Sarcospan reduces dystrophic pathology: stabilization of the utrophin–glycoprotein complex

Mutations in the dystrophin gene cause Duchenne muscular dystrophy and result in the loss of dystrophin and the entire dystrophin–glycoprotein complex (DGC) from the sarcolemma. We show that sarcospan (SSPN), a unique tetraspanin-like component of the DGC, ameliorates muscular dystrophy in dystrophin-deficient mdx mice. SSPN stabilizes the sarcolemma by increasing levels of the utrophin–glycoprotein complex (UGC) at the extrasynaptic membrane to compensate for the loss of dystrophin. Utrophin is normally restricted to the neuromuscular junction, where it replaces dystrophin to form a functionally analogous complex. SSPN directly interacts with the UGC and functions to stabilize utrophin protein without increasing utrophin transcription. These findings reveal the importance of protein stability in the prevention of muscular dystrophy and may impact the future design of therapeutics for muscular dystrophies

Sarcospan Regulates Cardiac Isoproterenol Response and Prevents Duchenne Muscular Dystrophy-Associated Cardiomyopathy.

BackgroundDuchenne muscular dystrophy is a fatal cardiac and skeletal muscle disease resulting from mutations in the dystrophin gene. We have previously demonstrated that a dystrophin-associated protein, sarcospan (SSPN), ameliorated Duchenne muscular dystrophy skeletal muscle degeneration by activating compensatory pathways that regulate muscle cell adhesion (laminin-binding) to the extracellular matrix. Conversely, loss of SSPN destabilized skeletal muscle adhesion, hampered muscle regeneration, and reduced force properties. Given the importance of SSPN to skeletal muscle, we investigated the consequences of SSPN ablation in cardiac muscle and determined whether overexpression of SSPN into mdx mice ameliorates cardiac disease symptoms associated with Duchenne muscular dystrophy cardiomyopathy.Methods and resultsSSPN-null mice exhibited cardiac enlargement, exacerbated cardiomyocyte hypertrophy, and increased fibrosis in response to β-adrenergic challenge (isoproterenol; 0.8 mg/day per 2 weeks). Biochemical analysis of SSPN-null cardiac muscle revealed reduced sarcolemma localization of many proteins with a known role in cardiomyopathy pathogenesis: dystrophin, the sarcoglycans (α-, δ-, and γ-subunits), and β1D integrin. Transgenic overexpression of SSPN in Duchenne muscular dystrophy mice (mdx(TG)) improved cardiomyofiber cell adhesion, sarcolemma integrity, cardiac functional parameters, as well as increased expression of compensatory transmembrane proteins that mediate attachment to the extracellular matrix.ConclusionsSSPN regulates sarcolemmal expression of laminin-binding complexes that are critical to cardiac muscle function and protects against transient and chronic injury, including inherited cardiomyopathy

Family-Focused Treatment for Adolescents and Young Adults at High Risk for Psychosis: Results of a Randomized Trial

Objective: Longitudinal studies have begun to clarify the phenotypic characteristics of adolescents and young adults at clinical high risk for psychosis. This 8-site randomized trial examined whether a 6-month program of family psychoeducation was effective in reducing the severity of attenuated positive and negative psychotic symptoms and enhancing functioning among individuals at high risk. Method: Adolescents and young adults (mean age 17.4 +/- 4.1 years) with attenuated positive psychotic symptoms, brief and intermittent psychosis, or genetic risk with functional deterioration were randomly assigned to 18 sessions of family-focused therapy for individuals at clinical high risk (FFT-CHR) in 6 months or 3 sessions of family psychoeducation (enhanced care [EC]. FFT-CHR included psychoeducation about early signs of psychosis, stress management, communication training, and problem-solving skills training, whereas EC focused on symptom prevention. Independent evaluators assessed participants at baseline and 6 months on positive and negative symptoms and social-role functioning. Results: Of 129 participants, 102 (79.1%) were followed up at 6 months. Participants in FFT-CHR showed greater improvements in attenuated positive symptoms over 6 months than participants in EC (F-1,F-97 = 5.49, p = .02). Negative symptoms improved independently of psychosocial treatments. Changes in psychosocial functioning depended on age: participants more than 19 years of age showed more role improvement in FFT-CHR, whereas participants between 16 and 19 years of age showed more role improvement in EC. The results were independent of concurrent pharmacotherapy. Conclusion: Interventions that focus on improving family relationships may have prophylactic efficacy in individuals at high risk for psychosis. Future studies should examine the specificity of effects of family intervention compared to individual therapy of the same duration and frequency. Clinical trial registration information-Prevention Trial of Family Focused Treatment in Youth at Risk for Psychosis

Development of a primary care pandemic plan informed by in-depth policy analysis and interviews with family physicians across Canada during COVID-19: A qualitative case study protocol

Introduction Given the recurrent risk of respiratory illness-based pandemics, and the important roles family physicians play during public health emergencies, the development of pandemic plans for primary care is imperative. Existing pandemic plans in Canada, however, do not adequately incorporate family physicians\u27 roles and perspectives. This policy and planning oversight has become increasingly evident with the emergence of the novel coronavirus disease, COVID-19, pandemic. This study is designed to inform the development of pandemic plans for primary care through evidence from four provinces in Canada: British Columbia, Newfoundland and Labrador, Nova Scotia, and Ontario. Methods and analysis We will employ a multiple-case study of regions in four provinces. Each case consists of a mixed methods design which comprises: (1) a chronology of family physician roles in the COVID-19 pandemic response; (2) a provincial policy analysis; and (3) qualitative interviews with family physicians. Relevant policy and guidance documents will be identified through targeted, snowball and general search strategies. Additionally, these policy documents will be analysed to identify gaps and/or emphases in existing policies and policy responses. Interviews will explore family physicians\u27 proposed, actual and potential roles during the pandemic, the facilitators and barriers they have encountered throughout and the influence of gender on their professional roles. Data will be thematically analysed using a content analysis framework, first at the regional level and then through cross-case analyses. Ethics and dissemination Approval for this study has been granted by the Research Ethics of British Columbia, the Health Research Ethics Board of Newfoundland and Labrador, the Nova Scotia Health Authority Research Ethics Board and the Western University Research Ethics Board. Findings will be disseminated via conferences and peer-reviewed publications. Evidence and lessons learnt will be used to develop tools for government ministries, public health units and family physicians for improved pandemic response plans for primary care

ABCC Multidrug Transporters in Childhood Neuroblastoma: Clinical and Biological Effects Independent of Cytotoxic Drug Efflux

Background Although the prognostic value of the ATP-binding cassette, subfamily C (ABCC) transporters in childhood neuroblastoma is usually attributed to their role in cytotoxic drug efflux, certain observations have suggested that these multidrug transporters might contribute to the malignant phenotype independent of cytotoxic drug efflux. Methods A v-myc myelocytomatosis viral related oncogene, neuroblastoma derived (MYCN)-driven transgenic mouse neuroblastoma model was crossed with an Abcc1-deficient mouse strain (658 hMYCN1/−, 205 hMYCN+/1 mice) or, alternatively, treated with the ABCC1 inhibitor, Reversan (n = 20). ABCC genes were suppressed using short interfering RNA or overexpressed by stable transfection in neuroblastoma cell lines BE(2)-C, SH-EP, and SH-SY5Y, which were then assessed for wound closure ability, clonogenic capacity, morphological differentiation, and cell growth. Real-time quantitative polymerase chain reaction was used to examine the clinical significance of ABCC family gene expression in a large prospectively accrued cohort of patients (n = 209) with primary neuroblastomas. Kaplan-Meier survival analysis and Cox regression were used to test for associations with event-free and overall survival. Except where noted, all statistical tests were two-sided. Results Inhibition of ABCC1 statistically significantly inhibited neuroblastoma development in hMYCN transgenic mice (mean age for palpable tumor: treated mice, 47.2 days; control mice, 41.9 days; hazard ratio [HR] = 9.3, 95% confidence interval [CI] = 2.65 to 32; P < .001). Suppression of ABCC1 in vitro inhibited wound closure (P < .001) and clonogenicity (P = .006); suppression of ABCC4 enhanced morphological differentiation (P < .001) and inhibited cell growth (P < .001). Analysis of 209 neuroblastoma patient tumors revealed that, in contrast with ABCC1 and ABCC4, low rather than high ABCC3 expression was associated with reduced event-free survival (HR of recurrence or death = 2.4, 95% CI = 1.4 to 4.2; P = .001), with 23 of 53 patients with low ABCC3 expression experiencing recurrence or death compared with 31 of 155 patients with high ABCC3. Moreover, overexpression of ABCC3 in vitro inhibited neuroblastoma cell migration (P < .001) and clonogenicity (P = .03). The combined expression of ABCC1, ABCC3, and ABCC4 was associated with patients having an adverse event, such that of the 12 patients with the "poor prognosis” expression pattern, 10 experienced recurrence or death (HR of recurrence or death = 12.3, 95% CI = 6 to 27; P < .001). Conclusion ABCC transporters can affect neuroblastoma biology independently of their role in chemotherapeutic drug efflux, enhancing their potential as targets for therapeutic interventio

Joint Inference in Weakly-Annotated Image Datasets via Dense Correspondence

We present a principled framework for inferring pixel labels in weakly-annotated image datasets. Most previous, example-based approaches to computer vision rely on a large corpus of densely labeled images. However, for large, modern image datasets, such labels are expensive to obtain and are often unavailable. We establish a large-scale graphical model spanning all labeled and unlabeled images, then solve it to infer pixel labels jointly for all images in the dataset while enforcing consistent annotations over similar visual patterns. This model requires significantly less labeled data and assists in resolving ambiguities by propagating inferred annotations from images with stronger local visual evidences to images with weaker local evidences. We apply our proposed framework to two computer vision problems, namely image annotation with semantic segmentation, and object discovery and co-segmentation (segmenting multiple images containing a common object). Extensive numerical evaluations and comparisons show that our method consistently outperforms the state-of-the-art in automatic annotation and semantic labeling, while requiring significantly less labeled data. In contrast to previous co-segmentation techniques, our method manages to discover and segment objects well even in the presence of substantial amounts of noise images (images not containing the common object), as typical for datasets collected from Internet search

Determining molecular orientation via single molecule SERS in a plasmonic nano-gap

In this work, plasmonic nano-gaps consisting of a silver nanoparticle coupled to an extended silver film have been fully optimized for single molecule Surface-Enhanced Raman Scattering (SERS) spectroscopy. The SERS signal was found to be strongly dependent on the particle size and the molecule orientation with respect to the field inside the nano-gap. Using Finite Difference Time Domain (FDTD) simulations to complement the experimental measurements, the complex interplay between the excitation enhancement and the emission enhancement of the system as a function of particle size were highlighted. Additionally, in conjunction with Density Functional Theory (DFT), the well-defined field direction in the nano-gap enables to recover the orientation of individual molecules

Comparability: manufacturing, characterization and controls, report of a UK Regenerative Medicine Platform Pluripotent Stem Cell Platform Workshop, Trinity Hall, Cambridge, 14–15 September 2015

This paper summarizes the proceedings of a workshop held at Trinity Hall, Cambridge to discuss comparability and includes additional information and references to related information added subsequently to the workshop. Comparability is the need to demonstrate equivalence of product after a process change; a recent publication states that this ‘may be difficult for cell-based medicinal products’. Therefore a well-managed change process is required which needs access to good science and regulatory advice and developers are encouraged to seek help early. The workshop shared current thinking and best practice and allowed the definition of key research questions. The intent of this report is to summarize the key issues and the consensus reached on each of these by the expert delegates