Perioperative elafin for ischaemia-reperfusion injury during coronary artery bypass graft surgery: a randomised-controlled trial.

BACKGROUND: Elafin is a potent endogenous neutrophil elastase inhibitor that protects against myocardial inflammation and injury in preclinical models of ischaemic-reperfusion injury. We investigated whether elafin could inhibit myocardial ischaemia-reperfusion injury induced during coronary artery bypass graft (CABG) surgery. METHODS AND RESULTS: In a randomised double-blind placebo-controlled parallel group clinical trial, 87 patients undergoing CABG surgery were randomised 1:1 to intravenous elafin 200 mg or saline placebo administered after induction of anaesthesia and prior to sternotomy. Myocardial injury was measured as cardiac troponin I release over 48 h (area under the curve (AUC)) and myocardial infarction identified with MRI. Postischaemic inflammation was measured by plasma markers including AUC high-sensitive C reactive protein (hs-CRP) and myeloperoxidase (MPO). Elafin infusion was safe and resulted in >3000-fold increase in plasma elafin concentrations and >50% inhibition of elastase activity in the first 24 h. This did not reduce myocardial injury over 48 h (ratio of geometric means (elafin/placebo) of AUC troponin I 0.74 (95% CI 0.47 to 1.15, p=0.18)) although post hoc analysis of the high-sensitive assay revealed lower troponin I concentrations at 6 h in elafin-treated patients (median 2.4 vs 4.1 µg/L, p=0.035). Elafin had no effect on myocardial infarction (elafin, 7/34 vs placebo, 5/35 patients) or on markers of inflammation: mean differences for AUC hs-CRP of 499 mg/L/48 h (95% CI -207 to 1205, p=0.16), and AUC MPO of 238 ng/mL/48 h (95% CI -235 to 711, p=0.320). CONCLUSIONS: There was no strong evidence that neutrophil elastase inhibition with a single-dose elafin treatment reduced myocardial injury and inflammation following CABG-induced ischaemia-reperfusion injury. TRIAL REGISTRATION NUMBER: (EudraCT 2010-019527-58, ISRCTN82061264).

Towards an understanding of the control of 'crumbly' fruit in red raspberry.

The genetic disorder known as 'crumbly' fruit is becoming a serious problem in the European raspberry industry. The study set out to examine the crumbly phenotype in a red raspberry mapping population under two environments (field and polytunnel) across six seasons in an effort to understand variability of the syndrome and to examine whether genetic factors were important and if so, whether QTL associated with the phenotype could be identified. This highlighted that seasonal, environmental (field or polytunnel) and genetic factors all influence the condition. Two QTL that are important for the genetic control of the condition have been located on linkage groups one and three, and an association with ripening time has been identified.

Nosocomial outbreak of staphyloccocal scalded skin syndrome in neonates in England, December 2012 to March 2013.

Staphylococcal scalded skin syndrome (SSSS) is a blistering skin condition caused by exfoliative toxin-producing strains of Staphylococcus aureus. Outbreaks of SSSS in maternity settings are rarely reported. We describe an outbreak of SSSS that occurred among neonates born at a maternity unit in England during December 2012 to March 2013. Detailed epidemiological and microbiological investigations were undertaken. Eight neonates were found to be infected with the outbreak strain of S. aureus, of spa type t346, representing a single pulsotype. All eight isolates contained genes encoding exfoliative toxin A (eta) and six of them contained genes encoding toxin B (etb). Nasal swabs taken during targeted staff screening yielded a staphylococcal carriage rate of 21% (17/80), but none contained the outbreak strain. Mass screening involving multi-site swabbing and pooled, enrichment culture identified a healthcare worker (HCW) with the outbreak strain. This HCW was known to have a chronic skin condition and their initial nasal screen was negative. The outbreak ended when they were excluded from work. This outbreak highlights the need for implementing robust swabbing and culture methodswhen conventional techniques are unsuccessful in identifying staff carrier(s). This study adds to the growing body of evidence on the role of HCWs in nosocomial transmission of S. aureus.

Campylobacteriosis outbreak associated with consumption of undercooked chicken liver pâté in the East of England, September 2011: identification of a dose-response risk.

A foodborne outbreak with 49 cases (22 culture positive for Campylobacter sp.) following a wedding party in the East of England was investigated. A retrospective cohort study identified an association between consumption of chicken liver pâté and infection with Campylobacter jejuni/coli. There was a statistically significant association between dose (amount of chicken liver pâté eaten) and the risk of disease ['tasted': odds ratio (OR) 1·5, 95% confidence interval (CI) 0·04-∞; 'partly eaten': OR 8·4, 95% CI 1·4-87·5; 'most or all eaten': OR 36·1, 95% CI 3·3-2119). The local authority found evidence that the preparation of chicken livers breached Food Standards Agency's guidelines. This epidemiological investigation established a clear dose-response relationship between consumption of chicken liver pâté and the risk of infection with Campylobacter. The continuing need to raise public awareness of the risk to human health posed by undercooked chicken liver is evident.

Streptococcus pyogenes cluster in a care home in England April to June 2010.

Two fatal cases of Streptococcus pyogenes emm st22.6 bacteraemia occurred in a care home in England during April and June 2010, initiating a cluster investigation. The first case had left the home 13 days before the second case took up residence. We sought further cases and carriers. We swabbed throat and chronic skin lesions from residents and staff and examined these specimens for the presence of S. pyogenes. 61 specimens were taken from 18 of 19 residents and 39 of 39 staff. All results from swabbing were culture negative. We observed infection control practices and the environment at the care home for deficiencies. Issues were identified relating to the correct use of personal protective equipment, hand hygiene, clinical waste and laundry. Infection control practices were improved and training given. Infection control practices and the environment at a care home should be examined as part of the investigation of a S. pyogenes cluster. Screening for carriage of S. pyogenes should be done before antibiotic chemoprophylaxis is issued to care home residents and staff.

Five hepatitis B outbreaks in care homes in the UK associated with deficiencies in infection control practice in blood glucose monitoring.

Healthcare-associated hepatitis B virus (HBV) outbreaks have been reported in the USA and from several countries in Europe. Patient-to-patient transmission of HBV in these settings has been linked to several different types of exposure but one of the most common exposures implicated is the use of 'finger-stick' lancet devices for blood glucose testing. This article is an account of the investigations into a series of HBV outbreaks linked to the use of lancing devices in community healthcare settings in the UK. Between February 2004 and December 2006, nine individuals with acute HBV infection were reported to five local units of the Health Protection Agency. Investigations identified a further 12 individuals with HBV infection in residents in these settings. The epidemiological and environmental evidence suggests that HBV transmission occurred mostly from a significant breakdown in infection control measures in blood glucose testing. The occurrence of these outbreaks has highlighted the confusion that exists and the need for clear recommendations regarding the use of such devices in the UK.

Acute and long-term effects of exposure to sodium monofluoroacetate (1080) in sheep.

Acute and long-term effects of a single, relatively high oral dose (0.25 and 0.30 mg/kg) of sodium monofluoroacetate (1080) on the survival and productivity of sheep were evaluated to establish a better understanding of 1080 poisoning and identify more specific changes diagnostic of toxicosis. In survivors, clinical signs of acute 1080 toxicosis such as salivation and lethargy were generally very mild. Fasted animals were more prone to 1080 toxicity. In animals that died, more severe signs, including tachypnoea, dyspnoea, and tremors occurred for 15-20 min prior to death. 1080 concentrations were highest in the blood > heart > skeletal muscle > liver. 1080 could not be detected in any of these organs of the animals that survived. Serum citrate concentrations were elevated for 4 days after dosing. No clinical or biochemical abnormalities were found in any animal after 4 days. Histopathological lesions were most marked in the heart and lung with inflammation, necrosis, and scattered foci of fibrous tissue in the myocardium, pulmonary oedema and inflammation of the lung. No adverse long-term effects on general health or reproductive performance were observed in any sheep that survived the first 4 days following exposure to 1080. The most reliable diagnostic indicators of 1080 exposure in sheep were measurement of its residues in blood, skeletal muscle and ruminal contents, increased serum citrate concentration, elevated heart rate, and characteristic electrocardiograph changes (up to 4 days after exposure). Death from 1080 is most likely to occur within 96 h, and animals that survived this period appeared normal.

Outbreak of Salmonella Enteritidis phage type 13a: case-control investigation in Hertsmere, United Kingdom.

Cases of illness were reported to Hertsmere Borough Council among attendees of a children's charity event in June 2006. Initial laboratory investigation identified Salmonella Enteritidis PT13a as a possible cause of the outbreak. We carried out an unmatched case-control investigation. The population at risk included all individuals who attended the event. Self-completion questionnaires were sent to 53 presumptive cases and 212 randomly selected potential controls. Information was available for 49 cases and 128 controls (overall response rate=75%). We calculated odds ratios from single and multivariable analysis and tested for all two-way interactions. Risk factors for diarrhoea were eating egg mayonnaise bagels (OR=34.1, 95%CI 10.5 - 111.3) and drinking apple juice (OR=16.1, 95% CI 3.5 - 74.2). There was weak statistical evidence to suggest that the risk of diarrhoea after eating egg mayonnaise bagels was greater in the afternoon. No food samples were available to confirm which food item might have caused this outbreak. Eggs from Spain were used by the caterer. The ecology of salmonella, experience from previous outbreaks and epidemiological findings from this case-control investigation suggest that the most likely cause of the outbreak was contaminated eggs.

Behavioural, biochemical, and pathological responses of possums (Trichosurus vulpecula) poisoned with phosphorus paste.

AIM: To investigate the behavioural, biochemical and pathological responses of possums following poisoning with phosphorus paste, in order to assess the implications for the welfare of possums. METHODS: After ingestion of phosphorus paste by wild-caught possums (18 high dose, nine low dose, and 12 non-poisoned controls), behavioural observations were made at 15-min intervals for 24 h or until death. Serum biochemistry, and gross and microscopic pathology were assessed at 3-hourly intervals in a further 21 possums. RESULTS: Possums that ingested phosphorus paste developed an abnormal posture (high incidence of crouching after 4-8 h), mild congestion of the gastric mucosa, and elevated levels of creatine kinase (CK) in serum after 3-6 h. Retching was observed in 67% possums, and 44% vomited at least once. Possums were prostrate from about 18 h after eating the poison, and the response to handling, an indicator of consciousness, was lost at about 24 h, followed by death at 25 h. CONCLUSION: The main welfare concern was the possibility of discomfort or pain caused by the congestion of the gastric mucosa, as indicated by the crouched posture adopted by poisoned possums. Retching and vomiting may also have caused pain and distress. The degree of pain or discomfort would depend on the degree of congestion of the gastric mucosa, which was typically mild, and on the duration and severity of retching and vomiting, which were typically short and mild. Possums remained conscious until 1 h before death, implying that they were able to experience pain and distress from the effects of ingestion of phosphorus for almost the entire period of illness, which lasted for approximately one day.

Coenzyme Q differentially modulates phospholipid hydroperoxide glutathione peroxidase gene expression and free radicals production in malignant and non-malignant prostate cells.

The aim of this study was to investigate the role of coenzyme Q on the mRNA abundance of PHGPx and the reactive oxygen species (ROS) production in two different cell lines from human prostate, a line of non cancer cells (PNT2) and a line of cancer cells (PC3). Results showed that malignant cells markedly differ in their response to coenzyme Q compared to non-malignant cells, with no changes in PHGPx expression and greater ROS production. Furthermore coenzyme Q supplementation significantly lowered cell growth of the PC3 cancer line without affecting the PNT2. If these results are confirmed with additional experiments, it could represent a novel and interesting approach on the biomedical use of coenzyme Q10 in cancer therapy.

The biological activities of new polyamine derivatives as potential therapeutic agents.

A number of polyamine derivatives have demonstrated potential as therapeutic agents. For example, 1,12-bisethylspermine and bisnaphthalimide (elinafide) are currently in phase I clinical trials for the treatment of certain cancers. Here, the biological activities of two new groups of polyamine derivative, namely the oxa-polyamines and the bisnaphthalimides, are presented. The most active compounds in the oxa-polyamine and bisnaphthalimido series possessed IC(50) values of 2.93 and 1.38 microM, respectively, against MCF7 cells after 48 h of exposure. The structure-relationship activities of each group of compounds are discussed. Bisnaphthalimido compounds are DNA-binding agents. Addition of the bisnaphthalimides PK3, PK4, PK5, PK6 and PK7, at a concentration of 10 microM, to the calf thymus DNA duplex increased the T (m) of DNA by 11.55+/-0.56, 14.545+/-1.59, 6.23+/-2.45, 12.56+/-1.84 and 16.45+/-0.39 degrees C respectively. With the exception of PK5, all compounds bind to DNA by intercalation as judged by effect of compounds on DNA mobility. Ethidium bromide displacement assay showed that all the compounds have significant affinity for calf thymus DNA (the drug concentration required to reduce the fluorescence of initially DNA-bound ethidium bromide by 50%, C(50), was 1.21-17.33 microM). The order of DNA-binding strength was PK4 > PK3 > PK7 > PK6 > PK5. In HL-60 promyelocytic leukaemia cells, oxa-polyamine and bisnaphthalimido treatment resulted in a decline in cell proliferation and viability. The assays performed suggested that apoptosis was not the principal cell death mechanism involved in oxa-polyamine cytotoxicity. In contrast, HL-60 cell death induced by the bisnaphthalimido series was characterized by early exposure of phosphatidylserine exclusive from membrane damage, elevated caspase-3 activity, increased DNA instability and, ultimately, DNA fragmentation. Thus the principal cytotoxic members of the bisnaphthalimido series appear to induce apoptosis.

Trichinellosis acquired in the United Kingdom.

An outbreak of trichinellosis that occurred in the United Kingdom is described. Members of four households consumed pork salami from northern Serbia, the Federal Republic of Yugoslavia. Eight cases of trichinellosis occurred. Clinical and laboratory features of the cases were typical with myalgia (7 cases), fever (6), headache (5), periorbital oedema (4), non-specific ST/T wave changes on electrocardiogram (3), Trichinella antibodies (6), eosinophilia (7) and raised serum creatine kinase (3). All recovered. Trichinella larvae were detected in the salami. During pre-travel counselling, travellers should be advised about possible risk from cured pork products which have been produced locally in Trichinella endemic areas.

Predictors of depression in female adolescents.

Depression during adolescence has been associated with a number of factors, including failure to individuate (Blos, 1968), insecure attachments (Armsden, McCauley, Greenberg, Burke, & Mitchell, 1990), negative parental representations, and object relations that lack self-other differentiation (Blatt, Wein, Chevron, & Quinlan, 1979). The present study examined factors associated with symptoms of depression in 59 nonclinical female adolescents. Specifically, the relationship between a number of theoretically related measures-separation-individuation, interpersonal concerns, self-critical concerns, attachment style, parental representations-and symptoms of depression was investigated. The model developed was able to explain the interrelationships of the variables involved in the psychological process of adolescence, and their demonstrated ability to predict symptoms of depression in normal female adolescents.

Cryptosporidium meleagridis from humans: molecular analysis and description of affected patients.

OBJECTIVES: To genetically characterize an unusual genotype of Cryptosporidium from the stools of humans with diarrhoea and to identify risk factors in the affected patients. METHODS: DNA was extracted from human faeces where Cryptosporidium oocysts were detected by light microscopy. Cryptosporidial gene fragments from six different loci were analysed by PCR alone, PCR/RFLP and by DNA sequencing. Oocysts were characterized by light and immunofluorescence microscopy and epidemiological data was collected from the affected patients. RESULTS: Analysis of the Cryptosporidium oocyst wall protein (COWP) gene amplified from > 2000 human faecal samples identified 19 patients all of which produced an unusual RFLP profile. Subsequent DNA sequence analysis of this and an additional four genetic loci (including 18S rRNA sequences) confirmed these as a homogeneous group which was genetically distinct from Cryptosporidium parvum. The isolates were identified as Cryptosporidium meleagridis since the gene sequences were identical to those from this species recovered from birds. Conventional microscopy showed oocysts indistinguishable from C. parvum and reacted strongly with two different commercially available anti-oocyst monoclonal antibodies. None of the patients showed risk factors unusual for cryptosporidiosis; however, ten of the cases occurred during the summer/autumn, six had a history of foreign travel, four were co-infected with Giardia, two were HIV positive, and six were without identifiable immunocompromising factors. CONCLUSIONS: This study further confirms that C. meleagridis, in addition to C. parvum, is involved in human disease. The study also highlights the lack of basic information on the host range of this genus of parasites, the complexity of the transmission routes involved in human cryptosporidiosis, and the value of molecular techniques in identify hitherto unrecognised differences in Cryptosporidium from human faeces.

Quercetin modifies reactive oxygen levels but exerts only partial protection against oxidative stress within HL-60 cells.

Quercetin may contribute to the protection afforded by fruit- and vegetable-rich diets against diseases for which excess production of reactive oxygen species (ROS) has been implicated as a causal or contributory factor. We examine the effect of short term (90 min) quercetin (1-100 microM) exposure on the progress of menadione induced oxidative stress within HL-60 cells. 2',7'-dichlorofluorescein and rhodamine-123 fluorescence, resulting from oxidation of the ROS-sensitive dyes dichlorodihydrofluorescein and dihydrorhodamine-123 respectively, were utilised as indicators of general ROS levels. Ethidium fluorescence, resulting from oxidation of dihydroethidium, was used as a potentially more specific indicator of O(2)(-). Exposure to quercetin alone induced a decrease in DCF and rhodamine fluorescence. Conversely, ethidium fluorescence was enhanced by treatment with >or=40 microM quercetin. Incubation with 1-100 microM quercetin reduced the extent of menadione-induced increase in DCF and rhodamine fluorescence but the menadione-induced increase in ethidium fluorescence was further elevated for cells treated with >or=25 microM quercetin. Exposure to >or=10 microM quercetin abrogated menadione-induced DNA single-strand breaks but, paradoxically, quercetin exacerbated membrane damage and failed to enhance the viability of menadione-challenged cells. In conclusion, quercetin exerts only site-specific protection against oxidative stress.

Alteration of culture regime modifies antioxidant defenses independent of intracellular reactive oxygen levels and resistance to severe oxidative stress within confluent Caco-2 "intestinal cells".

Reactive oxygen species are implicated in the development of gastrointestinal pathologies. Caco-2 monolayers are routinely used to study intestinal oxidative stress and its potential amelioration by pharmacological agents or dietary micronutrients. Little is known of the plasticity of Caco-2 antioxidant defenses with changes in culture conditions. We examined whether the frequency of culture media renewal alters the antioxidant-prooxidant status and integrity of Caco-2 monolayers. In comparison to monolayers subject to daily media renewal, increasing periods between media exchange resulted in varying degrees of suppression of catalase, glutathione reductase, and glutathione-S-transferase activity. No significant changes to superoxide dismutase activity, total glutathione, or intracellular ROS profiles were observed. Alkaline phosphatase activity, as a marker of differentiation, and mean monolayer cell population size were also unaffected. We suggest that Caco-2 antioxidant enzyme activities are differentially sensitive to changes in culture conditions. Studies employing this cell line for antioxidant-oxidative stress interactions will need to evaluate responses with respect to culture regime.

A simple, highly sensitive and improved method for the measurement of bleomycin-detectable iron: the 'catalytic iron index' and its value in the assessment of iron status in haemochromatosis.

In the presence of ferrous ions (Fe(2+)), the anti-tumour agent bleomycin will induce DNA degradation. Degradation of DNA into substances detectable by the thiobarbituric acid test has been used previously for the detection of iron in a form that is capable of catalysing the formation of the potentially harmful hydroxyl free radical. In the present paper, we describe the application of the ethidium-binding assay of DNA damage to the measurement of bleomycin-detectable iron, comparing its performance with the conventional method in the assessment of iron standard solutions and plasma samples from haemochromatosis patients. The ethidium-binding assay proved to be more responsive than the thiobarbituric acid test in the detection of DNA damage induced by very low concentrations of iron, but became saturated at higher iron concentrations. Agreement between the two versions of the assay in the identification of plasma samples containing bleomycin-detectable iron was good, but agreement on the actual concentrations of such iron in the positive samples was poor. This discrepancy is believed to be due to interference with the thiobarbituric acid assay by plasma. Consequently, it was not possible to obtain reliable estimates of free iron concentrations in plasma when using the conventional version of the bleomycin assay. We have devised a parameter of iron status called the catalytic iron index. For healthy, non-haemochromatotic individuals, the mean value of this parameter was found to be 0.81 (range 0.78-0.84; n=20). Elevated values were observed in some plasma samples from haemochromatosis patients, but these showed no correlation with serum ferritin levels. In contrast, correlations were seen with both serum iron and transferrin saturation levels, but only when these were above the normal range.

Oligoribonucleotide-based gene-specific transcription inhibitors that target the open complex.

We have demonstrated that oligoribonucleotides that lack a 3'-OH group and cannot be extended by RNA polymerase can hybridize to the single-stranded DNA formed inside the transcription initiation bubble (or open complex) and inhibit transcription. Using the lacUV5/Escherichia coli RNA polymerase or trpEDCBA/E. coli RNA polymerase transcription system as a model, we have found that effective inhibitors are five nucleotides in length and must be complementary to the DNA template strand in the region from -5 to +2 about the transcription start site (designated +1). We have used the DNA cleavage activity of 1,10-phenanthroline-copper to confirm that the mechanism of inhibition is via oligoribonucleotide hybridization to the open complex and have used this cleavage chemistry to demonstrate that these oligonucleotide inhibitors hybridize in an antiparallel orientation to their DNA target. Systematic modification of the parent phosphodiester oligoribonucleotide pentamer revealed that the phosphorothioate backbone-containing analogs have increased open complex binding affinity and are more effective transcription inhibitors than their phosphodiester counterparts.

Artificial nucleases.

The oxidation of DNA and RNA provides a facile approach for investigating the interaction of nucleic acids with proteins and oligonucleotides. In this article, we have outlined our understanding of the mechanism of DNA scission by 1,10-phenanthroline-copper(I) in the presence of hydrogen peroxide. We also discuss results obtained by using 1,10-phenanthroline-oligonucleotide conjugates in probing the size of the transcriptionally active open complex. Finally, we outline an effective method for converting DNA-binding proteins into site-specific modification agents by using 1,10-phenanthroline-copper(I).