Clinical and Biochemical Aspects of Trichopoliodystrophy Warren D. Grover, MD, Waine C . Johnson, MD, and Robert I. Henkin, M D ~
~~
~
The clinical and biochemical evaluation of 6 patients with trichopoliodystrophy indicates that the disease process can begin in utero and is related to a selective abnormality in copper metabolism. Examination of 2 infants on the first day of life revealed abnormal neurological signs, a characteristic hair abnormality, and elevated levels of copper and ceruloplasmin. Decreased hepatic copper levels and increased urinary copper excretion were documented during the first week. The 2 neonates demonstrated a progressive decrease in blood copper levels in the first month of life. Four infants identified at ages 2 to 11 months had low values for blood copper and ceruloplasmin. All infants had progressive neurological dysfunction, and 4 of the 6 died at ages ranging from 2$(2 months to 5v2 years. Parenteral copper therapy achieved normal blood and hepatic copper levels in 1 patient, but the copper values in the cerebral cortex and white matter were significantly decreased compared to control specimens. Grover WD, Johnson WC, Henkin RI: Clinical and biochemical aspects of trichopoliodystrophy. Ann Neurol 5:65-7 1, 1979
Trichopoliodystrophy (TPD, Menkes disease, kinky hair disease, steely hair disease) is a sex-linked neurodegenerative disorder that was first described by Menkes et a1 in 1762 [171 and has been extensively studied by several groups of investigators [2, 3 , 7, 12, 18, 25, 261, who have described neurological, integumental, vascular, genitourinary, and biochemical abnormalities. Over the past five years we have evaluated 6 patients with the disorder, and in this paper we discuss their clinical and biochemical findings. O u r data suggest that the disease can begin in utero and may be recognized at birth. T h e biochemical rationale for this suggestion is based o n selective abnormalities in copper metabolism found in these children.
Materials and Methods The subjects of the study were 6 patients with T P D (Table 1). Five were white and 1 was black. All had birth weights in normal ranges for gestational age. All were males, and 3 were firstborn. Children from five different families were studied; in one family 2 successive male children (Patients 2 and 3) exhibited the disease. Fetal wastage, sudden infant death, and mental retardation were observed in the remaining siblings in two families, and in the other families the index case was the firstborn child. The age at the time diagnosis was established varied from birth to 11 months. Two patients were the product of complicated gestational courses, and all had exhibited some abnormalities during the perinatal period. Each of the 4 patients whose disorder From S t . Christopher's Hospital for Children, Temple University School of Medicine, and the Skin and Cancer Hospital, Temple University School of Medicine, Philadelphia, PA, and the Center for Molecular Nutrition and Sensory Disorders and Georgetown University Medical Center, Washington, DC.
was detected after birth had a history of major motor seizures at the time of diagnosis. Four of the 6 children exhibited icterus in the first 48 hours of life (see Table 1). Before an exchange transfusion was performed, Patient 1 had a total serum bilirubin level of 19.0 m d d l with a direct bilirubin of 2 mgldl. In Patient 2, total serum bilirubin was 14.0 mg/dl with a direct bilirubin of 2 mg/dl at 48 hours of age; this level gradually decreased to a normal value over a ten-day period. Blood and urine levels of copper and zinc were obtained from our 6 patients and from 4 additional patients with TPD who were evaluated at other institutions. Blood, urine, and cerebrospinal fluid were obtained from 12 children who were age-matched with the 10 patients who had TPD. T h e control specimens were obtained from children hospitalized at St. Christopher's Hospital for Children with a variety of conditions, including seizures of unknown cause ( 6 patients), progressive spinomuscular atrophy, suspected central nervous system infections, static neurological dysfunction of undefined cause, and failure to thrive of unknown cause. Liver copper was measured in biopsy specimens obtained with a Vim-Silverman needle in 3 patients with TPD; approximately 10 mg of tissue was obtained in each biopsy. Copper values were measured in specimens of central nervous system obtained at the autopsies of Patient 3 and an infant 8 months of age who died with the sudden infant death syndrome. Control values for liver copper were measured in specimens obtained at autopsy from 3 infants aged 2 to 7 weeks; 2 had died of congenital heart disease without congestive heart failure and the third of sudden infant death syndrome. Autopsy samples of brain and liver conAccepted for publication May 30, 1978. Address reprint requests Dr Grover, 2600 Philadelphia, PA 19133,
Lawrence St,
0364-5134/79/010065-07$01.25 @ 1978 by Warren D . Grover 65
Table 1 . Clinical Status of Patients with Trirhopoliodystrophy ~~
~~
Diagnostic Features Serum Values Patient No.
Age (mo)
Neurologcal Signs
1
Binh
2
Blnh
3
2
4
8
5
2
Hypotoma. hypothermia, absent head control Hypotonla. increased DTR, absent head control Seizures. hypotonia with increased M R , absent head control Seizures. spasticity. retardation, strabismus Seizures. spastictry. retardation
6
11
n = normd (mean
Seizures, retardation k
Hepatic Cu (gg/gm dry art)
Roent&nographic Signs
Stigmata
Cu (ctddb
Ceruloplumin (mpjdl)
Skull
Long Bones
Bladder
Kinky hair, taliper equinovuur
84 (n, 29 z 3)
30 (n, 5
?
2)
14
Wormian bones
None
Dweniculi
Kinky hair
60 (n. 29 5 3)
26 (n, 5
?
2)
7
Wormian boner
None
Diveniculi
Kinky hair
12 (n, 65
12 (n, 15-30)
12
Worminn bones
Metaphysed spurring. humerus
N o chaues
Kinky hair
30 (n,90- 1 15)
19(after IV Cu for 12 mo)
Worm~an bones
None
Dlveruculi
Kinky hair
35 (n. 65
13 (n, 15-30)
NE
Wormian bones
None
NE
Kinky hair
56 (n, 130 2 1 0 )
10 (n, 25-60)
NE
Wormian
Metaphysed spurring. femur
NE
SFM or range); DTR
?
?
5)
5)
4 (n. 25-50)
h W S
=
deep tendon reflexes. NE = not examined.
sisted of approximately 100 mg of tissue obtained by excision with a dry stainless steel knife previously washed with EDTA and distilled water. Serum, urine, and cerebrospinal fluid copper and zinc levels were measured by flame-aspiration atomic absorption spectrophotometry by a method previously described 1201. Liver copper concentrations were measured by
flame-aspiration atomic absorption spectrophotometry after the tissue had been dried in an oven at 75°C for 18 hours and digested with nitric acid [ 111. Ceruloplasmin was measured by the copper oxidase method as modified for automatic analysis [ 2 3 ] . Skin and hair were obtained from 3 patients with TPD and from control subjects of similar age. The methods used included examination of histological sections of skin and hair stained with hematoxylin-eosin and examination of hairs using polarized light, dark-field illumination. and dark-field autofluorescence.
Results Clinical Findings At the time of diagnosis, each patient had physical findings and laboratory data indicating TPD to the alert physician. Each infant had a similar appearance, characterized by cherubism, a depressed nasal bridge, and decreased facial movement. All patients, including the 2 newborns, had steely, depigmented hair characteristic of the disease. All had definite neurological signs, including hypotonia, poor head con66 Annals of Neurology Vol 5 No 1 January 1979
trol, and abnormalities of deep tendon reflexes. Roentgenographic changes included wormian bones in the skull in all 6 patients, metaphyseal spurring of the long bones in 2, and large, lobular diverticula in the bladder in 3 [ 121. A cerebral angiogram and computerized axial tomogram performed on Patient 1 at 6 weeks and 18 months, respectively, were normal. A computerized axial tomogram obtained on Patient 4 at 2 years of age revealed decreased cortical mass and generalized ventricular dilatation (Fig 1). Electroencephalograms recorded from Patient 1 from 2 weeks to 5 years of age showed no abnormality. Electroencephalograms obtained on Patient 4 at 18 months and 2 years of age demonstrated only mild slow activity. Recordings from scalp electrodes on Patients 2 and 3 revealed diffuse slow-wave activity with admixed paroxysmal features. Normal computerized averaged responses were recorded from electroretinograms of Patient 3 at 10 months of age. Study of specimens of hair and scalp from 3 patients (Nos. 2, 3, and 4) revealed similar abnormalities consistent with a defect in the hair protein matrix. Sections of the scalp stained with hematoxylin-eosin demonstrated anagen (growing)-phase follicles with a slight to moderate hair curvature and a number of follicles that did not contain a well-formed hair. Examination of scalp hair mounted
Race
Family History
Birth Weight (kg)-
White
Sib died suddenly at 8 mo
2.9
None
Icterus (exchange transfusion)
Living, 48 m o
White
Sib with T P D (Patient 3)
2.6
None
Icterus, hypotonia, hypothermia
Died, 2% m o
White
Sib with T P D (Patient 2)
2.7
None
Icterus
Died, 14 mo
White
None
2.9
None
Hypotonia
Living, 25 mo
White
Sib retarded, sib stillborn, and 6 sibs living and well
3.5
Premature rupture of membranes
Icterus (phototherapy)
Died, 5% yi
Black
2 paternal nephews with friable, kinky hair
2.2
Hypertension; bleeding, last trimester
Muconium staining; hypothermia
Died, 3% yr
on slides revealed intertwined, tortuous, thin, fragile-appearing structures with occasional fracturing and twisting that produced a bead-like effect. Inspection of hair with fluorescent and dark-field microscopy demonstrated replacement of the normal pink to red fluorescence with a whitish color. With polarized light, hair from the patients also appeared white instead of the light brown observed in controls. After administration of copper salts for a nine-month period in 1 patient (No. l), partial return to a normal hair color was observed. Four patients died at 295 months to 5% years of age. At postmortem examination, bilateral bronchopneumonia was observed in each child. Neuropathological abnormalities were found in 2 of them. These included small brain size and weight with cerebral and cerebellar atrophy. Histological findings were similar to abnormalities reported by other investigators [71 and included loss of subcortical myelin, reactive gliosis, and neuron loss in the cerebral hemispheres and in the molecular, Purkinje, and granular cell layers of the cerebellum. The spinal cord revealed bilateral spinocerebellar tract degeneration with varying degrees of gliosis. The cerebral and cerebellar vessels showed areas of splitting and beading of elastic tissue of the intima and of the internal elastic membrane.
Complications Gestational
Perinatal
Course
Both patients surviving at this time demonstrate neurological dysfunction. Patient 1, at 4 years of age, has marked hypotonia, no language, and adaptive abilities at a 1-year level. Patient 4 , at age 3 years, has spastic quadriparesis, bilateral esotroyia, and a social smile. The steely hair persists in each patient.
Biochemical Findings Measurements of trace metals in each patient with T P D studied beyond the first week of life revealed serum copper and ceruloplasmin concentrations below normal levels (Tables 1, 2). Mean serum copper and ceruloplasmin concentrations in 10 patients with TPD ranging in age from 1 week to 5 years were significantly lower (p < 0.001) than values in control specimens (Table 2). No difference in serum zinc concentration was observed between patients and unaffected infants and children. In each untreated patient with TPD, urinary copper excretion was significantly greater than control levels (p < 0.001); this abnormality was found in the first specimen from each patient. No differences were observed in urinary zinc excretion measured simultaneously (Table 2). Cerebrospinal fluid copper and zinc concentrations were not significantly different in the two groups of children. Hepatic copper concentrations were measured in 3
Grover, Johnson, and Henkin: Trichopoliodystrophy
67
patients before treatment (Table 3). The liver copper values in the patients were significantly lower than in age-matched controls (p < 0.001). Copper metabolism was also studied in 2 patients (Nos. 1 and 2) before and immediately after delivery (Table 4). At term, blood and amniotic fluid from the mother of Patient 2 were analyzed for copper, ceruloplasmin, and zinc concentrations. The amniotic fluid copper level was over four times higher than the mean level measured in mothers of normal infants at term, whereas the zinc level in the amniotic fluid was within the wide range of normal values. There were no differences in serum copper, ceruloplasmin, or zinc concentration between the mother of Patient 2 and the mothers of normal children. Initial copper concentrations obtained in cord blood in both patients were greater than twice the upper limit measured in normal term infants: 60 pg/dl in Patient l and 8 4 p d d l in Patient 2 (normal, 29 ? 3 pg/dl) [ 151. In Patient 2 the serum copper concentration fell rapidly over the first week of life, in contrast to the increase in serum copper concentration that occurs in unaffected newborns [ 161 (Fig 2). The serum copper concentration in Patient 1 was 19 pg/dl at 21 days of age, a time when the mean copper concentration in normal infants is approximately 80 pg/dl. Serum ceruloplasmin values paralleled the copper levels. Repeated measurements of urinary copper excretion during the first two weeks of life averaged 7.0 pgl24 hrlkg body weight (normal, < 1.0 pg/kg124 hr) with zinc excretion at 4.0 pg/kg/24 hour (normal, 3.8 pgj kgJ24 hr).
F i g 1 . Compziterized axial tomograni in Patient 4 at 2 years of age detnomtrutes mild i*entrirzilardilatation and an increase in .culci ronsiJtent wirh atrophy.
Tdble 2. Capper atid Zinc Leieh in Serum, Urine, and Cerebrospinal Fluid in 10 Patient.r with Untreated TPD" ~~~~~~
~
~~
Serum Values
Patient No.
cu (ddl)
Zn (pg/dl)
Ceruloplasrnin (mddl)
1
16
80
4
2
43
97
12
3
25 32 30 20 13 27 20
... ...
4 14 8 4 3 5
4 5 6 7
8 9 10 Mean
134 102 73 92 68 106
39 &
SEM
Controls (N=12), age matched
... ...
cu
Zn
cu
Zn
3.8 7.4 4.7
13.7
4.8 ... 4.1
4.5 ...
...
4.0 ... ...
4.6 4.8
7.3 15.0
7
7& 1
5.0
100 t 5
80
?
7
32 & 7
1.1
"All values after first week of life.
68 Annals of Neurology Vol 5 N o 1 January 1979
0.4
8.0 2 2.1
3.4
2
1
0.3
9.8
4.0
&
1
4.0 ...
&
&
.o
...
4.0
5.4 ... ...
94
2
... ... 1
2.0 2.5
... ... ... ... ...
4.1
3
26
CSF Values (pddl)
Urine Values (pg/24 hdkg body wt)
&
3.2
... ... ... ... ...
4.4 2 1
Tdble in
.j
.$. Lher Capper Conrentration Plrtients u i t h IJntredted T P D
Hepatic Cu Patient No.
(&gm dry wt)
1
14
7
7 12
.3
Mcan
?
1 SEM
Controls tN =3), age matched
I
11 2 2
226
&
57
45A
3s
Discussion Synthesis of the biochemical findings and clinical data obtained from the 6 patients with TPD indicates a possible mechanism for thc &case process that has not been previously emphasized (Fig 3 ) . Important facts include the documentation of elevated levels of copper in amniotic tluid, cord blood, and urinc of
I
j
a
25
FiK 2. Serial blood copper detrrininutio)is in Patient 2. Poimti repre.ceni ropper lei,elc I p,gidt).Dotted ciiri'e oiitlines nort t d ICI e l l .
newborn infants but hepatic copper values only 1057
of control levels. Abnormalities in neurological function, hair structure, bone formation, and hepatic function were present in the imme'iiate perinatal period and indicated organ dysfunction in utero. Untreated patients beyonci the newborn period also exhibited low concentrations of serum copper, ccruloplasmin, and hepatic copper but increased urinary copper excretion. T h e zinc lcvcls in all fluids and tissucs studied were within the average range, in&cating normal zinc metabolism. These data suggest the presence of a selective defect in tissue uptake o r rctcntion of copper or both. Correlative evidence has been obtained by H o r n et al, w h o measured tissue copper concentrations in a fetus with TPD and compared the results with those from control patients of a similar age [ 1'1. They
found that tissue levcls of copper in kidney, spleen. pancreas, placenta, brain, lung, muscle, and skin were at least twice control values. They also noted that the liver copper value in the patient was one-thin1 of control level. In the normal newborn, copper levels in liver are several times higher than values from normal adults. H e p i t i c copper in the newborn is rcportedly sequestered in lysosomes [9] and in mitochondria in the form of a protein, hcpatomitochon~lriocuprcin[ 2 11. We Actermined that blood levcls of coppcr and ceruloplasmin in newborn patients with TPD were at least twice normal values and that the copper level in amniotic fluid in thc mother of 1 of thcse patients was four times the normal mean value at tcrm. O u r studies indicate that liver copper in the
?'able 4. Comparison of Srrilni Copper, Cerzilaplasmin. and Zinc Conmitrations and Aniniotir Fluid Copper and Zinr Vdlues in Meii,horm with TPD. the Mother nf'riti Aflicted 1nJ;itit. cind Control Siikjtrt,
Ncwborns
Morhcrs 'L'PD Determination
Normal" ( N = 15)
TPD"
Normal" ( N = 15)
221 2 14 91 2 4 48 2 3
245
29
5
90
10
5
55
83
5
3
0 2 3 2 12
20 12
Patient 1
Patient 2
3
00
2
23 05
84 30
Blood
cu ( p d d l ) Ceruloplasmin (rnddl) Zn (pgldl) Amniotic fluid CU
(pgldl)
Zn (pd.11)
32
"Mean 2 1 SEM. 'The mother of Patient 2.
Grover, Johnson, and Hcnkin: lrichopoliodysrrophy 69
ANTENATAL
[Kidney
I
TISSUES MA T E R N A L ~ F E T A L
Brain AMNIOTIC FLUID
NEONATAL TISSUES -*URINARY .)6INDING & STORAGE .)GI ABSORPTION
INFANCY
Cu
TISSUE
‘TOTAL
BODY
Fig 3. Postulatedschema of copper metabolism i n T P D . The transplacental kinetics of copper appears intact, but abnormal copper transport or binding (or both) a t the cellular level results in a n increase in copper concentrations in alltissues except the liver. The excess copper is eventually excreted by the fetal kidney and causes an increase i n amniotic copper levels. A t birth, the defect in copper binding or transport causes a persistent loss of copper from cells anddecreased absorption of copperfrom the gastrointestinal tract, resulting in low blood and tissue copper levels w i t h increased urinary excretion.
patients is one-tenth of normal levels of age-matched controls at a time when serum copper concentration is more than twice comparable levels in normal newborns. These data suggest that copper is available to all tissues of the fetus but that transport and retention vary with individual cell systems. Other data suggesting a defect in copper transport or binding (or both) was obtained by Goka et al [8], who demonstrated that cultured fibroblasts from patients with TPD accumulated more copper than cells from normal subjects. The copper values in cultured fibroblasts from 3 patients in this study (Nos. 1, 2, and 3) were approximately four times the copper content of fibroblasts from normal subjects. These data are consistent with the hypothesis of a selective tissue defect in the uptake or retention (or both) of copper in tissues of patients with TPD. The concept of a selective tissue defect in the uptake or retention of copper in this disease has a correlate in some forms of copper deficiency previously observed in animals. In enzootic ataxia or swayback, ewes are anemic but have normal wool; their lambs exhibit ataxia, blindness, depigmentation, and loss of crimp of their wool [b]. Cattle suffering from copper deficiency commonly die suddenly from “falling disease,” attributed to sudden heart failure after exercise or excitement [l]. In our patients and in those reported by others, there is no anemia in spite of an apparent severe deficiency of copper. We have identified no patient who died of cardiovascular causes. Our data and information provided by other in-
70 Annals of Neurology Vol 5
No 1 January 1979
vestigators indicate a spectrum of activity of copperdependent enzyme systems in patients with TPD. We have documented renal tubular acidosis with normal tubular clearance of phosphorus and normal amino acid excretion in 2 of our patients (Nos. 1 and 3). Renal dysfunction was not reversed following therapy with copper that increased serum, hepatic, and renal levels of copper. We have also documented defective oxidative metabolism of mitochondria [ 113 obtained from muscle and low values of serum dopamine /3-hydroxylase [ 131. However, Zelkowitz et al 1271 have noted normal utilization of oxygen by muscle mitochondria and normal values for superoxide dismutase, which suggest genetic heterogeneity resulting in variable biochemical abnormali ties. Selective changes in tissue copper metabolism occurred after treatment of these patients. Parenteral therapy with copper salts or oral therapy with copper-nitriloacetic acid produced increases in serum copper and ceruloplasmin [4, 5, 10, 11, 14,261, urine copper [ l o , 141, and hepatic and renal copper, improved function of some copper-dependent oxidative enzymes, and increased cerebrospinal fluid copper levels [ 113 but did not raise copper values in brain [lo, 11, 141. In 3 untreated patients with TPD, copper in cerebral white matter and cortex ranged from 4 to 9.5 &gm dry tissue [22, 261. In our Patient 3 [ 101, even following therapy that achieved a return of normal copper levels in liver and elevated copper levels in kidney, the copper concentration in cerebral white matter was less than 1 p g / p dry tissue. A comparable specimen from an age-matched control patient contained 18 &gm dry tissue. However, copper levels in the medulla of both the patient and the control specimen were similar, implying that the defect may be selective even within the brain. Because the brain is the only organ in humans in which increasing concentrations of copper occur with age [24], this organ has an important need for copper, particularly in the first years of life when active myelination occurs.
Supported in part by HEW Training Grant I TO1 NSI0072-OIA2, NSRA, Grant RR-75 from the General Clinical Research Center, and Grant PO1 CA11536. Presented in part at the 1974 Annual Meeting of the Child Neurology Society, Madison, WI, and the 1976 meetings of the American Pediatric Society, the Society for Pediatric Research, and the American Society of Clinical Investigators. The authors wish to acknowledge the following contributions: Cyril D’Cruz, MD, and Guillermo A. deleon, MD, for pathological examinations; Hope Punnett, PhD, and Nancy Brodsky, PhD, for fibroblast cultures; and the Pennridge Pediatric Group for the identification of the 2 afflicted newborns.
References 1. Bennetts HW, Beck AB, Hurley R: Pathogenesis of "falling disease." Aust Vet J 24237-244, 1948 2. Bucknall WE, Haslam RH, Holtzmann NA: Kinky hair syndrome: response to copper therapy. Pediatrics 52:653-657, 1973 3. Danks DM, Campbell PE, Stevens BJ, et al: Menkes kinky hair syndrome: an inherited defect in copper absorption with widespread effects. J Pediatr 50:188-201, 1972 4. Danks DM. Campbell PE, Walker-Smith J, et al: Menkes' kinky hair syndrome. Lancet 1:llOO-1102, 1972 5 . Dekaban AS: Menkes' kinky hair disease treated with subcutaneous copper sulfate. Lancet 2:1523, 1974 6. Dick AT: Preliminary observations on the effect of high in. take of inorganic sulfate on blood copper & fleece character in cross bred sheep. Aust Vet J 30:196-202, 1968 7. French JH, Sherrard ES, Lubell H , et al: Trichopoliodystrophy: report of a case and biochemical studies. Arch Neurol 26:229-244, 1972 8. Goka TJ, Stevenson RE, Herreran PM, et al: Menkes disease: a biochemical abnormality in cultured human fibroblasts. Proc Natl Acad Sci USA 73:604-606, 1976 9. Goldfischer S, Bernstein JJ: Lipofuscin (aging) pigment granules of the newborn human liver. J Cell Biol42:253-261, I969 10. Grover WD, Henkin RI: Trichopoliodystrophy (TPD): a fetal disorder of copper (Cu) metabolism. Pediatr Res 10:448. 1976 11. Grover WD, Scrurton MC: Copper therapy in trichopoliodystrophy. J Pediatr 86:2 16-220, 1975 12. Harcke T H , Capitanio MA, Grover WD, et al: Bladder diverticuli and Menkes' syndrome. Radiology 124:459-461, 1977 13. Henkin RI, Grover WD: Trichopoliodystrophy (T'PD): new aspects of pathology and treatment in trace element merabolism in man and animals 111, in Kirchgessner M (ed): Arbeitskreis fur Tierernahrungsforschung. Weihenstephan. 1978. pp 405-408 14. Henkin RI, Grover WD: Trichopoliodystrophy (TPD): a genetic defect in selective Cu binding and storage as well as transpon. Clin Res 24:435A, 1976
15. Henkin RI, Marshall J, Meret S: Maternal-fetal metabolism of copper and zinc at term. Am J Obstet Gynecol 110: 131- 134, 1971 16. Henkin RI, Schulman JD, Schulman CB, et al: Changes in total nondiffusible and diffusible plasma zinc and copper during infancy. J Pediatr 82:831-837, 1973 17. Horn N , Mikkelsen M, Heydorn K, et al: Copper and steely hair. Lancet 1:1236. 1975 18. Lott IT, DiPaolo R, Schwanz D, et al: Copper metabolism in the steely hair syndrome. N Engl J Med 292:197-199, 1975 19. Menkes JH. Alter M, Steigleder GK, et al: A sex-linked recessive disorder with retardation, peculiar hair and focal cerebral and cerebellar degeneration. Pediatrics 29:764-779, 1962 20. Meret S. Henkin RI: Simultaneous direct estimation by atomic absorption spectrophotometry of copper and zinc in serum, urine and cerebrospinal fluid. Clin Chem 17:369-373, 197 1 2 1. Porter H: Neonatal hepatic mitochondrocuprein. 3: Solubilization of the copper and protein from mitochondria of newborn liver by reduction with mercaproerhanol. Biochim Biophys Acta lS4:236-238, 1968 22. Reske-Nielson E, Loy HOC, Andersen P, et al: Brain-copper concentration in Menkes disease. Lancet 1:613, 1973 23. Scheinberg IH, Morel1 AG: Exchange ceruloplasmin copper with ionic CU'" with reference to Wilson's disease. J Clin Invest 36:1193-1201, 1957 24. Schroeder HA, Nason AP, Tipton IH, et al: Essential trace metals in man: copper. J Chronic Dis 19:1007-1034, 1966 25. Singh S, Bresnan MJ: Menkes kinky hair syndrome. Am J Dis Child 125:572-578. 1973 26. Walker-Smith JA, Turner B, Blomfield J, et al: Therapeutic implications of copper deficiency in Menkes' steely hair syndrome. Arch Dis Child 48958-962, 1973 27. Zelkowitz M, Cote L. Miranda A, et al: Trichopoliodystrophy: ultracrructure and biochemical studies in muscle. Presented at the Annual Meeting of the Child Neurology Society, Monterey, CA 1976
Grover, Johnson, and Henkin: Trichopoliodystrophy
71
~~
~
The clinical and biochemical evaluation of 6 patients with trichopoliodystrophy indicates that the disease process can begin in utero and is related to a selective abnormality in copper metabolism. Examination of 2 infants on the first day of life revealed abnormal neurological signs, a characteristic hair abnormality, and elevated levels of copper and ceruloplasmin. Decreased hepatic copper levels and increased urinary copper excretion were documented during the first week. The 2 neonates demonstrated a progressive decrease in blood copper levels in the first month of life. Four infants identified at ages 2 to 11 months had low values for blood copper and ceruloplasmin. All infants had progressive neurological dysfunction, and 4 of the 6 died at ages ranging from 2$(2 months to 5v2 years. Parenteral copper therapy achieved normal blood and hepatic copper levels in 1 patient, but the copper values in the cerebral cortex and white matter were significantly decreased compared to control specimens. Grover WD, Johnson WC, Henkin RI: Clinical and biochemical aspects of trichopoliodystrophy. Ann Neurol 5:65-7 1, 1979
Trichopoliodystrophy (TPD, Menkes disease, kinky hair disease, steely hair disease) is a sex-linked neurodegenerative disorder that was first described by Menkes et a1 in 1762 [171 and has been extensively studied by several groups of investigators [2, 3 , 7, 12, 18, 25, 261, who have described neurological, integumental, vascular, genitourinary, and biochemical abnormalities. Over the past five years we have evaluated 6 patients with the disorder, and in this paper we discuss their clinical and biochemical findings. O u r data suggest that the disease can begin in utero and may be recognized at birth. T h e biochemical rationale for this suggestion is based o n selective abnormalities in copper metabolism found in these children.
Materials and Methods The subjects of the study were 6 patients with T P D (Table 1). Five were white and 1 was black. All had birth weights in normal ranges for gestational age. All were males, and 3 were firstborn. Children from five different families were studied; in one family 2 successive male children (Patients 2 and 3) exhibited the disease. Fetal wastage, sudden infant death, and mental retardation were observed in the remaining siblings in two families, and in the other families the index case was the firstborn child. The age at the time diagnosis was established varied from birth to 11 months. Two patients were the product of complicated gestational courses, and all had exhibited some abnormalities during the perinatal period. Each of the 4 patients whose disorder From S t . Christopher's Hospital for Children, Temple University School of Medicine, and the Skin and Cancer Hospital, Temple University School of Medicine, Philadelphia, PA, and the Center for Molecular Nutrition and Sensory Disorders and Georgetown University Medical Center, Washington, DC.
was detected after birth had a history of major motor seizures at the time of diagnosis. Four of the 6 children exhibited icterus in the first 48 hours of life (see Table 1). Before an exchange transfusion was performed, Patient 1 had a total serum bilirubin level of 19.0 m d d l with a direct bilirubin of 2 mgldl. In Patient 2, total serum bilirubin was 14.0 mg/dl with a direct bilirubin of 2 mg/dl at 48 hours of age; this level gradually decreased to a normal value over a ten-day period. Blood and urine levels of copper and zinc were obtained from our 6 patients and from 4 additional patients with TPD who were evaluated at other institutions. Blood, urine, and cerebrospinal fluid were obtained from 12 children who were age-matched with the 10 patients who had TPD. T h e control specimens were obtained from children hospitalized at St. Christopher's Hospital for Children with a variety of conditions, including seizures of unknown cause ( 6 patients), progressive spinomuscular atrophy, suspected central nervous system infections, static neurological dysfunction of undefined cause, and failure to thrive of unknown cause. Liver copper was measured in biopsy specimens obtained with a Vim-Silverman needle in 3 patients with TPD; approximately 10 mg of tissue was obtained in each biopsy. Copper values were measured in specimens of central nervous system obtained at the autopsies of Patient 3 and an infant 8 months of age who died with the sudden infant death syndrome. Control values for liver copper were measured in specimens obtained at autopsy from 3 infants aged 2 to 7 weeks; 2 had died of congenital heart disease without congestive heart failure and the third of sudden infant death syndrome. Autopsy samples of brain and liver conAccepted for publication May 30, 1978. Address reprint requests Dr Grover, 2600 Philadelphia, PA 19133,
Lawrence St,
0364-5134/79/010065-07$01.25 @ 1978 by Warren D . Grover 65
Table 1 . Clinical Status of Patients with Trirhopoliodystrophy ~~
~~
Diagnostic Features Serum Values Patient No.
Age (mo)
Neurologcal Signs
1
Binh
2
Blnh
3
2
4
8
5
2
Hypotoma. hypothermia, absent head control Hypotonla. increased DTR, absent head control Seizures. hypotonia with increased M R , absent head control Seizures. spasticity. retardation, strabismus Seizures. spastictry. retardation
6
11
n = normd (mean
Seizures, retardation k
Hepatic Cu (gg/gm dry art)
Roent&nographic Signs
Stigmata
Cu (ctddb
Ceruloplumin (mpjdl)
Skull
Long Bones
Bladder
Kinky hair, taliper equinovuur
84 (n, 29 z 3)
30 (n, 5
?
2)
14
Wormian bones
None
Dweniculi
Kinky hair
60 (n. 29 5 3)
26 (n, 5
?
2)
7
Wormian boner
None
Diveniculi
Kinky hair
12 (n, 65
12 (n, 15-30)
12
Worminn bones
Metaphysed spurring. humerus
N o chaues
Kinky hair
30 (n,90- 1 15)
19(after IV Cu for 12 mo)
Worm~an bones
None
Dlveruculi
Kinky hair
35 (n. 65
13 (n, 15-30)
NE
Wormian bones
None
NE
Kinky hair
56 (n, 130 2 1 0 )
10 (n, 25-60)
NE
Wormian
Metaphysed spurring. femur
NE
SFM or range); DTR
?
?
5)
5)
4 (n. 25-50)
h W S
=
deep tendon reflexes. NE = not examined.
sisted of approximately 100 mg of tissue obtained by excision with a dry stainless steel knife previously washed with EDTA and distilled water. Serum, urine, and cerebrospinal fluid copper and zinc levels were measured by flame-aspiration atomic absorption spectrophotometry by a method previously described 1201. Liver copper concentrations were measured by
flame-aspiration atomic absorption spectrophotometry after the tissue had been dried in an oven at 75°C for 18 hours and digested with nitric acid [ 111. Ceruloplasmin was measured by the copper oxidase method as modified for automatic analysis [ 2 3 ] . Skin and hair were obtained from 3 patients with TPD and from control subjects of similar age. The methods used included examination of histological sections of skin and hair stained with hematoxylin-eosin and examination of hairs using polarized light, dark-field illumination. and dark-field autofluorescence.
Results Clinical Findings At the time of diagnosis, each patient had physical findings and laboratory data indicating TPD to the alert physician. Each infant had a similar appearance, characterized by cherubism, a depressed nasal bridge, and decreased facial movement. All patients, including the 2 newborns, had steely, depigmented hair characteristic of the disease. All had definite neurological signs, including hypotonia, poor head con66 Annals of Neurology Vol 5 No 1 January 1979
trol, and abnormalities of deep tendon reflexes. Roentgenographic changes included wormian bones in the skull in all 6 patients, metaphyseal spurring of the long bones in 2, and large, lobular diverticula in the bladder in 3 [ 121. A cerebral angiogram and computerized axial tomogram performed on Patient 1 at 6 weeks and 18 months, respectively, were normal. A computerized axial tomogram obtained on Patient 4 at 2 years of age revealed decreased cortical mass and generalized ventricular dilatation (Fig 1). Electroencephalograms recorded from Patient 1 from 2 weeks to 5 years of age showed no abnormality. Electroencephalograms obtained on Patient 4 at 18 months and 2 years of age demonstrated only mild slow activity. Recordings from scalp electrodes on Patients 2 and 3 revealed diffuse slow-wave activity with admixed paroxysmal features. Normal computerized averaged responses were recorded from electroretinograms of Patient 3 at 10 months of age. Study of specimens of hair and scalp from 3 patients (Nos. 2, 3, and 4) revealed similar abnormalities consistent with a defect in the hair protein matrix. Sections of the scalp stained with hematoxylin-eosin demonstrated anagen (growing)-phase follicles with a slight to moderate hair curvature and a number of follicles that did not contain a well-formed hair. Examination of scalp hair mounted
Race
Family History
Birth Weight (kg)-
White
Sib died suddenly at 8 mo
2.9
None
Icterus (exchange transfusion)
Living, 48 m o
White
Sib with T P D (Patient 3)
2.6
None
Icterus, hypotonia, hypothermia
Died, 2% m o
White
Sib with T P D (Patient 2)
2.7
None
Icterus
Died, 14 mo
White
None
2.9
None
Hypotonia
Living, 25 mo
White
Sib retarded, sib stillborn, and 6 sibs living and well
3.5
Premature rupture of membranes
Icterus (phototherapy)
Died, 5% yi
Black
2 paternal nephews with friable, kinky hair
2.2
Hypertension; bleeding, last trimester
Muconium staining; hypothermia
Died, 3% yr
on slides revealed intertwined, tortuous, thin, fragile-appearing structures with occasional fracturing and twisting that produced a bead-like effect. Inspection of hair with fluorescent and dark-field microscopy demonstrated replacement of the normal pink to red fluorescence with a whitish color. With polarized light, hair from the patients also appeared white instead of the light brown observed in controls. After administration of copper salts for a nine-month period in 1 patient (No. l), partial return to a normal hair color was observed. Four patients died at 295 months to 5% years of age. At postmortem examination, bilateral bronchopneumonia was observed in each child. Neuropathological abnormalities were found in 2 of them. These included small brain size and weight with cerebral and cerebellar atrophy. Histological findings were similar to abnormalities reported by other investigators [71 and included loss of subcortical myelin, reactive gliosis, and neuron loss in the cerebral hemispheres and in the molecular, Purkinje, and granular cell layers of the cerebellum. The spinal cord revealed bilateral spinocerebellar tract degeneration with varying degrees of gliosis. The cerebral and cerebellar vessels showed areas of splitting and beading of elastic tissue of the intima and of the internal elastic membrane.
Complications Gestational
Perinatal
Course
Both patients surviving at this time demonstrate neurological dysfunction. Patient 1, at 4 years of age, has marked hypotonia, no language, and adaptive abilities at a 1-year level. Patient 4 , at age 3 years, has spastic quadriparesis, bilateral esotroyia, and a social smile. The steely hair persists in each patient.
Biochemical Findings Measurements of trace metals in each patient with T P D studied beyond the first week of life revealed serum copper and ceruloplasmin concentrations below normal levels (Tables 1, 2). Mean serum copper and ceruloplasmin concentrations in 10 patients with TPD ranging in age from 1 week to 5 years were significantly lower (p < 0.001) than values in control specimens (Table 2). No difference in serum zinc concentration was observed between patients and unaffected infants and children. In each untreated patient with TPD, urinary copper excretion was significantly greater than control levels (p < 0.001); this abnormality was found in the first specimen from each patient. No differences were observed in urinary zinc excretion measured simultaneously (Table 2). Cerebrospinal fluid copper and zinc concentrations were not significantly different in the two groups of children. Hepatic copper concentrations were measured in 3
Grover, Johnson, and Henkin: Trichopoliodystrophy
67
patients before treatment (Table 3). The liver copper values in the patients were significantly lower than in age-matched controls (p < 0.001). Copper metabolism was also studied in 2 patients (Nos. 1 and 2) before and immediately after delivery (Table 4). At term, blood and amniotic fluid from the mother of Patient 2 were analyzed for copper, ceruloplasmin, and zinc concentrations. The amniotic fluid copper level was over four times higher than the mean level measured in mothers of normal infants at term, whereas the zinc level in the amniotic fluid was within the wide range of normal values. There were no differences in serum copper, ceruloplasmin, or zinc concentration between the mother of Patient 2 and the mothers of normal children. Initial copper concentrations obtained in cord blood in both patients were greater than twice the upper limit measured in normal term infants: 60 pg/dl in Patient l and 8 4 p d d l in Patient 2 (normal, 29 ? 3 pg/dl) [ 151. In Patient 2 the serum copper concentration fell rapidly over the first week of life, in contrast to the increase in serum copper concentration that occurs in unaffected newborns [ 161 (Fig 2). The serum copper concentration in Patient 1 was 19 pg/dl at 21 days of age, a time when the mean copper concentration in normal infants is approximately 80 pg/dl. Serum ceruloplasmin values paralleled the copper levels. Repeated measurements of urinary copper excretion during the first two weeks of life averaged 7.0 pgl24 hrlkg body weight (normal, < 1.0 pg/kg124 hr) with zinc excretion at 4.0 pg/kg/24 hour (normal, 3.8 pgj kgJ24 hr).
F i g 1 . Compziterized axial tomograni in Patient 4 at 2 years of age detnomtrutes mild i*entrirzilardilatation and an increase in .culci ronsiJtent wirh atrophy.
Tdble 2. Capper atid Zinc Leieh in Serum, Urine, and Cerebrospinal Fluid in 10 Patient.r with Untreated TPD" ~~~~~~
~
~~
Serum Values
Patient No.
cu (ddl)
Zn (pg/dl)
Ceruloplasrnin (mddl)
1
16
80
4
2
43
97
12
3
25 32 30 20 13 27 20
... ...
4 14 8 4 3 5
4 5 6 7
8 9 10 Mean
134 102 73 92 68 106
39 &
SEM
Controls (N=12), age matched
... ...
cu
Zn
cu
Zn
3.8 7.4 4.7
13.7
4.8 ... 4.1
4.5 ...
...
4.0 ... ...
4.6 4.8
7.3 15.0
7
7& 1
5.0
100 t 5
80
?
7
32 & 7
1.1
"All values after first week of life.
68 Annals of Neurology Vol 5 N o 1 January 1979
0.4
8.0 2 2.1
3.4
2
1
0.3
9.8
4.0
&
1
4.0 ...
&
&
.o
...
4.0
5.4 ... ...
94
2
... ... 1
2.0 2.5
... ... ... ... ...
4.1
3
26
CSF Values (pddl)
Urine Values (pg/24 hdkg body wt)
&
3.2
... ... ... ... ...
4.4 2 1
Tdble in
.j
.$. Lher Capper Conrentration Plrtients u i t h IJntredted T P D
Hepatic Cu Patient No.
(&gm dry wt)
1
14
7
7 12
.3
Mcan
?
1 SEM
Controls tN =3), age matched
I
11 2 2
226
&
57
45A
3s
Discussion Synthesis of the biochemical findings and clinical data obtained from the 6 patients with TPD indicates a possible mechanism for thc &case process that has not been previously emphasized (Fig 3 ) . Important facts include the documentation of elevated levels of copper in amniotic tluid, cord blood, and urinc of
I
j
a
25
FiK 2. Serial blood copper detrrininutio)is in Patient 2. Poimti repre.ceni ropper lei,elc I p,gidt).Dotted ciiri'e oiitlines nort t d ICI e l l .
newborn infants but hepatic copper values only 1057
of control levels. Abnormalities in neurological function, hair structure, bone formation, and hepatic function were present in the imme'iiate perinatal period and indicated organ dysfunction in utero. Untreated patients beyonci the newborn period also exhibited low concentrations of serum copper, ccruloplasmin, and hepatic copper but increased urinary copper excretion. T h e zinc lcvcls in all fluids and tissucs studied were within the average range, in&cating normal zinc metabolism. These data suggest the presence of a selective defect in tissue uptake o r rctcntion of copper or both. Correlative evidence has been obtained by H o r n et al, w h o measured tissue copper concentrations in a fetus with TPD and compared the results with those from control patients of a similar age [ 1'1. They
found that tissue levcls of copper in kidney, spleen. pancreas, placenta, brain, lung, muscle, and skin were at least twice control values. They also noted that the liver copper value in the patient was one-thin1 of control level. In the normal newborn, copper levels in liver are several times higher than values from normal adults. H e p i t i c copper in the newborn is rcportedly sequestered in lysosomes [9] and in mitochondria in the form of a protein, hcpatomitochon~lriocuprcin[ 2 11. We Actermined that blood levcls of coppcr and ceruloplasmin in newborn patients with TPD were at least twice normal values and that the copper level in amniotic fluid in thc mother of 1 of thcse patients was four times the normal mean value at tcrm. O u r studies indicate that liver copper in the
?'able 4. Comparison of Srrilni Copper, Cerzilaplasmin. and Zinc Conmitrations and Aniniotir Fluid Copper and Zinr Vdlues in Meii,horm with TPD. the Mother nf'riti Aflicted 1nJ;itit. cind Control Siikjtrt,
Ncwborns
Morhcrs 'L'PD Determination
Normal" ( N = 15)
TPD"
Normal" ( N = 15)
221 2 14 91 2 4 48 2 3
245
29
5
90
10
5
55
83
5
3
0 2 3 2 12
20 12
Patient 1
Patient 2
3
00
2
23 05
84 30
Blood
cu ( p d d l ) Ceruloplasmin (rnddl) Zn (pgldl) Amniotic fluid CU
(pgldl)
Zn (pd.11)
32
"Mean 2 1 SEM. 'The mother of Patient 2.
Grover, Johnson, and Hcnkin: lrichopoliodysrrophy 69
ANTENATAL
[Kidney
I
TISSUES MA T E R N A L ~ F E T A L
Brain AMNIOTIC FLUID
NEONATAL TISSUES -*URINARY .)6INDING & STORAGE .)GI ABSORPTION
INFANCY
Cu
TISSUE
‘TOTAL
BODY
Fig 3. Postulatedschema of copper metabolism i n T P D . The transplacental kinetics of copper appears intact, but abnormal copper transport or binding (or both) a t the cellular level results in a n increase in copper concentrations in alltissues except the liver. The excess copper is eventually excreted by the fetal kidney and causes an increase i n amniotic copper levels. A t birth, the defect in copper binding or transport causes a persistent loss of copper from cells anddecreased absorption of copperfrom the gastrointestinal tract, resulting in low blood and tissue copper levels w i t h increased urinary excretion.
patients is one-tenth of normal levels of age-matched controls at a time when serum copper concentration is more than twice comparable levels in normal newborns. These data suggest that copper is available to all tissues of the fetus but that transport and retention vary with individual cell systems. Other data suggesting a defect in copper transport or binding (or both) was obtained by Goka et al [8], who demonstrated that cultured fibroblasts from patients with TPD accumulated more copper than cells from normal subjects. The copper values in cultured fibroblasts from 3 patients in this study (Nos. 1, 2, and 3) were approximately four times the copper content of fibroblasts from normal subjects. These data are consistent with the hypothesis of a selective tissue defect in the uptake or retention (or both) of copper in tissues of patients with TPD. The concept of a selective tissue defect in the uptake or retention of copper in this disease has a correlate in some forms of copper deficiency previously observed in animals. In enzootic ataxia or swayback, ewes are anemic but have normal wool; their lambs exhibit ataxia, blindness, depigmentation, and loss of crimp of their wool [b]. Cattle suffering from copper deficiency commonly die suddenly from “falling disease,” attributed to sudden heart failure after exercise or excitement [l]. In our patients and in those reported by others, there is no anemia in spite of an apparent severe deficiency of copper. We have identified no patient who died of cardiovascular causes. Our data and information provided by other in-
70 Annals of Neurology Vol 5
No 1 January 1979
vestigators indicate a spectrum of activity of copperdependent enzyme systems in patients with TPD. We have documented renal tubular acidosis with normal tubular clearance of phosphorus and normal amino acid excretion in 2 of our patients (Nos. 1 and 3). Renal dysfunction was not reversed following therapy with copper that increased serum, hepatic, and renal levels of copper. We have also documented defective oxidative metabolism of mitochondria [ 113 obtained from muscle and low values of serum dopamine /3-hydroxylase [ 131. However, Zelkowitz et al 1271 have noted normal utilization of oxygen by muscle mitochondria and normal values for superoxide dismutase, which suggest genetic heterogeneity resulting in variable biochemical abnormali ties. Selective changes in tissue copper metabolism occurred after treatment of these patients. Parenteral therapy with copper salts or oral therapy with copper-nitriloacetic acid produced increases in serum copper and ceruloplasmin [4, 5, 10, 11, 14,261, urine copper [ l o , 141, and hepatic and renal copper, improved function of some copper-dependent oxidative enzymes, and increased cerebrospinal fluid copper levels [ 113 but did not raise copper values in brain [lo, 11, 141. In 3 untreated patients with TPD, copper in cerebral white matter and cortex ranged from 4 to 9.5 &gm dry tissue [22, 261. In our Patient 3 [ 101, even following therapy that achieved a return of normal copper levels in liver and elevated copper levels in kidney, the copper concentration in cerebral white matter was less than 1 p g / p dry tissue. A comparable specimen from an age-matched control patient contained 18 &gm dry tissue. However, copper levels in the medulla of both the patient and the control specimen were similar, implying that the defect may be selective even within the brain. Because the brain is the only organ in humans in which increasing concentrations of copper occur with age [24], this organ has an important need for copper, particularly in the first years of life when active myelination occurs.
Supported in part by HEW Training Grant I TO1 NSI0072-OIA2, NSRA, Grant RR-75 from the General Clinical Research Center, and Grant PO1 CA11536. Presented in part at the 1974 Annual Meeting of the Child Neurology Society, Madison, WI, and the 1976 meetings of the American Pediatric Society, the Society for Pediatric Research, and the American Society of Clinical Investigators. The authors wish to acknowledge the following contributions: Cyril D’Cruz, MD, and Guillermo A. deleon, MD, for pathological examinations; Hope Punnett, PhD, and Nancy Brodsky, PhD, for fibroblast cultures; and the Pennridge Pediatric Group for the identification of the 2 afflicted newborns.
References 1. Bennetts HW, Beck AB, Hurley R: Pathogenesis of "falling disease." Aust Vet J 24237-244, 1948 2. Bucknall WE, Haslam RH, Holtzmann NA: Kinky hair syndrome: response to copper therapy. Pediatrics 52:653-657, 1973 3. Danks DM, Campbell PE, Stevens BJ, et al: Menkes kinky hair syndrome: an inherited defect in copper absorption with widespread effects. J Pediatr 50:188-201, 1972 4. Danks DM. Campbell PE, Walker-Smith J, et al: Menkes' kinky hair syndrome. Lancet 1:llOO-1102, 1972 5 . Dekaban AS: Menkes' kinky hair disease treated with subcutaneous copper sulfate. Lancet 2:1523, 1974 6. Dick AT: Preliminary observations on the effect of high in. take of inorganic sulfate on blood copper & fleece character in cross bred sheep. Aust Vet J 30:196-202, 1968 7. French JH, Sherrard ES, Lubell H , et al: Trichopoliodystrophy: report of a case and biochemical studies. Arch Neurol 26:229-244, 1972 8. Goka TJ, Stevenson RE, Herreran PM, et al: Menkes disease: a biochemical abnormality in cultured human fibroblasts. Proc Natl Acad Sci USA 73:604-606, 1976 9. Goldfischer S, Bernstein JJ: Lipofuscin (aging) pigment granules of the newborn human liver. J Cell Biol42:253-261, I969 10. Grover WD, Henkin RI: Trichopoliodystrophy (TPD): a fetal disorder of copper (Cu) metabolism. Pediatr Res 10:448. 1976 11. Grover WD, Scrurton MC: Copper therapy in trichopoliodystrophy. J Pediatr 86:2 16-220, 1975 12. Harcke T H , Capitanio MA, Grover WD, et al: Bladder diverticuli and Menkes' syndrome. Radiology 124:459-461, 1977 13. Henkin RI, Grover WD: Trichopoliodystrophy (T'PD): new aspects of pathology and treatment in trace element merabolism in man and animals 111, in Kirchgessner M (ed): Arbeitskreis fur Tierernahrungsforschung. Weihenstephan. 1978. pp 405-408 14. Henkin RI, Grover WD: Trichopoliodystrophy (TPD): a genetic defect in selective Cu binding and storage as well as transpon. Clin Res 24:435A, 1976
15. Henkin RI, Marshall J, Meret S: Maternal-fetal metabolism of copper and zinc at term. Am J Obstet Gynecol 110: 131- 134, 1971 16. Henkin RI, Schulman JD, Schulman CB, et al: Changes in total nondiffusible and diffusible plasma zinc and copper during infancy. J Pediatr 82:831-837, 1973 17. Horn N , Mikkelsen M, Heydorn K, et al: Copper and steely hair. Lancet 1:1236. 1975 18. Lott IT, DiPaolo R, Schwanz D, et al: Copper metabolism in the steely hair syndrome. N Engl J Med 292:197-199, 1975 19. Menkes JH. Alter M, Steigleder GK, et al: A sex-linked recessive disorder with retardation, peculiar hair and focal cerebral and cerebellar degeneration. Pediatrics 29:764-779, 1962 20. Meret S. Henkin RI: Simultaneous direct estimation by atomic absorption spectrophotometry of copper and zinc in serum, urine and cerebrospinal fluid. Clin Chem 17:369-373, 197 1 2 1. Porter H: Neonatal hepatic mitochondrocuprein. 3: Solubilization of the copper and protein from mitochondria of newborn liver by reduction with mercaproerhanol. Biochim Biophys Acta lS4:236-238, 1968 22. Reske-Nielson E, Loy HOC, Andersen P, et al: Brain-copper concentration in Menkes disease. Lancet 1:613, 1973 23. Scheinberg IH, Morel1 AG: Exchange ceruloplasmin copper with ionic CU'" with reference to Wilson's disease. J Clin Invest 36:1193-1201, 1957 24. Schroeder HA, Nason AP, Tipton IH, et al: Essential trace metals in man: copper. J Chronic Dis 19:1007-1034, 1966 25. Singh S, Bresnan MJ: Menkes kinky hair syndrome. Am J Dis Child 125:572-578. 1973 26. Walker-Smith JA, Turner B, Blomfield J, et al: Therapeutic implications of copper deficiency in Menkes' steely hair syndrome. Arch Dis Child 48958-962, 1973 27. Zelkowitz M, Cote L. Miranda A, et al: Trichopoliodystrophy: ultracrructure and biochemical studies in muscle. Presented at the Annual Meeting of the Child Neurology Society, Monterey, CA 1976
Grover, Johnson, and Henkin: Trichopoliodystrophy
71
