Lipofuscinosis Ceroidea Neuronal - II

7/30/2019 Lipofuscinosis Ceroidea Neuronal - II

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Seizure 2004; 13: 235240

doi:10.1016/S1059-1311(03)00163-8

Neuronal ceroid lipofuscinosis:

a clinicopathological study

SANJIB SINHA , P. SATISHCHANDRA , VANI SANTOSH , N. GAYATRI & S. K. SHANKAR

Departments of Neurology and Neuropathology, National Institute of Mental Health &Neurosciences (NIMHANS), Bangalore, India

Correspondence to: Dr P. Satishchandra, Professor and Head, Department of Neurology, National Institute ofMental Health & Neurosciences (NIMHANS), Bangalore, India. E-mail: [email protected]

We report the clinical, electrophysiological, radiological and morphological features in a series of 12 patients of histopatho-

logically confirmed cases (infantile, juvenile and adult onset) of neuronal ceroid lipofuscinosis (NCL) observed from 1979to 1998 at National Institute of Mental Health & Neurosciences (NIMHANS), Bangalore (South India). The commonest type

of NCL was juvenile (n = 8, 67%) while infantile and adult forms were two each (n = 2, 16.8%). The age at presentation

ranged from 2 to 45 years (mean12.6, 14.3 years; median7 years; M:F ratio of 2:1). Four patients (33%) had positive

family history and five patients had history of consanguineous parentage (41.6%). The commonest presenting symptoms were

regression of milestones (83.3%) and/or seizures, myoclonus (83.8%) followed by involuntary choreiform movements (50%),

visual loss (41.6%), ataxia (33.3%) and abnormal behaviour (16.6%). Neuro-ophthalmological abnormalities like optic atrophy

(50%), maculardegeneration (33.3%) and retinitis pigmentosa (8.3%) were seen in two thirds. Nerve conduction studies (n = 4)

revealed abnormalities in two, suggestive of sensorimotor neuropathy. Scalp EEG (n = 9) showed slowing of background

activity (BGA) of varying degrees with paroxysmal bursts of seizure discharges in majority. Cranial CT scan (n = 4) revealed

varying degrees of diffuse atrophy. Diagnostic brain biopsy was carried out in 11 and brain was examined at autopsy in 1 case.

Histological examination revealed characteristic PAS and Luxol Fast Blue (LFB) positive, autofluorescent (AF) intracellular

ceroid material, both in neurons and astrocytes in the grey matter. Electron microscopy (n = 5) revealed curvilinear (n = 4),

lamellar (n = 2) and electron dense (n = 2) inclusions in neurons, astrocytes and vascular endothelial cells. To conclude,

this neurodegenerative disease had varied but characteristic clinical presentations and required histopathological confirmation

of diagnosis.

2003 BEA Trading Ltd. Published by Elsevier Ltd. All rights reserved.

Key words: neuronal ceroid lipofuscinosis (NCL); infantile NCL; juvenile NCL; Kufs variant.

INTRODUCTION

The neuronal ceroid lipofuscinosis (NCL) represents

one of the common progressive neurodegenerativedisorders during childhood in which the exact chem-

ical nature of storage material and primary enzyme

defect are still being defined1, 3. The term NCL was

coined by Zeman et al. to distinguish the familial cere-

bromacular degeneration clinically and pathologically

distinctive from gangliosidoses3. The disease group

has an autosomal recessive pattern of inheritance and

is commonly characterised by progressive myoclonic

epilepsy with visual failure and accumulation of an

autofluorescent lipopigment in the neurons and glial

elements1, 35. The recognised clinico-pathological

forms of NCL are (1) infantile, (2) late infantile, (3)

early juvenile, (4) juvenile, (5) adult, and (6) Finnish

variant of late infantile form57.The diagnosis is established by characteristic

histopathological and ultrastructural features in brain

and various other extracerebral tissues1 ,3 ,8. Though it

has characteristic clinico-pathological features, there

are only two published case reports from India14,15.

The aim of the present study is to review the pre-

senting manifestations and clinical features and to

review the histopathological characteristics in a series

of neuronal ceroid lipofuscinosis cases seen at one

center in South India.

10591311/$30.00 + 0.00 2003 BEA Trading Ltd. Published by Elsevier Ltd. All r ights reserved.


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236 S. Sinha et al.

MATERIALS AND METHODS

The case records of 12 patients of neuronal ceroid lipo-

fuscinosis (NCL) confirmed pathologically obtained

from the medical records were reviewed to evaluate the

clinical, electrophysiological and pathological charac-

teristics. The family history and the clinical manifes-

tations of the disease were carefully noted. CSF re-

sults were available in 10 patients. Scalp EEG (n =9), nerve conduction studies (n = 4), cranial CT scan

(n = 4) findings were noted.

Histopathological study of brain

Eleven patients underwent diagnostic right frontal

brain biopsy after obtaining the informed consent.

In addition partial autopsy confined to the brain was

performed in one patient who succumbed to the ill-

ness. One half of the tissue was fixed in 10% of

buffered neutral formalin for light microscopy (LM)and the other half in 3% gluteraldehyde for elec-

tron microscopic (EM) studies. Paraffin embedded

sections from each case were routinely stained with

Haematoxylin and Eosin (H & E), Luxol Fast Blue

(LFB) and Periodic Acid Schiff (PAS) to identify

the stored lipofuscin pigments. The unstained sec-

tions were examined under fluorescent microscopy by

epi-illumination (HBO50, DMRB, Leica) to record

autofluorescent (AF) nature of the intracellular ad-

ventitious material. For EM study, the grey and white

matter from the biopsied tissue, (fixed earlier) were

processed separately. The ultra-thin sections were

stained with uramyl acetate/lead citrate and viewed

under JEOL 100 CX at 60 kV16.

RESULTS

The age of the patients at the time of presentation

ranged from 2 to 45 years (mean12.6 14.3 years,

median7 years). There were eight male and four fe-

Table 1: Clinical manifestations in NCL.

Features INCL (n = 2) JNCL (n = 8) ANCL (n = 2) Total (n = 12) (%)

Regressed milestones 2 8 1 11 (91.6)

Seizures 3 7 10 (83.3)

Myoclonus 2 7 5 (41.6)

GTCS 1 3 4 (33.3)

Partial seizures 1 1 (8.3)

Involuntary movements 2 5 2 9 (75)

Visual abnormalities 1 7 8 (66.6)

Pyramidal signs 1 5 1 7 (58.3)

Ataxia 3 1 4 (33.3)

Dementia 1 1 2 (16.6)

Abnormal behaviour 2 2 (16.6)

male patients. The mean age of onset of symptoms was

10.814.5 years (median4.75 years). The duration

of illness ranged from 1 to 60 months (21.8 18.3

months). Five patients (33%) had positive family his-

tory with autosomal recessive pattern of Mendelian

inheritance. Consanguineous parentage was noted in

five (41.6%) and two among them had positive family

history.

Based on the age of onset of illness, three clinicalforms could be categorisedinfantile NCL = 2 cases

(mean age1.250.4 years), juvenile NCL= 8 cases

(mean age5.6 9 years) and adult NCL = 2 cases

(mean age41.52.1 years). The presenting clinical

features in infantile NCL group were regressed mile-

stones, chorea, seizures and myoclonus, while juvenile

NCL patients had regression of acquired milestones

(100%) followed by chorea and choreoathetosis (3/8),

seizure and myoclonus (3/8) and ataxia (3/8). The two

cases with adult variant of NCL presented with ab-

normal behaviour and extra pyramidal features, one of

them in addition had pyramidal signs, ataxia and de-mentia (Table 1). Only one case of infantile NCL had

primary optic atrophy. The patients in juvenile group

had ophthalmic abnormalities in the form of primary

optic atrophy (6/8), macular degeneration (3/8) and re-

tinitis pigmentosa (1/8). The adult forms did not have

any ophthalmic abnormality.

Cranial CT scan (n = 4) revealed severe diffuse at-

rophy in three patients and mild to moderate diffuse

atrophy in one. CSF examinations carried out in 10 pa-

tients revealed mean cell count of 2 mm3; the CSF pro-

tein was raised in one case of juvenile NCL (220 mg%)

and in one adult NCL case (114 mg%). Scalp EEG

was abnormal in majority of the patients (8/9). The

background activity (BGA) revealed paucity of alpha

waves (n = 8) with varying degree of diffuse slow-

ing. Paroxysmal generalised bursts of spike and wave

(n = 4), polyspikes and wave (n = 3) and or or

bursts of slow waves (n = 2) were observed. Nerve

conduction studies carried out in four were abnormal

in two (50%). One had decreased compound muscle

action potential (CMAP) when recorded over right


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NCL 237

Fig. 1: Microphotograph showing granules within the neurons and astrocytes exhibiting autofluorescence; this feature isdiagnostic (AF 480).

extensor digitorum brevis (EDB) and decreased sen-

sory nerve action potential (SNAP) when recorded

over right sural nerve while the second patient had

increased distal latency of 9.0 millliseconds (ankle to

EDB).

On histopathological examination, the brunt of the

disease was noted in the grey matter with relatively

unaffected white matter. There was evidence of vari-

able degree of focal neuronal loss in the superficial

layers, while in the deeper layer neuronal swelling

was noted. In the zones of neuronal depletion and

swelling, mild to moderate reactive astrocytosis with

prominent processes was noted. Most of the neurons

and reactive astrocytes have intracytophasic LFB

positive, autofluorescent ceroid lipofuscin material

displacing the Nissls substance (Fig. 1). Similar ma-

terial was also noted in the neuropil as free granules,infiltrating granules and occasionally in endothelial

cells. Ultrastructural studies carried out in five cases

revealed characteristic inclusions within the neuronal

cytoplasm. Such inclusions were also noted in as-

trocytes and endothelial cells. They were curvilinear

inclusion in four, lamellar inclusions in two and elec-

tron dense inclusions in another two cases (Fig. 2).

Four patients were followed up regularly from 3

months to 4 years (18.719.9 months). Patients with

seizure and myoclonus were treated with antiepileptic

drugs. One adult onset NCL patient who had a pro-

tracted course died in the hospital after 3 months of

illness. One of the juvenile NCL patient had a follow

up of 4 years during which period his jerks were not

controlled with anticonvulsants and developed pro-

gressive loss of acquired milestones and optic atrophy.

DISCUSSION

NCL was more frequently seen among males (M:F

2:1). The mean duration of illness prior to presenta-

tion was 22 months. Similar data had been reported in

literature35, 7. Three clinical forms of NCL could be

identified in the present study i.e. infantile NCL (n =

2), juvenile NCL (n = 8) and adult NCL (n = 2).

Most of the literature revealed juvenile NCL to be thecommonest3. However, Sjogren (1931) and Raynes

(1962) had reported equal occurrence of juvenile NCL

and late infantile NCL3. The adult variety of NCL re-

marks to be a rare entity in most series as in the present

one3 ,4 ,7. To the best of our knowledge, this is the first

series of NCL from India.

In the present series neuro-ophthalmic abnormali-

ties were seen in two thirds of cases with primary

optic atrophy as the commonest abnormality4, 7 fol-

lowed by macular degeneration (n = 4) and retinitis


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238 S. Sinha et al.

Fig. 2: Electron micrograph showing characteristic curvilinear inclusions (marked as C) within the neuronal cytoplasm (30 000).

pigmentosa (n = 1). None of the adult NCL patients

had any ophthalmological findings. Zeman et al. had

found pigmentary changes in 25, macular degenera-

tion in 12 and optic atrophy in 5 patients giving clue

to the diagnosis when associated with regression of

milestones and myoclonus3.

Both adult NCL patients in this study had mani-

fested in early part of the fifth decade. One of these

adult NCL had been reported earlier17. The oldest pub-

lished case of NCL in literature was at the age of 63

years18. The two Kufs adult variant cases in our series

could be categorised into type B with predominant

behavioural changes, dementia, ataxia and rigidity as

described in 19975,19.

Since long-term follow up is not available in our

series, it is difficult to comment on the natural history.

Generally the infantile and late infantile NCL have arapid course and child usually dies within 510 and

12 years, respectively. In juvenile NCL death occurs

by 1525 years, while the adult group has a variable

course35. The response to anticonvulsants has been

unsatisfactory3 ,4 ,7.

The juvenile NCL may have circulating vacuolated

lymphocytes in 1050% of smears3 ,4 ,7. The periph-

eral blood film did not reveal any vacuolated lympho-

cytes in this series. The significance of raised CSF

protein noted in two of our cases remains unexplained

and has not been reported in the literature. MRI of

brain in these patients reveals diffuse cerebral and

cerebellar atrophy and it tends to increase with dis-

ease progression21. MRI could not be performed in

this study. The cerebral atrophy noted in the cranial

CT scan had been observed to worsen as the disease

progresses on serial neuroimaging studies2022. Nerve

conduction studies are usually within normal limits

though subclinical peripheral neuropathy had been

reported2, 3. Nerve conduction performed in our pa-

tient showed features of neuropathy in two4, 7. Photic

response in EEG is characteristically reported with

low frequency stimulation in NCL3,21,25. However,

this was not noticed in any of our cases in this series as

in any other form of progressive myoclonic epilepsy

reported from India8.

The cornerstone in the diagnosis of NCL is histo-pathological examination of the brain tissues with

demonstration of autofluorescence exhibited by the

lipopigments3, 5. This was evident in all of our cases.

Apart from brain, extra cerebral tissues may also

show inclusions, for example, lymphocytes, skin, rec-

tal mucosa, peripheral nerves1, 5,2128. The sensitivity

of finding inclusions in these extracerebral tissues,

which is easily accessible, is variable. We did not ex-

amine these peripheral tissues in our study and hence

cannot be commented. Electron microscopy in our


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NCL 239

series revealed that the inclusions were in neuronal

cytoplasm (100%) and endothelial cell (80%). The

commonest type of inclusion material was curvilin-

ear inclusion seen mainly in juvenile NCL patients.

Recently, based on the occurrence of morphological

forms of lymphocytic inclusions in electro microscopy

(EM), NCL had been further categorised as (1) infan-

tile NCLgranular bodies/GRODs, (2) late infantile

NCLcurvilinear bodies, (3) juvenile NCLfingerprint bodies, and (4) adult onset NCL with varied

forms and combination of inclusions1, 5. However, in

the present study characteristic fingerprint or GRODs

inclusions were not seen and instead curvilinear in-

clusions were seen in JNCL cases. A combination of

fingerprint and curvilinear inclusions in JNCL cases

had been documented in other studies29. Overlapping

of morphological features of inclusions, formation of

inclusions of similar morphologies in different stages

of disease can lead to difficulties in diagnosis by

ultrastructural studies30.

Recently, the genotypes and the chromosomal locushad been determined in the various subtypes of NCL

and in some of them the gene products have also been

identified911 (Table 2). Attempts are made to under-

stand the pathogenesis by finding the primary abnor-

mality and assist in the prenatal and heterozygote de-

tection for counselling1,12,13. Genetic analysis could

not be done in the present series, as facilities for this

are not available in India. However, in future if ge-

netic analysis is done the genetic classification can

become the method of choice along with the possible

gene therapy.

To conclude, a clinical diagnosis of NCL should be

considered in all children with characteristic clinical

features of progressive myoclonic epilepsy and visual

failure and could be confirmed by histopathological

examination. Less invasive extracerebral tissue biop-

sies of skin, rectal mucosa, peripheral nerves could

be helpful and its role needs to be further evaluated

in larger number of cases. Electron microscopic stud-

Table 2: Genetic classification of NCLa.

Clinical

types

Genetic

types

Chromosomal

loci

Gene product

INCL CLN1 Ip32 Lysosomal palmitoyl

Protein thioesterase

LINCL CLN2 11p15 Lysosomal pepstatin

Insensitive peptidase

LINCL CLN5 13q31-32 Not known

JNCL CLN3 16p12 Transmembranous (?)

ANCL CLN4 Not known Not known

EJNCL CLN6 15q21-23 Not known

INCL = infantile NCL, LINCL = late infantile NCL,

JNCL = juvenile NCL, ANCL = adult NCL, EJNCL = early

juvenile NCL.a Adapted from Goebel and Sharp1.

ies are definitely complementary to further classify

NCL according to the characteristic forms of inclu-

sions. However, the main focus should be on the ge-

netic studies of the NCL, which is also useful in pre-

natal diagnosis, genetic counselling and possible gene

therapy in near future.

REFERENCES

1. Goebel, H. H. and Sharp, J. D. The neuronal ceroid lipofusci-

nosesrecent advances. Brain Pathology 1998; 8: 151

162.

2. Gutteridge, J. M. C., Westermarck, T. and Santavuori, P.

Iron and oxygen radicals in tissue damageimplication for

the NCL. Acta Neurologica Scandinavica 1983; 6: 365

370.

3. Zeman, W., Donanne, S., Dyken, P. and Green, J. The NCL

(Batten-Vogt syndrome). In: Handbook of Neurology, Vol.

10 (Eds P. J. Vinken and G. W. Bruyn). Amsterdam, North

Holland Publishers, 1970: pp. 588679.

4. Swick, H. M. Disease of grey matter. In: Pediatric Neurology,

Vol. 2, 1st edn. (Ed. K. F. Swaiman). USA, Mosby, 1988:

pp. 772794.5. Lake, B. D. Lysosomal and peroxisomal disorders. In:

Greenfields Neuropathology, 6th edn. (Eds J. H. Adams and

L. W. Duchen). London, Edward Arnold, 1997: pp. 668676.

6. Boustany, R. M., Alroy, J. and Kolodny, E. M. Clinical clas-

sification of NCL subtypes. American Journal of Human Ge-

netics 1988; 5 (Suppl.): 283289.

7. Brett, E. M. and Lake, B. D. Progressive neurometabolic brain

disease. In: Pediatric Neurology, 3rd edn. (Ed. E. M. Brett).

London, Churchill Livingstone, 1997: pp. 143200.

8. Acharya, J. N., Satishchandra, P. and Shankar, S. K. PME

and related disorders, a clinical and pathological appraisal.

Neurology (India) 1992; 40: 145153.

9. Williams, R., Vesa, J., Jarvel, I. and Gardiner, R. H. Genetic

heterogeneity in NCLevidence that LINCL (CLN2) is not

an allelic form of juvenile or infantile subtypes. American

Journal of Human Genetics 1993; 53: 931935.

10. Williams, R., Santavuori, P., Peltonen, L., Gardiner, R. H. and

Jarvela, I. Variants of LINCL (CLN5) is not an allelic form of

batten disease (CLN3)exclusion of linkage to CLN3 region

of chromosome 16. Genomics 1994; 20: 289296.

11. Gardiner, M., Sandord, A., Deadman, M. et al. Batten dis-

ease (Spielmeyer Vogt disease). Juvenile onset NCL gene

(CLN 3) maps to chromosome 16. Genomics 1990; 8: 387

390.

12. Conradi, N. G., Uvebrart, P., Hakegard, K. M., Wahlstrom,

J. and Mellqvist, L. First trimester diagnosis of JNCL by

demonstration of finger print inclusions in chorionic villi.

Prenatal Diagnosis 1989; 9: 283287.

13. Macleod, P. M., Dolman, C. L., Nickel, R. E., Chang, E.,

Zonana, J. and Silvey, K. Prenatal diagnosis of NCL. New

England Journal of Medicine 1984;310

: 595.14. Nadkarni, S., Despande, D. H., Mondkao, V. D. and Bharucha,

E. P. Neuronal ceroid lipofuscinosisclinical and histochem-

ical observations in 2 cases. Journal of Neurological Sciences

1979; 43: 395.

15. Radhakrishna, K., Bannerjee, A. K., Deve, S. R. and Chopra,

J. S. Late infantile NCL. Neurology (India) 1978; 26: 21

23.

16. Frasca, J. M. and Parks, V. R. A routine technique for double

staining ultrathin sections using uramyl and lead salts. Journal

of Cell Biology 1960; 25: 157162.

17. Shankar, S. K., Despande, D. H., Srinivas, H. and Kalyana-

sundaram, S. Dementiaa pathological study of 10 cases.

Neurology (India) 1982; 30 (2): 93103.


6/6

240 S. Sinha et al.

18. Constantinidis, J., Wishniewski, K. E. and Wisniewski, T. M.

The adult onset and a new late adult form of NCL. Acta

Neuropathologica (Berlin) 1992; 83 (5): 461468.

19. Berkovic, S. F., Carpenter, S., Andermann, F., Anderman, E.

and Wolfe, L. S. Kufs diseasea critical appraisal. Brain

1988; 11: 2762.

20. Savolaine, E. R., Voeller, K., Gunning, K. and Smith, R. R. CT

in NCL4 case reports. Journal of Computed Tomography

1987; 11: 7378.

21. Machen, B. S., Williams, J. P., Lum, G. B., Joslyn, J. W.,

Harper, M. D. and Dotson, P. MRI in NCL. Journal ofComputed Tomography 1987; 1: 160166.

22. Carpenter, S., Karpati, G. and Andermann, F. Specific in-

volvement of muscle, nerve and skin in late infantile

and juvenile amaurotic idiocy. Neurology 1973; 22: 170

174.

23. Pampiglione, D. and Harden, A. S. So called NCL

neurophysiological studies in 60 children. Journal of Neurol-

ogy, Neurosurgery, and Psychiatry 1977; 40: 323330.

24. Goebel, H. H. Fingerprint inclusion in non-vacuolated lym-

phocytes in JNCL. Clinical Neuropathology 1985; 4: 210

213.

25. Markesbury, W. R., Shield, L. K., Engel, R. T. and Jameson,

H. D. Late infantile NCL. Archives of Neurology 1976; 33:

630635.

26. Brod, R. D., Packer, A. J. and Van Dyk, H. J. Diagnosis

of NCL by ultra structural examination of peripheral blood

lymphocytes. Archives of Ophthalmology 1987; 105: 1388

1393.

27. Ceuterick, C. and Martin, J. J. Diagnostic role of skin or

conjunctiva biopsies in neurological disorders. Journal of the

Neurological Sciences 1984; 65: 170191.

28. Arsenio-Nunes, M. L., Goutieres, F. and Aicardi, J. An ultramicroscopic study of skin in the diagnosis of infantile and late

infantile types of NCL. Journal of Neurology, Neurosurgery,

and Psychiatry 1975; 38: 994999.

29. Carpenter, S., Karpati, G. and Andermann, F. The ul-

trastructural characteristics of the abnormal cytosomes in

Batten-Kufs disease. Brain 1977; 100: 137156.

30. Shankar, S. K., Gayathri, N., Yasha, T., Santosh, V. and

Ramamohan, Y. Electron microscopy in characterizing some

inherited metabolic neurologic diseases. In: Electron Mi-

croscopy in Medicine and Biology. Science Publishers, 2000:

pp. 131151.

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