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TRANSMISSION OF CREUTZFELDT-JAKOB DISEASE FROM MAN TO SQUIRREL MONKEY
435
TRANSMISSION OF CREUTZFELDTJAKOB DISEASE FROM MAN TO SQUIRREL MONKEY D. P. GRANT
I. ZLOTNIK
Microbiological Research Establishment, Porton, near Salibury SP4 0JG, Wiltshire A. D. DAYAN * C. J. EARL National Hospitals for Nervous Diseases, London WC1N 3BG
Brain material from a patient with Creutzfeldt-Jakob disease was inoculated intracerebrally into four squirrel monkeys. Subsequently three out of the four inoculated monkeys developed clinical signs and a syndrome similar to the human disease. The incubation period from inoculation until the appearance of first signs varied from 20 to 23 months. Histological examination of the brains of the above monkeys revealed severe spongy degeneration and astrogliosis, lesions that resembled those seen in the brain of the patient. The findings not only confirm the fact that Creutzfeldt-Jakob disease is transmissible, but also draw attention to the usefulness of the squirrel monkey, a small and easily handled New World monkey, as a laboratory animal for the transmission of human subacute spongiform encephalopathies. Summary
Fig. 2-T.S.H.
response to T.R.H. in
depressive and normal
female patients.
Discussion
.
,
We have demonstrated that T.R.H. does not have any therapeutic effect in depressive patients either in a single dose or in multiple doses given with a tricyclic antidepressant over a 3-week period. Our patients all had acute depression which subsequently responded well to conventional treatment. The T.S.H. response was impaired in a proportion of our patients. These patients had no specific clinical features. Patients retested after 3 weeks’ therapy with amitriptyline, who showed.some clinical improvement, demonstrated no significant change in T.s.H. response. Takahashi et a1.7 also reported a significantly impaired response to T.R.H. in depressive patients. This relative lack of response to T.R.H. is difficult to explain since our patients were euthyroid and a previous careful examination of the thyroid state of depressive patients showed no evidence of thyroid abnormality 11 This abnormality may be analogous to the reduced growthhormone response to hypoglycsemia reported by Sachar et a112 in depressed patients. We thank Miss Janet Smith, St. Luke’s Hospital, Guildford, for her help in estimating thyroid-function tests. We are grateful to Hoechst Pharmaceuticals for the supply of T.R.H. REFERENCES 1. 2.
Prange, A. J., Jr., Wilson, I. C. Psychopharmacologia, 1972, 26, 82. Prange, A. J., Jr., Wilson, I. C., Lara, P. P., Alltop, L. B., Breese,
G. R. Lancet, 1972, ii, 999. Kastin, A. J., Ehrensing, R. H., Schalch, D. S., Anderson, M. S. ibid. p. 740. 4. Van der Vis-Melsen, M. J. E., Wiener, J. D. ibid. p. 1415. 5. Dimitrikoudi, M., Hanson-Norty, E., Jenner, F. A. ibid., 1974, i, 3.
456. 6.
7. 8. 9.
10. 11. 12
Mountjoy, C. Q., Price, J. S., Weller, M., Hunter, P., Hall, R., Dewar, J. H. ibid. p. 958. Takahashi, S., Kondo, H., Yoshimura, M., Ochi, Y. Folia psychiat. neurol. Jap. 1973, 27, 305. Hamilton, M. J. Neurol. Neurosurg. Psychiat. 1960, 23, 56. Zealley, A. K., Aitken, R. C. B. Proc. R. Soc. Med. 1969, 62, 993. Hall, R., Amos, J., Ormston, B. J. Br. med. J. 1971, i, 582. Whybrow, P. C., Coppen, A., Prange, A. J., Jr., Noguera, R., Bailey, J. E. Archs gen. Psychiat. 1972, 26, 242. Sachar, E. J., Finkelstein, J., Hellman, L. ibid. 1971, 25, 263.
Introduction
Creutzfeldt-Jakob disease is a subacute spongiform encephalopathy, characterised by progressive dementia with pyramidal and extrapyramidal signs. The aetiology of the disease was unknown until 1968, when it was shown that it could be transmitted from man to chimpanzees by the inoculation of brain homogenates.1 Since then further transmissions have been carried out from fifteen more cases of the disease 2 However, chimpanzees are by no means the only susceptible animals, since the disease has now been transmitted to a number of New World monkeys such as the spider, squirrel, capucin, and woolly monkeys, and to two Old World monkeys, the bushbabies and the mangabeys. Of the non-primates CreutzfeldtJakob disease was transmitted only to the domestic cat.3 However, despite the fact that Creutzfeldt-Jakob disease was shown to be transmissible4 and that it could be transmitted to New World monkeys,5 further evidence from a different team of workers would be of immense value to confirm the aetiology of this disease. Hitherto all transmissions from patients to animals have been carried out in the same laboratory, where the original infection of chimpanzees with material from kuru patients was achieved.6,7 Although the evidence presented by Gajdusek, Gibbs, and their colleagues is very convincing, the remote possibility of accidental contamination with kuru material or contact infection from experimental kuru in animals could not be completely exciu.ed.8 We felt, therefore, that a transmission of Creuizfeldi-Jakob disease from a patient to primates in a laboratory that had never * Present address: Wellcome Research Laboratories, Beckenham, Kent BR3 3BS.
436 worked with slow-virus infections would add great support to the theory of infectious or viral aetiology of Creutzfeldt-Jakob disease. Materials and Methods Pieces of cerebral cortex were obtained at necropsy of a patient with Creutzfeldt-Jakob disease, whose casehistory is given below. Unfixed pieces ofthe brain were stored at -20°C but were later transferred to -70°C, awaiting the availability of monkeys. The inoculum consisted of a 10% homogenate of the patient’s brain prepared in borate buffer pH 9. After centrifugation for 5 minutes at 800 g, a sufficient quantity of a solution of ’Crystamycin’ (Glaxo) was added so that each ml. of supernate contained 200 units of benzylpenicillin sodium and 0-2 mg. of streptomycin. Four squirrel monkeys (Saimiri sciureus) were inoculated intracerebrally with 0’5 ml. of the brain supernate under general ansesA similar homogenate was prepared from the thesia. same brain 10 months later, when spider monkeys also became available, and two such monkeys received intracerebral inoculations of 1-0 ml. each. The inoculated monkeys remained under observation, and those developing clinical signs received extra care. Moribund monkeys were killed by’Nembutal’ general anaesthesia and brains and spinal cords were removed immediately after death. After selecting a few pieces of brain for subinoculations the remainder of the brain Paraffin sections 5 im. was fixed in 10% formol saline. in thickness were stained with hxmatoxylin and eosin or luxol fast-blue and frozen sections were stained with Cajal’s gold chloride for the demonstration of astrocytes.’
Fig. I-Cerebral cortex of the patient with Creutzfeldt-}akob disease, whose brain was used for transmission experiments. (H. & E. ; ;" 225.)
Case-history patient whose brain was used for transmission experiments was a 50-year-old British woman who had no family history of neurological disease. 6 years before her death she had had a clinically typical attack of poliomyelitis which left her with weakness of the trunk and such wasting of the right leg that she was able to walk only with callipers and crutches. Her illness began with the onset over many days of progressive confusion, loss of memory, and depression. The
On examination, after about 8 weeks from the onset of clinical signs, she was disorientated and behaved ab-
normally ; conjugate gaze was defective, her eye movements were slow and fragmented, and there was horizontal nystagmus
to
the left.
Both
arms
and the left
leg
Fig. 2-Astrocytic reaction in the cerebral patient. (Cajal; Y 225.)
fluid: no cells were seen, but protein 45 mg. per 100 ml. and sugar 62 mg. per 100 ml. were noted. Repeated showed electroencephalograms repetitive complexes strongly suggestive of subacute spongiform encephalopathy. Her condition deteriorated further; myoclonic jerking was detected in the arms and she became totally unresponsive before dying after an illness that had lasted for about 4 months. The necropsy, the first of a case of Creutzfeldt-Jakob disease to be carried out in the National Hospitals for a few decades, was started 30 minutes after death, and the only macroscopic abnormality was fibrosis of muscles of the trunk and right leg. The brain weighed 1230 g. Histologically there was extensive loss of neurons, proliferation of astrocytes and gliosis in the cerebral cortex, accompanied by typical spongy vacuolation both of cells and neuropil (figs. 1 and 2). Similar lesions were present in some thalamic and brainstem nuclei and to a lesser extent in the cerebellum. Argyrophilic plaques and neurofibrillary tangles were not seen. There were
of the above
active lesions in the spinal cord. Electron microscopy of the brain confirmed the presence of empty vacuoles in cell processes, apparent redundant profiles of trilaminar membranes, and other less specific degenerative features. The pathological diagnosis was subacute spongiform encephalopathy. Tissue-cultures of the patient’s brain were established and the extensive virological investigations will be reported elsewhere.1O
no
Results
were
ataxic, the right leg and trunk being wasted. The right plantar and left flexor responses were absent. Laboratory investigations included examination of the cerebrospinal
cortex
No signs of disease were noticed in any of the inoculated squirrel monkeys during the first 19 months. However, between the 20th and 21st month a general slowing down of movements and activity became obvious in three monkeys; at the same time the animals began losing weight. The loss of weight was slow and gradual in two monkeys but very rapid in one, leading to emaciation and wasting. To begin with, appetite was maintained in all animals. Two of the monkeys, the wasted one and a second animal, also developed a severe itch with constant scratching and rubbing of the body against the bars of the cage. Loss of hair became striking along the tail, especially the base, where the sacral region was covered with excoriations and scabs. Very soon the highly wasted monkey developed neurological signs in the form of tremors along the body and extremities and occasional twitching of the facial muscles. The back of the animal was continuously arched and the hind legs were lifted very high during progression. The animal at
437
rigid, lost appetite, and his vision became affected, and he finally collapsed and was killed in extremis 2 years and 22 days after the intracerebral
became
inoculation. The second
monkey
to
develop neurological signs
that never showed any evidence of itch The first signs began with visual disor rubbing. turbances, ataxia, rigidity of the body, tremors, and partial paralysis of the right upper limb. He kept his back arched and walked with an obvious limp (fig. 3). He frequently fell and had great difficulty in getting up. Soon, however, he was unable completely to lift himself up and his extremities became flaccid. He became moribund and was killed 2 years and 2 months after the experimental inoculation. Although the neurological signs were late to appear in this monkey, once they became obvious they. progressed very quickly and the animal had to be killed within 4 weeks of the appearance of definite neurological signs. was
the
one
Fig. 3-Squirrel monkey in Note arched back and rigid
an
Fig. 4-Spongy degeneration in the cerebral cortex of an affected monkey. (F3. & E. ; x225.)
however, revealed
severe lesions throughout the of the less same pattern in the three brain, squirrel monkeys. The most striking change consisted of widespread spongy degeneration and astrocytosis throughout the grey-matter of the central nervous system. The most severe spongy degeneration was seen in the corpus striatum, while in the cerebral cortex lesions were of a slightly lesser intensity but uniformly spread throughout (fig. 4). Similar lesions but of a patchy character were present in the thalamus, while somewhat less striking changes were seen in the midbrain and the brainstem. In the cerebellum, the molecular was affected, while in the only layer cord mild affected the whole greyspinal changes In addition to the above changes neuronal matter. degeneration was very obvious, especially in the cerebral cortex, while vacuolation of neurons was very pronounced in the corpus striatum. Neuronal
very
more or
advanced stage of the disease. (Approximately ’/. life size.)
posture.
The third monkey had a clinical history somewhat similar to the first one with an itch, rubbing, and emaciation, but the clinical signs were spread over a much longer period (4 months) and he lived for 2 This years and 2 months after the inoculation. animal, apart from rigidity, tremors, and ataxia, also showed a variety of peculiar signs; he either pressed his forehead against the bars of the cage and held his head with both hands, or assumed an unnatural posture where he would be standing for long periods in an arched position with the head down on the floor of the cage and holding the head tightly with his hands. The fourth squirrel monkey has not yet shown any definite signs of disease, 2 years and 3 months after inoculation, neither has any abnormality been noticed in the two spider monkeys (1 year and 6 months after
inoculation). Post-mortem
examination
did
not
reveal
any
definite changes in the internal organs, but there was a slight decrease in the size of the brains and an increase in the amount of the cerebrospinal fluid.
Histological examination of brains
and
spinal cords,
Fig. 5-Neuronal vacuolation in the globus paHidus of an affected monkey. (H. & E. ; x 480.)
438 vacuolation affected the larger neurons of the globus pallidus and the medium-size neurons of the putamen and caudate nucleus. The vacuoles were confined, as
Preliminary Communication
rule, to the cytoplasm (fig. 5). They were small, usually more than one vacuole was found in the cell; however, in many neurons large numbers of a
REYE’S SYNDROME DUE TO A NOVEL PROTEIN-TOLERANT VARIANT OF ORNITHINE-TRANSCARBAMYLASE DEFICIENCY
and
vacuoles often became confluent and sometimes displaced the nucleus. Astrocytic proliferation and hypertrophy affected the whole grey-matter, but in the thalamus, corpus striatum, medial geniculate body, and the olives, apart from severe hypertrophy, there was also a considerable increase in the number of astrocytic processes which often formed a dense network (fig. 6).
M. MICHAEL THALER
Department of Pediatrics, University of California, San Francisco, California 94143 MARILYN BOSWELL J. HOOGENRAAD* Division of Developmental Medicine, Department of Pediatrics, Stanford University School of Medicine, Palo Alto, California 94305
NICHOLAS
A novel variant of ornithine-transcarbamylase (O.T.C.) deficiency has been discovered in a patient with encephalopathy and fatty visceral degeneration (Reye’s syndrome). Hepatic ornithine transcarbamylase activity was 20·7% of normal, with extremely low affinity of the enzyme for ornithine, and substrate inhibition by carbamyl phosphate and ornithine. The findings reveal a link between Reye’s syndrome and heritable O.T.C. deficiency, and suggest that administration of ornithine or arginine may be specific therapy for this condition.
Summary
Fig. 6-Astrocytosis in the cerebral cortex of an affected monkey. (Cajal; x 225.)
INTRODUCTION
Discussion
.
THE
clinicopathological entity reported in 1963 by Reye et al 1 as encephalopathy and fatty degeneration of the viscera" has since been recognised as a major cause of mortality and morbidity among infants and children. Reye’s syndrome is endemic to most parts of the world, and frequently occurs in association "
findings provide unequivocal evidence of the transmissibility of Creutzfeldt-Jakob disease- and confirm the work of Gibbs et al.and Gajdusek and Gibbs." Of special significance is the comparative ease with which the disease was transmitted, since These
this has been our first attempt to transmit CreutzfeldtJakob disease. At the same time it draws attention to the great value of the squirrel monkey as a laboratory animal for work on the transmissibility of subacute spongiform encephalopathies of man. The squirrel monkey is a small friendly animal which is easily handled and can therefore be kept without difficulty in a laboratory. It is much more readily obtainable and considerably cheaper than the large apes such as
chimpanzees.
It is remarkable that both the clinical picture and the histological changes in the central nervous system of the squirrel monkeys resembled those of the patient from whom the disease was transmitted. However, the intensity of lesions was much greater in the animals than in the patient from whom the disease was transmitted, a fact common with other transmissible spongiform encephalopathies of animals, such as scrapie in mice, mouse scrapie in the goat, or transmissible mink encephalopathy in goats,u-15 We thank Miss M. I. Stokes and Mr J. A. Mills from the National Hospitals for their help with necropsy specimens. Requests for reprints should be addressed to 1. Z. REFERENCES 1.
Gibbs, C. J., Jr., Gajdusek, D. C., Asher, D. M., Alpers, M. P., Beck, E., Daniel, P. M., Matthews, W. B. Science, 1968, 161, 388.
viral infections of childhood such as and influenza &bgr;.2 The disorder progresses chickenpox mild from a prodromal viral illness to vomiting and abdominal pain which signal the sequential appearance of cerebral dysfunction and coma. Mortality is at least 40 %; survivors recover rapidly. The biochemical abnormalities most frequently observed in Reye’s syndrome include raised serum-transaminase levels, prolonged prothrombin-time, and hypoglycxmia. Blood-
with
*
common
Present address: Department of Biochemistry, LaTrobe University, Bandoora, 3083 Victoria, Australia. DR ZLOTNIK AND OTHERS:
REFERENCES—continued
D. C., Gibbs, C. J., Jr. Brain, 1973, 96, 1. J., Jr., Gajdusek, D. C. Science, 1973, 182, 67. J., Jr., Gajdusek, D. C. ibid. 1969, 165, 1023. D. C., Gibbs, C. J., Jr. Nature, 1971, 230, 588. Gajdusek, D. C., Gibbs, C. J., Jr., Alpers, M. ibid. 1966, 209, 794 Gajdusek, D. C., Gibbs, C. J., Jr., Alpers, M. Science, 1967, 155, 212. Gajdusek, D. C., Gibbs, C. J., Jr. in Biohazards in Biological Research (edited by A. Helman, M. N. Oxman, and R. Pollock); p. 288. New York, 1973. 9. Zlotnik, I. Br. J. exp. Path. 1968, 49, 555. 10. Slack, P. M., Dayan, A. D., Slavin, G., Tyrell, D. A J
2. 3. 4. 5. 6. 7. 8.
11. 12. 13. 14.
15.
Roos, R., Gibbs, C. Gibbs, C. Gajdusek,
Gajdusek,
Unpublished. Zlotnik, I., Rennie, J. C. J. comp. Path. 1962, 72, 360. Zlotnik, I., Rennie, J. C. ibid. 1963, 73, 150. Zlotnik, I., Rennie, J. C. ibid. 1965, 75, 147. Zlotnik, I. National Institute for Nervous Diseases and Blindness monograph no. 2. U.S. Public Health Service publication 1378, p. 237. U.S. Government Printing Office, 1965. Zlotnik, I., Barlow, R. M. Vet. Rec. 1967, 81, 55.
TRANSMISSION OF CREUTZFELDTJAKOB DISEASE FROM MAN TO SQUIRREL MONKEY D. P. GRANT
I. ZLOTNIK
Microbiological Research Establishment, Porton, near Salibury SP4 0JG, Wiltshire A. D. DAYAN * C. J. EARL National Hospitals for Nervous Diseases, London WC1N 3BG
Brain material from a patient with Creutzfeldt-Jakob disease was inoculated intracerebrally into four squirrel monkeys. Subsequently three out of the four inoculated monkeys developed clinical signs and a syndrome similar to the human disease. The incubation period from inoculation until the appearance of first signs varied from 20 to 23 months. Histological examination of the brains of the above monkeys revealed severe spongy degeneration and astrogliosis, lesions that resembled those seen in the brain of the patient. The findings not only confirm the fact that Creutzfeldt-Jakob disease is transmissible, but also draw attention to the usefulness of the squirrel monkey, a small and easily handled New World monkey, as a laboratory animal for the transmission of human subacute spongiform encephalopathies. Summary
Fig. 2-T.S.H.
response to T.R.H. in
depressive and normal
female patients.
Discussion
.
,
We have demonstrated that T.R.H. does not have any therapeutic effect in depressive patients either in a single dose or in multiple doses given with a tricyclic antidepressant over a 3-week period. Our patients all had acute depression which subsequently responded well to conventional treatment. The T.S.H. response was impaired in a proportion of our patients. These patients had no specific clinical features. Patients retested after 3 weeks’ therapy with amitriptyline, who showed.some clinical improvement, demonstrated no significant change in T.s.H. response. Takahashi et a1.7 also reported a significantly impaired response to T.R.H. in depressive patients. This relative lack of response to T.R.H. is difficult to explain since our patients were euthyroid and a previous careful examination of the thyroid state of depressive patients showed no evidence of thyroid abnormality 11 This abnormality may be analogous to the reduced growthhormone response to hypoglycsemia reported by Sachar et a112 in depressed patients. We thank Miss Janet Smith, St. Luke’s Hospital, Guildford, for her help in estimating thyroid-function tests. We are grateful to Hoechst Pharmaceuticals for the supply of T.R.H. REFERENCES 1. 2.
Prange, A. J., Jr., Wilson, I. C. Psychopharmacologia, 1972, 26, 82. Prange, A. J., Jr., Wilson, I. C., Lara, P. P., Alltop, L. B., Breese,
G. R. Lancet, 1972, ii, 999. Kastin, A. J., Ehrensing, R. H., Schalch, D. S., Anderson, M. S. ibid. p. 740. 4. Van der Vis-Melsen, M. J. E., Wiener, J. D. ibid. p. 1415. 5. Dimitrikoudi, M., Hanson-Norty, E., Jenner, F. A. ibid., 1974, i, 3.
456. 6.
7. 8. 9.
10. 11. 12
Mountjoy, C. Q., Price, J. S., Weller, M., Hunter, P., Hall, R., Dewar, J. H. ibid. p. 958. Takahashi, S., Kondo, H., Yoshimura, M., Ochi, Y. Folia psychiat. neurol. Jap. 1973, 27, 305. Hamilton, M. J. Neurol. Neurosurg. Psychiat. 1960, 23, 56. Zealley, A. K., Aitken, R. C. B. Proc. R. Soc. Med. 1969, 62, 993. Hall, R., Amos, J., Ormston, B. J. Br. med. J. 1971, i, 582. Whybrow, P. C., Coppen, A., Prange, A. J., Jr., Noguera, R., Bailey, J. E. Archs gen. Psychiat. 1972, 26, 242. Sachar, E. J., Finkelstein, J., Hellman, L. ibid. 1971, 25, 263.
Introduction
Creutzfeldt-Jakob disease is a subacute spongiform encephalopathy, characterised by progressive dementia with pyramidal and extrapyramidal signs. The aetiology of the disease was unknown until 1968, when it was shown that it could be transmitted from man to chimpanzees by the inoculation of brain homogenates.1 Since then further transmissions have been carried out from fifteen more cases of the disease 2 However, chimpanzees are by no means the only susceptible animals, since the disease has now been transmitted to a number of New World monkeys such as the spider, squirrel, capucin, and woolly monkeys, and to two Old World monkeys, the bushbabies and the mangabeys. Of the non-primates CreutzfeldtJakob disease was transmitted only to the domestic cat.3 However, despite the fact that Creutzfeldt-Jakob disease was shown to be transmissible4 and that it could be transmitted to New World monkeys,5 further evidence from a different team of workers would be of immense value to confirm the aetiology of this disease. Hitherto all transmissions from patients to animals have been carried out in the same laboratory, where the original infection of chimpanzees with material from kuru patients was achieved.6,7 Although the evidence presented by Gajdusek, Gibbs, and their colleagues is very convincing, the remote possibility of accidental contamination with kuru material or contact infection from experimental kuru in animals could not be completely exciu.ed.8 We felt, therefore, that a transmission of Creuizfeldi-Jakob disease from a patient to primates in a laboratory that had never * Present address: Wellcome Research Laboratories, Beckenham, Kent BR3 3BS.
436 worked with slow-virus infections would add great support to the theory of infectious or viral aetiology of Creutzfeldt-Jakob disease. Materials and Methods Pieces of cerebral cortex were obtained at necropsy of a patient with Creutzfeldt-Jakob disease, whose casehistory is given below. Unfixed pieces ofthe brain were stored at -20°C but were later transferred to -70°C, awaiting the availability of monkeys. The inoculum consisted of a 10% homogenate of the patient’s brain prepared in borate buffer pH 9. After centrifugation for 5 minutes at 800 g, a sufficient quantity of a solution of ’Crystamycin’ (Glaxo) was added so that each ml. of supernate contained 200 units of benzylpenicillin sodium and 0-2 mg. of streptomycin. Four squirrel monkeys (Saimiri sciureus) were inoculated intracerebrally with 0’5 ml. of the brain supernate under general ansesA similar homogenate was prepared from the thesia. same brain 10 months later, when spider monkeys also became available, and two such monkeys received intracerebral inoculations of 1-0 ml. each. The inoculated monkeys remained under observation, and those developing clinical signs received extra care. Moribund monkeys were killed by’Nembutal’ general anaesthesia and brains and spinal cords were removed immediately after death. After selecting a few pieces of brain for subinoculations the remainder of the brain Paraffin sections 5 im. was fixed in 10% formol saline. in thickness were stained with hxmatoxylin and eosin or luxol fast-blue and frozen sections were stained with Cajal’s gold chloride for the demonstration of astrocytes.’
Fig. I-Cerebral cortex of the patient with Creutzfeldt-}akob disease, whose brain was used for transmission experiments. (H. & E. ; ;" 225.)
Case-history patient whose brain was used for transmission experiments was a 50-year-old British woman who had no family history of neurological disease. 6 years before her death she had had a clinically typical attack of poliomyelitis which left her with weakness of the trunk and such wasting of the right leg that she was able to walk only with callipers and crutches. Her illness began with the onset over many days of progressive confusion, loss of memory, and depression. The
On examination, after about 8 weeks from the onset of clinical signs, she was disorientated and behaved ab-
normally ; conjugate gaze was defective, her eye movements were slow and fragmented, and there was horizontal nystagmus
to
the left.
Both
arms
and the left
leg
Fig. 2-Astrocytic reaction in the cerebral patient. (Cajal; Y 225.)
fluid: no cells were seen, but protein 45 mg. per 100 ml. and sugar 62 mg. per 100 ml. were noted. Repeated showed electroencephalograms repetitive complexes strongly suggestive of subacute spongiform encephalopathy. Her condition deteriorated further; myoclonic jerking was detected in the arms and she became totally unresponsive before dying after an illness that had lasted for about 4 months. The necropsy, the first of a case of Creutzfeldt-Jakob disease to be carried out in the National Hospitals for a few decades, was started 30 minutes after death, and the only macroscopic abnormality was fibrosis of muscles of the trunk and right leg. The brain weighed 1230 g. Histologically there was extensive loss of neurons, proliferation of astrocytes and gliosis in the cerebral cortex, accompanied by typical spongy vacuolation both of cells and neuropil (figs. 1 and 2). Similar lesions were present in some thalamic and brainstem nuclei and to a lesser extent in the cerebellum. Argyrophilic plaques and neurofibrillary tangles were not seen. There were
of the above
active lesions in the spinal cord. Electron microscopy of the brain confirmed the presence of empty vacuoles in cell processes, apparent redundant profiles of trilaminar membranes, and other less specific degenerative features. The pathological diagnosis was subacute spongiform encephalopathy. Tissue-cultures of the patient’s brain were established and the extensive virological investigations will be reported elsewhere.1O
no
Results
were
ataxic, the right leg and trunk being wasted. The right plantar and left flexor responses were absent. Laboratory investigations included examination of the cerebrospinal
cortex
No signs of disease were noticed in any of the inoculated squirrel monkeys during the first 19 months. However, between the 20th and 21st month a general slowing down of movements and activity became obvious in three monkeys; at the same time the animals began losing weight. The loss of weight was slow and gradual in two monkeys but very rapid in one, leading to emaciation and wasting. To begin with, appetite was maintained in all animals. Two of the monkeys, the wasted one and a second animal, also developed a severe itch with constant scratching and rubbing of the body against the bars of the cage. Loss of hair became striking along the tail, especially the base, where the sacral region was covered with excoriations and scabs. Very soon the highly wasted monkey developed neurological signs in the form of tremors along the body and extremities and occasional twitching of the facial muscles. The back of the animal was continuously arched and the hind legs were lifted very high during progression. The animal at
437
rigid, lost appetite, and his vision became affected, and he finally collapsed and was killed in extremis 2 years and 22 days after the intracerebral
became
inoculation. The second
monkey
to
develop neurological signs
that never showed any evidence of itch The first signs began with visual disor rubbing. turbances, ataxia, rigidity of the body, tremors, and partial paralysis of the right upper limb. He kept his back arched and walked with an obvious limp (fig. 3). He frequently fell and had great difficulty in getting up. Soon, however, he was unable completely to lift himself up and his extremities became flaccid. He became moribund and was killed 2 years and 2 months after the experimental inoculation. Although the neurological signs were late to appear in this monkey, once they became obvious they. progressed very quickly and the animal had to be killed within 4 weeks of the appearance of definite neurological signs. was
the
one
Fig. 3-Squirrel monkey in Note arched back and rigid
an
Fig. 4-Spongy degeneration in the cerebral cortex of an affected monkey. (F3. & E. ; x225.)
however, revealed
severe lesions throughout the of the less same pattern in the three brain, squirrel monkeys. The most striking change consisted of widespread spongy degeneration and astrocytosis throughout the grey-matter of the central nervous system. The most severe spongy degeneration was seen in the corpus striatum, while in the cerebral cortex lesions were of a slightly lesser intensity but uniformly spread throughout (fig. 4). Similar lesions but of a patchy character were present in the thalamus, while somewhat less striking changes were seen in the midbrain and the brainstem. In the cerebellum, the molecular was affected, while in the only layer cord mild affected the whole greyspinal changes In addition to the above changes neuronal matter. degeneration was very obvious, especially in the cerebral cortex, while vacuolation of neurons was very pronounced in the corpus striatum. Neuronal
very
more or
advanced stage of the disease. (Approximately ’/. life size.)
posture.
The third monkey had a clinical history somewhat similar to the first one with an itch, rubbing, and emaciation, but the clinical signs were spread over a much longer period (4 months) and he lived for 2 This years and 2 months after the inoculation. animal, apart from rigidity, tremors, and ataxia, also showed a variety of peculiar signs; he either pressed his forehead against the bars of the cage and held his head with both hands, or assumed an unnatural posture where he would be standing for long periods in an arched position with the head down on the floor of the cage and holding the head tightly with his hands. The fourth squirrel monkey has not yet shown any definite signs of disease, 2 years and 3 months after inoculation, neither has any abnormality been noticed in the two spider monkeys (1 year and 6 months after
inoculation). Post-mortem
examination
did
not
reveal
any
definite changes in the internal organs, but there was a slight decrease in the size of the brains and an increase in the amount of the cerebrospinal fluid.
Histological examination of brains
and
spinal cords,
Fig. 5-Neuronal vacuolation in the globus paHidus of an affected monkey. (H. & E. ; x 480.)
438 vacuolation affected the larger neurons of the globus pallidus and the medium-size neurons of the putamen and caudate nucleus. The vacuoles were confined, as
Preliminary Communication
rule, to the cytoplasm (fig. 5). They were small, usually more than one vacuole was found in the cell; however, in many neurons large numbers of a
REYE’S SYNDROME DUE TO A NOVEL PROTEIN-TOLERANT VARIANT OF ORNITHINE-TRANSCARBAMYLASE DEFICIENCY
and
vacuoles often became confluent and sometimes displaced the nucleus. Astrocytic proliferation and hypertrophy affected the whole grey-matter, but in the thalamus, corpus striatum, medial geniculate body, and the olives, apart from severe hypertrophy, there was also a considerable increase in the number of astrocytic processes which often formed a dense network (fig. 6).
M. MICHAEL THALER
Department of Pediatrics, University of California, San Francisco, California 94143 MARILYN BOSWELL J. HOOGENRAAD* Division of Developmental Medicine, Department of Pediatrics, Stanford University School of Medicine, Palo Alto, California 94305
NICHOLAS
A novel variant of ornithine-transcarbamylase (O.T.C.) deficiency has been discovered in a patient with encephalopathy and fatty visceral degeneration (Reye’s syndrome). Hepatic ornithine transcarbamylase activity was 20·7% of normal, with extremely low affinity of the enzyme for ornithine, and substrate inhibition by carbamyl phosphate and ornithine. The findings reveal a link between Reye’s syndrome and heritable O.T.C. deficiency, and suggest that administration of ornithine or arginine may be specific therapy for this condition.
Summary
Fig. 6-Astrocytosis in the cerebral cortex of an affected monkey. (Cajal; x 225.)
INTRODUCTION
Discussion
.
THE
clinicopathological entity reported in 1963 by Reye et al 1 as encephalopathy and fatty degeneration of the viscera" has since been recognised as a major cause of mortality and morbidity among infants and children. Reye’s syndrome is endemic to most parts of the world, and frequently occurs in association "
findings provide unequivocal evidence of the transmissibility of Creutzfeldt-Jakob disease- and confirm the work of Gibbs et al.and Gajdusek and Gibbs." Of special significance is the comparative ease with which the disease was transmitted, since These
this has been our first attempt to transmit CreutzfeldtJakob disease. At the same time it draws attention to the great value of the squirrel monkey as a laboratory animal for work on the transmissibility of subacute spongiform encephalopathies of man. The squirrel monkey is a small friendly animal which is easily handled and can therefore be kept without difficulty in a laboratory. It is much more readily obtainable and considerably cheaper than the large apes such as
chimpanzees.
It is remarkable that both the clinical picture and the histological changes in the central nervous system of the squirrel monkeys resembled those of the patient from whom the disease was transmitted. However, the intensity of lesions was much greater in the animals than in the patient from whom the disease was transmitted, a fact common with other transmissible spongiform encephalopathies of animals, such as scrapie in mice, mouse scrapie in the goat, or transmissible mink encephalopathy in goats,u-15 We thank Miss M. I. Stokes and Mr J. A. Mills from the National Hospitals for their help with necropsy specimens. Requests for reprints should be addressed to 1. Z. REFERENCES 1.
Gibbs, C. J., Jr., Gajdusek, D. C., Asher, D. M., Alpers, M. P., Beck, E., Daniel, P. M., Matthews, W. B. Science, 1968, 161, 388.
viral infections of childhood such as and influenza &bgr;.2 The disorder progresses chickenpox mild from a prodromal viral illness to vomiting and abdominal pain which signal the sequential appearance of cerebral dysfunction and coma. Mortality is at least 40 %; survivors recover rapidly. The biochemical abnormalities most frequently observed in Reye’s syndrome include raised serum-transaminase levels, prolonged prothrombin-time, and hypoglycxmia. Blood-
with
*
common
Present address: Department of Biochemistry, LaTrobe University, Bandoora, 3083 Victoria, Australia. DR ZLOTNIK AND OTHERS:
REFERENCES—continued
D. C., Gibbs, C. J., Jr. Brain, 1973, 96, 1. J., Jr., Gajdusek, D. C. Science, 1973, 182, 67. J., Jr., Gajdusek, D. C. ibid. 1969, 165, 1023. D. C., Gibbs, C. J., Jr. Nature, 1971, 230, 588. Gajdusek, D. C., Gibbs, C. J., Jr., Alpers, M. ibid. 1966, 209, 794 Gajdusek, D. C., Gibbs, C. J., Jr., Alpers, M. Science, 1967, 155, 212. Gajdusek, D. C., Gibbs, C. J., Jr. in Biohazards in Biological Research (edited by A. Helman, M. N. Oxman, and R. Pollock); p. 288. New York, 1973. 9. Zlotnik, I. Br. J. exp. Path. 1968, 49, 555. 10. Slack, P. M., Dayan, A. D., Slavin, G., Tyrell, D. A J
2. 3. 4. 5. 6. 7. 8.
11. 12. 13. 14.
15.
Roos, R., Gibbs, C. Gibbs, C. Gajdusek,
Gajdusek,
Unpublished. Zlotnik, I., Rennie, J. C. J. comp. Path. 1962, 72, 360. Zlotnik, I., Rennie, J. C. ibid. 1963, 73, 150. Zlotnik, I., Rennie, J. C. ibid. 1965, 75, 147. Zlotnik, I. National Institute for Nervous Diseases and Blindness monograph no. 2. U.S. Public Health Service publication 1378, p. 237. U.S. Government Printing Office, 1965. Zlotnik, I., Barlow, R. M. Vet. Rec. 1967, 81, 55.