Effect of aerobic exercise training and cognitive behavioural therapy on reduction of chronic fatigue in patients with facioscapulohumeral dystrophy: protocol of the FACTS-2-FSHD trial

<p>Abstract</p> <p>Background</p> <p>In facioscapulohumeral dystrophy (FSHD) muscle function is impaired and declines over time. Currently there is no effective treatment available to slow down this decline. We have previously reported that loss of muscle strength contributes to chronic fatigue through a decreased level of physical activity, while fatigue and physical inactivity both determine loss of societal participation. To decrease chronic fatigue, two distinctly different therapeutic approaches can be proposed: aerobic exercise training (AET) to improve physical capacity and cognitive behavioural therapy (CBT) to stimulate an active life-style yet avoiding excessive physical strain. The primary aim of the FACTS-2-FSHD (acronym for Fitness And Cognitive behavioural TherapieS/for Fatigue and ACTivitieS in FSHD) trial is to study the effect of AET and CBT on the reduction of chronic fatigue as assessed with the Checklist Individual Strength subscale fatigue (CIS-fatigue) in patients with FSHD. Additionally, possible working mechanisms and the effects on various secondary outcome measures at all levels of the International Classification of Functioning, Disability and Health (ICF) are evaluated.</p> <p>Methods/Design</p> <p>A multi-centre, assessor-blinded, randomized controlled trial is conducted. A sample of 75 FSHD patients with severe chronic fatigue (CIS-fatigue ≥ 35) will be recruited and randomized to one of three groups: (1) AET + usual care, (2) CBT + usual care or (3) usual care alone, which consists of no therapy at all or occasional (conventional) physical therapy. After an intervention period of 16 weeks and a follow-up of 3 months, the third (control) group will as yet be randomized to either AET or CBT (approximately 7 months after inclusion). Outcomes will be assessed at baseline, immediately post intervention and at 3 and 6 months follow up.</p> <p>Discussion</p> <p>The FACTS-2-FSHD study is the first theory-based randomized clinical trial which evaluates the effect and the maintenance of effects of AET and CBT on the reduction of chronic fatigue in patients with FSHD. The interventions are based on a theoretical model of chronic fatigue in patients with FSHD. The study will provide a unique set of data with which the relationships between outcome measures at all levels of the ICF could be assessed.</p> <p>Trial registration</p> <p>Dutch Trial Register, NTR1447.</p

FSHD muscular dystrophy Region Gene 1 binds Suv4-20h1 histone methyltransferase and impairs myogenesis

Facioscapulohumeral Muscular Dystrophy (FSHD) is an autosomal dominant myopathy with a strong epigenetic component. It is associated with deletion of a macrosatellite repeat leading to over-expression of the nearby genes. Among them, we focused on FSHD Region Gene 1 (FRG1) since its over-expression in mice, X. laevis and C. elegans leads to muscular dystrophy-like defects, suggesting that FRG1 plays a relevant role in muscle biology. Here we show that, when overexpressed, FRG1 binds and interferes with the activity of the histone methyltransferase Suv4-20h1 both in mammals and Drosophila. Accordingly, FRG1 over-expression or Suv4-20h1 knockdown inhibits myogenesis. Moreover, Suv4-20h KO mice develop muscular dystrophy signs. Finally, we identify the FRG1/Suv4-20h1 target Eid3 as a novel myogenic inhibitor that contributes to the muscle differentiation defects. Our study suggests a novel role of FRG1 as epigenetic regulator of muscle differentiation and indicates that Suv4-20h1 has a gene-specific function in myogenesis

Artificial Intelligence, Mathematical Modeling and Magnetic Resonance Imaging for Precision Medicine in Neurology and Neuroradiology

La tesi affronta la possibilità di utilizzare metodi matematici, tecniche di simulazione, teorie fisiche riadattate e algoritmi di intelligenza artificiale per soddisfare le esigenze cliniche in neuroradiologia e neurologia al fine di descrivere e prevedere i patterns e l’evoluzione temporale di una malattia, nonché di supportare il processo decisionale clinico. La tesi è suddivisa in tre parti. La prima parte riguarda lo sviluppo di un workflow radiomico combinato con algoritmi di Machine Learning al fine di prevedere parametri che favoriscono la descrizione quantitativa dei cambiamenti anatomici e del coinvolgimento muscolare nei disordini neuromuscolari, con particolare attenzione alla distrofia facioscapolo-omerale. Il workflow proposto si basa su sequenze di risonanza magnetica convenzionali disponibili nella maggior parte dei centri neuromuscolari e, dunque, può essere utilizzato come strumento non invasivo per monitorare anche i più piccoli cambiamenti nei disturbi neuromuscolari oltre che per la valutazione della progressione della malattia nel tempo. La seconda parte riguarda l’utilizzo di un modello cinetico per descrivere la crescita tumorale basato sugli strumenti della meccanica statistica per sistemi multi-agente e che tiene in considerazione gli effetti delle incertezze cliniche legate alla variabilità della progressione tumorale nei diversi pazienti. L'azione dei protocolli terapeutici è modellata come controllo che agisce a livello microscopico modificando la natura della distribuzione risultante. Viene mostrato come lo scenario controllato permetta di smorzare le incertezze associate alla variabilità della dinamica tumorale. Inoltre, sono stati introdotti metodi di simulazione numerica basati sulla formulazione stochastic Galerkin del modello cinetico sviluppato. La terza parte si riferisce ad un progetto ancora in corso che tenta di descrivere una porzione di cervello attraverso la teoria quantistica dei campi e di simularne il comportamento attraverso l'implementazione di una rete neurale con una funzione di attivazione costruita ad hoc e che simula la funzione di risposta del modello biologico neuronale. E’ stato ottenuto che, nelle condizioni studiate, l'attività della porzione di cervello può essere descritta fino a O(6), i.e, considerando l’interazione fino a sei campi, come un processo gaussiano. Il framework quantistico definito può essere esteso anche al caso di un processo non gaussiano, ovvero al caso di una teoria di campo quantistico interagente utilizzando l’approccio della teoria wilsoniana di campo efficace.The thesis addresses the possibility of using mathematical methods, simulation techniques, repurposed physical theories and artificial intelligence algorithms to fulfill clinical needs in neuroradiology and neurology. The aim is to describe and to predict disease patterns and its evolution over time as well as to support clinical decision-making processes. The thesis is divided into three parts. Part 1 is related to the development of a Radiomic workflow combined with Machine Learning algorithms in order to predict parameters that quantify muscular anatomical involvement in neuromuscular diseases, with special focus on Facioscapulohumeral dystrophy. The proposed workflow relies on conventional Magnetic Resonance Imaging sequences available in most neuromuscular centers and it can be used as a non-invasive tool to monitor even fine change in neuromuscular disorders and to evaluate longitudinal diseases’ progression over time. Part 2 is about the description of a kinetic model for tumor growth by means of classical tools of statistical mechanics for many-agent systems also taking into account the effects of clinical uncertainties related to patients’ variability in tumor progression. The action of therapeutic protocols is modeled as feedback control at the microscopic level. The controlled scenario allows the dumping of uncertainties associated with the variability in tumors’ dynamics. Suitable numerical methods, based on Stochastic Galerkin formulation of the derived kinetic model, are introduced. Part 3 refers to a still-on going project that attempts to describe a brain portion through a quantum field theory and to simulate its behavior through the implementation of a neural network with an ad-hoc activation function mimicking the biological neuron model response function. Under considered conditions, the brain portion activity can be expressed up to O(6), i.e., up to six fields interaction, as a Gaussian Process. The defined quantum field framework may also be extended to the case of a Non-Gaussian Process behavior, or rather to an interacting quantum field theory in a Wilsonian Effective Field theory approach

ChIP analysis of the histone modifications at the D4Z4 repeats in facioscapulohumeral muscular dystrophy (FSHD)

Genomic DNA must exist in a particular chromatin configuration and modification of this structure is essential for the correct control of gene expression. There are several human genetic disorders that are caused by misregulation of epigenetic gene control. Facioscapulohumeral muscular dystrophy (FSHD) is a disease that may be caused by alterations in chromatin structure. FSHD is the third most common form of muscular dystrophy. The majority of FSHD cases show contraction of the D4Z4 repeats on the 4q35 chromosome (FSHD1). However, a small number of FSHD cases show no contraction at this region (FSHD2), but share epigenetic changes at the D4Z4 region with the FSHD1 patients. In 2009, Zeng et al. reported a specific loss of H3K9me3 histone modification at the D4Z4 repeats in FSHD patients. The main focus of this study was to verify the published data by Zeng et al (2009) and further investigate the histone modification changes at the D4Z4 array. Chromatin immunoprecipitation (ChIP) coupled with real-time quantitative PCR (qPCR) was employed to investigate the histone modifications within the D4Z4 array. The results obtained were in agreement with the previously published data on the reduction of H3K9me3 histone modification at the D4Z4 repeats in FSHD patients. However, contradictory to the previous data, the reduction of this histone modification was also observed on other genomic regions. A global reduction of H3K27me3 was also observed in FSHD patients

Facio-scapulo-humeral dystrophy. Role of exercise in its rehabilitation

La distrofia Facio-escápulo humeral o enfermedad de Landouzy-Dejerine es la tercera enfermedad neuromuscular hereditaria más común. Se trata de una miopatía lentamente progresiva, autosómica dominante, con una incidencia aproximada de 1:20.000. Una de sus características principales es el patrón de debilidad muscular, que además suele ser asimétrico. Por ello, los terapeutas suelen recomendar ejercicio como base de su tratamiento. Existiendo una gran variedad de clases de ejercicio, en este artículo se revisan los ensayos publicados al respecto para identificar el tipo de ejercicio que puede ser más beneficioso para estos pacientes.Facio-scapular humeral dystrophy or Landouzy-Dejerine disease is the third most common hereditary neuromuscular disease. It is a slowly progressive, autosomal dominant myopathy, with an approximate incidence of 1: 20,000. One of its main characteristics is the muscle weakness pattern, which is usually asymmetric. Therefore, therapists often recommend exercise as a basis for their treatment. Existing a great variety of exercise classes, this article reviews the trials published in this regard to identify the type of exercise that may be most beneficial for these patients

Facio-scapulo-humeral dystrophy: clinical follow-up and role of chromosome X inactivation in female patients - SHP2: a novel therapeutic target in MuSK-myasthenia

Part I. Facio-scapulo-humeral dystrophy: clinical follow-up and role of chromosome X inactivation in female patients. Facio-scapulo-humeral dystrophy (FSHD) is an autosomal dominant muscular dystrophy characterized by high prevalence and clinical variability. It involves facial muscles, shoulder girdle and, in most severe cases, the lower limbs. Over 95% of patients have a contraction of D4Z4 repeated units on chromosome 4q35 that causes hypomethylation of the region and the overexpression of a toxic factor, DUX4. In about 5% of patients, called FSHD2, D4Z4 hypomethylation is due to mutations of SMCHD1, a gene involved in the inactivation of chromosome X during embryotic development. The aim of this study was to evaluate the clinical progression of FSHD over a 5-year follow-up using the Clinical Severity Score (CSS), a scale previously validated by the Italian Network for FSHD. Furthermore, considering the role of SMCHD1 in the epigenetic control of the chromosome X and the differences in severity between genders, the inactivation pattern of the chromosome X was analysed in order to test whether it could represent a genetic modifier of the disease. The sample consisted of 55 FSHD patients (29 males and 26 females) from the Neuromuscular Centre of Padova. All subjects carried a pathological D4Z4 fragment between 17-40 Kb. Patients who had been evaluated with CSS at T0 were re-evaluated after an average period of 5 years (T1). The score of each muscle region, the total CSS and the MRC score were recorded and compared between T0 and T1. Chromosome X inactivation was analysed in 38 FSHD1 and 4 FSHD2 females measuring the degree of methylation of the CAG repeated sequence of the androgen receptor gene. Genomic DNA was digested with methylation-sensitive restriction enzymes (HpAII and HhAI), amplified by PCR and finally genotyped. 48 healthy individuals were studied as controls and X inactivation patterns were correlated to muscle impairment measured by CSS. After 6 years of follow-up, the mean CSS difference between T0 and T1, and MRC score at biceps, triceps and tibialis anterior reached significance only in the probands group. There was no difference at shorter follow-up times or between relatives. Disease progression appeared independent of the size of D4Z4 fragment and no differences were found between genders. X-inactivation pattern was normally distributed in patients and controls. There was a moderate linear correlation between the percentage of X-inactivation and the severity of the shoulder girdle involvement, but not with any other muscle groups. In conclusion, FSHD symptoms progress slowly over time and, although CSS represents a valuable tool for patient assessment, it lacks sensitivity for the detection of subtle clinical modifications even over a five-year period. Therefore, its use in follow-up appears to be limited. Moreover, the X inactivation pattern in FSHD patients mirrors the normal distribution observed in healthy females and correlates only modestly with the severity of the disease. These latter findings suggest that different genetic regulators are involved in the full phenotypic expression of the disease, and make evaluation of potential therapies difficult. Part II. SHP2: a novel therapeutic target in MuSK-myasthenia. Muscle Specific Kinase antibody myasthenia gravis (MuSK-MG) is an autoimmune disease that impairs neuromuscular transmission leading to widespread muscle weakness and fatigability. Under physiological conditions, agrin activates the LRP4-MuSK complex, initiating a phosphorylation cascade that culminates with the clustering of acetylcholine receptors (AChRs). SH2 domain-containing phosphatase (SHP2) is a negative regulator of AChR clustering that inhibits MuSK phosphorylation. In MuSK-MG, (auto)antibodies against MuSK, mainly of the IgG4 subclass, block MuSK interaction with LRP4 and, therefore, its activation. The smaller population of MuSK IgG1-3 appear to act by a different mechanism. Although MuSK-MG is a treatable disease, a therapy that targets specifically its pathogenic mechanisms is still not available. The aim of this study was to confirm and extend preliminary findings that demonstrated the in vitro effects of a specific SHP2 inhibitor, NSC-87877, as a potential specific treatment for MuSK-MG. Total IgG and IgG subclasses (IgG1-3, IgG4) were purified from plasma of 3 MuSK-MG patients. MuSK-Ab titres were measured by radioimmunoassay. To test the effects of NSC-87877 on MuSK phosphorylation and AChR clustering, C2C12 myotubes were used. The myotubes were incubated with increasing concentrations of NSC-87877 at different time intervals (from 15 to 360 minutes) using agrin and DMEM as positive and negative controls respectively. To test whether the drug was able to reverse the pathogenic effects of MuSK-Abs, myotubes were then exposed to agrin either with MuSK total IgG, IgG1-3 or IgG4, in the presence or absence of NSC-87877. MuSK expression and tyrosine phosphorylation were detected by western blotting, and the phosphorylation expressed as the ratio of the densitometry values of phospho-tyrosine MuSK to total MuSK. For AChR cluster quantification, myotubes were labelled with α-bungataroxin-594 followed by image acquisition and analysis with ImageJ software. For all experiments 20 images were scanned, coded, and the number of clusters > 5 μm counted using ImageJ. In the absence of agrin, NSC-87877 caused a dose-dependent increase in both MuSK phosphorylation and AChR clustering, reaching a maximum at 100 μM, after 40 minutes of incubation. 100 μM NSC-87877 enhanced MuSK phosphorylation in the presence of MuSK total IgG and purified IgG4 while no significant change was observed with purified IgG1-3. Nevertheless, MuSK total IgG and both subclass fractions caused the dispersal of AChR clusters (see also Koneczny et al, 2013), and NSC-87877 was able to reverse their pathological effects in all samples. SHP2 inhibition by NSC-87877 induced MuSK phosphorylation and increased AChR clustering regardless of direct stimulation by agrin. Moreover, NSC-87877 effectively induced MuSK activation despite the inhibitory effects of MuSK IgG4 antibodies, and enhanced AChR clustering in the presence of all the different IgG subclasses. Therefore, irrespectively of the MuSK IgG subclass, SHP2 inhibition represents a potential therapeutic strategy in MuSK myasthenia and further studies should access its efficacy and reliability in vivo

Understanding Neuromuscular Health and Disease: Advances in Genetics, Omics, and Molecular Function

This compilation focuses on recent advances in the molecular and cellular understandingof neuromuscular biology, and the treatment of neuromuscular disease.These advances are at the forefront of modern molecular methodologies, oftenintegrating across wet-lab cell and tissue models, dry-lab computational approaches,and clinical studies. The continuing development and application ofmultiomics methods offer particular challenges and opportunities in the field,not least in the potential for personalized medicine

ChIP analysis of the histone modifications at the D4Z4 repeats in facioscapulohumeral muscular dystrophy (FSHD)

Genomic DNA must exist in a particular chromatin configuration and modification of this structure is essential for the correct control of gene expression. There are several human genetic disorders that are caused by misregulation of epigenetic gene control. Facioscapulohumeral muscular dystrophy (FSHD) is a disease that may be caused by alterations in chromatin structure. FSHD is the third most common form of muscular dystrophy. The majority of FSHD cases show contraction of the D4Z4 repeats on the 4q35 chromosome (FSHD1). However, a small number of FSHD cases show no contraction at this region (FSHD2), but share epigenetic changes at the D4Z4 region with the FSHD1 patients. In 2009, Zeng et al. reported a specific loss of H3K9me3 histone modification at the D4Z4 repeats in FSHD patients. The main focus of this study was to verify the published data by Zeng et al (2009) and further investigate the histone modification changes at the D4Z4 array. Chromatin immunoprecipitation (ChIP) coupled with real-time quantitative PCR (qPCR) was employed to investigate the histone modifications within the D4Z4 array. The results obtained were in agreement with the previously published data on the reduction of H3K9me3 histone modification at the D4Z4 repeats in FSHD patients. However, contradictory to the previous data, the reduction of this histone modification was also observed on other genomic regions. A global reduction of H3K27me3 was also observed in FSHD patients

Empathy and executive functions in neuromuscular diseases

Objective: the relationship between Executive Functions (EFs) and Empathy, in particular Theory of Mind (ToM), is well known, considering neuropsychological performances and the neural correlates underpinning each one. Evidences regarding deficits on ToM skills related to impairment in EFs are nowadays well known in Amyotrophic Lateral Sclerosis (ALS) and Myothonic dystrophy (DM1). The aim of the present study was the evaluation of ToM and its association with EFs in four neuromuscular diseases in which EFs can be impaired, and so: Duchenne muscular dystrophy (DMD), Becker muscular dystrophy (BMD), Facioscapulohumeral dystrophy (FSHD) and X-linked Spinal and Bulbar Muscular Atrophy (X-SBMA). Moreover, also abilities related to Affective Empathy have been investigated. Methods: the study involved 25 DMD patients, 21 BMD patients, 21 FSHD patients, 64 X-SBMA patients and 145 healthy controls matched for age, education and gender. Both patients and controls completed tests and questionnaires with the aim to assess EFs, ToM and Affective Empathy abilities. Results: deficits in EFs have been found only in DMD patients. EFs impairment resulted to be associated with deficits in ToM ability but also with problems related to Affective Empathy. Conclusions: DMD patients without mental retardation manifest problems on EFs. These deficits are related to impairments in ToM abilities and Affective Empathy skills. These results can give an important contribution to clinical practice: know the Empathic abilities of patients may be useful to clinicians in order to create a good alliance which is extremely useful in the compliance of treatments and therapeutic choices

The use of new technology to improve genetic testing

Molecular confirmation of a clinical diagnosis of an inherited disease or of congenital malformations is of paramount importance for patients and their families. It is the conclusion of the differential diagnostic process, and provides information on the prognosis, in some cases on the therapeutic options, and on the recurrence risk. Currently, targeted sequencing of gene (s) of interest is the preferred approach for searching for small pathogenic mutations. The work presented in this thesis describes the application of new techniques for detecting small variations (mutations) in genomic DNA that underlie various disorders. These techniques include High Resolution Melting Curve Analysis (HR-MCA) followed by Sanger sequencing, targeted, X-exome and whole exome capture followed by Next Generation Sequencing (NGS). We have optimized, tested and applied the different new molecular techniques mentioned above to 1) faci litate the detection of disease causing mutations in several disorders with suspected Mendelian inheritance, 2) to speed up the identification of disease genes, 3) to provide a systematic tool for classifying previously intractable genetic diseasesUBL - phd migration 201