Challenges of antithrombotic therapy in the management of cardiovascular disease in patients with inherited bleeding disorders: A single-centre experience.

INTRODUCTION: Cardiovascular events in patients with inherited bleeding disorders are challenging to manage. The risk of bleeding secondary to antithrombotic treatment must be balanced against the risk of thrombosis secondary to haemostatic therapy. METHODS: Patients with inherited bleeding disorders with coronary artery bypass grafting (CABG), percutaneous coronary intervention (PCI) or atrial fibrillation (AF) from a single centre (2010-2018) are included. RESULTS: A total of 11 patients undergoing CABG (n = 3), PCI (n = 5) or with AF (n = 3) and a diagnosis of haemophilia A (n = 8), haemophilia B (n = 1), factor XI deficiency (n = 1) and von Willebrand disease (n = 1) managed by a multidisciplinary team are reported. In patients undergoing CABG, factor levels were normalized for 7-10 days with trough levels of 70-80% with severe patients continuing high-dose factor prophylaxis (trough 20-30%) three weeks post-operatively with daily aspirin. In a patient with mild haemophilia A and an inhibitor, recombinant factor VIIa dosing was monitored with thromboelastometry. For PCI, a 3rd-generation drug-eluting stent with one month of dual antiplatelet therapy in addition to high-dose prophylaxis as needed was preferred. Patients with AF and severe haemophilia did not receive antithrombotic treatment, and a thrombin generation assay was used to guide heparin dosing in mild haemophilia. CONCLUSION: Our experience demonstrates the importance of interdisciplinary communication to identify strategies that decrease the risk of bleeding and thrombosis. The use of extended, increased intensity prophylaxis facilitated antiplatelet therapy. Global assays may help balance the intensity of haemostatic and antithrombotic treatment.

Radiotherapy and Conservative Surgery in the Locoregional Management of Merkel Cell Carcinoma: The British Columbia Cancer Agency Experience.

BACKGROUND: Merkel cell carcinoma (MCC) is an uncommon aggressive skin malignancy. Published series mainly focus on wide excision, which can be difficult at some sites (e.g., face) and in patients with comorbidities. In British Columbia, an approach of conservative surgery followed by radiotherapy is common. MATERIALS AND METHODS: This is a retrospective review of 179 patients treated for MCC with curative intent in British Columbia. RESULTS: Totals of 68, 63, and 37 patients underwent narrow excision of primary, attempted wide excision, and biopsy only, respectively. Adjuvant radiotherapy reduced local recurrence after narrow excision (<10 mm margin) from 25 to 4.9 % (p = .03) and was effective in the presence of microscopic positive margins. Local recurrence rate was 7.1 % if the margin was >10 mm irrespective of radiation use. Local RFS was improved by adjuvant radiation therapy (RT) (p = 0.04), and there was a trend to reduced nodal relapse after elective nodal RT (p = .07). Irradiation of macroscopic tumor at 37 primary and 33 nodal sites provided 5-year local and nodal RFS of 90 and 75 %, respectively. The 5-year cancer specific survival was 77 % and was not improved by the use of adjuvant radiotherapy. CONCLUSIONS: Local excision plus adjuvant RT is an effective treatment for MCC. Adjuvant radiation should be considered when the excision margin is <1 cm.

An assessment of clinical laboratory performance for the determination of manganese in blood and urine.

BACKGROUND: Proficiency testing or external quality assessment schemes (PT/EQASs) are an important method of assessing laboratory performance. As each scheme establishes assigned values and acceptable ranges for the analyte according to its own criteria, monitoring of participant performance varies according to the scheme and can lead to conflicting conclusions. METHODS: Standard deviations (SDs) for PT were derived from Thompson's and biological variation models applied to blood and urine manganese (Mn) robust data from four EQASs from North America and Europe. The fitness for purpose was verified by applying these SDs to individual results. RESULTS: Using Thompson characteristic function the relationship between SD and Mn concentration, expressed in nmol/L was the square root of [19.72+(0.07712×Mn concentration2)] for blood and the square root of [6.772+(0.09852×Mn concentration2)] for urine. While the biological variation model was not suitable for urine, it produced an acceptable range for blood as ±54.4 nmol/L (assigned value ≤320 nmol/L) or 17% (assigned value >320 nmol/L). For blood, individual performance evaluated by the two approaches led to similar conclusions. CONCLUSIONS: The biological variation model can be used to propose quality specifications for blood, however it could not be applied to urine. The Thompson characteristic function model could be applied to derive quality specifications for Mn in urine and, to a lesser extent in blood. The more lenient quality specifications for blood highlight the difficulty of determining Mn in this matrix. Further work is needed to harmonize PT, such as using assigned ranges for the specimens.

Feasibility of asymmetric flow field-flow fractionation coupled to ICP-MS for the characterization of wear metal particles and metalloproteins in biofluids from hip replacement patients.

Hip replacements are used to improve the quality of life of people with orthopaedic conditions, but the use of metal-on-metal (MoM) arthroplasty has led to poor outcomes for some patients. These problems are related to the generation of micro- to nanosized metal wear particles containing Cr, Co or other elements, but the current analytical methods used to investigate the processes involved do not provide sufficient information to understand the size or composition of the wear particles generated in vivo. In this qualitative feasibility study, asymmetric flow field-flow fractionation (AF(4)) coupled with inductively coupled plasma mass spectrometry (ICP-MS) was used to investigate metal protein binding and the size and composition of wear metal particles present in serum and hip aspirates from MoM hip replacement patients. A well-established HPLC anion exchange chromatography (AEC) separation system coupled to ICP-MS was used to confirm the metal-protein associations in the serum samples. Off-line single particle ICP-MS (spICP-MS) analysis was used to confirm the approximate size distribution indicated by AF(4) of the wear particles in hip aspirates. In the serum samples, AF(4) -ICP-MS suggested that Cr was associated with transferrin (Tf) and Co with albumin (Alb) and an unidentified species; AEC-ICP-MS confirmed these associations and also indicated an association of Cr with Alb. In the hip aspirate sample, AF(4)-ICP-MS suggested that Cr was associated with Alb and Tf and that Co was associated with Alb and two unidentified compounds; AEC analysis confirmed the Cr results and the association of Co with Alb and a second compound. Enzymatic digestion of the hip aspirate sample, followed by separation using AF(4) with detection by UV absorption (280 nm), multi-angle light scattering and ICP-MS, suggested that the sizes of the Cr-, Co- and Mo-containing wear particles in a hip aspirate sample were in the range 40-150 nm. Off-line spICP-MS was used to confirm these findings for the Co- and Cr-containing nanoparticles. Whilst limited in scope, the results are sufficient to show the interaction of ions with transport proteins and give an indication of particle size, providing useful pathological indices. As such, the methods indicate a new way forward for in vivo investigation of the processes which lead to tissue necrosis and hip loosening in patients with MoM hip replacements.

Adenovirus-associated virus vector-mediated gene transfer in hemophilia B.

BACKGROUND: Hemophilia B, an X-linked disorder, is ideally suited for gene therapy. We investigated the use of a new gene therapy in patients with the disorder. METHODS: We infused a single dose of a serotype-8-pseudotyped, self-complementary adenovirus-associated virus (AAV) vector expressing a codon-optimized human factor IX (FIX) transgene (scAAV2/8-LP1-hFIXco) in a peripheral vein in six patients with severe hemophilia B (FIX activity, <1% of normal values). Study participants were enrolled sequentially in one of three cohorts (given a high, intermediate, or low dose of vector), with two participants in each group. Vector was administered without immunosuppressive therapy, and participants were followed for 6 to 16 months. RESULTS: AAV-mediated expression of FIX at 2 to 11% of normal levels was observed in all participants. Four of the six discontinued FIX prophylaxis and remained free of spontaneous hemorrhage; in the other two, the interval between prophylactic injections was increased. Of the two participants who received the high dose of vector, one had a transient, asymptomatic elevation of serum aminotransferase levels, which was associated with the detection of AAV8-capsid-specific T cells in the peripheral blood; the other had a slight increase in liver-enzyme levels, the cause of which was less clear. Each of these two participants received a short course of glucocorticoid therapy, which rapidly normalized aminotransferase levels and maintained FIX levels in the range of 3 to 11% of normal values. CONCLUSIONS: Peripheral-vein infusion of scAAV2/8-LP1-hFIXco resulted in FIX transgene expression at levels sufficient to improve the bleeding phenotype, with few side effects. Although immune-mediated clearance of AAV-transduced hepatocytes remains a concern, this process may be controlled with a short course of glucocorticoids without loss of transgene expression. (Funded by the Medical Research Council and others; ClinicalTrials.gov number, NCT00979238.).

Outcomes of Merkel cell carcinoma treated with radiotherapy without radical surgical excision.

BACKGROUND: Achieving clear surgical margins in Merkel cell carcinoma (MCC) can be difficult due to tumor location or patient comorbidity. Clinical impression suggests that radiation treatment achieves good control of macroscopic disease. METHODS: A retrospective chart review was undertaken of all patients with pathological evidence of MCC and treated with curative intent at the BC Cancer Agency between 1979 and 2007. This is a report on the outcomes of those with gross disease treated with radiotherapy, without radical surgery. RESULTS: Fifty-seven patients received definitive radiotherapy to the primary and/or nodal disease. Median age was 75 years and median follow-up was 34 months (84.5 months for those alive at last follow-up). American Joint Committee on Cancer (AJCC) stage distribution was 23, 19, and 58 % for stages I, II, and III, respectively. Tumor control at sites treated for macroscopic disease was 88 % at 12 months and 82 % at 2 years, and 5-year local relapse-free survival (RFS) was 90 %. Five-year RFS, cancer-specific survival (CSS), and overall survival were 57, 68, and 39 %, respectively. On univariate and multivariate analyses, only male sex was associated with a worse RFS, and a radiotherapy dose >50 Gy was associated with a better CSS. LIMITATIONS: The retrospective nature of the study and small sample size limit the strength of the conclusions. CONCLUSIONS: Radical radiotherapy is effective in the curative treatment of MCC, especially in patients who would tolerate wide surgical excision poorly, or where it would cause significant cosmetic or functional deficits.

Review: Advances in the Accuracy and Traceability of Metalloprotein Measurements Using Isotope Dilution Inductively Coupled Plasma Mass Spectrometry.

Advances in inductively coupled plasma mass spectrometry and the methods used to prepare isotopically enriched standards, allow for the high accuracy measurement of metalloproteins by isotope dilution mass spectrometry. This technique has now reached a level of maturity whereby a step change in the accuracy, precision, and traceability of, in particular, clinical, and biomedical measurements is achievable. Current clinical measurements, which require low limits of detection in the presence of complex sample matrices, use indirect methods based on immunochemistry for the study of human disease. However, this approach suffers from poor traceability, requiring comparisons based on provision of matrix-based reference materials, used as analytical standards. This leads to difficulty when changes in the reference material are required, often resulting in a lack of interlaboratory and temporal comparability in clinical results and reference ranges. In this review, we focus on the most important metalloproteins for clinical studies, to illustrate how the attributes of chromatography coupled to inorganic mass spectrometry can be used for the direct measurement of metalloproteins such as hemoglobin, transferrin, and ceruloplasmin. By using this approach, we hope to demonstrate how isotope dilution analysis can be used as a reference method to improve traceability and underpin clinical, biomedical, and other biological measurements.

Rationale and design of a randomised trial of trientine in patients with hypertrophic cardiomyopathy.

AIMS: Hypertrophic cardiomyopathy (HCM) is characterised by left ventricular hypertrophy (LVH), myocardial fibrosis, enhanced oxidative stress and energy depletion. Unbound/loosely bound tissue copper II ions are powerful catalysts of oxidative stress and inhibitors of antioxidants. Trientine is a highly selective copper II chelator. In preclinical and clinical studies in diabetes, trientine is associated with reduced LVH and fibrosis, and improved mitochondrial function and energy metabolism. Trientine was associated with improvements in cardiac structure and function in an open-label study in patients with HCM. METHODS: The Efficacy and Mechanism of Trientine in Patients with Hypertrophic Cardiomyopathy (TEMPEST) trial is a multicentre, double-blind, parallel group, 1:1 randomised, placebo-controlled phase II trial designed to evaluate the efficacy and mechanism of action of trientine in patients with HCM. Patients with a diagnosis of HCM according to the European Society of Cardiology Guidelines and in New York Heart Association classes I-III are randomised to trientine or matching placebo for 52 weeks. Primary outcome is change in left ventricular (LV) mass indexed to body surface area, measured using cardiovascular magnetic resonance. Secondary efficacy objectives will determine whether trientine improves exercise capacity, reduces arrhythmia burden, reduces cardiomyocyte injury, improves LV and atrial function, and reduces LV outflow tract gradient. Mechanistic objectives will determine whether the effects are mediated by cellular or extracellular mass regression and improved myocardial energetics. CONCLUSION: TEMPEST will determine the efficacy and mechanism of action of trientine in patients with HCM. TRIAL REGISTRATION NUMBERS: NCT04706429 and ISRCTN57145331.

Metal(loid) levels in biological matrices from human populations exposed to mining contamination--Panasqueira Mine (Portugal).

Mining activities may affect the health of miners and communities living near mining sites, and these health effects may persist even when the mine is abandoned. During mining processes various toxic wastes are produced and released into the surrounding environment, resulting in contamination of air, drinking water, rivers, plants, and soils. In a geochemical sampling campaign undertaken in the Panasqueira Mine area of central Portugal, an anomalous distribution of several metals and arsenic (As) was identified in various environmental media. Several potentially harmful elements, including As, cadmium (Cd), chromium (Cr), manganese (Mn), nickel (Ni), lead (Pb), and selenium (Se), were quantified in blood, urine, hair, and nails (toe and finger) from a group of individuals living near the Panasqueira Mine who were environmentally and occupationally exposed. A group with similar demographic characteristics without known exposure to mining activities was also compared. Genotoxicity was evaluated by means of T-cell receptor (TCR) mutation assay, and percentages of different lymphocyte subsets were selected as immunotoxicity biomarkers. Inductively coupled plasma-mass spectrometry (ICP-MS) and inductively coupled plasma-atomic emission spectrometry (ICP-AES) analysis showed elevated levels of As, Cd, Cr, Mn, and Pb in all biological samples taken from populations living close to the mine compared to controls. Genotoxic and immunotoxic differences were also observed. The results provide evidence of an elevated potential risk to the health of populations, with environmental and occupational exposures resulting from mining activities. Further, the results emphasize the need to implement preventive measures, remediation, and rehabilitation plans for the region.

A Proposed Concept Model for Cancer Risk in Nigerian Electronic Waste Exposure - A Brief Report.

Aim: This report aims to render a proposed concept model for cancer risk in Nigerian electronic waste exposure by making deductions from data on the assessment of Nigerians' exposure to toxic metals in e-waste, using biomarkers of exposure and genotoxicity to evaluate the risk of cancer development. Material and methods: In the cross-sectional study, 632 consenting participants, consisting of 381 e-waste workers (EW) and 120 environmental e-waste exposed participants (EEEP), age-matched with 131 unexposed participants (controls), were enrolled from Benin, Lagos and Ibadan, Southwestern Nigeria. Levels of selected toxic metals in blood and essential metals in serum were determined using inductively coupled plasma-mass spectrometry. Oxidative stress biomarkers, including malondialdehyde and uric acid (UA), and activities of enzymatic antioxidants [catalase, superoxide dismutase (SOD), γ-glutamyltransferase (GGT) and glutathione peroxidase (GPx)], were determined in serum using standard methods like spectrophotometry. Genotoxicity biomarkers - wild-type tumour suppressor protein (wt-p53), 8-oxoguanine-DNA glycosylase (OGG1), and 8-hydroxy-2'-deoxyguanosine (8-OHdG); glutathione (GSH); and tumour markers [prostate-specific antigen (PSA) and alpha-fetoprotein] - were determined in serum using ELISA. Micronucleus assay was carried out using microscopy. Data were analysed using ANOVA and Pearson's correlation coefficient at α0.05. Results: There was evidence indicating elevated levels of genotoxic toxic metals, decreased levels of genome protective metals, increased oxidative stress markers as well as reduced cellular antioxidants in both EW and EEEP compared to controls. Additionally, the levels of wt-p53 in EW and EEEP were lower than controls, while OGG1 activity in EEEP was higher. The PSA and alpha-fetoprotein in EW were more elevated than EEEP and controls, respectively. The MnPCE/1000PCE in EW was higher than EEEP and controls. Conclusion: The proposed schematic model could be adopted to illustrate cancer risk in Nigerian population exposed to electronic waste.

Determination of cisplatin 1,2-intrastrand guanine-guanine DNA adducts in human leukocytes by high-performance liquid chromatography coupled to inductively coupled plasma mass spectrometry.

Platinum-containing drugs are widely used to treat cancer in a variety of clinical settings. Their mode of action involves the formation of DNA adducts, which facilitate apoptosis in cancer cells. Cisplatin binds to the N7 position of the purine DNA bases forming intrastrand cross-links between either two adjacent guanines [cis-Pt(NH(3))(2)d(pGpG), 1,2-GG] or an adjacent adenine and guanine [cis-Pt(NH(3))(2)d(pApG), 1,2-AG)]. The cytotoxic efficacy for each of the different types of DNA adducts and the relationship between adduct levels in tumor cells and blood are not well understood. By using these Pt-containing adduct species as biomarkers, information on a patient's response to chemotherapy would be directly related to the mode of action of the drug. This type of analysis requires the most sensitive and specific methods available, to facilitate detection limits sufficient to measure the DNA adduct in the limited sample quantities available from patients. This was achieved in the current study by coupling a highly specific enzyme-based adduct isolation method with a sensitive detection system based on HPLC coupled to inductively coupled plasma mass spectrometry to measure the 1,2-GG cisplatin adducts formed in DNA. The method was developed and validated using calf thymus DNA and two different adenocarcinoma cell lines. The values for the limit of detection (LOD) and the limit of quantitation determined for the 1,2-GG cisplatin adduct were 0.21 and 0.67 fmol per microg DNA, respectively. This corresponds to an absolute LOD of 0.8 pg as Pt for the 1,2-GG adduct. Cisplatin-sensitive (H23) and -resistant (A549) tumor cells were exposed to the drug, and the 1,2-GG adduct levels were measured over a 24 h time period. The results showed a statistically significant (P < 0.05) higher concentration in the sensitive cells as compared to the resistant cells after repair for 7 h. Although the adduct concentration present fell at subsequent time points (12 and 24 h), the levels in each cell line were broadly similar. The protocol was then applied to the analysis of patient samples taken before and then 1 h after treatment. The 1,2-GG cisplatin adduct was present in the range from 113 to 1245 fg Pt per microg DNA in all of the patient samples taken after treatment. Although the adduct was not present at levels greater than the LOD in the initial pretreatment samples, trace amounts were discernible in some patient samples on their third treatment cycle.

DNA damage in earthworms from highly contaminated soils: assessing resistance to arsenic toxicity by use of the Comet assay.

Earthworms native to the former mine site of Devon Great Consols (DGC), UK reside in soils highly contaminated with arsenic (As). These earthworms are considered to have developed a resistance to As toxicity. The mechanisms underlying this resistance however, remain unclear. In the present study, non-resistant, commercially sourced Lumbricus terrestris were exposed to a typical DGC soil in laboratory mesocosms. The earthworms bio-accumulated As from the soil and incurred DNA-damage levels significantly above those observed in the control mesocosm (assessed using the Comet assay). A dose response was observed between DNA damage (% tail DNA) and As concentration in soil (control, 98, 183, 236, 324 and 436mgkg(-1)). As-resistant earthworms (Lumbricus rubellus, Dendrodrilus rubidus and L. terrestris) collected from contaminated soils at DGC (203 to 9025mgkg(-1) As) had also bio-accumulated high levels of As from their host soils, yet demonstrated low levels of DNA damage compared with earthworms from uncontaminated sites. The results demonstrate that the As-contaminated soils at DGC are genotoxic to non-native earthworms and much less so to earthworms native to DGC, thus providing further evidence of an acquired resistance to As toxicity in the native earthworms.

Biomedical copper speciation in relation to Wilson's disease using strong anion exchange chromatography coupled to triple quadrupole inductively coupled plasma mass spectrometry.

Biomedical analytical methods often rely on indirect measurements, such as immunoassays, which can lack effective metrological traceability. In the nephelometric determination of ceruloplasmin (Cp), an important protein whose circulating level is altered in Wilson's disease (WD), the anti-Cp antibody used is not specific for the biologically active holoprotein so the assay can overestimate the concentration of Cp due to the presence of the apoprotein. By providing quantitation using elemental standards, the use of strong anion exchange chromatography (SAX) coupled to triple quadrupole inductively coupled plasma mass spectrometry (ICP-MS-MS) can overcome the drawbacks of methods for the measurement of metalloproteins reliant on immunoassays. In the current study, a SAX-ICP-MS-MS method for Cp quantification was designed and tested in samples of blood serum of WD patients and healthy controls. Using standards based on a copper-EDTA complex for calibration, the method provides relatively simple quantification of Cp with the limit of detection of 0.1 µg L-1 (limit of quantification 0.4 µg L-1). The method was also used to investigate the copper species separated by using a 30 kDa cut-off ultrafiltration device. The so-called "exchangeable" copper fraction is considered as an alternative clinical biomarker of WD. Using the designed speciation approach, it was shown that the ultrafiltration method can overestimate the "exchangeable" copper fraction due to a removal of copper from Cp. This was confirmed by comparing the enzymatic activity of the fractions. Thus, the specificity of the "exchangeable" copper test can be ensured only under strict maintenance of ultrafiltration conditions.

Human toenails as a biomarker of exposure to elevated environmental arsenic.

A pilot study was conducted to determine the applicability of toenails as a biomarker of exposure to elevated environmental arsenic (As) levels. A total of 17 individuals were recruited for the pilot study: 8 residents living near to a former As mine, Devon, UK, forming the exposed group, plus 9 residents from Nottinghamshire, UK, with no anticipated As exposure who were used for comparison as a control group. All toenail samples were thoroughly washed prior to analysis and the wash solutions retained for As determination via ICP-MS to provide an indication of the background environmental As levels for each group. Total As was determined in washed toenail samples via ICP-MS following microwave assisted acid digestion. Concentrations of total As in the toenails of the exposed group were elevated, ranging from 858 to 25 981 microg kg(-1) (geometric mean = 5406 microg kg(-1)), compared to the control group whose toenail As concentrations ranged from 73 to 273 microg kg(-1) (geometric mean = 122 microg kg(-1)). Higher levels of exogenous As contamination were present on the toenails of the exposed group (geometric mean = 506 microg kg(-1)) compared to the control group (geometric mean = 4.0 microg kg(-1)) providing evidence of higher environmental As levels in the exposed group. Total As concentrations in toenail samples were positively correlated to environmental As levels (r = 0.60, p < 0.001). HPLC-ICP-MS analysis of aqueous toenail extracts revealed inorganic arsenite (As(III)) to be the dominant species extracted ( approximately 83%) with lesser amounts of inorganic arsenate (As(V)) and organic dimethylarsinate (DMA(V)) at approximately 13% and approximately 8.5%, respectively. Arsenic speciation in analysed toenail extracts from the two groups was comparable. The only notable difference between groups was the presence of small amounts (<1%) of organic methylarsonate (MA(V)) in two toenail samples from the exposed group. Toenails are presented as a viable biomarker of exposure at sites with elevated environmental As, such as the former mining sites found throughout Devon and Cornwall, UK.

Arsenic biotransformation in earthworms from contaminated soils.

Two species of arsenic (As) resistant earthworm, Lumbricus rubellus and Dendrodrillus rubidus, their host soils and soil excretions (casts) were collected from 23 locations at a former As mine in Devon, UK. Total As concentrations, measured by ICP-MS, ranged from 255 to 13,080 mg kg(-1) in soils, 11 to 877 mg kg(-1) in earthworms and 284 to 4221 mg kg(-1) in earthworm casts from a sub-sample of 10 of the 23 investigated sites. The samples were also measured for As speciation using HPLC-ICP-MS to investigate potential As biotransformation pathways. Inorganic arsenate (As(V)) and arsenite (As(III)) were the only species detected in the soil. As(V) and As(III) were also the dominant species found in the earthworms and cast material together with lower proportions of the organic species methylarsonate (MA(V)), dimethylarsinate (DMA(V)), arsenobetaine (AB) and three arsenosugars. Whilst the inorganic As content of the earthworms increased with increasing As body burden, the concentration of organic species remained relatively constant. These results suggest that the biotransformation of inorganic arsenic to organic species does not contribute to As resistance in the sampled earthworm populations. Quantification of As speciation in the soil, earthworms and cast material allows a more comprehensive pathway for the formation of AB in earthworms to be elucidated.

Earthworms and in vitro physiologically-based extraction tests: complementary tools for a holistic approach towards understanding risk at arsenic-contaminated sites.

The relationship of the total arsenic content of a soil and its bioaccumulation by earthworms (Lumbricus rubellus and Dendrodrilus rubidus) to the arsenic fraction bioaccessible to humans, measured using an in vitro physiologically-based extraction test (PBET), was investigated. Soil and earthworm samples were collected at 24 sites at the former arsenic mine at the Devon Great Consols (DGC) in southwest England (UK), along with an uncontaminated site in Nottingham, UK, for comparison. Analysis of soil and earthworm total arsenic via inductively coupled plasma mass spectrometry (ICP-MS) was performed following a mixed acid digestion. Arsenic concentrations in the soil were elevated (204-9,025 mg kg(-1)) at DGC. The arsenic bioaccumulation factor (BAF) for both earthworm species was found to correlate positively with the human bioaccessible fraction (HBF), although the correlation was only significant (P < or = 0.05) for L. rubellus. The potential use of both in vitro PBETs and earthworms as complementary tools is explored as a holistic and multidisciplinary approach towards understanding risk at contaminated sites. Arsenic resistant earthworm species such as the L. rubellus populations at DGC are presented as a valuable tool for understanding risk at highly contaminated sites.

Isotopic labelling of peptides and isotope ratio analysis using LC-ICP-MS: a preliminary study.

Bradykinin and substance P have been derivatised with cyclic diethylenetriaminepentaacetic anhydride (cDTPA) and subsequently labelled with natural and isotopically enriched Eu(3+). This enabled the detection and relative quantitation of the peptides using element-selective detection by high-performance liquid chromatography inductively coupled plasma mass spectrometry (LC-ICP-MS). Relative quantitation was achieved by differentially labelling two peptide sources, after derivatisation with cDTPA, using natural and enriched (151)Eu respectively. The (151)Eu:(153)Eu isotope ratio was measured and used to calculate the original peptide ratio. The measured ratios came within 5.2% of the known ratio. Derivatisation and chelation reactions were additionally confirmed using LC-ESI-MS.

Quantitative arsenic speciation in two species of earthworms from a former mine site.

The relationship between the total arsenic concentration and the chemical speciation of arsenic in two species of earthworm (Lumbricus rubellus and Dendrodrilus rubidus) in relation to the host soil, was investigated for 13 sites of varying arsenic content, including a background level garden soil and a former mine site at the Devon Great Consols, UK. Earthworms were collected with the host soil (As soil concentration range 16-12, 466 mg kg(-1) dry weight) and measured for their total arsenic (concentration range 7-595 mg kg(-1) dry weight) using inductively coupled plasma mass spectrometry (ICP-MS). A methanol-water mixture was used to extract arsenic species from the earthworms prior to determination of the individual arsenic species by a combination of anion and cation exchange high performance liquid chromatography coupled to inductively coupled plasma mass spectrometry (HPLC-ICP-MS). A gradient elution anion exchange method is presented whereby nine arsenic species could be measured in one sample injection. Arsenic species were identified by comparison of retention times and sample spiking with known standards and a fully characterised seaweed extract. Arsenic was generally present in the earthworm as arsenate (As(V)) or arsenite (As(III)) and arsenobetaine (AB). Methylarsonate (MA), dimethylarsinate (DMA) and three arsenosugars (glycerol, phosphate, sulfate) were present as minor constituents. These results are discussed in relation to the mechanisms for coping with exposure to soil bound arsenic.

Mediastinal node staging by positron emission tomography-computed tomography and selective endoscopic ultrasound with fine needle aspiration for patients with upper gastrointestinal cancer: Results from a regional centre.

AIM: To investigate the impact of endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) and positron emission tomography-computed tomography (PET-CT) in the nodal staging of upper gastrointestinal (GI) cancer in a tertiary referral centre. METHODS: We performed a retrospective review of prospectively recorded data held on all patients with a diagnosis of upper GI cancer made between January 2009 and December 2015. Only those patients who had both a PET-CT and EUS with FNA sampling of a mediastinal node distant from the primary tumour were included. Using a positive EUS-FNA result as the gold standard for lymph node involvement, the sensitivity, specificity, positive and negative predictive values (PPV and NPV) and accuracy of PET-CT in the staging of mediastinal lymph nodes were calculated. The impact on therapeutic strategy of adding EUS-FNA to PET-CT was assessed. RESULTS: One hundred and twenty one patients were included. Sixty nine patients had a diagnosis of oesophageal adenocarcinoma (Thirty one of whom were junctional), forty eight had oesophageal squamous cell carcinoma and four had gastric adenocarcinoma. The FNA results were inadequate in eleven cases and the PET-CT findings were indeterminate in two cases, therefore thirteen patients (10.7%) were excluded from further analysis. There was concordance between PET-CT and EUS-FNA findings in seventy one of the remaining one hundred and eight patients (65.7%). The sensitivity, specificity, PPV and NPV values of PET-CT were 92.5%, 50%, 52.1% and 91.9% respectively. There was discordance between PET-CT and EUS-FNA findings in thirty seven out of one hundred and eight patients (34.3%). MDT discussion led to a radical treatment pathway in twenty seven of these cases, after the final tumour stage was altered as a direct consequence of the EUS-FNA findings. Of these patients, fourteen (51.9%) experienced clinical remission of a median of nine months (range three to forty two months). CONCLUSION: EUS-FNA leads to altered staging of upper GI cancer, resulting in more patients receiving radical treatment that would have been the case using PET-CT staging alone.