country), with EF3 values ranging from 0.08% (sheep urine on medium and steep slopes) to 0.98% (dairy cattle on flatland and low slopes). While the mechanism(s) causing differences in urine EF3 values for sheep and cattle are unknown, the 'slope effect' on urine EF3 is partly due to differences in soil chemical and physical characteristics, which influence soil microbial processes on the different slope classes. The revised EF3 values were used in an updated New Zealand inventory approach, resulting in 30% lower national N2O emissions for 2017 compared to using the current EF3 values. We recommend using the revised EF3 values in New Zealand's national greenhouse gas inventory to more accurately capture N2O emissions from livestock grazing.

Using alternative forage species to reduce emissions of the greenhouse gas nitrous oxide from cattle urine deposited onto soil.

Grazed pastures are a major contributor to emissions of the greenhouse gas nitrous oxide (N2O), and urine deposition from grazing animals is the main source of the emissions. Incorporating alternative forages into grazing systems could be an approach for reducing N2O emissions through mechanisms such as release of biological nitrification inhibitors from roots and increased root depth. Field plot and lysimeter (intact soil column) trials were conducted in a free draining Horotiu silt loam soil to test whether two alternative forage species, plantain (Plantago lanceolate L.) and lucerne (Medicago sativa L.), could reduce N2O emissions relative to traditional pasture species, white clover (Trifolium repens L.) and perennial ryegrass (Lolium perenne L.). The amounts of N2O emitted from the soil below each forage species, which all received the same cow urine at the same rates, was measured using an established static chamber method. Total N2O emissions from the plantain, lucerne and perennial ryegrass controls (without urine application) were generally very low, but emissions from the white clover control were significantly higher. When urine was applied in autumn or winter N2O emissions from plantain were lower compared with those from perennial ryegrass or white clover, but this difference was not found when urine was applied in summer. Lucerne had lower emissions in winter but not in other seasons. Incorporation of plantain into grazed pasture could be an approach to reduce N2O emissions. However, further work is required to understand the mechanisms for the reduced emissions and the effects of environmental conditions in different seasons.

A role for Rac1 activity in malignant progression of sebaceous skin tumors.

The small GTPase Rac1 is crucial for maintaining stem cells (SCs) in mammalian epidermis, and Rac1 activation leads to SC expansion. Loss or inhibition of Rac1 correlates with decreased frequency of skin cancer formation in a chemical carcinogenesis model. Here, we have addressed whether Rac1 activation would enhance carcinogenesis and result in tumor progression. We used K14ΔNLef1 mice, a model for differentiated sebaceous adenomas (SAs), and activated Rac1 in an epidermis-specific manner (K14L61Rac1). Surprisingly, Rac1 activation did not change the incidence and frequency of sebaceous tumors. However, tumors, which occurred exclusively in K14ΔNLef1/K14L61Rac1 double-transgenic mice, were poorly differentiated resembling malignant sebaceous tumors and were termed sebaceous carcinoma-like tumors (SCLTs). Compared with SAs, SCLTs showed an aberrant pattern of cell proliferation, invasive growth and less abundant expression of sebocyte differentiation markers, including stearoyl-CoA desaturase-1 and adipophilin. Interestingly, the adnexal SC marker Lrig1 was upregulated in SCLTs, showing that active Rac1 leads to the accumulation of sebocyte precursors in the context of K14ΔNLef1-induced skin tumors. In a search for targets of Rac1, we found cancer progression-related proteins, Dhcr24/Seladin1 and Nuclear protein 1/P8, to be strongly regulated in SCLTs. At last, Rac1 and Dhcr24/Seladin1 were detected in human sebaceous tumors demonstrating a potential high impact of our findings for human skin disease. This is the first study showing that Rac1 activity can lead to malignant progression of skin tumors.

Statistical analysis of nitrous oxide emission factors from pastoral agriculture field trials conducted in New Zealand.

Between 11 May 2000 and 31 January 2013, 185 field trials were conducted across New Zealand to measure the direct nitrous oxide (N2O) emission factors (EF) from nitrogen (N) sources applied to pastoral soils. The log(EF) data were analysed statistically using a restricted maximum likelihood (REML) method. To estimate mean EF values for each N source, best linear unbiased predictors (BLUPs) were calculated. For lowland soils, mean EFs for dairy cattle urine and dung, sheep urine and dung and urea fertiliser were 1.16 ± 0.19% and 0.23 ± 0.05%, 0.55 ± 0.19% and 0.08 ± 0.02% and 0.48 ± 0.13%, respectively, each significantly different from one another (p < 0.05), except for sheep urine and urea fertiliser. For soils in terrain with slopes >12°, mean EFs were significantly lower. Thus, urine and dung EFs should be disaggregated for sheep and cattle as well as accounting for terrain.

Losses at chromosome 4q are associated with poor survival in operable ductal pancreatic adenocarcinoma.

Here we tested the prognostic impact of genomic alterations in operable localized pancreatic ductal adenocarcinoma (PDAC). Fifty-two formalin-fixed and paraffin-embedded primary PDAC were laser micro-dissected and were investigated by comparative genomic hybridization after whole genome amplification using an adapter-linker PCR. Chromosomal gains and losses were correlated to clinico-pathological parameters and clinical follow-up data. The most frequent aberration was loss on chromosome 17p (65%) while the most frequent gains were detected at 2q (41%) and 8q (41%), respectively. The concomitant occurrence of losses at 9p and 17p was found to be statistically significant. Higher rates of chromosomal losses were associated with a more advanced primary tumor stage and losses at 9p and 18q were significantly associated with presence of lymphatic metastasis (chi-square: p = 0.03, p = 0.05, respectively). Deletions on chromosome 4 were of prognostic significance for overall survival and tumor recurrence (Cox-multivariate analysis: p = 0.026 and p = 0.021, respectively). In conclusion our data suggest the common alterations at chromosome 8q, 9p, 17p and 18q as well as the prognostic relevant deletions on chromosome 4q as relevant for PDAC progression. Our comprehensive data from 52 PDAC should provide a basis for future studies with a higher resolution to discover the relevant genes located within the chromosomal aberrations identified.

Prioritisation of farm scale remediation efforts for reducing losses of nutrients and faecal indicator organisms to waterways: a case study of New Zealand dairy farming.

The international competitiveness of the New Zealand (NZ) dairy industry is built on low cost clover-based systems and a favourable temperate climate that enables cows to graze pastures mostly all year round. Whilst this grazed pasture farming system is very efficient at producing milk, it has also been identified as a significant source of nutrients (N and P) and faecal bacteria which have contributed to water quality degradation in some rivers and lakes. In response to these concerns, a tool-box of mitigation measures that farmers can apply on farm to reduce environmental emissions has been developed. Here we report the potential reduction in nutrient losses and costs to farm businesses arising from the implementation of individual best management practices (BMPs) within this tool-box. Modelling analysis was carried out for a range of BMPs targeting pollutant source reduction on case-study dairy farms, located in four contrasting catchments. Due to the contrasting physical resources and management systems present in the four dairy catchments evaluated, the effectiveness and costs of BMPs varied. Farm managements that optimised soil Olsen P levels or used nitrification inhibitors were observed to result in win-win outcomes whereby nutrient losses were consistently reduced and farm profitability was increased in three of the four case study farming systems. Other BMPs generally reduced nutrient and faecal bacteria losses but at a small cost to the farm business. Our analysis indicates that there are a range of technological measures that can deliver substantial reductions in nutrient losses to waterways from dairy farms, whilst not increasing or even reducing other environmental impacts (e.g. greenhouse gas emissions and energy use). Their implementation will first require clearly defined environmental goals for the catchment/water body that is to be protected. Secondly, given that the major sources of water pollutants often differed between catchments, it is important that BMPs are matched to the physical resources and management systems of the existing farm businesses.

[From single disseminated tumor cells to metastasis insights from molecular genetic analyses of single cells]. / Von der einzelnen disseminierten Tumorzelle zur Metastase: Erkenntnisse aus der molekulargenetischen Einzelzellanalyse.

The prevailing models of cancer metastasis postulate that, after a series of accumulating genetic and epigenetic changes during transformation and invasive growth, the most advanced clone within a primary tumour acquires the critical cellular phenotype enabling dissemination and metastasis. This postulate is particularly based on observations that metastases usually display more genetic changes than the primary tumour. The development of several novel techniques is now enabling experimental testing of the model: The detection of single disseminated cancer cells by epithelial specific antibodies directed against cytokeratin in mesenchymal tissues has become possible even before manifestation of metastasis. After their isolation by micromanipulation comprehensive amplification of the single cell genomes allows the application of several molecular genetic methods for further characterisation. Our first data from single cytokeratin-positive cells that were isolated from the bone marrow of breast cancer patients indicate that the model of cancer progression should be revised. Cancer cells already disseminate in an early stage of genomic development and still have to acquire the critical chromosomal aberrations needed for metastatic outgrowth and full malignant potential. These observations apparently imply consequences for the development of novel adjuvant therapies. The early diversification of primary tumours and disseminated cancer cells precludes a simple extrapolation from local to systemic disease eventually necessitating increasing efforts for the direct analysis of single disseminated cancer cells as the cellular targets of adjuvant therapies.

Seven-fluorochrome mouse M-FISH for high-resolution analysis of interchromosomal rearrangements.

The mouse has evolved to be the primary mammalian genetic model organism. Important applications include the modeling of human cancer and cloning experiments. In both settings, a detailed analysis of the mouse genome is essential. Multicolor karyotyping technologies have emerged to be invaluable tools for the identification of mouse chromosomes and for the deciphering of complex rearrangements. With the increasing use of these multicolor technologies resolution limits are critical. However, the traditionally used probe sets, which employ 5 different fluorochromes, have significant limitations. Here, we introduce an improved labeling strategy. Using 7 fluorochromes we increased the sensitivity for the detection of small interchromosomal rearrangements (700 kb or less) to virtually 100%. Our approach should be important to unravel small interchromosomal rearrangements in mouse models for DNA repair defects and chromosomal instability.

[Direct molecular analysis of single disseminated cancer cells: a prerequisite for the development of adjuvant therapies?]. / Die direkte molekulare Analyse einzelner disseminierter Tumorzellen: Eine Voraussetzung für die Entwicklung adjuvanter Therapien?

Cancer mortality has only marginally decreased in the last decades despite huge diagnostic and therapeutic efforts. Early dissemination of cancer cells has to be blamed for this finding, because ectopically residing cells are necessarily left behind by the surgeon. Consequently, this cell population has been designated as minimal residual disease that may eventually lead to systemic relapse months or years after presumed curative surgery. Paradoxically, systemic adjuvant treatments are currently administered without any precise knowledge about the target population of such drugs. Here it is argued that the direct analysis of disseminated cells may be a prerequiste for the development of future therapies since first molecular genetic data of single disseminated cancer cells suggest an excessive intercellular heterogeneity and distant relationship to the primary tumor.

Early cancer cell dissemination and late metastatic relapse: clinical reflections and biological approaches to the dormancy problem in patients.

Two clinical entities, unknown-primary cancer and inadvertent transmission of cancer with organ transplants are reviewed and discussed in the context of early and occult tumor cell dissemination. Both entities are taken as chief witnesses for cell dissemination being an early event in tumor progression. The involuntary transmission of tumor by organ grafts directly supports the notion that very few quiescent cells lodging at improbable sites such as kidney or heart suffice to generate de novo metastatic disease in the organ recipient. As to the nature of the cells and their biological and clinical significance a short review is given on the detection of disseminated cells in bone marrow and their prognostic significance for a metastatic relapse in patients with resected primary tumors. A novel single-cell genomic analysis is described, that allows the detection of multiple chromosomal aberration in single tumor cells.

Transcriptome analysis reveals a role of interferon-gamma in human neointima formation.

The most effective immediate cure for coronary stenosis is stent-supported angioplasty. Restenosis due to neointima proliferation represents a major limitation. We investigated the expression of 2435 genes in atherectomy specimens and blood cells of patients with restenosis, normal coronary artery specimens, and cultured human smooth muscle cells (SMCs). Of the 223 differentially expressed genes, 37 genes indicated activation of interferon-gamma (IFN-gamma) signaling in neointimal SMCs. In cultured SMCs, IFN-gamma inhibited apoptosis. Genetic disruption of IFN-gamma signaling in a mouse model of restenosis significantly reduced the vascular proliferative response. Our data suggest an important role of IFN-gamma in the control of neointima proliferation.

Gene expression profiling of human stent-induced neointima by cDNA array analysis of microscopic specimens retrieved by helix cutter atherectomy: Detection of FK506-binding protein 12 upregulation.

BACKGROUND: Restenosis due to neointima formation is the major limitation of stent-supported balloon angioplasty. Despite abundant animal data, molecular mechanisms of neointima formation have been investigated on only a limited basis in patients. This study sought to establish a method for profiling gene expression in human in-stent neointima and to identify differentially expressed genes that may serve as novel therapeutic targets. METHODS AND RESULTS: We retrieved tissue specimens from patients with symptomatic in-stent restenosis using a novel helix cutter atherectomy device. cDNA samples prepared from neointima (n=10) and, as a control, from the media of normal arteries (n=14) were amplified using a novel polymerase chain reaction protocol and hybridized to cDNA arrays. Immunohistochemistry characterized the atherectomy material as neointima. cDNA arrays readily identified differentially expressed genes. Some of the differentially expressed genes complied with expected gene expression patterns of neointima, including downregulation of desmin and upregulation of thrombospondin-1, cyclooxygenase-1, and the 70-kDa heat shock protein B. Additionally, we discovered previously unknown gene expression patterns, such as downregulation of mammary-derived growth inhibitor and upregulation of FK506-binding protein 12 (FKBP12). Upregulation of FKBP12 was confirmed at the protein level in neointimal smooth muscle cells. CONCLUSIONS: Gene expression patterns of human neointima retrieved by helix-cutter atherectomy can be reliably analyzed by cDNA array technology. This technique can identify therapeutic targets in patients, as exemplified by the findings regarding FKBP12. FKBP12 is the receptor for Rapamycin (sirolimus), which in animal models reduced neointima formation. Our study thus yields a rationale for the use of Rapamycin to prevent restenosis in patients.