Genetic and pathophysiological insights from autopsied patient with primary familial brain calcification: novel MYORG variants and astrocytic implications.

Primary familial brain calcification (PFBC) is a genetic neurological disorder characterized by symmetric brain calcifications that manifest with variable neurological symptoms. This study aimed to explore the genetic basis of PFBC and elucidate the underlying pathophysiological mechanisms. Six patients from four pedigrees with brain calcification were enrolled. Whole-exome sequencing identified two novel homozygous variants, c.488G > T (p.W163L) and c.2135G > A (p.W712*), within the myogenesis regulating glycosidase (MYORG) gene. Cerebellar ataxia (n = 5) and pyramidal signs (n = 4) were predominant symptoms, with significant clinical heterogeneity noted even within the same family. An autopsy of one patient revealed extensive brainstem calcifications, sparing the cerebral cortex, and marked by calcifications predominantly in capillaries and arterioles. The pathological study suggested morphological alterations characterized by shortened foot processes within astrocytes in regions with pronounced calcification and decreased immunoreactivity of AQP4. The morphology of astrocytes in regions without calcification remains preserved. Neuronal loss and gliosis were observed in the basal ganglia, thalamus, brainstem, cerebellum, and dentate nucleus. Notably, olivary hypertrophy, a previously undescribed feature in MYORG-PFBC, was discovered. Neuroimaging showed reduced blood flow in the cerebellum, highlighting the extent of cerebellar involvement. Among perivascular cells constituting the blood-brain barrier (BBB) and neurovascular unit, MYORG is most highly expressed in astrocytes. Astrocytes are integral components of the BBB, and their dysfunction can precipitate BBB disruption, potentially leading to brain calcification and subsequent neuronal loss. This study presents two novel homozygous variants in the MYORG gene and highlights the pivotal role of astrocytes in the development of brain calcifications, providing insights into the pathophysiological mechanisms underlying PFBC associated with MYORG variants.

Microbleed clustering in thalamus sign in CADASIL patients with NOTCH3 R75P mutation.

Background and objective: Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is an inherited cerebral microvascular disease characterized by the development of vascular dementia and lacunar infarctions. This study aimed to identify the genetic and clinical features of CADASIL in Japan. Methods: We conducted genetic analysis on a case series of patients clinically diagnosed with CADASIL. Clinical and imaging analyses were performed on 32 patients with pathogenic mutations in the NOTCH3 gene. To assess the presence of cerebral microbleeds (CMBs), we utilized several established rating scales including the Fazekas scale, Scheltens rating scale, and Microbleed Anatomical Rating Scale, based on brain MRI images. Results: Among the 32 CADASIL patients, 24 cases were found carrying the R75P mutation in NOTCH3, whereas the remaining eight cases had other NOTCH3 mutations (R75Q, R110C, C134F, C144F, R169C, and R607C). The haplotype analysis of the R75P mutation uncovered the presence of a founder effect. A brain MRI analysis revealed that cases with the R75P mutation had a significantly higher total number of CMBs, particularly in the thalamus when compared to patients with other NOTCH3 mutations. Among 15 out of 24 cases with the R75P mutation, we observed a notable clustering of CMBs in the thalamus, termed microbleed clustering in thalamus sign (MCT sign). Conclusion: We propose that the MCT sign observed in NOTCH3 R75P-related CADASIL patients may serve as a potentially characteristic imaging feature. This finding offers further insights into the interactions between genotypes and phenotypes between NOTCH3 and CADASIL.

The natural history of hereditary motor and sensory neuropathy with proximal dominant involvement (HMSN-P) in 97 Japanese patients.

Hereditary motor and sensory neuropathy with proximal dominant involvement (HMSN-P) is a motor and sensory neuronopathy with autosomal dominant inheritance, adult onset, slowly progressive course, and is associated with TRK-fused gene (TFG) mutation. At advanced stages, respiratory failure and dysphagia becomes life-threatoning, and patients typically die by their 70s. Although there is currently no evidence for effective treatment, a therapy may be found by elucidation of the function of TFG. Recently its pathomechanism has been proposed to be associated with abnormalities in protein transfer from the endoplasmic reticulum. Such pathomechanisms might involve a similar process in amyotrophic lateral sclerosis; thus, its pathomechanisms and treatment strategy might make it a good model for neurodegenerative disorders. It is of great value to clarify the natural history of HMSN-P, in oder to judge the treatment effect. By evaluating 97 patients (79 out of 97 were examined and all confirmed with p.Pro 285 Leu mutation) in this study, it was confirmed that this disease follows a uniform course in the earlier stages, and there are individual differences in the onset between 20 and 30 years. Such uniformity might be due to the proposed single gene abnormality. At advanced stages, there are larger individual differences in the progression, but the reasons for these are unknown. Longer survival might be achieved with a better care for respiratory failure and dysphagia if such cares were undertaken at appropriate times.

[Clinical manifestations of 5 patients with idiopathic paroxysmal kinesigenic choreoathetosis].

Paroxysmal kinesigenic choreoathetosis (PKC) is a rare disorder characterized by recurrent and brief attacks of choreoathetoid and/or dystonic movements in trunk and limbs triggered by initiation of voluntary movement. Of 5 patients with idiopathic PKC in our hospital, four were men and one was with family history. Age of onset ranged from 8 to 15 years old. They were consistent with previous reports in the characteristics of involuntary movements, normal neurological findings, normal laboratory data, no abnormal findings of standard imaging studies, and good restraining effects on attacks with carbamazepine. Individual body parts where attacks often involved were different among 5 patients. Although previous reports which said the prognosis and outcome of PKC were good, neuropsychological examinations in our study revealed that 2 patients out of 5 had certain cortical dysfunction, one patient was with progressive deterioration, and the other was with underlying mild abnormalities. Detailed and serial neuropsychological examinations might be necessary for some PKC patients.

Three-tesla magnetic resonance neurography of the brachial plexus in cervical radiculopathy.

INTRODUCTION: There have been no reports of the use of 3-Tesla magnetic resonance neurography (3T MRN) to characterize cervical radiculopathy. In particular, there are no reports of MRN of brachial plexus involvement in patients with cervical radiculopathy. METHODS: We reviewed retrospectively 12 consecutive patients with cervical radiculopathy who underwent 3T MRN. RESULTS: The median age was 54.5 years. Eleven of 12 patients were men. The distribution of nerve-root signal abnormality was correlated with intervertebral foraminal stenosis and the presence of muscles that exhibited weakness and/or signs of denervation on electromyography. MRN abnormalities were found to extend into the distal part of the brachial plexus in 10 patients. CONCLUSION: This study demonstrates that MRN is potentially useful for diagnosis in patients with suspected cervical radiculopathy. Moreover, the finding of brachial plexus involvement on MRN may indicate a possible pathophysiological relationship between cervical radiculopathy and brachial plexopathy.

Persistent hiccups and vomiting with multiple cranial nerve palsy in a case of zoster sine herpete.

A 76-year-old man came to our hospital complaining of hiccups and vomiting lasting for five days. A neurological examination showed dysfunction of cranial nerves V, VII, VIII, IX and X on the left side. Cerebrospinal fluid polymerase chain reaction for varicella zoster virus-DNA was positive. The patient responded well to treatment with intravenous acyclovir and steroids. To the best of our knowledge, this is the first case report of zoster sine herpete presenting with persistent hiccups and vomiting. It is important to keep in mind that herpes zoster can present with symptoms that closely resemble those of intractable hiccups and nausea of neuromyelitis optica. Early detection of the virus is critical for making appropriate treatment decisions.

[Autopsy case of a patient with Charcot-Marie-Tooth disease type 1A and suspected chronic inflammatory demyelinating polyradiculoneuropathy, which was later diagnosed as amyotrophic lateral sclerosis].

We report an autopsy case of a 74-year-old man with late onset Charcot-Marie-Tooth disease type 1A (CMT1A) diagnosed by genetic screening, later associated with amyotrophic lateral sclerosis (ALS). At the age of 70 years, the patient was admitted to our hospital because of progressive weakness and dysesthesia in the right upper limb. In the early stages of the illness, he was diagnosed with chronic inflammatory demyelinating polyradiculoneuropathy (CIDP), and transient improvement was achieved with intravenous immunoglobulin. However, the symptoms progressively worsened and became refractory. Gene analysis revealed PMP22 gene duplication, which confirmed CMT1A. On sural nerve biopsy, severe demyelinating neuropathy and abundant onion-bulb formations with endoneurial infiltration of inflammatory cells were observed. Thereafter, pseudo-bulbar palsy and respiratory muscle weakness developed insidiously and progressed rapidly along with muscle weakness in the limbs and trunk. The patient died about four years after the onset of this disease. Postmortem examination showed moderate neuronal cell loss, Bunina bodies, and TDP-43-positive inclusions in the anterior horn cells. The spinal cord revealed axonal loss and extensive macrophage permeation in the corticospinal tracts. On the basis of these findings, the final neuropathological diagnosis was ALS. This is the first report of an autopsy case of CMT1A complicated with ALS. We here discuss the significant clinical and neuropathological findings of this case.

Clinical and genetic characterization of 16q-linked autosomal dominant spinocerebellar ataxia in South Kyushu, Japan.

16q-ADCA (OMIM no. 117210) is an autosomal dominant spinocerebellar ataxia (AD-SCA) characterized by late-onset pure cerebellar ataxia and -16C>T substitution of the puratrophin-1 gene. Recently, a series of single-nucleotide polymorphisms (haplotype block) were found to be specific to 16q-ADCA. We screened patients with ataxia and found 62 patients, including four homozygotes who carry the C-T substitution of the puratrophin-1 gene. By further analysis of the patients with the haplotype block, we observed a single-founder effect for 16q-ADCA, even in patients who are supposed to be sporadic late cortical cerebellar atrophy (LCCA). We also observed slippage mutations of microsatellite markers, GATA01 and 17 msm, in the pedigrees. We compared the clinical course of 16q-ADCA in heterozygotes and homozygotes with the haplotype block and observed no apparent gene dosage effect. 16q-ADCA accounts for 27% of AD-SCAs and is the most frequent AD-SCA in South Kyushu, Japan.

Molecular mechanism of rigid spine with muscular dystrophy type 1 caused by novel mutations of selenoprotein N gene.

Mutations of selenoprotein N, 1 gene (SEPN1) cause rigid spine with muscular dystrophy type 1 (RSMD1), multiminicore disease, and desmin-related myopathy. We found two novel SEPN1 mutations in two Japanese patients with RSMD1. To clarify the pathomechanism of RSMD1, we performed immunohistochemical studies using a newly developed antibody for selenoprotein N. Selenoprotein N was diffusely distributed in the cytoplasm of the control muscle, but was reduced and irregularly expressed in the cytoplasm of a patient with RSMD1. The expression pattern was very similar to that of calnexin, a transmembrane protein of the endoplasmic reticulum. Selenoprotein N seems to be an endoplasmic reticulum glycoprotein, and loss of this protein leads to disturbance of muscular function. One of the families had the SEPN1 homozygous mutation in the initiation codon 1_2 ins T in exon 1 and showed truncated protein expression. The other had a homozygous 20-base duplication mutation at 80 (80_99dup, frameshift at R27) which, in theory, should generate many nonsense mutations including TGA. These nonsense mutations are premature translation termination codons and they degrade immediately by the process of nonsense-mediated decay (NMD). However, truncated selenoprotein N was also expressed. A possible mechanism behind this observation is that SEPN1 mRNAs may be resistant to NMD. We report on the possible molecular mechanism behind these mutations in SEPN1. Our study clarifies molecular mechanisms of this muscular disorder.

Abnormal expression of proteoglycans in Ullrich's disease with collagen VI deficiency.

Patients with Ullrich's disease have generalized muscle weakness, multiple contractures of the proximal joints, and hyperextensibility of the distal joints. Recently we found a marked reduction of fibronectin receptors in the skin and cultured fibroblasts of two patients with Ullrich's disease with collagen VI deficiency, and speculated that an abnormality of cell adhesion may be involved in the pathogenesis of the disease. In this study, we investigated the expression of proteoglycans and adhesion molecules in Ullrich's disease and other muscle diseases. We found a reduction of NG2 proteoglycan in the membrane of skeletal muscle but not in the skin in Ullrich's disease. By contrast, we found the upregulation of tenascin C in the extracellular matrix of skeletal muscle in Ullrich's disease. Our findings suggest that abnormal expression of proteoglycans and adhesion molecules may be involved in the pathogenesis of the dystrophic muscle changes in Ullrich's disease.

Pathological characteristics of skeletal muscle in Ullrich's disease with collagen VI deficiency.

Patients with Ullrich's disease have generalized muscle weakness, multiple contractures of the proximal joints and hyperextensibility of the distal joints. Recently, we found a deficiency of collagen VI protein in skeletal muscle from two patients with Ullrich's disease. In this study, we investigated immunohistochemically the expression of extracellular matrix proteins and various proteins, which are markers for regenerating muscle fibers. Although we have detected the reduction of collagen VI in Ullrich's disease with the two kinds of monoclonal antibodies for the different domains of collagen VI, the remaining immunoreactive material was different between them. This might suggest the presence of incomplete collagen VI protein in the muscle fibers. Furthermore, we found that very small muscle fibers in the patients with Ullrich's disease showed marked expression of desmin, neural cell adhesion molecule and neonatal myosin heavy chain, which is a characteristic finding of regenerating fibers, however, they showed poor expression of developmental myosin heavy chain and thrombomodulin. The present findings suggest that abnormal regeneration or maturation processes are involved in the pathogenesis of dystrophic muscle changes at least in the advanced stage of Ullrich's disease.

Capillary changes in skeletal muscle of patients with Ullrich's disease with collagen VI deficiency.

We examined the capillaries in muscle biopsy specimens from two patients with Ullrich's disease with collagen VI deficiency by light and electron microscopy. Collagen VI plays an important role in platelet aggregation for binding von Willebrand factor. Using immunohistochemistry, collagen VI was shown to be absent on capillaries from patients with Ullrich's disease, while von Willebrand factor, collagen IV, and vascular endothelial growth factor were normally expressed. Electron microscopy revealed narrow lumens, large nuclei in endothelial cells, and fenestration of a capillary. The number of pinocytotic vesicles per unit endothelial cytoplasm was increased. The cytoplasm of endothelial cells was strongly stained with uranyl acetate and lead citrate. Replication of the capillary basement membrane was observed. On the other hand, easy bleeding and coagulation were not observed in the two patients. These findings suggested that the collagen VI deficiency might have caused the electron microscopic changes of capillaries, while the function of the capillaries is apparently retained.

Miyoshi myopathy patients with novel 5' splicing donor site mutations showed different dysferlin immunostaining at the sarcolemma.

We analyzed five clinically defined cases of Miyoshi myopathy both genetically and immunologically. Western blot of muscle specimens confirmed that all of these patients had dysferlin deficiency. Immunohistochemistry revealed that two of the five patients showed positive dysferlin immunostaining. Subsequent mutation analysis of the dysferlin gene in these two patients revealed that both had novel 5' splicing donor site mutations. One patient with a homozygous G to C substitution at nucleotide 1036+1 exon 6 splicing donor site showed patchy sarcolemmal dysferlin immunostaining. The second patient with both a heterozygous G to A substitution at nucleotide 1310+1 exon 10 splicing donor site and a heterozygous C to G substitution at nucleotide 1939 (which induces Tyr 522 Stop of exon 18) showed both patchy sarcolemmal and diffuse cytoplasmic dysferlin immunostaining. In contrast to Becker muscular dystrophy, the clinical course and severity of dysferlin staining positive patients was not clearly different from negative patients. These results suggest that a splicing mutation of the dysferlin gene may have the potential to cause decreased dysferlin expression but may not be related to the milder clinical phenotype.

Fibronectin receptor reduction in skin and fibroblasts of patients with Ullrich's disease.

Ullrich's disease is a congenital muscular dystrophy characterized clinically by generalized muscle weakness, multiple contractures of the proximal joints, and hyperextensibility of the distal joints. Recent studies have demonstrated that collagen VI is deficient in the muscles of patients with Ullrich's disease, and some cases result from recessive mutations of the collagen VIalpha2 gene (COL6A2). Fibronectin is one of the main components of the extracellular matrix (ECM) and associates with a variety of other matrix molecules including collagen. The behavior of fibronectin on cells is mediated by fibronectin receptors, members of the integrin family. We studied the expression of fibronectin receptors and fibronectin in patients with Ullrich's disease, and found a marked reduction of fibronectin receptors in the ECM of skin and cultured fibroblasts of these patients. These results suggest that collagen VI deficiency may lead to the reduction of fibronectin receptors and that an abnormality of cell adhesion may be involved in the pathogenesis of Ullrich's disease.

Electron microscopic abnormalities of skeletal muscle in patients with collagen VI deficiency in Ullrich's disease.

By electron microscopy, we examined the skeletal muscle from two patients with Ullrich's disease. One patient had a deletion in the collagen VI alpha 2 gene. The muscle biopsy specimens showed no collagen VI immunoreaction, while the expression of collagen IV was increased. Collagen VI is a microfibrillar protein in the extracellular matrix with cell adhesive properties, and collagen IV is a principal component of the basal lamina. Electron microscopy revealed unevenness, extension, and folding of the muscle plasma membrane, and showed thickening and overproduction of the basal lamina. The data show that type VI collagen is certainly one of the important extracellular matrix components maintaining the structural integrity of skeletal muscle, and a defect of the collagen VI protein causes abnormalities of the muscle plasma membrane, dystrophic muscle changes, and up-regulation of collagen IV.