Traumatic brain injury modifies synaptic plasticity in newly-generated granule cells of the adult hippocampus.

The hippocampus is vulnerable to traumatic brain injury (TBI), and hippocampal damage is associated with cognitive deficits that are often the hallmark of TBI. Recent studies have found that TBI induces enhanced neurogenesis in the dentate gyrus (DG) of the hippocampus, and this cellular response is related to innate cognitive recovery. However, cellular mechanisms of the role of DG neurogenesis in post-TBI recovery remain unclear. This study investigated changes in long-term potentiation (LTP) within the DG in relation to TBI-induced neurogenesis. Adult male rats received a moderate TBI or sham injury and were sacrificed for brain slice recordings at 30 or 60 days post-injury. Recordings were taken from the medial perforant path input to DG granule cells in the presence or absence of the GABAergic antagonist picrotoxin, reflecting activity of either all DG granule cells or predominately newborn granule cells, respectively. Measurements of LTP observed in the total granule cell population (with picrotoxin) showed a prolonged impairment which worsened between 30 and 60 days post-TBI. Under conditions which predominantly reflected the LTP elicited in newly born granule cells (no picrotoxin), a strikingly different pattern of post-TBI changes was observed, with a time-dependent cycle of functional impairment and recovery. At 30 days after injury this cell population showed little or no LTP, but by 60 days the capacity for LTP of the newly born granule cells was no different from that of sham controls. The time-frame of LTP improvements in the newborn cell population, comparable to that of behavioral recovery reported previously, suggests the unique functional properties of newborn granule cells enable them to contribute to restorative change following brain injury.

Inter-professional prescribing masterclass for medical students and non-medical prescribing students (nurses and pharmacists): a pilot study.

BACKGROUND AND AIMS: Prescribing errors cause significant patient morbidity and mortality. Current legislation allows prescribing by different health professions. Inter-professional collaboration and learning may result in safer prescribing practice. This study aimed to develop, pilot and test the feasibility of a simulated inter-professional prescribing masterclass for non-medical prescribing students, medical students and pharmacists. METHODS AND RESULTS: A three-scenario, simulated patient session was designed and implemented by an expert panel. Medical students, non-medical prescribing students and pharmacists worked together to formulate and implement evidence-based prescriptions. The Readiness for Inter-professional Learning Score (RIPLS) and a self-efficacy score were administered to the students and the Trust in Physician Score to the simulated patients. Overall, the RIPLS and self-efficacy scores increased. Pharmacists showed the highest rating in the Trust in Physician score. Post masterclass group discussions suggested that the intervention was viewed as a positive educational experience. CONCLUSION: An inter-professional prescribing masterclass is feasible and acceptable to students. It increases self-efficacy, readiness for inter-professional learning and allows students to learn from, about and with each other. A larger study is warranted and the use of feedback from simulated patients explored further.

Next-generation sequencing for typing and detection of resistance genes: performance of a new commercial method during an outbreak of extended-spectrum-beta-lactamase-producing Escherichia coli.

Next-generation sequencing (NGS) has the potential to provide typing results and detect resistance genes in a single assay, thus guiding timely treatment decisions and allowing rapid tracking of transmission of resistant clones. We evaluated the performance of a new NGS assay (Hospital Acquired Infection BioDetection System; Pathogenica) during an outbreak of sequence type 131 (ST131) Escherichia coli infections in a nursing home in The Netherlands. The assay was performed on 56 extended-spectrum-beta-lactamase (ESBL) E. coli isolates collected during 2 prevalence surveys (March and May 2013). Typing results were compared to those of amplified fragment length polymorphism (AFLP), whereby we visually assessed the agreement of the BioDetection phylogenetic tree with clusters defined by AFLP. A microarray was considered the gold standard for detection of resistance genes. AFLP identified a large cluster of 31 indistinguishable isolates on adjacent departments, indicating clonal spread. The BioDetection phylogenetic tree showed that all isolates of this outbreak cluster were strongly related, while the further arrangement of the tree also largely agreed with other clusters defined by AFLP. The BioDetection assay detected ESBL genes in all but 1 isolate (sensitivity, 98%) but was unable to discriminate between ESBL and non-ESBL TEM and SHV beta-lactamases or to specify CTX-M genes by group. The performance of the hospital-acquired infection (HAI) BioDetection System for typing of E. coli isolates compared well with the results of AFLP. Its performance with larger collections from different locations, and for typing of other species, was not evaluated and needs further study.

Characterization of a human monoclonal antibody against Shiga toxin 2 expressed in Chinese hamster ovary cells.

Shiga toxin-producing Escherichia coli infections can often lead to the development of hemolytic-uremic syndrome (HUS) in a small percentage of infected humans. Patients with HUS receive only supportive treatment as the benefit of antibiotic therapy remains uncertain. We have previously reported the generation and preclinical evaluation of neutralizing human monoclonal antibodies (HuMAbs) against the Shiga toxins (Stx). In this paper, we describe the expression in Chinese hamster ovary (CHO) cells of 5C12 HuMAb, which is directed against the A subunit of Stx2. The cDNAs of the light and heavy chain immunoglobulin (Ig) variable regions of 5C12 HuMAb were isolated and cloned into an expression vector containing human IgG1 constant regions. The vector was transfected into CHO cells, and transfectants secreting Stx2-specific antibody were screened by an Stx2-specific enzyme-linked immunosorbent assay. The CHO-produced recombinant 5C12 (r5C12) showed similar specificity and binding affinity to Stx2 as the parent hybridoma-produced 5C12. More significantly, the r5C12 displayed the same neutralizing activity as the parent 5C12 in vitro and in vivo. In the mouse toxicity model, both antibodies significantly and equally prolonged survival at a dose of 0.312 microg/mouse. The data showed that since r5C12, produced in CHO cells, was equally effective as the parent 5C12, it is our choice candidate as a potential prophylactic or therapeutic agent against hemolytic-uremic syndrome.

Pre-prandial vs. post-prandial capillary glucose measurements as targets for repaglinide dose titration in people with diet-treated or metformin-treated Type 2 diabetes: a randomized controlled clinical trial.

OBJECTIVE: Repaglinide is an oral anti-diabetic agent that has a short duration of action, and is suitable for preventing post-prandial rises in glucose levels. Targeting post-prandial glucose levels may lead to lower HbA(1c) levels and rates of hypoglycaemia than targeting pre-prandial glucose levels. RESEARCH DESIGN AND METHODS: In 42 centres, 193 drug-naive (n = 122) or metformin-treated (n = 71) individuals with Type 2 diabetes were randomly allocated to a 40-day period of repaglinide dose-titration (starting at 0.5 mg three times daily) based on either self-measured pre-prandial or post-prandial glucose levels. They were followed for a further 12 weeks and HbA(1c) and hypoglycaemia rates were recorded. RESULTS: Repaglinide reduced HbA(1c) by 1.25 and 1.07% in the post-prandial and pre-prandial groups, respectively (P for difference = 0.6), and achieved target glucose levels in 80.7 and 66.7%, respectively (P = 0.16). The effect of titration strategy differed by baseline drug therapy, and was more effective in the metformin-treated individuals who experienced a HbA(1c) fall of 0.6 vs. 1.10% with pre-prandial vs. post-prandial titration (P for metformin-allocated group interaction = 0.043). The rate of hypoglycaemia did not differ by group. CONCLUSIONS: In drug-naive individuals with Type 2 diabetes, similar HbA(1c) levels are achieved with repaglinide when dosing is adjusted according to either post-prandial or pre-prandial levels. Conversely, in metformin-treated individuals, repaglinide dosing according to post-prandial levels may lead to better glycaemic control than dosing according to pre-prandial levels.

Localization of the binding site for the oligosaccharide moiety of Gb3 on verotoxin 1 using NMR residual dipolar coupling measurements.

By use of NMR residual dipolar coupling measurements in a dilute liquid-crystalline solvent, the solution structure has been determined of the complex between the oligosaccharide moiety of globotriaosylceramide (Gb(3)-OS) and the B-subunit homopentamer of verotoxin 1 (VTB). The dipolar coupling data indicate that Gb(3)-OS binds in a single binding site per monomer, which is identical to one of three sites inferred from the X-ray structure of the same complex. We find no evidence within experimental error for occupancy at either of the two additional binding sites observed per monomer in the crystal structure.

Patterns of improvement after methadone treatment: 1 year follow-up results from the National Treatment Outcome Research Study.

One year outcomes for substance use behaviours, health and criminal behaviour, and variation in treatment response, are reported for patients recruited to methadone maintenance and methadone reduction treatment programmes as part of NTORS. Significant reductions in the use of all illicit target drugs were found at follow-up for patients recruited to the methadone maintenance and methadone reduction modalities. Because of similarities in the treatments received by clients in the two modalities we caution against interpreting these findings as showing that methadone maintenance and reduction treatments lead to similar outcomes. At this stage, it is suggested that these outcomes be regarded as reflective of exposure to some general methadone substitution treatment. Further investigation of the outcomes for the two modalities will be conducted. Cluster analyses were used to classify patients according to level of improvement in drug use. Four groups were identified. Two groups (59% of cases) showed substantial reductions in their illicit drug use and criminality as well as reduced physical and psychological symptoms. Twenty two percent of cases showed poor outcomes across a range of measures. Results for alcohol consumption were less satisfactory for patients in all groups. A majority of patients achieved widespread improvements across a range of outcome measures after treatment in existing methadone treatment services. These changes represent important clinical benefits to the individual clients, to their families and to society.

Patterns of drinking and drinking outcomes among drug misusers. 1-year follow-up results.

This article investigates patterns of drinking and drinking outcomes among 753 drug-misusing clients recruited to the National Treatment Outcome Research Study (NTORS). More than one third of those who were drinking at intake reported problematic or highly problematic patterns of alcohol consumption. About one third of the sample were abstinent from alcohol at intake and at follow-up. Some improvements in drinking behavior were found at 1-year follow-up, especially among the heaviest and most problematic drinkers. Improvements were specifically related to patterns of preintake drinking behavior. The majority of clients made little change to their pattern of pretreatment drinking behavior and the continued heavy drinking of many drinkers at follow-up is a disappointing finding. Drinking problems have been given insufficient attention in the treatment of illicit substance misuse problems, and efforts should be made to develop and strengthen the assessment and treatment of drinking problems among drug misusers.

Escherichia coli O157:H7 strains that express Shiga toxin (Stx) 2 alone are more neurotropic for gnotobiotic piglets than are isotypes producing only Stx1 or both Stx1 and Stx2.

Infection with Escherichia coli O157:H7 can lead to hemolytic uremic syndrome (HUS) in some children. Epidemiologic data suggest that Shiga toxin (Stx) 2-producing strains are more frequently associated with HUS than are Stx1-producing strains. Less clear is whether strains that express Stx2 alone are more frequently associated with HUS than strains that express Stx1 and Stx2. Isogenic mutants 933stx1- and 933stx2- were produced from strain 933 (Stx1 and Stx2 producer), and 86-24stx2- was produced from strain 86-24 (Stx2 producer). Neurologic lesions or symptoms developed in 18 (90%) of 20 gnotobiotic piglets orally infected with strain 86-24, in 15 (85%) of 18 infected with mutant 933stx1-, in 9 (31%) of 29 infected with strain 933, in 0 of 5 infected with mutant 86-24stx2-, and in 0 of 6 infected with mutant 933stx2-. It was concluded that strains expressing Stx2 alone are more neurotropic for piglets when fed orally than are those strains expressing Stx1 and 2, whereas Stx1-producing strains induce only diarrhea. It is also conceivable that strains that produce Stx2 may constitute a significant predictive risk factor for HUS in humans.

Reductions in acquisitive crime and drug use after treatment of addiction problems: 1-year follow-up outcomes.

The relationship between acquisitive crime and drug misuse problems was studied among 753 clients recruited to the National Treatment Outcome Research Study (NTORS). More than 17000 offences were reported during the 90-day period prior to treatment. Half of the clients committed no acquisitive crimes during this period whereas 10% committed 76% of the crimes. At 1-year follow-up, the number of crimes was reduced to one third of intake levels, and criminal involvement was reduced by about half. Reductions in regular heroin use were strongly associated with reductions in crime. The reduction in crime following treatment is of great importance and provides immediate benefit to society through the reduced economic costs of crime.

Psychiatric symptoms among clients seeking treatment for drug dependence. Intake data from the National Treatment Outcome Research Study.

BACKGROUND: Many people with substance use disorders are vulnerable to other psychiatric disorders and present to addiction treatment services with comorbid psychiatric symptoms. AIMS: To describe the prevalence of recent psychiatric treatment and current psychiatric symptoms and explore links between substance misuse, personal/social functioning and symptom severity. METHOD: Subjects were 1075 adults recruited to the National Treatment Outcome Research Study (NTORS), of whom 90% were opiate-dependent. Psychiatric symptoms at intake were recorded using sub-scales from the Brief Symptom Inventory. RESULTS: Recent psychiatric treatment was reported by one in five subjects. Psychiatric symptom levels were high and females had elevated scores on all scales. Symptoms were elevated among opiate users who were also frequent users of benzodiazepines, alcohol and, in particular, stimulants. Gender, physical health, drug dependence and personal relationship problems were more powerful predictors of psychiatric symptoms than substance use. CONCLUSIONS: Addictions service providers should be vigilant to psychiatric problems among their clients at intake to treatment. Psychiatric symptoms are more closely linked to polydrug use than to opiate use in this population.

Syntheses and immunologic properties of Escherichia coli O157 O-specific polysaccharide and Shiga toxin 1 B subunit conjugates in mice.

Escherichia coli O157 is the major cause of diarrhea-associated hemolytic uremic syndrome (HUS). Strains causing HUS contain either Shiga toxin 1 (Stx1) or Stx2, or both. In adult volunteers, conjugate vaccines of detoxified lipopolysaccharide (LPS) elicited bactericidal antibodies to E. coli O157. Here, the detoxified LPS was conjugated with improved schemes to the nontoxic B subunit of Stx1. Mice injected with these bivalent conjugates elicited both bactericidal antibodies to E. coli O157 and neutralization antibodies to Stx1.

Characterization of single-nucleotide polymorphisms in coding regions of human genes.

A major goal in human genetics is to understand the role of common genetic variants in susceptibility to common diseases. This will require characterizing the nature of gene variation in human populations, assembling an extensive catalogue of single-nucleotide polymorphisms (SNPs) in candidate genes and performing association studies for particular diseases. At present, our knowledge of human gene variation remains rudimentary. Here we describe a systematic survey of SNPs in the coding regions of human genes. We identified SNPs in 106 genes relevant to cardiovascular disease, endocrinology and neuropsychiatry by screening an average of 114 independent alleles using 2 independent screening methods. To ensure high accuracy, all reported SNPs were confirmed by DNA sequencing. We identified 560 SNPs, including 392 coding-region SNPs (cSNPs) divided roughly equally between those causing synonymous and non-synonymous changes. We observed different rates of polymorphism among classes of sites within genes (non-coding, degenerate and non-degenerate) as well as between genes. The cSNPs most likely to influence disease, those that alter the amino acid sequence of the encoded protein, are found at a lower rate and with lower allele frequencies than silent substitutions. This likely reflects selection acting against deleterious alleles during human evolution. The lower allele frequency of missense cSNPs has implications for the compilation of a comprehensive catalogue, as well as for the subsequent application to disease association.

Influence of RecA on in vivo virulence and Shiga toxin 2 production in Escherichia coli pathogens.

The enterohemorrhagic Escherichia coli (EHEC) O157:H7 strains 933 and 86-24 as well as the uropathogenic E. coli (UPEC) strain 536 were compared with their isogenic rec A mutants and rec A trans -complemented strains in intravenous lethality and lung toxicity assays in mice. While the wild-type EHEC strains were fully virulent, the virulence of the rec A mutants was strongly reduced. Complementation of the EHEC rec A mutants with the cloned E. coli recA gene restored their virulence capacity. The stx2EHEC mutant TUV86-2 as well as its isogenic rec A mutant were completely avirulent in both assays. In contrast, RecA had no influence on the virulence of UPEC strain 536. We conclude that the lethality observed with EHEC is presumably mainly due to Shiga toxin, which is severely down-regulated in the rec A mutants as a result of lacking spontaneous phage induction. Therefore, the EHEC rec A+strains 933 and 86-24 were compared for their Shiga toxin 2 (Stx2) production with the respective rec A-counterparts. The rec A mutants of the EHEC strains were significantly reduced in toxin synthesis and were devoid of Stx2 specific phage production. Complementation of the EHEC rec A mutants with the cloned rec A gene enabled the rec A mutants to restore toxin and phage production. These results suggest that the higher level of Stx2 synthesis in the EHEC strains is the result of a higher level of spontaneous Stx2 specific phage induction, which is controlled by RecA.

Antibody-based protection of gnotobiotic piglets infected with Escherichia coli O157:H7 against systemic complications associated with Shiga toxin 2.

Hemolytic-uremic syndrome (HUS) is a serious disease in children, attributable in the majority of cases to infection with Shiga toxin (Stx)-producing Escherichia coli. Using gnotobiotic piglets orally infected with E. coli O157:H7, which develop Stx-related cerebellar lesions and fatal neurological symptoms, we show that administration of Stx2-specific antiserum well after challenge protected, in a dose-response fashion, against these symptoms for at least 24 h after bacterial challenge. Twenty-six of 30 piglets given Stx2 antiserum survived the challenge, compared to only 4 of 16 animals given control serum or saline. Given our observations in piglets, Stx antibody of human origin may likewise prevent HUS in children.

Treatment retention and 1 year outcomes for residential programmes in England.

This paper reports changes in substance use behaviours at 1-year follow-up, and investigates the relationship between time in treatment and observed outcomes. A total of 408 clients were interviewed at intake to 23 residential treatment programmes, and 286 (70%) of these were interviewed at 1 year. Substantial improvements were found in terms of abstinence from opiates, psychostimulants and benzodiazepines. At 1 year, half of the clients were abstinent from heroin. Reductions in injecting, sharing injecting equipment, heavy drinking and criminal behaviour were found. Critical treatment thresholds were identified using multiple logistic regression analyses. Longer stays in treatment were predictive of better 1 year outcomes.

Pretreatment with intralesional hyaluronidase prior to excision of dermatofibrosarcoma protuberans.

Dermatofibrosarcoma protuberans (DFSPs) are an uncommon malignancy that commonly recur, but rarely metastasize. The origin of DFSPs is controversial; however, they stain with the progenitor marker CD34. DFSPs usually show increased stromal mucin, mainly hyaluronic acid (HA). HA increases cellular proliferation, delays differentiation and increases cellular motion. We evaluated the pretreatment of DFSPs with intralesional injections of hyaluronidase (HD) prior to the surgical excision. Five of nine cases of DFSPs were pretreated with HD. In HD-pretreated cases the margins for excision of the residual tumor were reduced. HD pretreatment also decreased CD34 staining and increased polarizable collagen in the residual tumor.

Solution structure of the complex between the B-subunit homopentamer of verotoxin VT-1 from Escherichia coli and the trisaccharide moiety of globotriaosylceramide.

We report the solution structure of the carbohydrate-binding B subunit of verotoxin VT-1 (VTB) from enterohemorrhagic Escherichia coli in association with the trisaccharide Galalpha1-4Galbeta1-4Glcbeta1-O-trimethylsilylethyl , determined by use of stable isotope-assisted NMR techniques. In contrast to the crystal structure of the complex which predicts three binding sites per monomer, only one of these sites is observed with substantial occupancy by the trisaccharide in solution.

Construction and characterization of an isogenic slt-ii deletion mutant of enterohemorrhagic Escherichia coli.

Enterohemorrhagic Escherichia coli (EHEC) produces Shiga-like toxins (SLT), potent protein synthesis inhibitors. To further dissect the role of SLT-II in the course of disease, we have constructed E. coli TUV86-2, an isogenic SLT-II-negative mutant of EHEC strain 86-24. The slt-ii gene was inactivated by suicide vector mutagenesis. We also isolated derivatives of strain 86-24 that were cured of the phage carrying the toxin genes.