Sagittal kinematics and imbalance of the spine and whole body during walking in late-onset Pompe disease.

Late-onset Pompe disease (LOPD) is characterized by postural abnormalities mainly due to involvement of paraspinal lumbar and abdominal-pelvic muscles. Previous studies quantitatively analyzed static upright posture, spatial-temporal parameters, and kinematics of the lower limbs and trunk, considered as single bone segment. Sagittal plane analysis of the spine and whole body during walking has never been investigated in patients with LOPD. The aim of the study was to evaluate sagittal kinematics and imbalance of the spine and whole body in patients with LOPD by three-dimensional (3-D)-motion analysis using an appropriate marker set protocol and introducing innovative kinematic parameters. Seven siblings with LOPD were assessed by 3-D-stereophotogrammetry using the DB-total protocol, which allows to analyze sagittal alignment of whole body. Fourteen age- and sex-matched healthy subjects were used as controls. LOPD group showed a flattening of the spinal curvatures, with a head and neck posteriorization with respect to sacrum, a significant increase of concavity in Heel-S2-Nasion/C7 angles, a rear-position of upper limbs with respect to pelvis, a shorter pendular activity, and a trend of elbow extension during ambulation. Moreover, a significant increase of excursion range in most of sagittal parameters was found. The present study highlighted a specific pathological postural pattern, resembling "man falling backwards," which reveals a biomechanical compensation strategy of patients with LOPD to maintain the balance against the instability of the spinopelvic region, kinematically verified by increase of the excursion ranges. DB-total kinematic parameters might be useful for functional evaluation and for monitoring response to enzyme replacement therapy, rehabilitation project, and disease progression.NEW & NOTEWORTHY This study is the first to quantitatively characterize the sagittal spine and whole body posture of patients with late-onset Pompe disease during walking, showing a pathological kinematic pattern defined "man falling backwards." 3-D-motion analysis, with a specific marker set (DB-total protocol) introducing new whole body kinematic parameters, may be useful for accurate functional evaluation and monitoring this rare disease.

Clinical, Genetic, and Histological Characterization of Patients with Rare Neuromuscular and Mitochondrial Diseases Presenting with Different Cardiomyopathy Phenotypes.

Cardiomyopathies are mostly determined by genetic mutations affecting either cardiac muscle cell structure or function. Nevertheless, cardiomyopathies may also be part of complex clinical phenotypes in the spectrum of neuromuscular (NMD) or mitochondrial diseases (MD). The aim of this study is to describe the clinical, molecular, and histological characteristics of a consecutive cohort of patients with cardiomyopathy associated with NMDs or MDs referred to a tertiary cardiomyopathy clinic. Consecutive patients with a definitive diagnosis of NMDs and MDs presenting with a cardiomyopathy phenotype were described. Seven patients were identified: two patients with ACAD9 deficiency (Patient 1 carried the c.1240C>T (p.Arg414Cys) homozygous variant in ACAD9; Patient 2 carried the c.1240C>T (p.Arg414Cys) and the c.1646G>A (p.Ar549Gln) variants in ACAD9); two patients with MYH7-related myopathy (Patient 3 carried the c.1325G>A (p.Arg442His) variant in MYH7; Patient 4 carried the c.1357C>T (p.Arg453Cys) variant in MYH7); one patient with desminopathy (Patient 5 carried the c.46C>T (p.Arg16Cys) variant in DES); two patients with mitochondrial myopathy (Patient 6 carried the m.3243A>G variant in MT-TL1; Patient 7 carried the c.253G>A (p.Gly85Arg) and the c.1055C>T (p.Thr352Met) variants in MTO1). All patients underwent a comprehensive cardiovascular and neuromuscular evaluation, including muscle biopsy and genetic testing. This study described the clinical phenotype of rare NMDs and MDs presenting as cardiomyopathies. A multidisciplinary evaluation, combined with genetic testing, plays a main role in the diagnosis of these rare diseases, and provides information about clinical expectations, and guides management.

Using Cluster Analysis to Overcome the Limits of Traditional Phenotype-Genotype Correlations: The Example of RYR1-Related Myopathies.

Thanks to advances in gene sequencing, RYR1-related myopathy (RYR1-RM) is now known to manifest itself in vastly heterogeneous forms, whose clinical interpretation is, therefore, highly challenging. We set out to develop a novel unsupervised cluster analysis method in a large patient population. The objective was to analyze the main RYR1-related characteristics to identify distinctive features of RYR1-RM and, thus, offer more precise genotype-phenotype correlations in a group of potentially life-threatening disorders. We studied 600 patients presenting with a suspicion of inherited myopathy, who were investigated using next-generation sequencing. Among them, 73 index cases harbored variants in RYR1. In an attempt to group genetic variants and fully exploit information derived from genetic, morphological, and clinical datasets, we performed unsupervised cluster analysis in 64 probands carrying monoallelic variants. Most of the 73 patients with positive molecular diagnoses were clinically asymptomatic or pauci-symptomatic. Multimodal integration of clinical and histological data, performed using a non-metric multi-dimensional scaling analysis with k-means clustering, grouped the 64 patients into 4 clusters with distinctive patterns of clinical and morphological findings. In addressing the need for more specific genotype-phenotype correlations, we found clustering to overcome the limits of the "single-dimension" paradigm traditionally used to describe genotype-phenotype relationships.

Spinal Nerve Roots Abnormalities on MRI in a Child with SURF1 Mitochondrial Disease.

Variants in SURF1, encoding an assembly factor of mitochondrial respiratory chain complex IV, cause Leigh syndrome (LS) and Charcot-Marie-Tooth type 4K in children and young adolescents. Magnetic resonance imaging (MRI) appearance of enlarged nerve roots with postcontrastographic enhancement is a distinctive feature of hypertrophic neuropathy caused by onion-bulb formation and it has rarely been described in mitochondrial diseases (MDs). Spinal nerve roots abnormalities on MRI are novel findings in LS associated with variants in SURF1. Here we report detailed neuroradiological and neurophysiologic findings in a child with LS and demyelinating neuropathy SURF1-related. Our case underlines the potential contributive role of spinal neuroimaging together with neurophysiological examination to identify the full spectrum of patterns in MDs. It remains to elucidate if these observations remain peculiar of SURF1 variants or potentially detectable in other MDs with peripheral nervous system involvement.

Atypical Clinical and Neuroradiological Findings in a Child With Bifacial Weakness With Paresthesias.

BACKGROUND: Guillain-Barré syndrome (GBS) is the broad term used to describe a number of related acute autoimmune neuropathies, which together form a continuous spectrum of variable and overlapping syndromes. Bifacial weakness with paresthesias (BFP) is a rare variant of GBS, characterized by isolated facial diplegia in the absence of ophthalmoplegia, ataxia, or limb weakness, and it is usually associated with distal limb paresthesias. CASE DESCRIPTION: An 8-year-old boy was brought to our attention; because 5 days before coming to the hospital, he noticed he could no longer smile. Bilateral facial droop and inability to close both eyes were evident along with slight paresthesias at the hands and feet and gait disturbances. He progressively developed hypophonia, dysarthria, dysphagia associated with dysmetria, and limb ataxia. Nerve conduction studies showed a demyelinating polyneuropathy. Brain and spine magnetic resonance imaging (MRI) revealed contrast enhancement of both facial nerves and cauda equina nerve roots along with a hyperintense signal of the periaqueductal gray matter, superior cerebellar peduncles, and pontine tegmentum. Because BFP is not typically associated with other cranial neuropathies or ataxia, these clinical features along with peculiar MRI findings supported the diagnosis of "BFP plus." Finally, it can be speculated that this case configures a rare overlap between BFP and the other GBS variants, such as Bickerstaff encephalitis. CONCLUSIONS: This atypical case underlines the potential role of MRI in contributing to refining the nosological classification of GBS spectrum and optimizing individual treatment, especially in children where unusual manifestations are not infrequent and neurological examination is more challenging.

The Diagnostic Approach to Mitochondrial Disorders in Children in the Era of Next-Generation Sequencing: A 4-Year Cohort Study.

Mitochondrial diseases (MDs) are a large group of genetically determined multisystem disorders, characterized by extreme phenotypic heterogeneity, attributable in part to the dual genomic control (nuclear and mitochondrial DNA) of the mitochondrial proteome. Advances in next-generation sequencing technologies over the past two decades have presented clinicians with a challenge: to select the candidate disease-causing variants among the huge number of data provided. Unfortunately, the clinical tools available to support genetic interpretations still lack specificity and sensitivity. For this reason, the diagnosis of MDs continues to be difficult, with the new "genotype first" approach still failing to diagnose a large group of patients. With the aim of investigating possible relationships between clinical and/or biochemical phenotypes and definitive molecular diagnoses, we performed a retrospective multicenter study of 111 pediatric patients with clinical suspicion of MD. In this cohort, the strongest predictor of a molecular (in particular an mtDNA-related) diagnosis of MD was neuroimaging evidence of basal ganglia (BG) involvement. Regression analysis confirmed that normal BG imaging predicted negative genetic studies for MD. Psychomotor regression was confirmed as an independent predictor of a definitive diagnosis of MD. The findings of this study corroborate previous data supporting a role for neuroimaging in the diagnostic approach to MDs and reinforce the idea that mtDNA sequencing should be considered for first-line testing, at least in specific groups of children.

Rare Variants in Autophagy and Non-Autophagy Genes in Late-Onset Pompe Disease: Suggestions of Their Disease-Modifying Role in Two Italian Families.

Pompe disease is an autosomal recessive disorder caused by a deficiency in the enzyme acid alpha-glucosidase. The late-onset form of Pompe disease (LOPD) is characterized by a slowly progressing proximal muscle weakness, often involving respiratory muscles. In LOPD, the levels of GAA enzyme activity and the severity of the clinical pictures may be highly variable among individuals, even in those who harbour the same combination of GAA mutations. The result is an unpredictable genotype-phenotype correlation. The purpose of this study was to identify the genetic factors responsible for the progression, severity and drug response in LOPD. We report here on a detailed clinical, morphological and genetic study, including a whole exome sequencing (WES) analysis of 11 adult LOPD siblings belonging to two Italian families carrying compound heterozygous GAA mutations. We disclosed a heterogeneous pattern of myopathic impairment, associated, among others, with cardiac defects, intracranial vessels abnormality, osteoporosis, vitamin D deficiency, obesity and adverse response to enzyme replacement therapy (ERT). We identified deleterious variants in the genes involved in autophagy, immunity and bone metabolism, which contributed to the severity of the clinical symptoms observed in the LOPD patients. This study emphasizes the multisystem nature of LOPD and highlights the polygenic nature of the complex phenotype disclosed in these patients.

Clinical and Genetic Heterogeneity in a Large Family with Pseudoxanthoma Elasticum: MTHFR and SERPINE1 Variants as Possible Disease Modifiers in Developing Ischemic Stroke.

BACKGROUND AND OBJECTIVES: Pseudoxanthoma elasticum (PXE) is a rare autosomal recessive disorder caused by pathogenic variants in the ABCC6 gene. The phenotypic spectrum of PXE is highly variable and includes principally three major features: skin lesions, eye and vascular manifestations, while brain manifestations are less common. To date about 400 different PXE associated variants in ABCC6 gene are described without any evident genotype-phenotype correlation. Herein, we report the clinical and molecular findings of a large PXE family with clinical and genetic intra-familial variability with significant cerebrovascular involvement. METHODS: The analysis of the ABCC6 gene was performed in the proband and her familiars for the definition of genetic background. Then, in order to determine why some affected individuals had more prominent brain involvement, we investigated classic thrombophilic gene variants. RESULTS: Molecular findings disclosed two different ABCC6 mutations, i.e., the recurrent p.(Arg518Gln) and the novel p.(Val1285Met) missense substitution responsible of a pseudo-dominant inheritance. The study of thrombophilic gene variants revealed the presence of 4G/4G SERPINE1 genotype in the proband and in her father, which both developed ischemic stroke. The proband carried also the C677T variant the MTHFR gene. CONCLUSION: We argue, for the first time, that the 4G/4G SERPINE1 genotype could represent an additional risk factor in PXE for developing ischemic stroke, which adds up to the already known predisposing conditions. Therapeutic implications are discussed, we also advise that PXE patients should be adequately screened for cerebral vasculopathy, even more if familial history is suggestive of brain complications.

Neuroacanthocytosis Syndromes in an Italian Cohort: Clinical Spectrum, High Genetic Variability and Muscle Involvement.

Neuroacanthocytosis (NA) syndromes are a group of genetically defined diseases characterized by the association of red blood cell acanthocytosis, progressive degeneration of the basal ganglia and neuromuscular features with characteristic persistent hyperCKemia. The main NA syndromes include autosomal recessive chorea-acanthocytosis (ChAc) and X-linked McLeod syndrome (MLS). A series of Italian patients selected through a multicenter study for these specific neurological phenotypes underwent DNA sequencing of the VPS13A and XK genes to search for causative mutations. Where it has been possible, muscle biopsies were obtained and thoroughly investigated with histochemical assays. A total of nine patients from five different families were diagnosed with ChAC and had mostly biallelic changes in the VPS13A gene (three nonsense, two frameshift, three splicing), while three patients from a single X-linked family were diagnosed with McLeod syndrome and had a deletion in the XK gene. Despite a very low incidence (only one thousand cases of ChAc and a few hundred MLS cases reported worldwide), none of the 8 VPS13A variants identified in our patients is shared by two families, suggesting the high genetic variability of ChAc in the Italian population. In our series, in line with epidemiological data, McLeod syndrome occurs less frequently than ChAc, although it can be easily suspected because of its X-linked mode of inheritance. Finally, histochemical studies strongly suggest that muscle pathology is not simply secondary to the axonal neuropathy, frequently seen in these patients, but primary myopathic alterations can be detected in both NA syndromes.

Novel autophagic vacuolar myopathies: Phenotype and genotype features.

BACKGROUND: Autophagic vacuolar myopathies (AVMs) are an emerging group of heterogeneous myopathies sharing histopathological features on muscle pathology, in which autophagic vacuoles are the pathognomonic morphologic hallmarks. Glycogen storage disease type II (GSDII) caused by lysosomal acid α-glucosidase (GAA) deficiency is the best-characterised AVM. AIMS: This study aimed to investigate the mutational profiling of seven neuromuscular outpatients sharing clinical, myopathological and biochemical findings with AVMs. METHODS: We applied a diagnostic protocol, recently published by our research group for suspected late-onset GSDII (LO-GSDII), including counting PAS-positive lymphocytes on blood smears, dried blood spot (DBS)-GAA, muscle biopsy histological and immunofluorescence studies, GAA activity assay and expression studies on muscle homogenate, GAA sequencing, GAA multiplex ligation-dependent probe amplification (MLPA) and whole exome sequencing (WES). RESULTS: The patients had a limb girdle-like muscular pattern with persistent hyperCKaemia; vacuolated PAS-positive lymphocytes, glycogen accumulation and impaired autophagy at muscle biopsy. Decreased GAA activity was also measured. While GAA sequencing identified no pathogenic mutations, WES approach allowed us to identify for each patient an unexpected mutational pattern in genes cooperating in lysosomal-autophagic machinery, some of which have never been linked to human diseases. CONCLUSIONS: Our data suggest that reduced GAA activity may occur in any condition of impaired autophagy and that WES approach is advisable in all genetically undefined cases of autophagic myopathy. Therefore, deficiency of GAA activity and PAS-positive lymphocytes should be considered as AVM markers together with LC3/p62-positive autophagic vacuoles.

"Borderline" idiopathic CD4+ T-cell lymphocytopenia presenting with atypical progressive multifocal leukoencephalopathy.

Idiopathic CD4+ lymphocytopenia (ICL) is a rare disorder characterized by low counts of CD4+ cells (<300/mm3) in absence of other known causes of immunosuppression. A few cases of progressive multifocal leukoencephalopathy (PML) were reported in association with ICL with variable outcome. We describe the case of a 40 year-old man diagnosed with PML, which showed a monophasic course. Causes of primary and secondary immunodeficiency were ruled out, only a "borderline" ICL was found. This case highlights that a severe immunodepression could not be an absolute prerequisite in developing PML and also points the attention on current definition of ICL.

Expanding the spectrum of SPTLC1-related disorders beyond hereditary sensory and autonomic neuropathies: A novel case of the distinct "S331 syndrome".

Hereditary sensory and autonomic neuropathies (HSAN) encompass a group of peripheral nervous system disorders characterized by remarkable heterogeneity from a clinical and genetic point of view. Mutations in SPTLC1 gene are responsible for HSAN type IA, which usually starts from the second to fourth decade with axonal neuropathy, sensory loss, painless distal ulcerations, and mild autonomic features, while motor involvement usually occur later as disease progresses. Beyond the classic presentation of HSAN type IA, an exceedingly rare distinct phenotype related to SPTLC1 mutations at residue serine 331 (S331) has recently been reported, characterized by earlier onset, prominent muscular atrophy, growth retardation, oculo-skeletal abnormalities, and possible respiratory complications. In this report, we describe clinical, instrumental, and genetic aspects of a 13-year-old Sri Lankan male carrying the rare de novo p.S331Y heterozygous mutation in SPTLC1 gene found by whole exome sequencing. Patient's phenotype partly overlaps with the first case previously reported, however with some additional features not described before. This work represent the second report about this rare mutation and our findings strongly reinforce the hypothesis of a clearly distinct "S331 syndrome", thus expanding the spectrum of SPTLC1-related disorders.

Atypical progressive multifocal leukoencephalopathy in a kidney transplant recipient with improving symptoms after immunocompetence recovery.

Progressive multifocal leukoencephalopathy (PML) is a viral infection due to John Cunningham Virus (JCV) resulting in progressive damage of brain white matter, mostly related to HIV infection or hemato-oncological malignancies. PML onset is usually multifocal with rapid neurological progression and poor prognosis. Here we report an atypical case of PML with monofocal onset and a good outcome in a 64-year-old man who received a kidney transplant for end-stage renal disease (ESRD). The applied antirejection immunosuppressive drug regimen included tacrolimus, prednisone and mycophenolic acid. Three years after the transplant, he complained of right-hand tremor and rapidly progressive right hemiparesis, with prominent involvement of the upper limb. Brain magnetic resonance imaging (MRI) showed a significant demyelinating area in the left frontal lobe, without mass effect and contrast enhancement. Real-time PCR analysis revealed the presence of JCV on cerebrospinal fluid. Consequent immunosuppressive drug suspension resulted in a global improvement of neurological symptoms and a favourable evolution of the neuroradiological findings. Subsequent eight-year follow-up MRI confirmed the stability of imaging findings over time. Therefore, early recognition of PML symptoms and MRI sign along with the rapid suspension of immunosuppressive drugs can modify the natural history of this disease after a kidney transplant.

A novel missense mutation in CAV3 gene in an Italian family with persistent hyperCKemia, myalgia and hypercholesterolemia: Double-trouble.

Caveolins are essential proteins in caveolae architecture, small plasma membrane invaginations that play a key role in a variety of cellular processes, including vesicular trafficking and signal transduction. Mutations in the gene encoding caveolin-3 (CAV3) cause a broad spectrum of clinical phenotypes, ranging from isolated hyperCKemia to most severe limb girdle muscular dystrophy and cardiomyopathy. We report a novel heterozygous p.Val44Met (c.130G > A) CAV3 mutation in two brothers presenting with persistent elevation of serum creatine kinase, myalgia and hypercholesterolemia. Immunofluorescence study with anticaveolin-3 antibodies on muscle biopsy of the proband confirmed a reduced immuno-reactivity of caveolin-3 on the sarcolemma. This findings support the pathogenic effect of this novel mutation and extend the genotypic and clinical spectrum of Caveolinopathies. Finally, we discuss the hypothesis that the association between CAV3 mutations and hypercholesterolemia may not be coincidental.

Intrafamilial "DOA-plus" phenotype variability related to different OMI/HTRA2 expression.

Dominant Optic Atrophy and Deafness (DOAD) may be associated with one or more of the following disorders such as myopathy, progressive external ophthalmoplegia, peripheral neuropathy, and cerebellar atrophy ("DOA-plus"). Intra- and interfamilial variability of the "DOA-plus" phenotype is frequently observed in the majority of the patients carrying the same mutation in the OPA1 gene. We are describing two familial cases of "DOA-plus" carrying the same c.1334G>A (p.Arg445His) mutation in OPA1 and disclosing different clinical, pathological and biochemical features. The two patients showed different expression levels of the mitochondrial OMI/HTRA2 molecule, which acts as a mitochondrial stress sensor and has been described to interplay with OPA1 in in vitro studies. Our data offer the cue to inquire the role of OMI/HTRA2 as a modifier gene in determining the "DOAplus" phenotype variability.