Characterization of inhalation exposure to gaseous elemental mercury during artisanal gold mining and e-waste recycling through combined stationary and personal passive sampling.

While occupational inhalation exposure to gaseous elemental mercury (GEM) has decreased in many workplaces as mercury is being removed from most products and processes, it continues to be a concern for those engaged in artisanal and small-scale gold mining or in recycling mercury-containing products. Recently, stationary and personal passive air samplers based on activated carbon sorbents and radial diffusive barriers have been shown to be suitable for measuring GEM concentrations across the range relevant for chronic health effects. Here, we used a combination of stationary and personal passive samplers to characterize the inhalation exposure to GEM of individuals living and working in two Ghanaian gold mining communities and working at a Norwegian e-waste recycling facility. Exposure concentrations ranging from <7 ng m-3 to >500 µg m-3 were observed, with the higher end of the range occurring in one gold mining community. Large differences in the GEM exposure averaged over the length of a workday between individuals can be rationalized by their activity and proximity to mercury sources. In each of the three settings, the measured exposure of the highest exposed individuals exceeded the highest concentration recorded with a stationary sampler, presumably because those individuals were engaged in an activity that generated or involved GEM vapors. High day-to-day variability in exposure for those who participated on more than one day, suggests the need for sampling over multiple days for reliable exposure characterization. Overall, a combination of personal and stationary passive sampling is a cost-effective approach that cannot only provide information on exposure levels relative to regulatory thresholds, but also can identify emission hotspots and therefore guide mitigation measures.

Development, characterization, and testing of a personal passive sampler for measuring inhalation exposure to gaseous elemental mercury.

Inhalation of gaseous elemental mercury (GEM) is an occupational exposure concern for workers handling elemental mercury or mercury-containing waste. GEM is also often present near historically mercury-contaminated sites, potentially resulting in low-level, chronic exposure of the wider population. Here we introduce a passive sampler for personal GEM monitoring which combines a radial porous diffusive barrier with an activated carbon sorbent. A total mercury analyzer is used to quantify GEM sorbed to the carbon by thermal decomposition, amalgamation, and atomic absorption spectroscopy. A sampling rate of 0.070 m3/day was determined by calibrating the sampler at low and high concentrations. Deployments lasting 8 h result in limits of quantification well below 200 ng/m3. The sampler has a measurement range of at least four orders of magnitude. Derived air concentrations were not statistically significantly different from those obtained by active air sampling but were more precise than those obtained using a personal pump. If properly stored, the sampler maintains low blank levels in high GEM environments. Affordability, sturdiness, simplicity, and the wide availability of total mercury analyzers make this sampler highly suited for monitoring GEM inhalation exposure, including in developing countries.

X-ray Characterization and Structure-Based Optimization of Striatal-Enriched Protein Tyrosine Phosphatase Inhibitors.

Excessive activity of striatal-enriched protein tyrosine phosphatase (STEP) in the brain has been detected in numerous neuropsychiatric disorders including Alzheimer's disease. Notably, knockdown of STEP in an Alzheimer mouse model effected an increase in the phosphorylation levels of downstream STEP substrates and a significant reversal in the observed cognitive and memory deficits. These data point to the promising potential of STEP as a target for drug discovery in Alzheimer's treatment. We previously reported a substrate-based approach to the development of low molecular weight STEP inhibitors with Ki values as low as 7.8 µM. Herein, we disclose the first X-ray crystal structures of inhibitors bound to STEP and the surprising finding that they occupy noncoincident binding sites. Moreover, we utilize this structural information to optimize the inhibitor structure to achieve a Ki of 110 nM, with 15-60-fold selectivity across a series of phosphatases.

Characterization of the complex formed between a potent neutralizing ovine-derived polyclonal anti-TNFα Fab fragment and human TNFα.

TNFα (tumour necrosis factor α) is an early mediator in the systemic inflammatory response to infection and is therefore a therapeutic target in sepsis. AZD9773 is an ovine-derived, polyclonal anti-TNFα Fab fragment derived from a pool of serum and currently being developed as a treatment for severe sepsis and septic shock. In the present study, we show that although AZD9773 has a modest affinity for TNFα in a binding assay, the Ki in a cell-based assay is approximately four orders of magnitude lower. We show using SEC (size exclusion chromatography) that the maximum size of the complex between AZD9773 and TNFα is consistent with approximately 12 Fabs binding to one TNFα trimer. A number of approaches were taken to map the epitopes recognized by AZD9773. These revealed that a number of different regions on TNFα are involved in binding to the polyclonal Fab. The data suggest that there are probably three epitopes per monomer that are responsible for most of the inhibition by AZD9773 and that all three can be occupied at the same time in the complex. We conclude that AZD9773 is clearly demonstrated to bind to multiple epitopes on TNFα and suggest that the polyclonal nature may account, at least in part, for the very high potency observed in cell-based assays.

Structure of IL-17A in complex with a potent, fully human neutralizing antibody.

IL-17A is a pro-inflammatory cytokine produced by the newly identified Th17 subset of T-cells. We have isolated a human monoclonal antibody to IL-17A (CAT-2200) that can potently neutralize the effects of recombinant and native human IL-17A. We determined the crystal structure of IL-17A in complex with the CAT-2200 Fab at 2.6 A resolution in order to provide a definitive characterization of the epitope and paratope regions. Approximately a third of the IL-17A dimer is disordered in this crystal structure. The disorder occurs in both independent copies of the complex in the asymmetric unit and does not appear to be influenced by crystal packing. The complex contains one IL-17A dimer sandwiched between two CAT-2200 Fab fragments. The IL-17A is a disulfide-linked homodimer that is similar in structure to IL-17F, adopting a cystine-knot fold. The structure is not inconsistent with the previous prediction of a receptor binding cavity on IL-17 family members. The epitope recognized by CAT-2200 is shown to involve 12 amino acid residues from the quaternary structure of IL-17A, with each Fab contacting both monomers in the dimer. All complementarity-determining regions (CDRs) in the Fab contribute to a total of 16 amino acid residues in the antibody paratope. In vitro affinity optimization was used to generate CAT-2200 from a parental lead antibody using random mutagenesis of CDR3 loops. This resulted in seven amino acid changes (three in VL-CDR3 and four in VH-CDR3) and gave an approximate 30-fold increase in potency in a cell-based neutralization assay. Two of the seven amino acids form part of the CAT-2200 paratope. The observed interaction site between CAT-2200 and IL-17A is consistent with data from hydrogen/deuterium exchange mass spectrometry and mutagenesis approaches.

Isolation by HPLC and characterisation of the bioactive fraction of New Zealand manuka (Leptospermum scoparium) honey.

Using HPLC a fraction of New Zealand manuka honey has been isolated, which gives rise to the non-peroxide antibacterial activity. This fraction proved to be methylglyoxal, a highly reactive precursor in the formation of advanced glycation endproducts (AGEs). Methylglyoxal concentrations in 49 manuka and 34 non-manuka honey samples were determined using a direct detection method and compared with values obtained using standard o-phenylenediamine derivatisation. Concentrations obtained using both the methods were similar and varied from 38 to 828 mg/kg.

Offspring produced from heterotopic ovarian allografts in male and female recipient mice.

Studies on human ovarian xenografts and mouse allografts indicate that the male hormonal milieu and exogenous gonadotrophin administration stimulate antral follicle growth. However, it is not known whether oocytes produced under these conditions are developmentally competent. The objective of our study was to evaluate the developmental competence of oocytes produced in heterotopic mouse ovarian grafts placed in male and female recipient mice. Gonadotrophins were 7.5 IU pregnant mare serum gonadotrophin (PMSG) alone or 7.5 IU PMSG and 7.5 IU human chorionic gonadotrophin or were not given prior to oocyte collection. The developmental competence of oocytes was assessed by performing in vitro fertilisation and embryo transfer to recipients. When no gonadotrophins were given the cleavage rate was similar for oocytes collected from ovarian grafts in male and female recipients. Gonadotrophin treatment significantly (P < 0.05) increased two-cell formation by oocytes grown in female graft recipients but not in male recipients. Implantation rates, fetal development and the birth of live young were unaffected by the sex of the graft recipient or gonadotrophin treatment. Live offspring were produced from oocytes collected from ovarian grafts in male and female recipients treated with or without gonadotrophins. In conclusion, this work has shown that the hormonal environment of male mice can support the growth of oocytes in ovarian allografts and that these oocytes can produce live offspring.

Xenotransplantation: a tool for reproductive biology and animal conservation?

The transplantation of reproductive organs, including ovaries and ovarian tissue, was pioneered over 100 years ago. In the 1960s, ovarian grafting was used as a tool to investigate ovarian function, but with the recent development of more effective cryopreservation protocols for ovarian tissue, germline preservation and propagation have now become realistic goals. This review describes progress in ovarian banking and ovarian tissue transplantation, with emphasis on how fresh and frozen ovarian tissue can be used in assisted reproduction for both humans and animals. This paper focuses most closely on the potential value of xenotransplantation, the transplantation of gonads from one species to another, to conserve rare and endangered species. Specific attention is drawn to the use of xenotransplantation as a strategy for generating viable gametes that can be used to produce live fertile offspring. Other upcoming xenogeneic technologies that may be of potential significance in animal conservation, such as transplantation of whole ovaries or isolated growing follicles, and even male germ cells, are discussed.