Compensating for central nervous system dysmyelination Females with a proteolipid protein gene duplication and sustained clinical improvement.код для вставкиСкачать
Compensating for Central Nervous System Dysmyelination: Females with a Proteolipid Protein Gene Duplication and Sustained Clinical Improvement Ken Inoue, MD, PhD,1 Hajime Tanaka, MD, PhD,3 Fernando Scaglia, MD,1 Akiko Araki, MD,3 Lisa G. Shaffer, PhD,1 and James R. Lupski, MD, PhD1,2 A submicroscopic duplication that contains the entire proteolipid protein gene is the major cause of PelizaeusMerzbacher disease, an X-linked central nervous system dysmyelinating disorder. Previous studies have demonstrated that carrier females for the duplication are usually asymptomatic. We describe 2 unrelated female patients who present with mild Pelizaeus-Merzbacher disease or spastic paraplegia. In 1 patient, clinical features as well as cranial magnetic resonance imaging and brainstem auditory evoked potential results have improved dramatically over a 10-year period. The other patient, who presented with spastic diplegia and was initially diagnosed with cerebral palsy, has also shown clinical improvement. Interphase fluorescent in situ hybridization identified a proteolipid protein gene duplication in both patients. Interphase fluorescent in situ hybridization analyses of the family members indicated that the duplication in both patients occurred as de novo events. Neither skewing of X inactivation in the peripheral lymphocytes nor proteolipid protein gene coding alterations were identified in either patient. These findings indicate that, occasionally, females with a proteolipid protein gene duplication can manifest an early-onset neurological phenotype. We hypothesize that the remarkable clinical improvement is a result of myelin compensation by oligodendrocytes expressing one copy of proteolipid protein gene secondary to selection for a favorable X inactivation pattern. These findings indicate plasticity of oligodendrocytes in the formation of central nervous system myelin and suggest a potential role for stem cell transplantation therapies. Ann Neurol 2001;50:747–754 Proteolipid protein (PLP) is the most abundant myelin protein in the central nervous system (CNS). Mutations in PLP are associated with a severe X-linked CNS dysmyelinating disorder, Pelizaeus-Merzbacher disease (PMD), and a clinically milder allelic disorder, spastic paraplegia type 2 (SPG2).1–3 Although most female carriers show no clinical symptoms, in some families carrier females manifest a late-onset spastic paraplegia phenotype with variable severity.4 –7 A gene dosage effect of PLP by submicroscopic genomic duplication (ie, not detectable by conventional G-banded chromosome analysis) accounts for 60 to 70% of PLP mutations among PMD patients.8 –11 Recent molecular studies revealed that the size and breakpoints of the duplicated genomic fragments are surprisingly variable, yet the underlying mechanism for the genomic recombination is likely to be common; that is, an unequal sister chromatid exchange in male meiosis.9,10 A few families with PLP duplication in which the additional gene copies were translocated to different regions in the X chromosome were recently reported.12 Patients with PLP duplication typically present with the classic form of PMD, whereas more severe connatal and transient forms are mostly associated with point mutations in PLP.10 No symptomatic female carrier of a PLP duplication has been reported in previous studies.10,13 We identified 2 independent female patients with From the Departments of 1Molecular and Human Genetics and 2 Pediatrics, Baylor College of Medicine, Houston, TX; and 3Department of Pediatrics, Asahikawa Habilitation Center for Disabled Children, Asahikawa, Japan. Published online Nov 1, 2001; DOI 10.1002/aon.10036 Received May 17, 2001, and in revised form Jul 10. Accepted for publication Aug 14, 2001. Address correspondence to Dr Lupski, Department of Molecular and Human Genetics, Baylor College of Medicine, One Baylor Plaza, Room 604B, Houston, TX 77030. E-mail: firstname.lastname@example.org © 2001 Wiley-Liss, Inc. 747 dysmyelinating disorders who showed progressive clinical improvement. Interphase fluorescent in situ hybridization (FISH) analyses indicated that both patients have duplications of PLP. We hypothesize that CNS myelin compensation may play a role in the mechanism of this unique phenomenon. Interphase FISH Patients and Methods Patient 1 Genomic DNA was extracted from peripheral blood cells. X inactivation status was evaluated at the trinucleotide repeat region in the androgen receptor gene by a PCR-based method,15 using fluorescently labeled primers and an automated sequencer (ABI377; Applied Biosystems, Foster City, CA). Each exon of the PLP gene was amplified by PCR and sequenced using dye primer chemistry and an ABI377 automated sequencer, as described.16 BAB1532 is an 11-year-old Japanese girl who was born weighing 3,126g at full-term (38-weeks) from nonconsanguineous parents (Family HOU612). She has no family history of neurological disorders. She had neonatal asphyxia with Apgar scores of 2 (1 minute) and 7 (5 minutes). Nystagmus began at 2 weeks of age. Although developmentally delayed, she has been catching up substantially in attaining developmental milestones (holding her head at 6 months, rolling over at 8 months, creeping at 18 months, speaking one word at 11 months and two words at 21 months of age). Her intelligence quotient (IQ) was verbal IQ of 70, performance IQ of 61, full-scale IQ of 59 at 4 years and 9 months of age (Wechsler Preschool and Primary Scales of Intelligence); and verbal IQ of 101, performance IQ of 90, full-scale IQ of 95 at 8 years of age (Wechsler Intelligence Scale for Children). Hypotonia and mild hyperreflexia in her upper limbs noted at 3 months were diminished by 5 years of age and mild difficulty in coordination in her upper extremities improved to normal. Hyperreflexia and spasticity in her lower limbs, noted at 3 months and 8 months of age, respectively, have kept her wheelchair bound. Now she can move by crawling and control her wheelchair by herself. She has exhibited no extrapyramidal or cerebellar symptoms during her clinical course. Serial head magnetic resonance imaging (MRI) studies and brainstem auditory evoked potentials (BAEPs) were obtained from 3 months to 11 years of age. Patient 2 BAB1718 is a 6-year-old Hispanic girl from nonconsanguineous parents (Family HOU671). One of her maternal cousins died at 8 days of life secondary to anencephaly. No abnormality was noticed during the pregnancy or after the birth by Cesarean section for breech presentation. She was healthy with normal developmental milestones until 14 months of age, when she showed spasticity in her lower extremities and started to walk on her toes and drag her feet. She showed neither cerebellar nor extrapyramidal tract signs. No nystagmus was noted. She is intellectually normal for her age. Despite the residual spasticity in her lower extremities, her gait has recently improved with milder toewalking in comparison with the previous 2 years. She still drags her right foot while walking; however, her gait is steadier and she no longer falls when tired. She still trips when she tries to run but she can walk at a brisk pace, which was not previously possible. MRI studies were performed at age 2 and 6 years. BAEPs were obtained at 6 years of age. 748 Annals of Neurology Vol 50 No 6 December 2001 White blood cells from the patients and other members of both families were cultured and harvested as described by Shaffer and colleagues.14 Interphase FISH was performed as described previously.10 X Inactivation and PLP Coding Region Sequencing Analyses Results MRI and BAEPs Improvement in Myelination The MRI studies of Patient 1 (BAB1532) from 4 months to 10 years of age demonstrated prominent delay in myelination, especially in her first 2 years of life. The myelination has been slowly improving; it reached to the corona radiata at 21 months and cortical white matter at 4 years of age (T1-weighted images in Fig 1 and Table). However, significant T2-hyperintensity regions in the cortical white matter remain at 10 years of age, indicating incomplete myelination (see Fig 1 and Table). The BAEPs of Patient 1 revealed significant improvement, which chronologically parallels the improvements of those seen in the MRI studies. Patient 2 (BAB1718) presented with a mild delay of myelination in the cerebral white matter at 2 years of age, which improved at 6 years of age (see Fig 1 and Table). She had normal BAEPs at 6 years of age. Interphase FISH Analyses Both patients have a normal 46,XX karyotype. Interphase FISH analyses identified three PLP signals with two control signals in both patients, indicating that they have a PLP duplication of one X chromosome (Fig 2). The PLP signals were observed on Xq22 for both normal and duplicated alleles by metaphase FISH indicating there was no translocation of PLP (data not shown). In family HOU612, both parents are negative for the PLP duplication by FISH analyses, suggesting that the duplication occurred as a de novo event. In the other family HOU671, both the mother and healthy sister produced normal FISH results, but the father’s sample was not available. Because the father has no phenotype related to PMD, he is unlikely to have PLP duplication as this results in a PMD phenotype in males. Therefore, the duplication event in this patient is also likely de novo. Random X Inactivation in Both Patients Patient BAB1532 revealed a ratio of 76 to 24 and patient BAB1718 showed a ratio of 48 to 52 for the X Fig 1. Magnetic resonance imaging (MRI) and brainstem auditory evoked potentials (BAEPs) revealed improvement in myelination. T1- and T2-weighted images of Patient 1 (BAB1532) at age 4 months (A and B), 21 months (C and D), and 10 years (E and F); T2-weighted images of Patient 2 (BAB1718) at age 2 years (G) and 6 years (H). (A and B) In Patient 1, MRI at 4 months showed initiation of myelination in the brainstem, thalamus, and internal capsule but no signs of myelination in the cerebral white matter (WM). Myelination appeared very slowly, barely reaching the basal ganglia at 9 months of age (data not shown) and cerebral WM at age 21 months (C). (D) Hyperintensity in T2-weighted images indicated that myelination in the cortical hemisphere was extensively delayed. At age 4 years, prominent progress of myelination was seen in cortical WM, although its patchy appearance suggested incomplete myelination (data not shown). (E) At age 10 years, myelination reached the subcortical WM, shown as hyperintensity in the T1-weighted image. (F) Irregular T2 hyperintensity was observed in the cortical WM and the contrast between gray matter (GM) and WM is unclear, suggesting that myelination is still incomplete in the cortical hemisphere. (G) In Patient 2, MRI at age 2 years revealed myelination up to the subcortical regions, but there were irregular patchy appearances in the WM; suggesting incomplete or mosaic myelination. (H) Similar findings are found in MRI at age 6 years, but the contrast between WM and GM had significantly improved in the comparative images. (I) BAEPs of Patient 1 obtained at age 3 months, 8 months, 1 year and 9 months, 5 years and 6 months, and 11 years. Wave II and subsequent waves were absent at age 3 months, but appeared, although still delayed, at 8 months of age. BAEP results gradually improved, parallel to the MRI findings, to almost normal range at age 11 years. Inoue et al: Compensating for CNS Dysmyelination 749 Table. MRI Findingsa at Different Ages in Female PLP Duplication Carriers Manifesting CNS Dysmyelination Patient 1 White matter Basis pontis Median leminiscus Decussation of the superior cerebellar peduncle Crura cerebri Anterior limb of internal capsule Medial posterior limb of internal capsule Lateral posterior limb of internal capsule Middle cerebellar peduncle Superior cerebellar peduncle Vermis Hilus of the dentate nucleus White matter of cerebellar hemisphere Central corona radiata Subcortical white matter Gray matter Tegmentum pontis Colliculi inferior Colliculi superior Red nucleus Substantia nigra Thalamus, ventrolateral Thalamus, not ventrolateral Globus pallidus, lateral Globus pallidus, not lateral Putamen, dorsolateral Putamen, not dorsolateral Dentate nucleus Pericentric cortex Hippocampus Patient 2 4 mo 9 mo 21 mo 4 yr 10 yr 2 yr 6 yr ⫹/⫺ ⫹/⫺ ⫹/⫺ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫺ ⫺/⫺ ⫹/⫺ ⫹/⫹⬃⫽ ⫹/⫺ ⫹/⫺ ⫹/⫹ ⫹/⫺ ⫹/⫺ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫺ ⫹/⫺ ⫹/⫺ ⫹/⫽ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫺ NA ⫹/⫽ ⫹/⫺ ⫽/⫺ ⫹/⫺ NA ⫹/⫽ ⫹/⫽ ⫹/⫽ ⫹/⫽ NA ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫺/⫺ ⫺/⫺ ⫺⬃⫽/⫺ ⫺/⫺ ⫹/⫺ ⫽⬃⫹/⫺ ⫹/⫺⬃⫽ ⫽⬃⫹/⫺ ⫹/⫽ ⫹/⫽ ⫹/⫽⬃⫹ ⫹/⫽⬃⫹ ⫹/⫽⬃⫹ ⫹/⫹ ⫹/⫺ ⫹/⫽ NA ⫹/⫽ NA ⫹/⫽ ⫺/⫺ ⫽/⫺ ⫺/⫺ ⫽/⫽ ⫺/⫺ ⫹/⫽ ⫺/⫺ ⫽/⫺ ⫹/⫹ ⫹/⫹ NA ⫹/⫽ ⫹/⫽ ⫹/⫹ ⫽⬃⫹/⫽ ⫽/⫽ ⫺/⫺ ⫹/⫹ ⫽/⫽ ⫹/⫺ ⫽/⫺ ⫽⬃⫹/⫽ ⫹/⫹ ⫹/⫹ NA ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫽⬃⫹ ⫹/⫽⬃⫹ ⫹/⫽ ⫹/⫹ ⫹/⫽ ⫹/⫽ ⫹/⫺⬃⫽ ⫽⬃⫹/⫽ ⫹/⫹ NA NA ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫽⬃⫹ ⫹/⫽⬃⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫽⬃⫹ ⫹/⫽⬃⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ ⫹/⫹ a Level of myelination compared with intensity of the caudate nucleus head.27 Each evaluation was presented as T1-weighted/T2-weighted images. ⫹ indicates myelinated regions, defined as hyperintensity in the T1-weighted images and hypointensity in T2-weighted images. ⫺ indicates nonmyelinated regions, defined as the reverse (hypointensity in T1-weighted images and hyperintensity in T2-weighted images). ⫽ indicates regions of uncertainty in determining degree of myelination, defined as isointensity in either T1- or T2-weighted images. ⬃ indicates transition between the two values. Dark gray indicates regions showing hypointensity in T1 and hyperintensity in T2-weighted images, thus indicating no myelination. Light gray indicates regions showing hyperintensity in either T1- or T2-weighted images, thus indicating incomplete or partial myelination. Significant progress in myelination is noted by diminishing unmyelinated or incompletely myelinated regions over time. PLP ⫽ proteolipid protein gene; CNS ⫽ central nervous system; NA ⫽ not available for evaluation. Fig 3. Schematic models show compensation of CNS myelin and distribution of two cell types during different time points in heterozygous females. Myelin generated by oligodendrocytes expressing normal proteolipid protein gene (PLP) (blue) and mutant PLP (red) with corresponding axon (pink) are shown at premyelination, mature myelination, and adult stage. A represents complete normalization observed in severe PLP mutations. In the premyelinating stage, equal numbers of cells express mutant or normal PLP allele. As the mutant cells are eliminated by subsequent apoptosis, cells expressing the normal allele predominate after preferential survival and myelinate all axons, resulting in normal myelination in the adult stage with selective skewing. B shows late-onset demyelination (dashed) as a result of the survival of mutant oligodendrocytes that have a mild mutation in PLP and form incomplete or fragile myelin (mutant myelin may be thinner than wild type myelin). In this case no compensation is observed. C presents two possibilities for the mechanism that best explains our results. Both mechanisms lead to a greater number of normal cells, but distinguishing between these two mechanisms is not yet possible. First, the upper group of cells shows the skewed X inactivation toward the direction of the normal X chromosome being inactivated (most oligodendrocytes express mutant PLP, shown in red), resulting in slow progression of myelin compensation because of the small number of normal oligodendrocyte precursors. Second, the lower group of cells shows the model for the “double-hit” hypothesis. Both mutant and normal oligodendrocyte precursors are distributed equally at the premyelination stage. Following apoptotic cell death of mutant cells, normal cells compensate with slow progression because of an unknown factor that diminishes the ability of normal oligodendrocyte precursors to proliferate and differentiate into mature myelin. On the right, diagrams represent changes of mutant and normal cell distribution over time. 750 Annals of Neurology Vol 50 No 6 December 2001 ‹ Fig 2. Interphase fluorescent in situ hybridization (FISH) analyses showing PLP duplications. Harvested cells were hybridized with a PLP-specific probe (cosmid u125A5 DNA) labeled by digoxigenin and an intrachromosomal control probe located at the BTK locus (PAC RP1-36B9 DNA) labeled with Biotin. Signals were visualized using a fluorescence microscope. Representative interphase nuclei are shown from Patient 1 (A) and Patient 2 (B). Note that two sets of signals are present; one normal allele that represents a single copy of PLP (red) and one control signal (green) on the same chromosome, and one duplicated allele that shows two PLP signals (red, arrowheads) and one control signal (green) on the other X chromosome. inactivation status in their peripheral blood cells, indicating that no skewing of X inactivation was evident in these cells in either patient (data not shown).17 No Mutations in PLP Coding Region No alterations in the coding regions or splice site junctions of the PLP gene were identified in either patient, Figure 3 Inoue et al: Compensating for CNS Dysmyelination 751 indicating that point mutations in the coding region of PLP are not likely to play roles in either patient and both carry three copies of wild type PLP (data not shown). Discussion An Improving Central Nervous System Dysmyelinating Disorder We describe 2 female patients with a PLP duplication who presented with CNS dysmyelinating disorders of different degrees of severity. One patient had developmental delay from infancy accompanied by mild intellectual impairment and spastic diplegia that later resolved, resulting in objectively documented normal intelligence and improved motor function. The initial clinical manifestation was similar to that of most male PMD patients with a PLP duplication but clearly differs by the improvement of clinical features. This degree of clinical improvement is not observed in male patients. Objective clinical studies, including MRI that revealed diffuse dysmyelination of the entire white matter as well as abnormal BAEPs, also dramatically improved and have almost normalized over time. The second patient presented with mild spastic diplegia in her lower extremities with neither developmental delay nor intellectual impairment but with minimal dysmyelinating MRI findings. Her clinical phenotype, as well as her MRI data, have been improving over a 3-year period. It is of note that her initial diagnosis was cerebral palsy. In fact, PMD is one of the differential diagnoses of cerebral palsy18 and a number of male patients with PMD have been diagnosed as having cerebral palsy. We emphasize that in females diagnosed with cerebral palsy a genetic abnormality of PLP should be also considered even if MRI shows only minimal dysmyelination. Compensation of Central Nervous System Myelin in Heterozygous Females Studies of more than 30 families with PLP duplication have shown that patients’ mothers are usually carriers for the duplication and present with no phenotype associated with PMD.10,11 Similarly, female carriers of point mutations that result in a severe form of PMD in males are usually asymptomatic. In contrast, PLP mutations that result in a mild PMD/SPG2 phenotype in affected males, including point mutations and null alleles of PLP, have been associated with adult-onset degenerative leukodystrophies among carrier females.4 –7,19,20 Female patients carrying PLP duplications presented with an early-onset dysmyelinating phenotype, suggesting that the underlying disease mechanism may be distinct. This paradoxical phenomenon that severe PLP mutations commonly result in asymptomatic carrier females, whereas the reciprocal mild alterations may result in symptomatic carriers, has been explained by 752 Annals of Neurology Vol 50 No 6 December 2001 normalization of the oligodendrocyte population.21 As the result of random X inactivation, a heterozygous female carrier has two distinct oligodendrocyte precursor populations: cells only expressing the normal PLP allele and cells only expressing the mutant PLP allele. Severe PLP mutations may affect differentiation of oligodendrocytes and lead to subsequent apoptosis. As a result, the remaining oligodendrocytes of mature myelin represent a population of cells expressing a normal PLP allele (Fig 3A). On the other hand, oligodendrocytes expressing mild PLP mutations may survive through development and form myelin, resulting in a mosaic population in mature myelin. Because the mutant myelin may be incomplete or unstable, and probably is subject to being degraded, subsequent demyelination and late-onset clinical manifestations may occur in carrier females (see Fig 3B). These hypotheses are supported by findings from studies on mutant Plp animal models. In heterozygote jimpy mice, a model for the severe form of PMD, myelin at birth is mosaic presenting as fully myelinated and barely myelinated axons.22,23 Subsequently, all axons become myelinated as normal oligodendrocytes replace mutant cells.22,23 In contrast, the heterozygote rumpshaker mice, a model of SPG2 in the hemizygote, maintain mutant oligodendrocytes survival with compact myelination.24 Neuropathological studies revealed the mosaic presence of axonal fibers with disproportionally thin myelin sheaths for the axon diameters, which is more prominent in older animals, as well as fibers with normal myelin. However, the animals continue to be phenotypically normal.24 Because the present clinical cases do not fit either of the models delineated from murine studies, we propose two possible models based on observations in our patients (see Fig 3C). First, if the X inactivation is skewed toward the unfortunate direction in which the normal X chromosome is selectively or predominantly inactivated, most oligodendrocyte precursor cells express the duplicated PLP allele and fail to differentiate further. This may result in a limited number of precursor cells capable of initiating myelin formation and, thus, the majority of axons are unmyelinated at this stage. As a result, carrier females may manifest a dysmyelinating pathological and clinical phenotype. However, as these axons are subsequently myelinated by compensating normal oligodendrocytes, the clinical symptoms may improve (see Fig 3C). A Plp mutant dog consistent with this model was reported recently.25 Skewing of X inactivation in the peripheral blood cells of our patients was not demonstrated. There is, however, a possibility that tissue-specific unfavorable skewing exists only in the CNS, although such evaluation was not possible in this study. Interestingly, asymptomatic female carriers of PLP duplication can have skewed X inactivation in their peripheral blood cells.13 Although it is unknown whether this is also the condition in the brain, a possible mechanism to escape phenotypic presentation in female carriers of PLP duplication is by selective X inactivation, which does not appear to be present in our patients. Alternatively, an additional factor may reduce the mitotic potential of normal precursor cells, resulting in suppressed or decreased myelin compensation (see Fig 3C). Such factors include PLP alterations in the normal allele or mutations in other genes in the CNS myelination pathway, which may not result in a dysmyelinating disorder by itself but reduce the potential to proliferate and compensate for dysmyelination. This “double-hit” hypothesis (PLP duplication and another mutation) is, however, less likely, and we have no evidence to support it. Furthermore, we did not find any alteration either in the PLP coding and minimal promoter region in the normal allele or in the duplicated allele of PLP in our patients. Implication for Stem Cell Treatment of PelizaeusMerzbacher Disease Although no effective treatment is available for PMD, stem cell transplantation may have great potential. Because PMD primarily affects oligodendrocyte development, transplantation of myelin lineage precursor cells may compensate the myelin deficiency. Despite the tight regulation of normal myelin development in the CNS, the dramatic recovery from clinical symptoms, as well as the improvement in MRI and BAEP findings, suggest sufficient plasticity of the oligodendrocyte lineage to proliferate and compensate the dysmyelination even years after birth. This implies the possibility of successful application of stem cell transplantation therapies for PMD, as preliminarily studies have shown in embryonic stem cell transplantation in mice.26 This study was supported in part by the National Institute for Neurological Disorders and Stroke (NIH R01 NS27042), the Muscular Dystrophy Association (to JRL), and the Baylor College of Medicine Mental Retardation Research Center (NIH P30 HD24064). K.I. is supported by fellowships from the Charcot-Marie-Tooth Association and Muscular Dystrophy Association. We greatly appreciate the participation of the patients and families. We thank Drs Kimiko Deguchi (Department of Pathology, Baylor College of Medicine), Noriko Aida (Division of Radiology, Kanagawa Children’s Medicain Center), and Cornelius Boerkoel (Department of Molecular and Human Genetics, Baylor College of Medicine) for their critical reviews and helpful advice. We thank the members of the Kleberg Cytogenetics Laboratory, Baylor College of Medicine, for their expert FISH and cell culturing on these cases. References 1. Yool DA, Edgar JM, Montague P, et al. The proteolipid protein gene and myelin disorders in man and animal models. Hum Mol Genet 2000;9:987–992. 2. Garbern J, Cambi F, Shy M, et al. The molecular pathogenesis of Pelizaeus-Merzbacher disease. Arch Neurol 1999;56: 1210 –1214. 3. Hodes ME, Pratt VM, Dlouhy SR. Genetics of PelizaeusMerzbacher disease. Dev Neurosci 1993;15:383–394. 4. Hodes ME, Blank CA, Pratt VM, et al. 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