Grace M. Hobson

Spastic paraplegia type 2 associated with axonal neuropathy and apparent PLP1 position effect

To report an association between spastic paraplegia type 2 with axonal peripheral neuropathy and apparent proteolipid protein gene (PLP1) silencing in a family.

Pelizaeus-Merzbacher Disease, Pelizaeus-Merzbacher-Like Disease 1, and Related Hypomyelinating Disorders

The purpose of this article is to present contemporary information on the clinical and molecular diagnosis and the treatment of Pelizaeus-Merzbachers disease (PMD) and related leukodystrophies. Various types of mutations of the X-linked proteolipid protein 1 gene (PLP1) that include copy number changes, point mutations, and insertions or deletions of a few bases lead to a clinical spectrum from the most severe connatal PMD, to the least severe spastic paraplegia 2 (SPG2). Signs of PMD include nystagmus, hypotonia, tremors, titubation, ataxia, spasticity, athetotic movements and cognitive impairment; the major findings in SPG2 are leg weakness and spasticity. A diffuse pattern of hypomyelination is seen on magnetic resonance imaging (MRI) of PMD/SPG2 patients. A similar constellation of signs and pattern of hypomyelination lead to the autosomal recessive disease called Pelizaeus-Merzbacher-like disease 1 (PMLD1) and the less-severe spastic paraplegia 44 (SPG44), caused by mutations of the gap junction protein, gamma-2 gene (GJC2), formerly known as the gap junction protein, α-12 gene (GJA12). Magnetic resonance spectroscopy (MRS) and brainstem auditory evoked potentials (BAEP) may assist with differential clinical diagnosis of PMD and PMLD1. Supportive therapy for patients with PMD/SPG2 and PMLD1/SPG44 includes medications for seizures and spasticity; physical therapy, exercise, and orthotics for spasticity management; surgery for contractures and scoliosis; gastrostomy for severe dysphagia; proper wheelchair seating, physical therapy, and orthotics to prevent or ameliorate the effects of scoliosis; special education; and assistive communication devices.

Myotonia and the muscle chloride channel Dominant mutations show variable penetrance and founder effect

The delayed relaxation or sustained contraction of skeletal muscle-myotonia--is frequently seen in myotonic dystrophy and sodium channelopathies (hyperkalemic periodic paralysis, paramyotonia congenita). Many cases of congenital myotonia without other clinical symptoms have been associated with mutations in the muscle chloride channel gene. Most cases reported to date show a recessive inheritance pattern, with loss of function of the corresponding protein. Six families have been reported with dominantly inherited myotonia and mutations of the chloride channel gene. Here we report clinical and molecular data on 38 family members from four new families with dominantly inherited myotonia congenita. Three families show a previously characterized G230E mutation, and we show that these three share a common affected ancestor despite living in different regions of the United States (linkage disequilibrium). One Italian family is shown to have a novel dominant mutation--I290M. This is the sixth mutation identified in Thomsens myotonia. Genotype/phenotype correlations in these four families showed that both of the dominant mutations resulted in a mild clinical picture in 90% of the patients, and no symptoms in 10% of mutation-positive patients. The EMG was the clinical feature that most closely correlated with mutation data; however, 3 of 16 (19%) mutation-positive patients tested negative by electromyography at least once, and 1 (6%) tested negative despite multiple tests. Only about half (55%) of the mutation-positive patients tested positive for percussion myotonia. Most of the clinically symptomatic individuals stated that cold temperatures and stress substantially worsened their myotonia. Our data show that dominantly inherited Thomsens myotonia is most often a very mild disorder that shows considerable clinical heterogeneity. NEUROLOGY 1996;47: 963-968

Mutations in noncoding regions of the proteolipid protein gene in Pelizaeus–Merzbacher disease

Background: Pelizaeus–Merzbacher disease (PMD) is an X-linked recessive dysmyelinating disorder of the CNS. Duplications or point mutations in exons of the proteolipid protein (PLP) gene are found in most patients. Objective: To describe five patients with PMD who have mutations in noncoding regions of the PLP gene. Methods: Quantitative multiplex PCR and Southern blot analyses were used to detect duplication of the PLP gene, and DNA sequence analysis, including exon-intron borders, was used to detect mutation of the PLP gene. Results: Duplication of the PLP gene was ruled out, and mutations were identified in noncoding regions of five patients in four families with PMD. In two brothers with a severe form of PMD, a G to T transversion at IVS6+3 was detected. This mutation resulted in skipping of exon 6 in the PLP mRNA of cultured fibroblasts. A patient who developed nystagmus at 16 months and progressive spastic ataxia at 18 months was found to have a 19–base pair (bp) deletion of a G-rich region near the 5′ end of intron 3 of the PLP gene. A patient with a T to C transition at IVS3+2 and a patient with an A to G transition at IVS3+4 have the classic form of PMD. These, like the 19-bp deletion, are in intron 3, which is involved in PLP/DM20 alternative splice site selection. Conclusions: Mutations in introns of the PLP gene, even at positions that are not 100% conserved at splice sites, are an important cause of PMD.

Neuronal loss in Pelizaeus–Merzbacher disease differs in various mutations of the proteolipid protein 1

Mutations affecting proteolipid protein 1 (PLP1), the major protein in central nervous system myelin, cause the X-linked leukodystrophy Pelizaeus–Merzbacher disease (PMD). We describe the neuropathologic findings in a series of eight male PMD subjects with confirmed PLP1 mutations, including duplications, complete gene deletion, missense and exon-skipping. While PLP1 mutations have effects on oligodendrocytes that result in mutation-specific degrees of dysmyelination, our findings indicate that there are also unexpected effects in the central nervous system resulting in neuronal loss. Although length-dependent axonal degeneration has been described in PLP1 null mutations, there have been no reports on neuronal degeneration in PMD patients. We now demonstrate widespread neuronal loss in PMD. The patterns of neuronal loss appear to be dependent on the mutation type, suggesting selective vulnerability of neuronal populations that depends on the nature of the PLP1 disturbance. Nigral neurons, which were not affected in patients with either null or severe misfolding mutations, and thalamic neurons appear particularly vulnerable in PLP1 duplication and deletion patients, while hippocampal neuronal loss was prominent in a patient with complete PLP1 gene deletion. All subjects showed cerebellar neuronal loss. The patterns of neuronal involvement may explain some clinical findings, such as ataxia, being more prominent in PMD than in other leukodystrophies. While the precise pathogenetic mechanisms are not known, these observations suggest that defective glial functions contribute to neuronal pathology.

Steroid-responsive neurologic relapses in a child with a proteolipid protein-1 mutation

A 10-year-old boy developed corticosteroid-responsive relapsing neurologic signs, including nystagmus and ataxia. MRI revealed multifocal T2 white matter hyperintensities; several were gadolinium-enhancing. CSF contained oligoclonal bands. Although the patient met criteria for multiple sclerosis (MS), the proteolipid protein-1 gene (PLP1) contained a mutation in exon 3B (c.409C>T), predicting a tryptophan-for-arginine substitution. This case raises questions about the role of inflammation in PLP1-related disorders and, conversely, PLP1 mutations in MS.

Deletion of a splicing enhancer disrupts PLP1/DM20 ratio and myelin stability.

PLP1 and DM20, major myelin proteins, are generated by developmentally regulated alternative splicing. In the post-natal brain, PLP1 is the predominant product. Deletion of a splicing enhancer in PLP1 intron 3 causes a mild form of Pelizaeus-Merzbacher disease and reduces PLP1 specific splicing in vitro (Hobson, G. M., Huang, Z., Sperle, K., Stabley, D. L., Marks, H. G., and Cambi, F., 2002. A PLP splicing abnormality is associated with an unusual presentation of PMD. Ann. Neurol. 52, 477-488). We sought to investigate the pathogenic role of the mutation and to determine the consequences on the developmental regulation of PLP1 alternative splicing and myelin stability and function in vivo. We have generated a knockin mouse that carries deletion of the intronic splicing enhancer and have characterized the PLP1/DM20 ratio by Real Time RT-PCR and Western blot analysis in the developing and mature brain and examined the clinical and pathological phenotype by motor testing and electron microscopy. The deletion impairs the increase in the PLP1/DM20 transcript and protein ratio at the time of myelination and in adulthood and results in a PLP1 hypomorph. Electron microscopy shows abnormal myelin wraps with fragmented myelin whorls, which are progressive with age, suggesting a defect in myelin stability. Phenotypic characterization of the knockin mouse shows a defect in motor coordination. The data indicate that the intronic splicing enhancer is necessary for the developmental increase in PLP1/DM20 ratio and that full PLP1 dosage is necessary for myelin stability and brain function. This knockin mouse represents a useful model to investigate the mechanisms of disease in human disorders in which PLP1 expression is reduced.

Neuroradiologic correlates of clinical disability and progression in the X-linked leukodystrophy Pelizaeus-Merzbacher disease.

OBJECTIVE To determine whether quantitative measure of magnetic resonance imaging data from patients with the inherited leukodystrophy, Pelizaeus-Merzbacher disease (PMD) correlates with clinical severity or progression. METHODS In our current work we have analyzed the clinical phenotypes and MRI scans of 51 male patients with PMD and 10 female carriers for whom the PLP1 genotype had been determined. In addition, we developed a 32-point functional disability scoring (FDS) system for PMD, and validated it for inter-rater reliability. Using conventional T1- and T2-weighted MRI images of the whole brain, we measured white matter and total brain volume (WMV and TBV), inter-caudate ratio (ICR), and corpus callosum area. RESULTS There was a significant positive correlation of FDS with white matter fraction (WMV/TBV) and corpus callosum area. Also, when applying a median split based on FDS, patients with lower FDS showed reduced white matter fraction and corpus callosum area, and increased ICR compared to patients with relatively higher FDS, regardless of age. CONCLUSION Although this patient population is heterogeneous, with multiple genetic and molecular mechanisms causing PMD, these data imply that white matter atrophy is a major pathological determinant of the clinical disability in most patients. Development of reliable non-invasive quantitative biomarkers of disease activity would be useful not only for following the natural history of the disease, but also raising the potential for evaluating future therapies.

A Large X-Chromosomal Deletion is Associated with Microphthalmia with Linear Skin Defects (MLS) and Amelogenesis Imperfecta (XAI)

A female patient is described with clinical symptoms of both microphthalmia with linear skin defects (MLS or MIDAS) and dental enamel defects, having an appearance compatible with X‐linked amelogenesis imperfecta (XAI). Genomic DNA was purified from the patients blood and semiquantitative multiplex PCR revealed a deletion encompassing the amelogenin gene (AMELX). Because MLS is also localized to Xp22, genomic DNA was subjected to array comparative genomic hybridization, and a large heterozygous deletion was identified. Histopathology of one primary and one permanent molar tooth showed abnormalities in the dental enamel layer, and a third tooth had unusually high microhardness measurements, possibly due to its ultrastructural anomalies as seen by scanning electron microscopy. This is the first report of a patient with both of these rare conditions, and the first description of the phenotype resulting from a deletion encompassing the entire AMELX gene. More than 50 additional genes were monosomic in this patient.

A case of complicated spastic paraplegia 2 due to a point mutation in the proteolipid protein 1 gene.

Pelizaeus-Merzbacher disease (PMD) is a rare X-linked dysmyelinating disorder resulting from mutation of the proteolipid protein gene (PLP1). Clinical features of PMD include progressive psychomotor developmental delay, nystagmus, spastic quadriplegia, dystonia, and cerebellar ataxia. PMD is clinically classified into three subtypes according to the severity of the disease: connatal, transitional, and classic forms. Patients with PMD have been identified with duplication, point mutations, and deletion of PLP1. In addition, spastic paraplegia 2 (SPG2) is allelic to PMD and typically caused by missense mutations in the second extracellular domain of PLP1 or in the PLP1-specific region that is spliced out during formation of the DM20 isoform. The authors describe a Korean boy diagnosed with SPG2 caused by a mutation that results in a Pro215Leu substitution in the second extracellular domain. Analysis of phenotypes resulting from mutations affecting PLP1 has been valuable in identifying functional domains of this still incompletely understood major myelin protein. Null mutations and mutations affecting the PLP1-specific domain cause peripheral neuropathy. The PLP1-specific domain also is important in the long-term maintenance of axonal integrity. This patients phenotype was relatively mild, in contrast with other mutations at position 215 of PLP1 that cause severe PMD. One of these severe mutations is also a missense mutation substituting an aliphatic residue, alanine, for proline. The distinct severity difference between the Pro215Leu and Pro215Ala substitutions suggests that this region of the protein is very sensitive to subtle structural changes and likely plays a critical role in PLP1 function.