Pathologic polyglutamine aggregation begins with a self-poisoning polymer crystal.

A long-standing goal of amyloid research has been to characterize the structural basis of the rate-determining nucleating event. However, the ephemeral nature of nucleation has made this goal unachievable with existing biochemistry, structural biology, and computational approaches. Here, we addressed that limitation for polyglutamine (polyQ), a polypeptide sequence that causes Huntington's and other amyloid-associated neurodegenerative diseases when its length exceeds a characteristic threshold. To identify essential features of the polyQ amyloid nucleus, we used a direct intracellular reporter of self-association to quantify frequencies of amyloid appearance as a function of concentration, conformational templates, and rational polyQ sequence permutations. We found that nucleation of pathologically expanded polyQ involves segments of three glutamine (Q) residues at every other position. We demonstrate using molecular simulations that this pattern encodes a four-stranded steric zipper with interdigitated Q side chains. Once formed, the zipper poisoned its own growth by engaging naive polypeptides on orthogonal faces, in a fashion characteristic of polymer crystals with intramolecular nuclei. We further show that self-poisoning can be exploited to block amyloid formation, by genetically oligomerizing polyQ prior to nucleation. By uncovering the physical nature of the rate-limiting event for polyQ aggregation in cells, our findings elucidate the molecular etiology of polyQ diseases.

Diseases that typically occur later in life, such as Alzheimer's, are often caused by specific proteins clumping together into structures known as amyloids. Once the process starts, amyloids will continue to form, leading to worse symptoms that cannot be cured. The best way to treat these diseases is therefore to stop amyloids from arising in the first place. Amyloids initially develop by proteins coming together to create an unstable structure referred to as the nucleus. The instability of the nucleus means it cannot be observed directly, making it hard to study this nucleation process. To overcome this, Kandola, Venkatesan et al. investigated the simplest protein known to form an amyloid ­ polyglutamine, which is made up of a chain of repeating building blocks known as amino acids. Polyglutamine forms only one type of amyloid which is associated with nine neurodegenerative diseases, including Huntington's disease. However, it only does this when its chain of amino acids exceeds a certain length, suggesting that a specific structure may be required for nucleation to begin. Kandola, Venkatesan et al. made alternative versions of the polyglutamine protein which each contained slightly different sequences of amino acids that will alter the way the protein folds. They then tested how well these different variants could form amyloids in yeast cells. This revealed that in order to join together into a nucleus, polyglutamine needs to be able to fold into a zipper shape made up of four interlocking strands. The length of the protein required to form this shape is also the same length that causes the amyloid associated with neurodegenerative diseases. Kandola, Venkatesan et al. also found that polyglutamine tends to bind to nuclei that have already formed in a way that hinders their growth. This 'self-poisoning' affect could potentially be exploited as a way to pre-emptively stop amyloids from initially arising. These findings have uncovered a potential therapeutic strategy for blocking amyloid formation that could eventually benefit people with or at risk of developing neurodegenerative diseases linked to polyglutamine. Additionally, this approach provides a blueprint for understanding how other proteins undergo amyloid nucleation, including those responsible for Alzheimer's, Parkinson's, and other diseases.

Student Pharmacist-Led Basic Life Support Training for High School Students Improves Knowledge and Skill Achievement.

Introduction: The performance of bystander cardiopulmonary resuscitation (CPR) improves survival among cardiac arrest victims. Near-peer teaching of Basic Life Support (BLS) may be an effective way to deliver resuscitation education. This article aims to assess the effectiveness of a student pharmacist-led American Heart Association (AHA) BLS course on high school students' knowledge and skill achievement. Methods: Student pharmacists were trained as AHA instructors and delivered BLS certification courses to high school students. Participants completed pre- and post-assessments adapted from the course learning objectives. Skills performance was evaluated using the AHA's standardized forms. Participants completed questions regarding their perceptions of the pharmacist's role in BLS and confidence in pursuing a career in healthcare. Results: There were 321 participants with the majority in 11th or 12th grade (86.6%) and attending public school (99.1%). After completing the training, the mean percentage of correct assessment responses increased from 41.2% to 89% (p <0.0001). All participants correctly performed BLS skills. Most participants strongly agreed or agreed that the course changed their perspective of the pharmacist's role during BLS (74.8%) and increased their confidence in their decision to pursue future careers in healthcare (61.7%). Conclusion: Student pharmacist-led BLS training, using near-peer delivery, improves high school students' knowledge and skill achievement. This strategy promotes high school students' positive perceptions regarding pharmacists and their role in BLS, as well as their confidence in pursuing careers in healthcare.

PICH regulates the abundance and localization of SUMOylated proteins on mitotic chromosomes.

Proper chromosome segregation is essential for faithful cell division and if not maintained results in defective cell function caused by the abnormal distribution of genetic information. Polo-like kinase 1-interacting checkpoint helicase (PICH) is a DNA translocase essential for chromosome bridge resolution during mitosis. Its function in resolving chromosome bridges requires both DNA translocase activity and ability to bind chromosomal proteins modified by the small ubiquitin-like modifier (SUMO). However, it is unclear how these activities cooperate to resolve chromosome bridges. Here, we show that PICH specifically disperses SUMO2/3 foci on mitotic chromosomes. This PICH function is apparent toward SUMOylated topoisomerase IIα (TopoIIα) after inhibition of TopoIIα by ICRF-193. Conditional depletion of PICH using the auxin-inducible degron (AID) system resulted in the retention of SUMO2/3-modified chromosomal proteins, including TopoIIα, indicating that PICH functions to reduce the association of these proteins with chromosomes. Replacement of PICH with its translocase-deficient mutants led to increased SUMO2/3 foci on chromosomes, suggesting that the reduction of SUMO2/3 foci requires the remodeling activity of PICH. In vitro assays showed that PICH specifically attenuates SUMOylated TopoIIα activity using its SUMO-binding ability. Taking the results together, we propose a novel function of PICH in remodeling SUMOylated proteins to ensure faithful chromosome segregation.

Evaluation of an individualized vs non-specific standardized patient activity in improving communication skills amongst pharmacy students.

BACKGROUND AND PURPOSE: To evaluate the impact of two standardized patient (SP) approaches on student communication skills and self-perceived confidence. EDUCATIONAL ACTIVITY AND SETTING: Second professional year pharmacy students were assessed on overall and area of improvement (AOI) communication skills and self-perceived confidence. Students were invited to participate in a SP intervention activity, following a fall and before a spring class assessment, where they were randomized to an individualized or non-specific SP group. Communication skills were assessed using a four component 64-point rubric; confidence was assessed using an 11-question Likert-type survey. FINDINGS: Nineteen students participated in the SP intervention activity; 11 students comprised the individualized group and eight students comprised the non-specific group. Students in the individualized group demonstrated a significantly higher median change in AOI communication scores compared to the non-specific group [five points vs. three points, respectively (p = 0.033)]. There was a significantly higher median change in overall communication skills in the individualized compared to the non-specific group of 12 points and 6.5 points, respectively (p = 0.017). Student self-perceived confidence in the individualized and non-specific group showed similar improvements overall from pre- to post assessment [seven points vs. eight points, respectively (p = 0.62)]. SUMMARY: The use of SPs in pharmacy curricula can improve student communication skills. However, tailoring the activity to students' needs significantly improves their communication AOI and overall communication skill scores more than a non-specific activity. Student self-perceived confidence improved similarly in both groups, although no statistically significant differences were found between groups.

Two-week wait symptoms are prevalent in screened patients with a positive faecal occult blood test but do not predict cancer.

AIM: Lower gastrointestinal (LGI) symptoms are prevalent in patients screened for bowel cancer yet do not predict a finding of cancer. This study evaluates symptoms in patients with these characteristics against the 2-week wait (2ww) criteria to determine whether they predicted cancer in these patients. METHOD: A prospective cohort study was performed. Patients with a positive faecal occult blood (FOB) test attending our unit over a 7-month period were included. Data on symptom prevalence, frequency and duration were collected and assessed against the 2ww criteria. Associations between symptom prevalence and patient outcome were investigated using the χ(2) test. RESULTS: Three hundred and ninety-seven patients were included and 37 (9%) were found to have colorectal cancer (CRC). The prevalence of undefined LGI symptoms was 71% and appeared comparable between those with and without CRC (65 vs 72%, P = 0.385). 2ww symptoms were reported in 147 (37%), with 2ww change in bowel habit in 10% and 2ww rectal bleeding in 31%. 2ww symptom prevalence was similar in those with and without cancer (38 vs 37%, P = 0.915). No significant differences in overall 2ww prevalence or prevalence of individual 2ww symptoms were demonstrated between those with a normal colonoscopy or one showing cancer, polyps or other pathology. Twenty nine per cent of patients with 2ww symptoms had reported these to their GP. CONCLUSION: Undefined LGI symptoms are prevalent in FOB-positive patients but do not predict CRC. 2ww symptoms are also highly prevalent, yet similarly fail to predict cancer. Further efforts to increase public awareness of cancer symptoms are required, whilst false reassurance from a negative FOB result should be discouraged.

Synthesis and biological evaluation of indenoisoquinolines that inhibit both tyrosyl-DNA phosphodiesterase I (Tdp1) and topoisomerase I (Top1).

Tyrosyl-DNA phosphodiesterase I (Tdp1) plays a key role in the repair of damaged DNA resulting from the topoisomerase I (Top1) inhibitor camptothecin and a variety of other DNA-damaging anticancer agents. This report documents the design, synthesis, and evaluation of new indenoisoquinolines that are dual inhibitors of both Tdp1 and Top1. Enzyme inhibitory data and cytotoxicity data from human cancer cell cultures were used to establish structure-activity relationships. The potencies of the indenoisoquinolines against Tdp1 ranged from 5 µM to 111 µM, which places the more active compounds among the most potent known inhibitors of this target. The cytotoxicity mean graph midpoints ranged from 0.02 to 2.34 µM. Dual Tdp1-Top1 inhibitors are of interest because the Top1 and Tdp1 inhibitory activities could theoretically work synergistically to create more effective anticancer agents.

Slipping through the bowel cancer screening programme.

AIM: Colorectal cancer is common and a leading cause of cancer death. Faecal occult blood screening has been shown to reduce mortality. The aim of this study was to identify patients in Gloucestershire with a new diagnosis of colorectal cancer who had previously been screened via the Bowel Cancer Screening Programme (BCSP). METHOD: Between 2006 and 2009, 1030 patients were diagnosed with colorectal cancer. Of these 237 (23%) had been invited to be screened via the BCSP. Their clinical notes were analysed. RESULTS: Fifty-seven (24%) of the 237 patients had previously had a negative faecal occult blood result. Thirty-three (14%) had their cancer discovered as part of the BCSP. Seventy (30%) had already been diagnosed with colorectal cancer prior to invitation, 62 (26%) did not respond to the invitation, nine (4%) were registered outside Gloucestershire and had therefore not been invited, and three (3%) had died before the invitation. Of the 57 patients with a negative faecal occult blood test, 47 (83%) had colorectal cancer staged Dukes B or C, and 34 (60%) had a rectal or sigmoid cancer. CONCLUSION: Patients will present with colorectal cancer despite having been invited to participate in the BCSP, with many having received a negative faecal occult blood test. This could be considered a high false negative rate.

Use of risedronate to prevent bone loss following a single course of glucocorticoids: findings from a proof-of-concept study in inflammatory bowel disease.

SUMMARY: We performed a randomised controlled trial (RCT) to determine whether risedronate 35 mg once weekly prevents bone loss following an 8-week reducing course of prednisolone given for an exacerbation of inflammatory bowel disease (IBD). The greatest change in bone mineral density (BMD) was at Ward's triangle (WT), which fell by 2.2% in the placebo group, compared with a reduction of 0.8% in the risedronate group. INTRODUCTION: Whether bisphosphonates can prevent bone loss associated with intermittent glucocorticoid (GC) therapy is unknown, reflecting the difficulty in performing RCTs in this context. METHOD: To explore the feasibility of RCTs to examine this question, lumbar spine (LS; L2-4) and hip dual X-ray absorptiometry (DXA) scans were performed in 78 patients commencing a GC therapy course for a relapse of IBD. They were then randomised to receive placebo or risedronate 35 mg weekly for 8 weeks, after which the DXA scan was repeated. RESULTS: For LS BMD, there was no change in the placebo group (0.1 +/- 0.4, p = 0.9), but there was an increase after risedronate (0.8 +/- 0.4, p = 0.04; mean% +/- SEM by paired Student's t test). There were small decreases in both groups at the total hip (-0.5 +/- 0.3, p = 0.04; -0.5 +/- 0.3, p < 0.05, placebo and risedronate, respectively). At WT, BMD fell after placebo (-2.2 +/- 0.5, p = 0.001) but not risedronate (-0.8 +/- 0.5, p = 0.09; p = 0.05 for between-group comparison). CONCLUSION: RCTs can be used to examine whether bisphosphonates prevent bone loss associated with intermittent GC therapy, providing metabolically active sites such as WT are employed as the primary outcome.

T-cell receptor repertoire in pyoderma gangrenosum: evidence for clonal expansions and trafficking.

BACKGROUND: The cause of pyoderma gangrenosum (PG) is unknown, but it is likely to be an immune-mediated disease because it is often associated with conditions such as inflammatory bowel disease and rheumatoid arthritis. T cells play an important role in these conditions and have been implicated in the pathogenesis of other skin diseases such as psoriasis. OBJECTIVES: We examined the T-cell receptor repertoire in PG in order to test the hypothesis that if the T cells were responding to antigen, there would be expanded T-cell clones in the skin and the circulation of these patients. PATIENTS AND METHODS: We studied five patients with PG and examined the T-cell receptor repertoire in cells taken from the peripheral blood and from biopsies of the ulcers, using complementarity determining region 3 spectratyping. RESULTS: We were able to demonstrate expanded clones in the peripheral blood lymphocyte population of each patient. Clonal expansions within the skin were found in four of the five patients. Most significantly, expanded clones that were shared between the blood and the skin were revealed in four of the five patients. CONCLUSIONS: These findings imply that T cells play an integral role in the development of PG and suggest that T cells are trafficking to the skin under the influence of an antigenic stimulus.

Infliximab for the treatment of pyoderma gangrenosum: a randomised, double blind, placebo controlled trial.

BACKGROUND: Pyoderma gangrenosum (PG) is a chronic ulcerating skin condition that often occurs in association with inflammatory bowel disease. There have been a number of reports of PG responding to infliximab, a monoclonal antibody against tumour necrosis factor alpha. AIM: In the first randomised placebo controlled trial of any drug for the treatment of PG, we have studied the role of infliximab in this disorder. SUBJECTS: Patients 18 years of age or older with a clinical diagnosis of PG were invited to take part. METHODS: Patients were randomised to receive an infusion of infliximab at 5 mg/kg or placebo at week 0. Patients were then assessed at week 2 and non-responders were offered open labelled infliximab. The primary end point was clinical improvement at week 2, with secondary end points being remission and improvement at week 6. RESULTS: Thirty patients were entered into the study. After randomisation, 13 patients received infliximab and 17 patients received placebo. At week 2, significantly more patients in the infliximab group had improved (46% (6/13)) compared with the placebo group (6% (1/17); p = 0.025). Overall, 29 patients received infliximab with 69% (20/29) demonstrating a beneficial clinical response. Remission rate at week 6 was 21% (6/29). There was no response in 31% (9/29) of patients. CONCLUSIONS: This study has demonstrated that infliximab at a dose of 5 mg/kg is superior to placebo in the treatment of PG. Open label treatment with infliximab also produced promising results. Infliximab treatment should be considered in patients with PG.

Churg-Strauss syndrome and granulomatous cholangiopathy.

Churg-Strauss syndrome is a vasculitis that is known to have gastroenterological manifestations. There have been no reports associating it with liver disease. We present a case of this syndrome in a man who was found to develop a co-existent cholangiopathy. It is likely that this represents an associated granulomatous reaction consequent to a hepatic manifestation of the vasculitis.