lysosomal storage disease in man and animals

1. LYSOSOMAL STORAGE DISEASES
Presented by
PRADEEP. M
Submitted to

2. LSD
• To date 51 naturally occurring LSD
• Most of them are autosomal recessive
gene traits
• 9 inherited and 2 acquired mechanisms of
LSD known
• No sex predilection
• Brain lesions are prevalent (2/3rd of LSD)

3. HISTORY OF LSD
• Symptoms of some LSDs were described as early
as the 1880s,
• Many had been described and classified before
the lysosome was discovered in 1955 and before
their biochemical and genetic basis was fully
understood
• This is why they received common names (i.e.:
Gaucher disease, name of discovering physician).
• Later, an additional, more clinically descriptive
name often came into use (glucocerebrosidase
deficiency)

4. HISTORY OF LSD
Ernest GAUCHER (1854-1919)
Gaucher cell 1882

5. LSD IN HUMAN IN INDIA
(Sheth et.al., 2014)
• 387 children (34.8 %)
• highest prevalence of
• glycolipid storage disorders in 48 %,
• mucopolysaccharide disorders in 22 %
• defective sulfatide degradation in 14 % of the children.
• glycogen degradation defect and protein degradation defect in
5 % each
• Lysosomal trafficking protein defect in 4 %,
• Transport defect in 3 % of the patients.
• Higher incidence of Gaucher disease (16 %) followed by GM2
gangliosidosis that includes Tay-Sachs disease (10 %) and
Sandhoff disease (7.8 %) and mucopolysaccharide disorders
among all LSDs

6. Pathways for degradation of cellular
materials

7. BIOCHEMICAL AND CELLULAR BASIS
OF LSDS
Futerman AH & van Meer G (2004) 5:554-
565
1 catalytic activity
2 activator
3 misfolding
4 multienzyme
complex
5 glycosylation
6 M-6-P targetting
7 other transport steps
8 membrane
transporters
9 membrane regulators

8. General outline of LSD dysfunction:
• Mutations arising in hydrolytic enzyme, co-factor or
factor essential of enzyme delivery to lysosome
• Factors essential for lysosome function and
biogenesis (membrane proteins, channels and proteins of
• Unknown function) plus factors for protein traffic to lysosome
• Material (substrate) continues to be delivered to lysosome
resulting in ‘stored’ material, usually ‘primary’ and ‘secondary’
leads to swollen lysosomes

9. Effect of lysosomal storage diseases
• Direct consequence of abnormal accumulation of substrates
or catabolites in the lysosomes
Severe impairment of cellular structures and functions and abnormal
extracellular matrix. They include alterations of
– Signalling pathways,
– Intracellular Ca2+ homeostasis,
– Intracellular trafficking,
– Dysmyelinogenesis
– Formations of ectopic dendrites,
– Retarded bone formation, and
– Cloudy cornea
• Secondary changes are due to disrupted recycling
– manifested in abnormal or shortage of cellular and extracellular
products
– In the brain, there is a compromised recycling of neurotransmitter
receptors, altered signalling and dysmylinogenesis

10. LYSOSOMAL STORAGE DISEASE- DAMAGING
EFFECTS IN NERVOUS SYSTEM

11. LSD- SUB-CATEGORIES
1. Sphingolipidoses
2. Glycoproteinoses
3. Mucopolysaccharidoses (variable nervous system involvement)
4. Mucolipidoses (originally considered an MPS)
5. Glycogen storage diseases
6. Lipid storage disorders
7. Multiple enzyme defects
8. Transport defects
9. Batten Disease (Ceroid lipofuscinoses)- proteinosis
(Blue = nervous system involvement)

12. 1. SPHINGOLIPIDOSES
• Sphingolipids- 1. Sphingomyelins ( a phospholipid)
2. Glycosphingolipids
(cerebroside, globoside, sulfatides and gangliosides)
Ceramide= sphingosine + FA
Glycosphingolipid = ceramide + oligosaccharide
Cerebroside = glycosphingolipid with one sugar as side chain (gluc or
gal)
Globoside = glycosphingolipid with more than one sugar.. (glucose,
galactose or GalNAc)
Ganglioside = glycosphingolipid with one or more sialic acid (NANA)
linked to sugar side chain

13. STRUCTURE AND CATABOLISM OF
SPHINGOLIPIDS
GM1 Gangliosidosis
GM2Gangliosidosis
(Tay-Sachs disease, Sandhoff,
variant AB)
Glucocerebrosidosis
(Gaucher disease)
Sphingomyelinosis
(Nieman-Pick typeA, B)
Galactosialidosis
Galactocerbrosidosis
(Krabbe disease)
sialidase
αGalactosidase A
Sandhoff
Fabry disease

14. 2. GLYCOPROTEINOSES
glycoprotein contain hybrid type oligosaccharides
Alpha-Mannosidosis Beta- mannosidosis
Alpha-L-fucosidosis

15. MPS I (Hurler, Hurler-Scheie, Scheie)
MPS II (Hunter)
MPS III (San filipo Types A,B,C and D)
MPS IV (Morquio type A and B)
MPS VI (Maroteaux-Lamy)
MPS VII (Sly)
MPS IX (Hyaluronidase deficiency)
Multiple Sulfatase deficiency
3. MUCOPOLYSACCHARIDOSES
Defective metabolism of glycosaminoglycans

16. MUCOPOLYSACCHARODOSIS-
CLASSIFICATION

17. 4. MUCOLIPIDOSES
• Features of both sphingolipidosis and
mucopolysaccharidoses
Disease Enzyme deficient signs Species
affected
Mucolipidoses I αNuraminidase Neurological signs,
organomegaly, and
skeletal abnormalities.
cats
ML II (I cell disease) phospho-
N-acetylglucosamine
transferase
-do- cat
Pseudo-Hurler
polydystrophy
(ML-III)
phospho-
N-acetylglucosamine
transferase
-do-

18. MUCOLIPIDOSES II – (I cell disease)
• Face; cat, 8 months of age. Left. Note the broad
face with hypertelorism, thickened eye lids, and
small ears. Right. Note frontal bossing

19. 5. GLYCOGENOSES
• Glycogen storage disease type II (Pompe
disease)
– α1,4-glucosidase deficiency
(Only lysosomal enzyme in glycogen catabolism others are metabolic
pathways
(type III- Amylo1,6 glucosidase defiency- Cori disease;
type IV- Polyglucosan body disease) )

20. OTHER LYSOSOMAL STORAGE
DISEASES
• Ceroid-lipofuscinoses
– Neuronal ceroid-lipofuscinoses (Batten disease)
– Autofluorescent neurons
• Laffora disease
– Storage of polyglucosan

21. GENETICS OF LSD
• X linked diseases in human
– Fabry disease (alpha- galactosidase dificiency)- affect both sex
– MPS II
– Adult lipofuscinoses
• All other are autosomal recessive

22. INDUCED STORAGE DISEASES
• Swainsonine toxicosis
– An indolizidine alkaloid
– Seen in plants- locoweeds, broom weed etc.
– Inhibit αMannosidase- αMannosidoses
– “locoism” , “pea-struck” – identical to genetic form
• Trachyandra poisoning
– Ingestion of Trachyandra sps plants
– Lipofuscinosis- central & peripheral neurones
– sheep, horse, goat, pig
– Reason not known
• Phalaris poisnoing
– By ingestion of Phalaris sp - grass
– Accumulation of granular pigment material
– Sudden death form- by cardio toxic compounds
– Stagger form

23. Phalaris poisoning
• A. Pigmentation in lateral geniculate body (arrow)
• B. Neuronal accumulation of granular pigmented material (arrow head)
B

24. Drug induced LSD
• Amiodarone and chloroquine,
• Induce phospholipidosis and MPS.

25. TAY-SACHS DISEASE
• Mutation- chromo 15- Hex A gene-
hexaminidase deficiency- accu of GM2
ganglioside

26. Typical signs and symptoms of Fabry disease
Typical time at onset Signs and symptoms
Childhood and adolescence (≤16
years)
Neuropathic pain(burning sensation)
Ophthalmological abnormalities
(cornea verticillata and tortuous retinal
blood vessels)
Hearing impairment
Dyshidrosis (hypohidrosis and
hyperhidrosis)
Hypersensitivity to heat and cold
Gastrointestinal disturbances and
abdominal pain
Lethargy and tiredness
Angiokeratomas
Onset of renal and cardiac signs, e.g.
microalbuminuria, proteinuria,
abnormal heart rate variability
Early adulthood (17–30 years)
Extension of any of the above
Proteinuria and progressive renal
failure
Cardiomyopathy
Transient ischaemic attacks, strokes
Facial dysmorphism
Later adulthood (age >30 years)
Worsening of any of the above
Heart disease (e.g. left ventricular
hypertrophy, angina, arrhythmia and
dyspnoea)
Stroke and transient ischaemic attacks
Osteopenia and osteoporosis

27. LSD IN DOGS
Abnormality Breeds affected (RARE) Special tests Clinical features
Ceroid lipofuscinosis (Batten
disease)
many breeds - Australian cattle
dog, border collie, chihuahua,
cocker spaniel, dalmatian, wire-
haired dachshund, English setter,
Tibetan terrier
none, diagnosis
confirmed on post-
mortem
signs usually start around 1 to 2 years of age, vary between
breeds (see resources below), may include diminished
eyesight, abnormal behaviour, incoordination, seizures
Fucosidosis English springer spaniel
measure enzyme
levels
signs by 6 to 12 months, include slow learning, anxiety and
behaviour changes, which gradually progress over the next
18 months or so, to severe incoordination, dementia, visual
problems
Glucocerebrosidosis
(Gaucher's disease)
Australian silky terrier
measure enzyme
levels
signs by 4 to 8 months, include incoordination, tremors,
hyperactivity, stiff gait
Glycogen storage disease type
III (Cori's disease)
Akita, German shepherd
measure enzyme
levels
signs by 6 to 12 weeks, muscle tremors, incoordination,
hypoglycemia (low blood sugar), seizures, death by 8
months
GM1 gangliosidosis
Portuguese water dog, English
springer spaniel
measure enzyme
levels
signs by 2 to 4 months, include vision problems, lethargy,
difficulty in walking, death occurs by 8 months.
GM2 gangliosidosis (type B -
Tay-Sachs disease, type O -
Sandhoff's disease)
German short-haired pointer
(rare)
measure enzyme
levels
signs by 6 to 9 months, include visual problems, abnormal
behavior, incoordination, stiff gait
Mucopolysaccharidosis I Plott hound (rare)
measure enzyme
levels
signs often develop later, in adulthood; this is a connective
tissue disorder and signs include musculoskeletal
abnormalities, and heart disease due to thickened valves.
Sphingomyelinosis (Niemann-
Pick disease)
very rare
measure enzyme
levels
signs by 2 to 5 months, include incoordination, exaggerated
gait, dullness

28. CLINICAL SIGNS 0F LSD
• Cerebellar or
cerebellovestibular signs
such as tremor, ataxia,
dysmetria, and nystagmus
with progression to
paresis and paralysis
• Visual impairment, ataxia,
hypermetria, tremors,
seizures, and behavioral
abnormalities. Differ from
other LSD by diffuse forebrain
disease and central blindness can
be among the earliest clinical signs
• Gangliosidoses in dogs and cats
• Niemann-Pick disease types A and C
• Globoid cell leukodystrophy
• Canine Gaucher disease
• Feline -mannosidosis
• Ceroid lipofuscinosis

29. Lysosomal storage diseases in dogs –
MRI STUDY (Hasegawa D, Tamura S, Nakamoto Y, Matsuki N, Takahashi K, et al. (2013) Magnetic
Resonance Findings of the Corpus Callosum in Canine and Feline Lysosomal Storage Diseases. PLoS ONE 8(12)
• The corpus callosum was ‘not visualized’ or ‘partially visualized’ in all cases of the juvenile-onset group, including the Shiba Inu
dogs. It was barely recognized on both the transverse and sagittal images (Figures 1C, D), and in the domestic shorthair cats
with GM1 gangliosidosis (data not shown) and GM2 gangliosidosis (Figures 1E, F). In the late juvenile-onset group,
visualization of the corpus callosum in the toy poodles (Figures 1G, H) and golden retriever (data not shown) with canine GM2
gangliosidoses was inconsistent (‘partially visualized’ or ‘visualized but atrophic’), although the sagittal plane was not obtained
from 2 of the 3 cases. In those 2 groups, if the corpus callosum was evaluated as ‘partially visualized’, the visualized portion
was significantly atrophic (Figure 1C) and shortened longitudinally (Figure 1H). On the other hand, the corpus callosum of the
early adult-onset group, comprised of those with canine neuronal ceroid lipofuscinosis, was recognized clearly and consistently,
although it was thinner than those of normal animals (Figures 1I, J).

30. Gross pathology
• Radial nerve enlargement- fucosidosis in
dogs

31. GM1-gangliosidosis - vertebrae
• lumbar vertebrae from a 3-month-old normal Portuguese water dog
(A) and his brother with GM1-gangliosidosis (B). In (B) the vertebrae
are shorter and the intervertebral disks are irregular and wide.

32. Ceroidlipofuscinosis- sheep

33. • Gross appearance of aMiniature Poodle lung with Niemann-
Pick disease, showing diffuse nodular appearance of lung on
pleural and cut surface

34. HISTOPATHOLOGY

35. SPHINGOLIPIDOSIS
• GM1 gangliosidosis, type I, II and III
• GM2 gangliosidosis (Tay-Sachs type I, II, III and Sandhoff)
• Acid sphingomyelinase deficiency (Niemann-Pick A & B)
• Fabry disease- (alpha-galactosidase A deficiency)
• Farber disease (Ceramidase deficiency)
• Gaucher disease, type I, II and III
• Krabbe disease (galactosylceramidase deficiency)

36. GM1 GANGLIOSIDOSIS- CEREBRUM
Diffusely, neurons are moderately swollen with finely granular to microvacuolated
cytoplasm. Multifocally, glial cells often contain large, discrete, clear vacuoles

37. GM1 GANGLIOSIDOSIS- CEREBRUM
Rarely within the white matter there are spheroids characterized
by swollen, hypereosinophilic axons (arrow) X400

38. GM1 GANGLIOSIDOSIS- CEREBRUM
Within the white matter there are rare digestions chambers characterized by swollen
axon sheaths containing gitter cells and cellular debris (arrow

39. GM1-gangliosides- Luxol Fast Blue stain
• Paraffin section of the brain of a 7-month-old English
Springer Spaniel dog stained with Luxol Fast Blue
demonstrating the storage of GM1-gangliosides.

40. GANGLIOSIDOSIS
High power electronmicrograph of membranous cytoplasmic bodies (open
arrow) and zebra bodies (closed arrow) within a brain stem neuronal cell
body

41. GM1 GANGLIOSIDOSIS- CEREBRUM
Intraneuronal concentric lamellar membranous whorls

42. Globoid cell leukodystrophy
(krabb’s disease)
PAS - Mulinucleated macrophages ("globoid cells") and loss of myelinated fibers in a
case of Krabbe's leukodystrophy (globoid cells are enlarged vacuolated macrophages
typically localized around blood vessels)

43. Krabbe disease
( globoid cell leukodystrophy or galactosylceramide lipidosis)
• Globoid cells with electron microscopical view(*) (enlarged lysosomes
packed with twisted tubules)

44. α-Mannosidosis -cow
Cow cerebrum with a-mannosidosis illustrating
• (A) enlarged neurons with vacuolated cytoplasm, hematoxylin and eosin (H&E).
• B, Staining with Con A demonstrating the storage of oligosaccharides with terminal a-
mannosyl residues

45. MANNOSIDOSIS- CALF
neuronal cell bodies with vacuolated cytoplasm (arrows)

46. CEROID LIPOFUSCINOSIS- SPINAL
CORD
• Frozen section of spinal cord of a 11=2-year-old Australian Blue Heeler dog
with neuronal ceroid lipofuscinosis in spinal cord stained with Sudan black
revealing the storage of ceroid lipofuscin within the neurons.

47. DIAGNOSIS
• Hematological and routine biochemical assessment is
usually unremarkable
• Blood smear- storage vacuoles in leukocytes
• Lymph node aspirates or biopsies can be
diagnostically helpful
• Radiograph- bone deformities
• CSF analyisis- vacuoles in macrophages /
lymphocytes (eg. globoid cell leukodystrophy and fucosidosis)
• Peripheral nerve biopsy- Globoid cell leukodystrophyl, g
Niemann-Pick type A,47 -mannosidosis,2 and fucosidosis,
fucosidosis
• Muscle biopsy- glycogen storage disease
• MRI

48. Blood smear examination
• Blood smear of a patient with multiple sulfatase deficiency showing
a lymphocyte and neutrophil stained with Wright-Giemsa. The
cytoplasm of both cells contains metachromatic granules.

49. • Urine analysis (in MPS)
– Fucosidosis - excrete fucoglycoconjugates
– Cats with mannosidosis - mannose-rich
oligosaccharides
– MPS spot test- staining urine on filter paper
with toluidine blue- give a crude indication of
increased concentrations of glycosaminoglycans
in urine

50. Confirming diagnosis- live animal
• Enzyme activity analysis-
– homozygotes- 0-5% of normal activity
– Heterzygotes- 50% of normal activity
(dried blood sample analysis using fluorescent
substrate; Sewell et.al., 2012)
• 2o increase of activities of other enzyme
when one is deficient

51. ANTI-NIEMANN-PICK DISEASE, TYPE C1
(NPC1) (C-TERM) ANTIBODY
• Rabbit antimouse antibody

52. Flow diagram- LSD diagnosis

53. Treatment:
• BMT (membrane proteins)
• Enzyme replacement (BBB?)
• Gene therapy
• Substrate reduction- Miglustat (monosaccharide
• mimetic-imino sugar)
• Neuronal stem cells (membrane proteins?)
• Chemical chaperone therapy

54. conclusion
• Lysosomal storage diseases are one of
the less identified animal diseases in India
mainly due to lack of awareness. As these
are mainly a genetic disease, early
identification will help in taking proper
preventive measures like excluding the
related animals from breeding. More study
needed to be conducted to evaluate the
extend of the disease in the animal
population

55. THANK YOU

56. • Sheth, Mehul Mistri, Frenny Sheth, Raju Shah, Ashish Bavdekar, Koumudi Godbole, Nidhish Nanavaty, Chaitanya
Datar, Mahesh Kamate, Nrupesh Oza, Chitra Ankleshwaria, Sanjeev Mehta, andMarie Jackson (2014), Burden of
Lysosomal Storage Disorders in India: Experience of 387 Affected Children from a Single Diagnostic Facility
JIMD Rep. 2014; 12: 51–63.