Estimating product and energy substitution benefits in national‐scale mitigation analyses for Canada

The potential of forests and the forest sector to mitigate greenhouse gas (GHG) emissions is widely recognized, but challenging to quantify at a national scale. Mitigation benefits through the use of forest products are affected by product life cycles, which determine the duration of carbon storage in wood products and substitution benefits where emissions are avoided using wood products instead of other emissions‐intensive building products and energy fuels. Here we determined displacement factors for wood substitution in the built environment and bioenergy at the national level in Canada. For solid wood products, we compiled a basket of end‐use products and determined the reduction in emissions for two functionally equivalent products: a more wood‐intensive product vs. a less wood‐intensive one. Avoided emissions for end‐use products basket were weighted by Canadian consumption statistics to reflect national wood uses, and avoided emissions were further partitioned into displacement factors for sawnwood and panels. We also examined two bioenergy feedstock scenarios (constant supply and constrained supply) to estimate displacement factors for bioenergy using an optimized selection of bioenergy facilities which maximized avoided emissions from fossil fuels. Results demonstrated that the average displacement factors were found to be similar: product displacement factors were 0.54 tC displaced per tC of used for sawnwood and 0.45 tC tC^?1 for panels; energy displacement factors for the two feedstock scenarios were 0.47 tC tC^?1 for the constant supply and 0.89 tC tC^?1 for the constrained supply. However, there was a wide range of substitution impacts. The greatest avoided emissions occurred when wood was substituted for steel and concrete in buildings, and when bioenergy from heat facilities and/or combined heat and power facilities was substituted for energy from high‐emissions fossil fuels. We conclude that (1) national‐level substitution benefits need to be considered within a systems perspective on climate change mitigation to avoid the development of policies that deliver no net benefits to the atmosphere, (2) the use of long‐lived wood products in buildings to displace steel and concrete reduces GHG emissions, (3) the greatest bioenergy substitution benefits are achieved using a mix of facility types and capacities to displace emissions‐intensive fossil fuels.     

Macrophage-specific responses to human- and animal-adapted tubercle bacilli reveal pathogen and host factors driving multinucleated cell formation

The Mycobacterium tuberculosis complex (MTBC) is a group of related pathogens that cause tuberculosis (TB) in mammals. MTBC species are distinguished by their ability to sustain in distinct host populations. While Mycobacterium bovis (Mbv) sustains transmission cycles in cattle and wild animals and causes zoonotic TB, M. tuberculosis (Mtb) affects human populations and seldom causes disease in cattle. The host and pathogen determinants underlying host tropism between MTBC species are still unknown. Macrophages are the main host cell that encounters mycobacteria upon initial infection, and we hypothesised that early interactions between the macrophage and mycobacteria influence species-specific disease outcome. To identify factors that contribute to host tropism, we analysed blood-derived primary human and bovine macrophages (hMϕ or bMϕ, respectively) infected with Mbv and Mtb. We show that Mbv and Mtb reside in different cellular compartments and differentially replicate in hMϕ whereas both Mbv and Mtb efficiently replicate in bMϕ. Specifically, we show that out of the four infection combinations, only the infection of bMϕ with Mbv promoted the formation of multinucleated giant cells (MNGCs), a hallmark of tuberculous granulomas. Mechanistically, we demonstrate that both MPB70 from Mbv and extracellular vesicles released by Mbv-infected bMϕ promote macrophage multinucleation. Importantly, we extended our in vitro studies to show that granulomas from Mbv-infected but not Mtb-infected cattle contained higher numbers of MNGCs. Our findings implicate MNGC formation in the contrasting pathology between Mtb and Mbv for the bovine host and identify MPB70 from Mbv and extracellular vesicles from bMϕ as mediators of this process.     

Use of within-array replicate spots for assessing differential expression in microarray experiments

MOTIVATION: Spotted arrays are often printed with probes in duplicate or triplicate, but current methods for assessing differential expression are not able to make full use of the resulting information. The usual practice is to average the duplicate or triplicate results for each probe before assessing differential expression. This results in the loss of valuable information about genewise variability. RESULTS: A method is proposed for extracting more information from within-array replicate spots in microarray experiments by estimating the strength of the correlation between them. The method involves fitting separate linear models to the expression data for each gene but with a common value for the between-replicate correlation. The method greatly improves the precision with which the genewise variances are estimated and thereby improves inference methods designed to identify differentially expressed genes. The method may be combined with empirical Bayes methods for moderating the genewise variances between genes. The method is validated using data from a microarray experiment involving calibration and ratio control spots in conjunction with spiked-in RNA. Comparing results for calibration and ratio control spots shows that the common correlation method results in substantially better discrimination of differentially expressed genes from those which are not. The spike-in experiment also confirms that the results may be further improved by empirical Bayes smoothing of the variances when the sample size is small. AVAILABILITY: The methodology is implemented in the limma software package for R, available from the CRAN repository http://www.r-project.org     

Effects of Boron Deficiency on Rubidium Uptake and Photosynthesis in the Diatom Cylindrotheca fusiformis

Culturing the diatom Cylindrotheca fusiformis under boron-deficient conditions leads to changes in ⁸⁶Rb uptake and photosynthesis prior to any effect on the rate of cell division. The influx rate of ⁸⁶Rb into boron-deficient cells was 79% of the control rate after 5 to 5.5 hours culture. Despite lowered ⁸⁶Rb influx, however, boron-deficient diatoms accumulated more ⁸⁶Rb than did control cells; this was due to the deficient cells' lower efflux rate. After 24 hours culture, boron-deficient cells had accumulated 30% more ⁸⁶Rb than had control cells, while releasing ⁸⁶Rb at only one-half the control rate. Increased photosynthetic rates were another effect of boron deficiency during this early stage of culture. Prior to 20 hours boron-deficient culture, diatoms had photosynthetic rates 37% greater than those of control cells. Corresponding to the increase in photosynthesis, boron-deficient diatoms had 12% more carbohydrate than control cells after 16 hours culture.     

A cytochrome b561 with ferric reductase activity from the parasitic blood fluke, Schistosoma japonicum

Background

Iron has an integral role in numerous cellular reactions and is required by virtually all organisms. In physiological conditions, iron is abundant in a largely insoluble ferric state. Ferric reductases are an essential component of iron uptake by cells, reducing iron to the soluble ferrous form. Cytochromes b561 (cyts-b561) are a family of ascorbate reducing transmembrane proteins found in most eukaryotic cells. The identification of the ferric reductase duodenal cytochrome b (dcytb) and recent observations that other cyts-b561 may be involved in iron metabolism have opened novel perspectives for elucidating their physiological function.

Methodology/Principal Findings

Here we have identified a new member of the cytochrome b561 (Sjcytb561) family in the pathogenic blood fluke Schistosoma japonicum that localises to the outer surface of this parasitic trematode. Heterologous expression of recombinant Sjcyt-b561 in a Saccharomyces cerevisiae mutant strain that lacks plasma membrane ferrireductase activity demonstrated that the molecule could rescue ferric reductase activity in the yeast.

Significance/Conclusions

This finding of a new member of the cytochrome b561 family further supports the notion that a ferric reductase function is likely for other members of this protein family. Additionally, the localisation of Sjcytb561 in the surface epithelium of these blood-dwelling schistosomes contributes further to our knowledge concerning nutrient acquisition in these parasites and may provide novel targets for therapeutic intervention.     

Gene-engineered T cells as a superior adjuvant therapy for metastatic cancer

The major limiting factor in the successful application of adjuvant therapy for metastatic disease is the lack of adjuvant specificity that leads to severe side effects. Reasoning that T cells of the immune system are highly specific, we generated tumor-specific T cells by genetic modification of mouse primary T cells with a chimeric receptor reactive with the human breast cancer-associated Ag erbB-2. These T cells killed breast cancer cells and secreted IFN-gamma in an Ag-specific manner in vitro. We investigated their use against metastatic breast cancer in mice in an adjuvant setting, and compared their effectiveness with the commonly applied adjuvants doxorubicin, 5-fluorouracil, and herceptin. Mice were inoculated orthotopically with the human erbB-2-expressing spontaneously metastatic mouse breast cancer 4T1.2 in mammary tissue, and the primary tumor was surgically removed 8 days later. Significant metastatic disease was demonstrated in lung and liver at the time of surgery on day 8 with increased tumor burden at later time points. T cell adjuvant treatment of day 8 metastatic disease resulted in dramatic increases in survival of mice, and this survival was significantly greater than that afforded by either doxorubicin, 5-fluorouracil, or herceptin.     

The dodecameric vanadium-dependent haloperoxidase from the marine algae Corallina officinalis: cloning, expression, and refolding of the recombinant enzyme

The dodecameric vanadium-dependent bromoperoxidase from Corallina officinalis has been cloned and over-expressed in Escherichia coli. However, the enzyme was found to be predominantly in the form of inclusion bodies. This protein presents a challenging target for refolding, both due to the size (768 kDa) and quaternary structure (12 × 64 kDa). Successful refolding conditions have been established which result in an increase in the final yield of active bromoperoxidase from 0.5 mg to 40 mg per litre of culture. The refolded protein has been characterised and compared to the native enzyme and was shown to be stable at temperatures of 80 °C, over a pH range 5.5–10 and in organic solvents such as ethanol, acetonitrile, methanol, and acetone. The novel refolding approach reported in this paper opens up the full potential of this versatile enzyme for use in large scale biotransformation studies.     

Heterodimerization of the alpha and beta isoforms of the human thromboxane receptor enhances isoprostane signaling

Isoprostanes are free radical catalyzed products of arachidonic acid that are elevated in pro-oxidant disease states. Two isoprostanes, 8-isoprostaglandin F(2alpha) (iPF(2alpha)III) and 8-isoprostaglandin E2 (iPE2III), act at the receptor for thromboxane A2 (the TP) to mediate pro-atherogenic effects in vivo. We confirmed dimerization of the human TP isoforms, TPalpha and TPbeta, and determined the impact on isoprostane signaling. No overt changes in ligand binding at the TP were observed as a result of TPalpha/TPbeta coexpression. The response to iPF(2alpha)III or iPE2III was enhanced in HEK293 cells stably coexpressing TPalpha and TPbeta, as measured by inositol phosphate generation or intracellular calcium mobilization, relative to cells expressing TPalpha or TPbeta individually. In contrast, the response to traditional thromboxane analogs was unaltered. Augmented isoprostane signaling was similarly observed in HEK 293 cell transiently transfected with TPalpha and TPbeta. These results indicate that TPalpha/TPbeta dimerization enhances isoprostane-mediated signal transduction.     

Dietary hydroxy fatty acids are absorbed in humans: implications for the measurement of 'oxidative stress' in vivo.

Lipid peroxidation products formed in vivo or originating from the diet may lead to atherosclerosis. However, little is known about the absorption of these products in man. We studied the absorption of fat (30 g) containing 14-15 mg [U-13C]-labeled hydroxy or dihydroxy triglycerides in two groups of six apparently healthy women aged 40 +/- 2 years. Post-prandial 13C-labeled hydroxy fatty acid concentration increased in a pattern somewhat different from that of plasma triglycerides, with peak levels being reached between 4 and 6 h. However, the amount of 13C-labeled oxidized fat absorbed (area under the curve of plasma concentrations from 0 to 8 h) was related to that of plasma triglycerides: 13C hydroxy vs TG (r = 0.88, p <.02), and 13C dihydroxy vs TG (r = 0.85, p <.05). 13C monohydroxy triglycerides appeared to be absorbed to a greater extent than those of 13C dihydroxy triglycerides. Although low levels of 13C hydroxy lipids could be detected in fasting plasma after 24 h, concentrations were very low. Dietary lipid oxidation products are absorbed. The measurement of hydroxy fatty acids in plasma total lipids may not be a valid marker of lipid peroxidation in vivo when subjects are not fasting.     

Mapping of sister-chromatid exchanges in human chromosomes using G-banding and autoradiography.

Lumphocytes were pulse-labelled with [3H] thymidine. Following G-banding, the cells were autoradiographed and 46 in their third post-labelling division selected. The locations of 611 sister-chromatid exchanges (SCE's) which had occurred in the previous two cell cycles were recorded as label discontinuities along identified chromosomes. Between particular chromosomes, SCE frequency was proportional to chromosome length. SCE frequency distributions within particular chromosomes fitted Poisson expectations. There was no over-representation of exchanges in centromeric regions, or in the C-banded regions of chromosomes 1, 9 and 16. A trend of increased frequency of SCE in darkly G-banded regions and in relatively darkly banded chromosomes was evident. The apparent excess of SCE in dark G-bands could be considered to be a consequence of the more condensed state of the DNA in these regions in the interphase nucleus relative to the DNA in pale G-band regions. Such compaction could result in an enhanced probability of SCE and a reduced probability of gross inter- or intra-change involving these regions. In contrast, the more extended interphase state of the DNA in pale G-banded regions would allow non-homologous exchange and account for the preferred location of X-ray-induced exchange events to pale G-bands.