Dietary n-3 polyunsaturated fatty acids alter the number, fatty acid profile and coagulatory activity of circulating and platelet-derived extracellular vesicles: a randomized, controlled crossover trial.

BACKGROUND: Extracellular vesicles (EVs) are proposed to play a role in the development of cardiovascular diseases (CVDs) and are considered emerging markers of CVDs. n-3 PUFAs are abundant in oily fish and fish oil and are reported to reduce CVD risk, but there has been little research to date examining the effects of n-3 PUFAs on the generation and function of EVs. OBJECTIVES: We aimed to investigate the effects of fish oil supplementation on the number, generation, and function of EVs in subjects with moderate risk of CVDs. METHODS: A total of 40 participants with moderate risk of CVDs were supplemented with capsules containing either fish oil (1.9 g/d n-3 PUFAs) or control oil (high-oleic safflower oil) for 12 wk in a randomized, double-blind, placebo-controlled crossover intervention study. The effects of fish oil supplementation on conventional CVD and thrombogenic risk markers were measured, along with the number and fatty acid composition of circulating and platelet-derived EVs (PDEVs). PDEV proteome profiles were evaluated, and their impact on coagulation was assessed using assays including fibrin clot formation, thrombin generation, fibrinolysis, and ex vivo thrombus formation. RESULTS: n-3 PUFAs decreased the numbers of circulating EVs by 27%, doubled their n-3 PUFA content, and reduced their capacity to support thrombin generation by >20% in subjects at moderate risk of CVDs. EVs derived from n-3 PUFA-enriched platelets in vitro also resulted in lower thrombin generation, but did not alter thrombus formation in a whole blood ex vivo assay. CONCLUSIONS: Dietary n-3 PUFAs alter the number, composition, and function of EVs, reducing their coagulatory activity. This study provides clear evidence that EVs support thrombin generation and that this EV-dependent thrombin generation is reduced by n-3 PUFAs, which has implications for prevention and treatment of thrombosis. CLINICAL TRIAL REGISTRY: This trial was registered at clinicaltrials.gov as NCT03203512.

MRI and Molecular Characterization of Pediatric High-Grade Midline Thalamic Gliomas: The HERBY Phase II Trial.

Background Diffuse midline gliomas (DMG) are characterized by a high incidence of H3 K27 mutations and poorer outcome. The HERBY trial has provided one of the largest cohorts of pediatric DMGs with available radiologic, histologic-genotypic, and survival data. Purpose To define MRI and molecular characteristics of DMG. Materials and Methods This study is a secondary analysis of a prospective trial (HERBY; ClinicalTrials.gov identifier, NCT01390948) undertaken between October 2011 and February 2016. Among 121 HERBY participants, 50 had midline nonpontine-based tumors. Midline high-grade gliomas were reclassified into DMG H3 K27 mutant, H3 wild type with enhancer of zest homologs inhibitory protein overexpression, epidermal growth factor receptormutant, or not otherwise stated. The epicenter of each tumor and other radiologic characteristics were ascertained from MRI and correlated with the new subtype classification, histopathologic characteristics, surgical extent, and outcome parameters. Kaplan-Meier curves and log-rank tests were applied to determine and describe survival differences between groups. Results There were 42 participants (mean age, 12 years ± 4 [SD]; 23 girls) with radiologically evaluable thalamic-based DMG. Eighteen had partial thalamic involvement (12 thalamopulvinar, six anteromedial), 10 involved a whole thalamus, nine had unithalamic tumors with diffuse contiguous extension, and five had bithalamic tumors (two symmetric, three partial). Twenty-eight participants had DMG H3 K27 mutant tumors; there were no differences in outcome compared with other DMGs (n = 4). Participants who underwent major debulking or total or near-total resection had longer overall survival (OS): 18.5 months vs 11.4 months (P = .02). Enrolled participants who developed leptomeningeal metastatic dissemination before starting treatment had worse outcomes (event-free survival, 2.9 months vs 8.0 months [P = .02]; OS, 11.4 months vs 18.5 months [P = .004]). Conclusion Thalamic involvement of diffuse midline gliomas ranged from localized partial thalamic to holo- or bithalamic with diffuse contiguous spread and had poor outcomes, irrespective of H3 K27 subtype alterations. Leptomeningeal dissemination and less than 50% surgical resection were adverse risk factors for survival. Clinical trial registration no. NCT01390948 © RSNA, 2022 Online supplemental material is available for this article. See also the editorial by Widjaja in this issue.

A subset of pediatric-type thalamic gliomas share a distinct DNA methylation profile, H3K27me3 loss and frequent alteration of EGFR.

BACKGROUND: Malignant astrocytic gliomas in children show a remarkable biological and clinical diversity. Small in-frame insertions or missense mutations in the epidermal growth factor receptor gene (EGFR) have recently been identified in a distinct subset of pediatric-type bithalamic gliomas with a unique DNA methylation pattern. METHODS: Here, we investigated an epigenetically homogeneous cohort of malignant gliomas (n = 58) distinct from other subtypes and enriched for pediatric cases and thalamic location, in comparison with this recently identified subtype of pediatric bithalamic gliomas. RESULTS: EGFR gene amplification was detected in 16/58 (27%) tumors, and missense mutations or small in-frame insertions in EGFR were found in 20/30 tumors with available sequencing data (67%; 5 of them co-occurring with EGFR amplification). Additionally, 8 of the 30 tumors (27%) harbored an H3.1 or H3.3 K27M mutation (6 of them with a concomitant EGFR alteration). All tumors tested showed loss of H3K27me3 staining, with evidence of overexpression of the EZH inhibitory protein (EZHIP) in the H3 wildtype cases. Although some tumors indeed showed a bithalamic growth pattern, a significant proportion of tumors occurred in the unilateral thalamus or in other (predominantly midline) locations. CONCLUSIONS: Our findings present a distinct molecular class of pediatric-type malignant gliomas largely overlapping with the recently reported bithalamic gliomas characterized by EGFR alteration, but additionally showing a broader spectrum of EGFR alterations and tumor localization. Global H3K27me3 loss in this group appears to be mediated by either H3 K27 mutation or EZHIP overexpression. EGFR inhibition may represent a potential therapeutic strategy in these highly aggressive gliomas.

ALK2 inhibitors display beneficial effects in preclinical models of ACVR1 mutant diffuse intrinsic pontine glioma.

Diffuse intrinsic pontine glioma (DIPG) is a lethal childhood brainstem tumour, with a quarter of patients harbouring somatic mutations in ACVR1, encoding the serine/threonine kinase ALK2. Despite being an amenable drug target, little has been done to-date to systematically evaluate the role of ACVR1 in DIPG, nor to screen currently available inhibitors in patient-derived tumour models. Here we show the dependence of DIPG cells on the mutant receptor, and the preclinical efficacy of two distinct chemotypes of ALK2 inhibitor in vitro and in vivo. We demonstrate the pyrazolo[1,5-a]pyrimidine LDN-193189 and the pyridine LDN-214117 to be orally bioavailable and well-tolerated, with good brain penetration. Treatment of immunodeprived mice bearing orthotopic xenografts of H3.3K27M, ACVR1R206H mutant HSJD-DIPG-007 cells with 25 mg/kg LDN-193189 or LDN-214117 for 28 days extended survival compared with vehicle controls. Development of ALK2 inhibitors with improved potency, selectivity and advantageous pharmacokinetic properties may play an important role in therapy for DIPG patients.

Euphol, a tetracyclic triterpene, from Euphorbia tirucalli induces autophagy and sensitizes temozolomide cytotoxicity on glioblastoma cells.

Glioblastoma (GBM) is the most frequent and aggressive type of brain tumor. There are limited therapeutic options for GBM so that new and effective agents are urgently needed. Euphol is a tetracyclic triterpene alcohol, and it is the main constituent of the sap of the medicinal plant Euphorbia tirucalli. We previously identified anti-cancer activity in euphol based on the cytotoxicity screening of 73 human cancer cells. We now expand the toxicological screening of the inhibitory effect and bioactivity of euphol using two additional glioma primary cultures. Euphol exposure showed similar cytotoxicity against primary glioma cultures compared to commercial glioma cells. Euphol has concentration-dependent cytotoxic effects on cancer cell lines, with more than a five-fold difference in the IC50 values in some cell lines. Euphol treatment had a higher selective cytotoxicity index (0.64-3.36) than temozolomide (0.11-1.13) and reduced both proliferation and cell motility. However, no effect was found on cell cycle distribution, invasion and colony formation. Importantly, the expression of the autophagy-associated protein LC3-II and acidic vesicular organelle formation were markedly increased, with Bafilomycin A1 potentiating cytotoxicity. Finally, euphol also exhibited antitumoral and antiangiogenic activity in vivo, using the chicken chorioallantoic membrane assay, with synergistic temozolomide interactions in most cell lines. In conclusion, euphol exerted in vitro and in vivo cytotoxicity against glioma cells, through several cancer pathways, including the activation of autophagy-associated cell death. These findings provide experimental support for further development of euphol as a novel therapeutic agent for GBM, either alone or in combination chemotherapy.

Novel Quinazolinone Inhibitors of ALK2 Flip between Alternate Binding Modes: Structure-Activity Relationship, Structural Characterization, Kinase Profiling, and Cellular Proof of Concept.

Structure-activity relationship and crystallographic data revealed that quinazolinone-containing fragments flip between two distinct modes of binding to activin receptor-like kinase-2 (ALK2). We explored both binding modes to discover potent inhibitors and characterized the chemical modifications that triggered the flip in binding mode. We report kinase selectivity and demonstrate that compounds of this series modulate ALK2 in cancer cells. These inhibitors are attractive starting points for the discovery of more advanced ALK2 inhibitors.

The impact of neuromuscular electrical stimulation on recovery after intensive, muscle damaging, maximal speed training in professional team sports players.

OBJECTIVES: During congested fixture periods in team sports, limited recovery time and increased travel hinder the implementation of many recovery strategies; thus alternative methods are required. We examined the impact of a neuromuscular electrical stimulation device on 24-h recovery from an intensive training session in professional players. DESIGN: Twenty-eight professional rugby and football academy players completed this randomised and counter-balanced study, on 2 occasions, separated by 7 days. METHODS: After baseline perceived soreness, blood (lactate and creatine kinase) and saliva (testosterone and cortisol) samples were collected, players completed a standardised warm-up and baseline countermovement jumps (jump height). Players then completed 60 m × 50 m maximal sprints, with 5 min recovery between efforts. After completing the sprint session, players wore a neuromuscular electrical stimulation device or remained in normal attire (CON) for 8 h. All measures were repeated immediately, 2 and 24-h post-sprint. RESULTS: Player jump height was reduced from baseline at all time points under both conditions; however, at 24-h neuromuscular electrical stimulation was significantly more recovered (mean±SD; neuromuscular electrical stimulation -3.2±3.2 vs. CON -7.2±3.7%; P<0.001). Creatine kinase concentrations increased at all time points under both conditions, but at 24-h was lower under neuromuscular electrical stimulation (P<0.001). At 24-h, perceived soreness was significantly lower under neuromuscular electrical stimulation, when compared to CON (P=0.02). There was no effect of condition on blood lactate, or saliva testosterone and cortisol responses (P>0.05). CONCLUSIONS: Neuromuscular electrical stimulation improves recovery from intensive training in professional team sports players. This strategy offers an easily applied recovery strategy which may have particular application during sleep and travel.

Targeted Physician Education Positively Affects Delivery of Nutrition Therapy and Patient Outcomes: Results of a Prospective Clinical Trial.

BACKGROUND: Malnutrition is a continuing epidemic among hospitalized patients. We hypothesize that targeted physician education should help reduce caloric deficits and improve patient outcomes. MATERIALS AND METHODS: We performed a prospective trial of patients (n = 121) assigned to 1 of 2 trauma groups. The experimental group (EG) received targeted education consisting of strategies to increase delivery of early enteral nutrition. Strategies included early enteral access, avoidance of nil per os (NPO) and clear liquid diets (CLD), volume-based feeding, early resumption of feeds postprocedure, and charting caloric deficits. The control group (CG) did not receive targeted education but was allowed to practice in a standard ad hoc fashion. Both groups were provided with dietitian recommendations on a multidisciplinary nutrition team per standard practice. RESULTS: The EG received a higher percentage of measured goal calories (30.1 ± 18.5%, 22.1 ± 23.7%, P = .024) compared with the CG. Mean caloric deficit was not significantly different between groups (-6796 ± 4164 kcal vs -8817 ± 7087 kcal, P = .305). CLD days per patient (0.1 ± 0.5 vs 0.6 ± 0.9), length of stay in the intensive care unit (3.5 ± 5.5 vs 5.2 ± 6.8 days), and duration of mechanical ventilation (1.6 ± 3.7 vs 2.8 ± 5.0 days) were all reduced in the EG compared with the CG (P < .05). EG patients had fewer nosocomial infections (10.6% vs 23.6%) and less organ failure (10.6% vs 18.2%) than did the CG, but these differences did not reach statistical significance. CONCLUSION: Implementation of specific educational strategies succeeded in greater delivery of nutrition therapy, which favorably affected patient care and outcomes.

Challenges in keeping pace with immunogenicity advances throughout the clinical development of a biological therapeutic.

Progress in the assessment of immunogenicity over the past decade has been rapid; therapeutics in late-phase clinical studies today are subject to antidrug antibody analyses that have evolved considerably since the start of their clinical development. Data from bridging and direct ELISA methods used for the analysis of antidrug antibodies generated in response to an antibody therapeutic will be presented. The case study will highlight some of the challenges faced and the learning points gained from supporting immunogenicity analysis for these therapeutics.

Evolution of bone and plasma concentration of lanthanum in dialysis patients before, during 1 year of treatment with lanthanum carbonate and after 2 years of follow-up.

BACKGROUND: Lanthanum carbonate (LC) has been proposed as a new phosphate binder. Presented here are the results from one centre that participated in a multicentre trial to assess the effect of treatment with LC and calcium carbonate (CC) on the evolution of renal osteodystrophy in dialysis patients. Bone biopsies were performed at baseline, after 1 year of treatment and after a further 2-year follow-up period to assess the lanthanum concentration in bone and plasma. METHODS: Twenty new dialysis patients were randomized to receive LC (median dose 1250 mg) for 1 year (n = 10), followed by 2 years of CC treatment or CC (n = 10) during the whole study period (3 years). RESULTS: After 36 weeks of treatment, steady state was reached with plasma lanthanum levels varying around 0.6 ng/ml. Six weeks after cessation of 1 year of treatment, the plasma lanthanum levels declined to a value of 0.17 +/- 0.12 ng/ml (P < 0.05) and after 2 years to 0.09 +/- 0.03 ng/ml. Plasma and bone lanthanum levels did not correlate with the average lanthanum dose at any time point. The mean bone concentration in patients receiving LC increased from 0.05 +/- 0.03 to 2.3 +/- 1.6 microg/g (P < 0.05) after 1 year and slightly decreased at the end of the study to 1.9 +/- 1.6 microg/g (P < 0.05). CONCLUSIONS: Bone deposition after 1 year of treatment with LC is low (highest concentration: 5.5 microg/g). There is a slow release of lanthanum from its bone deposits 2 years after the discontinuation of the treatment and no association with aluminium-like bone toxicity.

Pessary treatment of pelvic relaxation: factors affecting successful fitting and continued use.

OBJECTIVES: The aims of this study were to determine predictors of successful pessary fitting and continued pessary use in patients with pelvic relaxation. DESIGN: Retrospective observational study. SETTING AND SUBJECTS: The medical records of 130 consecutive patients evaluated for pessary treatment of pelvic relaxation by a single specially trained nurse practitioner (CRJ) at Kaiser Foundation Hospital, Harbor City, Calif, between May 1, 1997, and June 30, 2002, were retrospectively reviewed. INSTRUMENTS: Voiding diaries, data collection sheet, and questionnaires. METHODS: The medical records of the 130 patients were retrospectively reviewed, and data were recorded on data collection sheets. Patients using pessaries completed a questionnaire to assess treatment effectiveness. RESULTS: Coexisting stress urinary incontinence and previous prolapse and cystocele/rectocele repairs were each found to be independent predictors of unsuccessful pessary fitting. Fifty percent of successfully fitted patients had discontinued pessary use by 24 months. Current pessary users were more likely to have undergone prior pelvic reconstructive surgery (37% vs 13%, P = .02), less likely to require a space-filling pessary (13% vs 37%, P = .03), and more likely to recommend pessary to their friends or family (87% vs 50%, P = .007) compared to patients who discontinued pessary use. CONCLUSIONS: Prior pelvic reconstructive surgery is associated with an increased risk of unsuccessful pessary fitting; however, those patients who are successfully fitted tend to continue pessary use.

Engaging precontemplative dual diagnosis clients.

It can be difficult to engage with clients with co-existing substance misuse and mental health problems, known as dual diagnosis. This article describes a project that used music to engage service users with dual diagnosis problems.

Infection with Basidiobolus ranarum in two dogs.

Basidiobolus ranarum is a saprophytic fungus in the environment that also is a part of the endogenous microflora in the gastrointestinal tract of several vertebrates. These organisms may penetrate skin or muscosa of humans and other animals, causing granulomatous inflammation. Two dogs infected with B. ranarum had prolonged or repeated exposure to water or soil in their environment. One dog had progressive subcutaneous infection of all the limbs, and the other dog had recurrent coughing and dyspnea caused by tracheobronchitis. In both dogs, secondary bacterial infection of the lesions was evident. Treatment of the dog with subcutaneous infection involved cutaneous dressings and sequential use of enrofloxacin and itraconazole; however, this resulted in suspected liver damage without clinical improvement. Subsequent treatment with potassium iodide and a lipid formulation of amphotericin B was also unsuccessful, and the dog was euthanatized. The other dog was treated alternately with enrofloxacin and itraconazole. When the clinical signs and infection returned, combination treatment with both drugs was more effective; however, the dog developed liver damage. Subsequent treatment with enrofloxacin on an intermittent basis controlled the dog's coughing during a 3-year period.