BRIEF COMMUNICATIONS
Congenital Hypomyelination Neuropathy with Arthrogryposis Multiplex Congenita Kevin B. Boylan, MD,” Donna M. Ferriero, MD,t Claudia M. Greco, MD,$ R. Ann Sheldon, BA,t and Michael Dew, MD§
A term male infant is described with a n isolated disorder of peripheral myelination. A t necropsy, the great majority of medium-to-large axons were unmyelinated. Electron microscopy showed normal axons and redundant lamination of basement membrane, suggestive of early onion bulb pathology. Immunohistochemistry of peripheral nerve showed deficiency of the myelin proteins P2 and Po, myelin basic protein, and myelin-associated glycoprotein. Arrest of peripheral myelination at the promyelin stage appears to be the origin of myelin deficiency. Boylan IU3, Ferriero DM, Greco CM, Sheldon RA, Dew M. Congenital hypomyelination neuropathy with arthrogryposis multiplex congenita. Ann Neurol 1992;31:337-340
Deficiency or absence of peripheral myelin in newborns [ 1, 21, generally referred t o as congenital hypomyelinating neuropathy, is a rare cause of infantile neuromuscular weakness. Five patients with arthrogryposis muitiplex congenita with deficient peripheral myelin have been reported, b u t all differ from o u r patient in that 3 had associated central nervous system defects or congenital abnormalities apart from t h e arthrogryposis [3-51, and 2 had complete [ l ] or virtually complete [6J absence of peripheral myelin.
Patient Report The patient’s clinical features have been previously reported 171. Briefly, the infant was delivered at 38 weeks’ gestation,
was ventilator-dependent, had large joint flexion contractures with diffuse hypotonia, severe generalized weakness, and areflexia. Spinal fluid, karyotype, and neonatal inborn errors of metabolism tests were normal. Electroencephalography and brain and spine magnetic resonance images showed normal patterns for age. Sensory action potentials were unobtainable; motor conduction velocity was extremely slow at 1 to 2 m/sec (normal, 2 1 m/sec). Electromyography showed fibrillations and positive waves with absent motor unit potentials. Biopsy of vastus medialis at age 8 days showed variation in muscle fiber size and type 2 predominance in a pattern consistent with neurogenic change; neither grouped atrophy nor hypertrophy were present. Results of sural nerve biopsy are described. Death occurred at 5 weeks of age following withdrawal of ventilatory support. Autopsy was performed within 1 hour of death.
Materials and Methods Morphometric Analysis Morphometry was performed on biopsied sural nerve (fixed in 2.5% glutaraldehyde in 0.1 M cacodylate buffer, p H 7.4) and compared with published control data [Z, 83. Fiber counts were performed manually o n nonoverlapping electron micrographs of transverse sections (final enlargement, x 5,000 for myelinated axons and x 10,000 for unmyelinated axons). Axons in a one-to-one relationship with Schwann cells were counted as myelinated type axons even if the Schwann cell had not actually produced myelin membranes. Schwann cells with axons in this relationship would normally be expected to have begun myelination in a term infant f81. If the Schwann cell cytoplasm contained two or more axons, the axons were counted as unmyelinated. Axonal diameters were calculated using the “arithmetic” method of Dyck [9].
Immunoqytochemisytry Lumbosacral cord and roots from the patient were compared with control spinal cord from a similar-age infant without nervous system disease who died of hypoplastic left heart syndrome. Tissue was incubated overnight at 4°C using one of the following antibodies: antimyelin basic protein (MBP) at 1:1,000, anti-P2 at 1:500, anti-Po at 1:500, and antimyelin-associated glycoprotein (MAG) at 1:500. Antibody was visualized with the avidin-biotin-peroxidase method using the Vectastain ABC Elite kit (Vector Labs, Burlingame, CA). Diaminobenzidine (Sigma) was used as the chromagen.
Results From rhe *Deparrmenr of Neurology, Johns Hopkins Universiry, Balrimorc; rhe tDepdrrmenrs of Neurology and Pediatrics, University of California, and the tDeparrment of Neuropathology, Children’s Hospital, San Francisco, CA; and the $Departments of Psychiarry and Neurology, Dwight Eisenhower Army Medical Cenrer, Fort Gordon, G A 30905. Received Mar 6 , 1991, and in revised form Aug 8. Accepred for publication Sep 14, 1991. Address correspondence to D r Boylan, Deparrmenr of Neurology, Mayo Clinic Jacksonville, 4500 San Pablo Road, Jacksonville, FL 32224.
Premortem Studies Few fibers in t h e sural nerve biopsy w e r e myelinated (Fig 1A). Myelination, w h e n present, tended t o be thin relative to axon diameter, but myelin- membrane lamination was normal, as w e r e associated axons (Fig lB,C). Small unmyelinated fibers appeared normal. Lysosomal inclusion bodies w e r e n o t present. Redundant basal laminae surrounding nonmyelinated axons were c o m m o n (see Fig lB,C). Endoneurial collagen was not increased.
Copyright 0 1992 by the American Neurological Association
337
PERCENT OF AXONS
40
30
F i g I . Sural newe biopsy. (A) One pm-thick section of resin.embedded newe shouis striking diminution of myeiinated fibers. Schu~anncell nuclei appear increased in number. (Tohidine blue: x 120 before j292 reduction.) (B) Multiple single. nonmyelinated axons are surrounded by redundant bmal laminae and occasional extensions of Schwann cell cytoplasm. (Tohidine blue; x 7.500 before 525% reduction.) (C) A single, nonmyelinated axon, J p n in diameter, surrounded by two Schwann cell nuclei, u6ich may indicate Schwann cell mitotic ‘tctivity. This actizitj uJouldbe expected during fetal peripheral nerve development but would be abnormal in a term infant for this s h e axon. (Toluidine blue; x 7,500 before 52% reduction.) I n B and C. open arrows indicate redundant basal laminae thought to remain .from abortive attempts at myelination. and dotted arrows indicate fibroblast processes, distinguishable fkom Schuiann cell extensions because of the absence of basal laminae. Morphometric Data
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In surd nerve, 642 axons in a one-to-one relationship with Schwann cells were counted in a 2.52 x lo-’ mm’ cross-sectional area. Diameters ranged from 1 to 10 pm (Fig 2A). Twenty-six axons with diameters from 3 to 8 pm (4.0%’)were actually myelinated; the remainder were enwrapped with Schwann cell cytoplasm without elaboration of myelin membranes. The ratios of axon to fiber diameter (g ratio) [S] for the 26 myelinated axons were mostly higher than those reported for similar-age control infants (Fig 2C). In a crosssectional area of 5.24 x lo-’ mm’, 988 unmyelinated s o n s were counted. Diameters ranged from 0.2 to 2.6 pm (Fig 2B). Axon densities for myelinated type (1.94 x lo4 mm’) and unmyelinated axons (1.90 x lo5 mm’) were both normal [2, S}. In each distribution a subset of axons ( 5 % for myelinated arid 10% for unmyelinated) were up to 60% larger in diameter than the upper limit reported for control nerve, and the proportion of small diameter fibers was increased. Postmortem Studies
Brain and spinal cord were grossly normal. Motor neurons and dorsal root ganglia were histologically normal in appearance and number. Resin-embedded cross secdons of cervical and lumbar cord showed decreased myelinization of dorsal roots. There was a mild decrease in the number of myelinated fibers in ventral roots at both levels, and only rare, thinly myelinated fibers in sections of cauda equina. Distal median nerve showed severely delayed myelination compared with C7 dorsal and ventral roots (Fig 3). 338 Annals of Neurology Vol 31 No 3
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F i g 2. Diameter distributions for myelinated (A) and unmyelinated (Bi axons. Percentage of axons is plotted versus axon dimieter. Control data were redrawn (with permission from the authors and pzlbhher) from 181. iC) Twenty-six myelinatrd axons; g ratio is plotted versus axon diameter. The g ratio ues should generally be between 0.7 and 0.8 for children unu’er 3 years of age {8}. Z
J
~
A
B
C
Fig 3. C7 sensory root iA) and motor root (B) show slightly diminished myelination for age. Distal median nerve (C) shows a markedly decreased number of myelinated fibers. Also, axon diameters are decreased. Arrowhead in (B) and ICi shows an axon associated with t w o Schwann cell nuclei. The small c in (B) denotes a capillary. Filled arrows indicate unmyelinated large axons, and open arrows point t o myelinated axons. All sections are from resin-embedded autopsy tissue. IToluidine blue, x 480 before 52% reduction.)
lmmunocytocbemistly Sections through lumbosacral cord revealed a normal pattern of myelin MBP-like irnmunoreactivity. Transition areas through root entry zones revealed a markedly decreased imrnunostaining of P2, Po, and MBP in the peripheral nerve root, but only moderate deficiency of MAG (data not shown). The decreased immunostaining of the myelin proteins corresponded to the loss of myelinated nerve fibers (see Figs 1, 3) with no selective dropout of any of the myelin proteins tested.
Discussion This infant’s neuropathic disorder appeared to represent failure or severe delay of myelination at the promyelin stage. The promyelin stage is the phase of myelination in which Schwann cells have enwrapped single axons but have not yet begun myelin formation { 101. For axons of all diameters, the g ratios were relatively high; hypomyelination did not appear to be confined to any subset of axons distinguishable by size criteria. Redundant basal lamina and extensions of Schwann cell cytoplasm were relatively common in electron micrographs of sural nerve, which are similar to changes seen early in onion bulb formation in both human and animal forms of hereditary hypertrophic neuropathy [l l}. A rostral-caudal gradient in spinal root myelination is normal {12), but the degree of myelin deficiency in the roots, and especially in median and sural nerve, was quite abnormal. In addition, Figure 2 shows a marked difference in axon diameters between the spinal roots (which seemed normal) and the distal median nerve.
We found no published infant control data on this point, but in adults axon diameters in distal peripheral nerve are not substantially less than in corresponding spinal roots {9}. Similar to these observations in median nerve, a relative excess of smaller diameter axons was present in surd nerve (see Fig 2A,B). An increased proportion of smaller diameter axons therefore seems likely to have been a generalized condition. MAG is expressed by Schwann cells during the promyelin stage before full expression of Po, P2, and MBP [13}. There was no selective dropout of any of these proteins, so it seems unlikely that deficiency of any one protein led to diffuse peripheral hypomyelination. The moderate deficiency of MAG with severe dehciency of the remaining proteins supports the suggestion that hypomyelination resulted from developmental arrest at the promyelin stage, particularly given the presence of relatively normal myelination of some axons. Presented in part at the 42nd Annual Meeting of the American Academy of Neuroiogy, Miami Beach, FL, April 1990. The opinions or assertions contained herein are the private views of the authors and are not to be construed as official or as reflecting the views of the Army or the Department of Defense. We would like to thank Dr John Whitaker for the generous gift of anti-MBP; Dr Richard Quarles for the generous gift of anti-MAG; and D r Bruce Trapp for the generous gift of anti-Po and anti-P2. We also thank D r John Griffin and Dr Yuen So for helpful discussions regarding the manuscript.
References 1. Charnas L, Trapp B, Griffin J. Congenital absence of peripheral myelin: abnormal Schwann cell development causes lethal arthrogryposis multiplex congenita. Neurology 1988; 38:
966-974 2. Ouvrier R, McLeod JG, Pollard J, eds. Pevipherd neuroputhjj 292 rhildbood. New York: Raven, 1990:13-20 3. Hakamada S, Kumagai T, Hara K, et al. Congenital hypomyelinating neuropathy in a newborn. Neuropediatrics 1983;14: 182-183 4. Pleasure JR, Shuman S, Rorke LB. Congenital absence of peripheral nervous system (PNS) myelin presenting as arthrogryposis multiplex congenita (Part 11). Pediatr Res 1986;20: 465A 5. Seitz RJ, Wechsler W, Mosny DS, Lenard HG. Hypomyelination neuropathy in a female newborn presenting as arthrogryposis multiplex congenita. Neuropediatrics 1986;17: 132-136 6. Palix C, Coignet J. Un cas de polyneuropathie peripherique neonatale par myelination. Pediatrie 1978;33:201-207 7. Boylan K, Dew M, Greco C, Ferriero D. Hypomyelination neuropathy: a rare form of neonatal neuromuscular weakness. Neurology 1990;40:409A 8. Jacobs JM, Love S. Qualitative and quantitative morphology of human sural nerve at different ages. Brain 1985;108:837-924 9. Dyck PJ. Parhologic alterations of the peripheral nervous system in humans: In: Dyck PJ, Thomas PK, Lamberr EH, Bunge R, eds. Peviphevuf Neuroputhjj. Philadelphia: W.B. Saunders, 19841760-870
Brief Communication: Boylan et al: Congenital Hypomyelination Neuropathy
339
10. Friede RL, Samorajski T. Myelin formation in the sciatic: nerve of the rat. A quantitative electron microscopic and radioauto-
graphic study. J Neuropathol Exp Neurol 1968;27:546--570 11. Low PA. The evolution of “onion bulbs” in the hereditary hy-
pertrophic neuropathy of the trembler mouse. Neuropathol Appl Neurobiol 1977;3:81-92 12. Dobosz-Niebrol I, Fidzianska A, RafalowskaJ, Sawicka E. Correlative biochemical and morphological studies of myelination in human ontogenesis. 11. Myelination of the nerve roots. Acta Neuropathol 1980;49:153-158 13. Willison HJ, Ilyas AL. OShannessy DJ, et al. Myelin-associated glycoprotein and related glycoconjugates in developing cat peripheral nerve: a correlative biochemical and morphometric study. J Neurochern 1987;49:1853-1862
Familial Amyloidotic Polyneuropathy: Report of Patients Heterozygous for the Transthyretin Gly42Gene Tatsufumi Murakami, MD,” Shigehiro Yi, MD,” Kenji Yamamoto, MDJ Shoichi Maruyama, MD,t and Shukuro Araki. MD”
We studied 2 patients from aJapanese family with familial amyloidotic polyneuropathy (FAP). Their clinical features are similar to type 1 FAP, and the proband’s rectal tissue contained amyloid that stained with antihuman transthyretin (TTR) antiserum. Direct DNA sequencing of the proband’s T T R gene revealed a guanine-for-adenine substitution in the second base of codon 42, producing a glycine for glutamate substitution in the plasma protein. Murakami T, Yi S, Yamamoto K, Maruyama S, Araki S. Familial amyloidotic polyneuropathy: report of patients hererozygous for the transthyretin Gly”” gene. Ann Neurol 1992;31.340-342
Familial amyloidotic polyneuropathy (FAP) is an autosomal, dominantly inherited disorder characterized by the extracellular deposition of fibrillar amyloid pro-
tein and peripheral nerve involvement. The amyloid proteins of most types of FAP have been identified as variant types of transthyretin (?TR). The amyloid deposits derived from patients with FAP who are of Portuguese, Japanese, and Swedish origin were found to consist of a variant I T R with a single amino acid substitution of methionine for valine in position 30 of normal TTR El-31. Moreover, a number of FAP family groups throughout the world have genetic variants of ‘ITR E4- 101. Recently, Ueno and colleagues { 1 1, 121 identified three different variants of TTR (Glui2 to Gly, Ser5’ to Arg, and Tyrl’* to Cys) in Japanese patients with FAP by a combination of genomic amplification followed by cloning and sequencing. We studied 2 Japanese patients whose eldest brother was reported to have the variant GIu4’ to Gly and characterized this family with FAP.
Patient Reports The proband (111-3 in Fig 1) is a 38-year-old man whcise symptoms began at age 34 with chronic diarrhea followed several years later by orthostatic hypotension, as well as scnsory disturbance and weakness in the lower extremities. H e was admitted to a hospital in 1989, and amyloid was detected in a sural nerve biopsy specimen. Later that year, he was admitted to Tokyo Women’s Medical College Hospital. Clinical neurological examination revealed signs of polyneuropathy and bilateral Argyll Robertson pupils. His visual acuity was 0.7 in the right eye and 0.9 in the left. N o vitreous opacities were detected by ophthalmoscopic and slit-lamp examination. The elder brother is a 41-year-old man (111-2 in Fig I ) whose symptoms began at age 39 with muscle weakness in his lower extremities. In early 1980, he was admitted to Takeda Hospital in Kyoto, where clinical neurological examination revealed muscle weakness in the upper and lower extremities, sensory disturbance in the lower extremities, and orthostatic hypotension. His chest radiograph showed cardiac enlargement, and his echocardiogram showed diffuse hypertrophy and hypokinesis. No vitreous opacities were seen. Family history revealed that the proband’s mother (11-l), uncle (11-5), and maternal grandmother (1-2) had similar symptoms. The maternal grandmother was in Toyama Prefecture in Japan. His mother had died at age 57, and his uncle at age 48. Medical records were not available for review. The proband’s eldest brother (111-1) was reported to be 42 year!; old with symptoms of lower limb neuropathy and chronic: diarrhea. He was heterozygous for the ‘ITR Gly“’ gene 111.
Methods From the *First Department of Internal Medicine, Kurnamoto University Medical School, Kumamoto; and the ?Department of Neurology, Neurological Institute, Tokyo Women’s Medical College, Tokyo,Japan. Received Jun 28, 1991, and in revised form Aug 19 and Sep 9. Accepted for publicarion Sep 14, 1991. Address correspondence to Dr Murakami, the First Department of Internal Medicine, Kumamoto University Medical School, 1-1- I Honjo, Kumamoro 860, Japan.
340
A rectal biopsy specimen from the proband (111-3) was exam.ined histologically. Congo red stain, treatment with potassium permanganate (KMnO,,) by Wright’s staining method, and polarization microscopy were performed. For immunohistochemistry, the avidin-biotin complex method was employed, using antihuman TTR antiserum (Behrinperkc:, Marburg, Germany). Sections of kidney from a patient in Japan with FAP type 1 were also examined as control specimens.
Copyright 0 1092 by the American Neurological Association
Congenital Hypomyelination Neuropathy with Arthrogryposis Multiplex Congenita Kevin B. Boylan, MD,” Donna M. Ferriero, MD,t Claudia M. Greco, MD,$ R. Ann Sheldon, BA,t and Michael Dew, MD§
A term male infant is described with a n isolated disorder of peripheral myelination. A t necropsy, the great majority of medium-to-large axons were unmyelinated. Electron microscopy showed normal axons and redundant lamination of basement membrane, suggestive of early onion bulb pathology. Immunohistochemistry of peripheral nerve showed deficiency of the myelin proteins P2 and Po, myelin basic protein, and myelin-associated glycoprotein. Arrest of peripheral myelination at the promyelin stage appears to be the origin of myelin deficiency. Boylan IU3, Ferriero DM, Greco CM, Sheldon RA, Dew M. Congenital hypomyelination neuropathy with arthrogryposis multiplex congenita. Ann Neurol 1992;31:337-340
Deficiency or absence of peripheral myelin in newborns [ 1, 21, generally referred t o as congenital hypomyelinating neuropathy, is a rare cause of infantile neuromuscular weakness. Five patients with arthrogryposis muitiplex congenita with deficient peripheral myelin have been reported, b u t all differ from o u r patient in that 3 had associated central nervous system defects or congenital abnormalities apart from t h e arthrogryposis [3-51, and 2 had complete [ l ] or virtually complete [6J absence of peripheral myelin.
Patient Report The patient’s clinical features have been previously reported 171. Briefly, the infant was delivered at 38 weeks’ gestation,
was ventilator-dependent, had large joint flexion contractures with diffuse hypotonia, severe generalized weakness, and areflexia. Spinal fluid, karyotype, and neonatal inborn errors of metabolism tests were normal. Electroencephalography and brain and spine magnetic resonance images showed normal patterns for age. Sensory action potentials were unobtainable; motor conduction velocity was extremely slow at 1 to 2 m/sec (normal, 2 1 m/sec). Electromyography showed fibrillations and positive waves with absent motor unit potentials. Biopsy of vastus medialis at age 8 days showed variation in muscle fiber size and type 2 predominance in a pattern consistent with neurogenic change; neither grouped atrophy nor hypertrophy were present. Results of sural nerve biopsy are described. Death occurred at 5 weeks of age following withdrawal of ventilatory support. Autopsy was performed within 1 hour of death.
Materials and Methods Morphometric Analysis Morphometry was performed on biopsied sural nerve (fixed in 2.5% glutaraldehyde in 0.1 M cacodylate buffer, p H 7.4) and compared with published control data [Z, 83. Fiber counts were performed manually o n nonoverlapping electron micrographs of transverse sections (final enlargement, x 5,000 for myelinated axons and x 10,000 for unmyelinated axons). Axons in a one-to-one relationship with Schwann cells were counted as myelinated type axons even if the Schwann cell had not actually produced myelin membranes. Schwann cells with axons in this relationship would normally be expected to have begun myelination in a term infant f81. If the Schwann cell cytoplasm contained two or more axons, the axons were counted as unmyelinated. Axonal diameters were calculated using the “arithmetic” method of Dyck [9].
Immunoqytochemisytry Lumbosacral cord and roots from the patient were compared with control spinal cord from a similar-age infant without nervous system disease who died of hypoplastic left heart syndrome. Tissue was incubated overnight at 4°C using one of the following antibodies: antimyelin basic protein (MBP) at 1:1,000, anti-P2 at 1:500, anti-Po at 1:500, and antimyelin-associated glycoprotein (MAG) at 1:500. Antibody was visualized with the avidin-biotin-peroxidase method using the Vectastain ABC Elite kit (Vector Labs, Burlingame, CA). Diaminobenzidine (Sigma) was used as the chromagen.
Results From rhe *Deparrmenr of Neurology, Johns Hopkins Universiry, Balrimorc; rhe tDepdrrmenrs of Neurology and Pediatrics, University of California, and the tDeparrment of Neuropathology, Children’s Hospital, San Francisco, CA; and the $Departments of Psychiarry and Neurology, Dwight Eisenhower Army Medical Cenrer, Fort Gordon, G A 30905. Received Mar 6 , 1991, and in revised form Aug 8. Accepred for publication Sep 14, 1991. Address correspondence to D r Boylan, Deparrmenr of Neurology, Mayo Clinic Jacksonville, 4500 San Pablo Road, Jacksonville, FL 32224.
Premortem Studies Few fibers in t h e sural nerve biopsy w e r e myelinated (Fig 1A). Myelination, w h e n present, tended t o be thin relative to axon diameter, but myelin- membrane lamination was normal, as w e r e associated axons (Fig lB,C). Small unmyelinated fibers appeared normal. Lysosomal inclusion bodies w e r e n o t present. Redundant basal laminae surrounding nonmyelinated axons were c o m m o n (see Fig lB,C). Endoneurial collagen was not increased.
Copyright 0 1992 by the American Neurological Association
337
PERCENT OF AXONS
40
30
F i g I . Sural newe biopsy. (A) One pm-thick section of resin.embedded newe shouis striking diminution of myeiinated fibers. Schu~anncell nuclei appear increased in number. (Tohidine blue: x 120 before j292 reduction.) (B) Multiple single. nonmyelinated axons are surrounded by redundant bmal laminae and occasional extensions of Schwann cell cytoplasm. (Tohidine blue; x 7.500 before 525% reduction.) (C) A single, nonmyelinated axon, J p n in diameter, surrounded by two Schwann cell nuclei, u6ich may indicate Schwann cell mitotic ‘tctivity. This actizitj uJouldbe expected during fetal peripheral nerve development but would be abnormal in a term infant for this s h e axon. (Toluidine blue; x 7,500 before 52% reduction.) I n B and C. open arrows indicate redundant basal laminae thought to remain .from abortive attempts at myelination. and dotted arrows indicate fibroblast processes, distinguishable fkom Schuiann cell extensions because of the absence of basal laminae. Morphometric Data
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In surd nerve, 642 axons in a one-to-one relationship with Schwann cells were counted in a 2.52 x lo-’ mm’ cross-sectional area. Diameters ranged from 1 to 10 pm (Fig 2A). Twenty-six axons with diameters from 3 to 8 pm (4.0%’)were actually myelinated; the remainder were enwrapped with Schwann cell cytoplasm without elaboration of myelin membranes. The ratios of axon to fiber diameter (g ratio) [S] for the 26 myelinated axons were mostly higher than those reported for similar-age control infants (Fig 2C). In a crosssectional area of 5.24 x lo-’ mm’, 988 unmyelinated s o n s were counted. Diameters ranged from 0.2 to 2.6 pm (Fig 2B). Axon densities for myelinated type (1.94 x lo4 mm’) and unmyelinated axons (1.90 x lo5 mm’) were both normal [2, S}. In each distribution a subset of axons ( 5 % for myelinated arid 10% for unmyelinated) were up to 60% larger in diameter than the upper limit reported for control nerve, and the proportion of small diameter fibers was increased. Postmortem Studies
Brain and spinal cord were grossly normal. Motor neurons and dorsal root ganglia were histologically normal in appearance and number. Resin-embedded cross secdons of cervical and lumbar cord showed decreased myelinization of dorsal roots. There was a mild decrease in the number of myelinated fibers in ventral roots at both levels, and only rare, thinly myelinated fibers in sections of cauda equina. Distal median nerve showed severely delayed myelination compared with C7 dorsal and ventral roots (Fig 3). 338 Annals of Neurology Vol 31 No 3
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F i g 2. Diameter distributions for myelinated (A) and unmyelinated (Bi axons. Percentage of axons is plotted versus axon dimieter. Control data were redrawn (with permission from the authors and pzlbhher) from 181. iC) Twenty-six myelinatrd axons; g ratio is plotted versus axon diameter. The g ratio ues should generally be between 0.7 and 0.8 for children unu’er 3 years of age {8}. Z
J
~
A
B
C
Fig 3. C7 sensory root iA) and motor root (B) show slightly diminished myelination for age. Distal median nerve (C) shows a markedly decreased number of myelinated fibers. Also, axon diameters are decreased. Arrowhead in (B) and ICi shows an axon associated with t w o Schwann cell nuclei. The small c in (B) denotes a capillary. Filled arrows indicate unmyelinated large axons, and open arrows point t o myelinated axons. All sections are from resin-embedded autopsy tissue. IToluidine blue, x 480 before 52% reduction.)
lmmunocytocbemistly Sections through lumbosacral cord revealed a normal pattern of myelin MBP-like irnmunoreactivity. Transition areas through root entry zones revealed a markedly decreased imrnunostaining of P2, Po, and MBP in the peripheral nerve root, but only moderate deficiency of MAG (data not shown). The decreased immunostaining of the myelin proteins corresponded to the loss of myelinated nerve fibers (see Figs 1, 3) with no selective dropout of any of the myelin proteins tested.
Discussion This infant’s neuropathic disorder appeared to represent failure or severe delay of myelination at the promyelin stage. The promyelin stage is the phase of myelination in which Schwann cells have enwrapped single axons but have not yet begun myelin formation { 101. For axons of all diameters, the g ratios were relatively high; hypomyelination did not appear to be confined to any subset of axons distinguishable by size criteria. Redundant basal lamina and extensions of Schwann cell cytoplasm were relatively common in electron micrographs of sural nerve, which are similar to changes seen early in onion bulb formation in both human and animal forms of hereditary hypertrophic neuropathy [l l}. A rostral-caudal gradient in spinal root myelination is normal {12), but the degree of myelin deficiency in the roots, and especially in median and sural nerve, was quite abnormal. In addition, Figure 2 shows a marked difference in axon diameters between the spinal roots (which seemed normal) and the distal median nerve.
We found no published infant control data on this point, but in adults axon diameters in distal peripheral nerve are not substantially less than in corresponding spinal roots {9}. Similar to these observations in median nerve, a relative excess of smaller diameter axons was present in surd nerve (see Fig 2A,B). An increased proportion of smaller diameter axons therefore seems likely to have been a generalized condition. MAG is expressed by Schwann cells during the promyelin stage before full expression of Po, P2, and MBP [13}. There was no selective dropout of any of these proteins, so it seems unlikely that deficiency of any one protein led to diffuse peripheral hypomyelination. The moderate deficiency of MAG with severe dehciency of the remaining proteins supports the suggestion that hypomyelination resulted from developmental arrest at the promyelin stage, particularly given the presence of relatively normal myelination of some axons. Presented in part at the 42nd Annual Meeting of the American Academy of Neuroiogy, Miami Beach, FL, April 1990. The opinions or assertions contained herein are the private views of the authors and are not to be construed as official or as reflecting the views of the Army or the Department of Defense. We would like to thank Dr John Whitaker for the generous gift of anti-MBP; Dr Richard Quarles for the generous gift of anti-MAG; and D r Bruce Trapp for the generous gift of anti-Po and anti-P2. We also thank D r John Griffin and Dr Yuen So for helpful discussions regarding the manuscript.
References 1. Charnas L, Trapp B, Griffin J. Congenital absence of peripheral myelin: abnormal Schwann cell development causes lethal arthrogryposis multiplex congenita. Neurology 1988; 38:
966-974 2. Ouvrier R, McLeod JG, Pollard J, eds. Pevipherd neuroputhjj 292 rhildbood. New York: Raven, 1990:13-20 3. Hakamada S, Kumagai T, Hara K, et al. Congenital hypomyelinating neuropathy in a newborn. Neuropediatrics 1983;14: 182-183 4. Pleasure JR, Shuman S, Rorke LB. Congenital absence of peripheral nervous system (PNS) myelin presenting as arthrogryposis multiplex congenita (Part 11). Pediatr Res 1986;20: 465A 5. Seitz RJ, Wechsler W, Mosny DS, Lenard HG. Hypomyelination neuropathy in a female newborn presenting as arthrogryposis multiplex congenita. Neuropediatrics 1986;17: 132-136 6. Palix C, Coignet J. Un cas de polyneuropathie peripherique neonatale par myelination. Pediatrie 1978;33:201-207 7. Boylan K, Dew M, Greco C, Ferriero D. Hypomyelination neuropathy: a rare form of neonatal neuromuscular weakness. Neurology 1990;40:409A 8. Jacobs JM, Love S. Qualitative and quantitative morphology of human sural nerve at different ages. Brain 1985;108:837-924 9. Dyck PJ. Parhologic alterations of the peripheral nervous system in humans: In: Dyck PJ, Thomas PK, Lamberr EH, Bunge R, eds. Peviphevuf Neuroputhjj. Philadelphia: W.B. Saunders, 19841760-870
Brief Communication: Boylan et al: Congenital Hypomyelination Neuropathy
339
10. Friede RL, Samorajski T. Myelin formation in the sciatic: nerve of the rat. A quantitative electron microscopic and radioauto-
graphic study. J Neuropathol Exp Neurol 1968;27:546--570 11. Low PA. The evolution of “onion bulbs” in the hereditary hy-
pertrophic neuropathy of the trembler mouse. Neuropathol Appl Neurobiol 1977;3:81-92 12. Dobosz-Niebrol I, Fidzianska A, RafalowskaJ, Sawicka E. Correlative biochemical and morphological studies of myelination in human ontogenesis. 11. Myelination of the nerve roots. Acta Neuropathol 1980;49:153-158 13. Willison HJ, Ilyas AL. OShannessy DJ, et al. Myelin-associated glycoprotein and related glycoconjugates in developing cat peripheral nerve: a correlative biochemical and morphometric study. J Neurochern 1987;49:1853-1862
Familial Amyloidotic Polyneuropathy: Report of Patients Heterozygous for the Transthyretin Gly42Gene Tatsufumi Murakami, MD,” Shigehiro Yi, MD,” Kenji Yamamoto, MDJ Shoichi Maruyama, MD,t and Shukuro Araki. MD”
We studied 2 patients from aJapanese family with familial amyloidotic polyneuropathy (FAP). Their clinical features are similar to type 1 FAP, and the proband’s rectal tissue contained amyloid that stained with antihuman transthyretin (TTR) antiserum. Direct DNA sequencing of the proband’s T T R gene revealed a guanine-for-adenine substitution in the second base of codon 42, producing a glycine for glutamate substitution in the plasma protein. Murakami T, Yi S, Yamamoto K, Maruyama S, Araki S. Familial amyloidotic polyneuropathy: report of patients hererozygous for the transthyretin Gly”” gene. Ann Neurol 1992;31.340-342
Familial amyloidotic polyneuropathy (FAP) is an autosomal, dominantly inherited disorder characterized by the extracellular deposition of fibrillar amyloid pro-
tein and peripheral nerve involvement. The amyloid proteins of most types of FAP have been identified as variant types of transthyretin (?TR). The amyloid deposits derived from patients with FAP who are of Portuguese, Japanese, and Swedish origin were found to consist of a variant I T R with a single amino acid substitution of methionine for valine in position 30 of normal TTR El-31. Moreover, a number of FAP family groups throughout the world have genetic variants of ‘ITR E4- 101. Recently, Ueno and colleagues { 1 1, 121 identified three different variants of TTR (Glui2 to Gly, Ser5’ to Arg, and Tyrl’* to Cys) in Japanese patients with FAP by a combination of genomic amplification followed by cloning and sequencing. We studied 2 Japanese patients whose eldest brother was reported to have the variant GIu4’ to Gly and characterized this family with FAP.
Patient Reports The proband (111-3 in Fig 1) is a 38-year-old man whcise symptoms began at age 34 with chronic diarrhea followed several years later by orthostatic hypotension, as well as scnsory disturbance and weakness in the lower extremities. H e was admitted to a hospital in 1989, and amyloid was detected in a sural nerve biopsy specimen. Later that year, he was admitted to Tokyo Women’s Medical College Hospital. Clinical neurological examination revealed signs of polyneuropathy and bilateral Argyll Robertson pupils. His visual acuity was 0.7 in the right eye and 0.9 in the left. N o vitreous opacities were detected by ophthalmoscopic and slit-lamp examination. The elder brother is a 41-year-old man (111-2 in Fig I ) whose symptoms began at age 39 with muscle weakness in his lower extremities. In early 1980, he was admitted to Takeda Hospital in Kyoto, where clinical neurological examination revealed muscle weakness in the upper and lower extremities, sensory disturbance in the lower extremities, and orthostatic hypotension. His chest radiograph showed cardiac enlargement, and his echocardiogram showed diffuse hypertrophy and hypokinesis. No vitreous opacities were seen. Family history revealed that the proband’s mother (11-l), uncle (11-5), and maternal grandmother (1-2) had similar symptoms. The maternal grandmother was in Toyama Prefecture in Japan. His mother had died at age 57, and his uncle at age 48. Medical records were not available for review. The proband’s eldest brother (111-1) was reported to be 42 year!; old with symptoms of lower limb neuropathy and chronic: diarrhea. He was heterozygous for the ‘ITR Gly“’ gene 111.
Methods From the *First Department of Internal Medicine, Kurnamoto University Medical School, Kumamoto; and the ?Department of Neurology, Neurological Institute, Tokyo Women’s Medical College, Tokyo,Japan. Received Jun 28, 1991, and in revised form Aug 19 and Sep 9. Accepted for publicarion Sep 14, 1991. Address correspondence to Dr Murakami, the First Department of Internal Medicine, Kumamoto University Medical School, 1-1- I Honjo, Kumamoro 860, Japan.
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A rectal biopsy specimen from the proband (111-3) was exam.ined histologically. Congo red stain, treatment with potassium permanganate (KMnO,,) by Wright’s staining method, and polarization microscopy were performed. For immunohistochemistry, the avidin-biotin complex method was employed, using antihuman TTR antiserum (Behrinperkc:, Marburg, Germany). Sections of kidney from a patient in Japan with FAP type 1 were also examined as control specimens.
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