Fazli R. Awan

Characterization of a Mouse Model of Sandhoff Disease by Magnetic

Resonance Methods

Supervisors

Dr Fran M Platt

Prof Peter Styles

Sandhoff Disease

• A type of Glycolipid (i.e. Glycosphinoglipid) lysosomal storage disorder

• Defect in the -gene of the enzyme - Hexosaminidase A () and B ()

• A neurodegenerative disease

Knockout mouse model for Sandhoff disease

Head tremor

Muscle weakness

Ataxic gait

Motor dysfunction

Hind limb paralysis

Immobile unresponsive state

Humane end point

Natural death

Age in Weeks

1817161514131211109876543210

Disease Progression in Sandhoff Mice

2-Month 3-Month 4-Month

What mouse models could tell us about the diseases?

Much about the biochemistry, pathology and clinical course of the disease

BY…….

Various biochemical and immunohistology methods

In Vitro or Ex Vivo

(a main limitation)

BUT ?

To find out disease related changes in

Sandhoff mice by in vivo MR methods

Aim

400 MHz Vertical Bore Magnet

Mouse head

Mouse here

Anaesthesia and Scavenging

tubes

Portion of the Probe showing coil region

Anaesthesia RF cable

Data acquisition

Magnet

RF coil

Heating blanket

Water pipe

Body temperature

Heart rate / ECG

Mouse

IMAGING PROTOCOL

MRI Scans

• Brain water diffusion studies by ADC mapping

• T2 and T1 Relaxation Times

• Magnetization Transfer Ratios (MTRs)

• Cerebral blood flow (CBF) by ASSIST-MRI

MRI Data AnalysisIn whole brain

Horizontal Sagittal

MRI Data Analysis

In specific regions

STUDY DESIGN

(Disease stages studied in Sandhoff mice)

Presymptomatic (2 month)

Early symptomatic (3 month)

Terminal stage(4 month)

Apparent Diffusion Coefficient (ADC) Decreases in Sandhoff Mice

0

20

40

60

80

100

Time point (months)

AD

C (

in x

10-5

mm

2/se

c)

Control 78 70 71

Sandhoff 71 67 65

2M 3M 4M

*

Whole-brain8% ↓

Diffusion of water in Sandhoff mouse brain is low

slice 1

0

100

200

300

400

20 35 50 65 80 95 110 125 140

Fre

qu

en

cy

slice 2

050

100150200250300

20 35 50 65 80 95 110 125 140

Fre

qu

en

cy

slice 3

0100200300

400500600

20 35 50 65 80 95 110 125 140

Fre

qu

en

cy

slice 4

0

200

400

600

800

20 35 50 65 80 95 110 125 140

Fre

qu

en

cy

slice 5

0

50

100

150

200

250

20 35 50 65 80 95 110 125 140

Fre

qu

en

cy

whole brain

0

500

1000

1500

2000

2500

20 35 50 65 80 95 110 125 140

Fre

qu

en

cy

(SH = 65, C = 70)(SH = 64, C = 64)

(SH = 64, C = 70)(SH = 66, C = 71)

(SH = 67, C = 73)(SH = 64, C = 71)

Sandhoff Control

Bins on x-axis in all of the above histograms represent ADC (10-5 mm2/sec)

ADC Histograms for 4-Month-Old Mice

0

20

40

60

80

100

2 Month 3 Month 4 Month

Time points

ADC

(10

-5 m

m/s

ec)

Control (n=13) Sandhoff (n=10)

*

Decrease in ADC in the cerebral cortex of Sandhoff mouse brain

ADC Analysis in Specific Regions of Sandhoff Mouse Brain

Water diffusion in Sandhoff mice was low at all the three ages, in almost all of the brain regions investigated.

11% ↓

T2 (Spin-Spin) Relaxation Time

(Reduced in Sandhoff Mouse Brain ! )

0

10

20

30

40

50

Time point (months)

Tim

e (i

n m

sec)

Control 36 36 38

Sandhoff 40 37 35

2M 3M 4M

**

In whole-brain8% ↓

Histograms for T2 Relaxation Time in 4-Month-Old Mice

slice 1

050

100150200

10 15 20 25 30 35 40 45 50 55 60 65 70 75 80

Fre

qu

en

cy

slice 2

050

100150200

10 15 20 25 30 35 40 45 50 55 60 65 70 75 80

Fre

qu

en

cy

slice 3

0

100

200

300

10 15 20 25 30 35 40 45 50 55 60 65 70 75 80

Fre

qu

en

cy

slice 4

0

100

200

300

10 15 20 25 30 35 40 45 50 55 60 65 70 75 80

Fre

qu

en

cyslice 5

0

50

100

10 15 20 25 30 35 40 45 50 55 60 65 70 75 80

Fre

qu

en

cy

whole brain

0

500

1000

10 14 18 22 26 30 34 38 42 46 50 54 58 62 66 70

Fre

qu

enc

y

(SH = 35, C = 38) (SH = 36, C = 38)

(SH = 34, C = 38)(SH = 36, C = 39)

(SH = 34, C = 37)

(SH = 35, C = 38)

Sandhoff Control

Bins on x-axis in all of the above histograms represent T2 relaxation time (msec)

T2 Relaxation Time Decreases with disease progression

20

25

30

35

40

45

50

2 month 3 month 4 month

Olfactory bulb

Tim

e (m

sec)

Control Sandhoff

*

20

25

30

35

40

45

50

2 month 3 month 4 month

Brain stem

Tim

e (m

sec)

Control Sandhoff

**

(A)

(B)

T2 relaxation time:

• Increases in controls from 2-4 month

• Decrease from 2-4 month in Sandhoff mice

T1 (Spin-lattice) Relaxation Time

(Decreases in Sandhoff Mouse Brain ! )

0.0

0.5

1.0

1.5

2.0

Cerebral cortex (s2) Hippocampus (s2) Thalamus (s2) Cerebral cortex (s3)

Tim

e (s

eco

nd

s)

Control Sandhoff

**

**

4-month-old Sandhoff and control mice

Magnetization Transfer Ratios (MTRs) (in Sandhoff and Control Mouse Brain)

Lower MTRs in 3 month Sandhoff mice

Higher MTRs in 4 month Sandhoff mice

BUT

NOT SIGNIFICANTLY

Cerebral Blood Flow (CBF)

(Measured by ASSIST-MRI)

0.00

0.02

0.04

0.06

0.08

0.10

0.12

0.14(A

rbit

rary

un

its)

2m 3m 4m

2m 0.048 0.040 0.044 0.030 0.055 0.051

3m 0.053 0.068 0.042 0.056 0.054 0.073

4m 0.052 0.083 0.040 0.062 0.047 0.090

Control Sandhoff Control Sandhoff Control Sandhoff

Cerebral cortex Hippocampus Thalamus

*

*

CBF increases in Sandhoff mice with disease progression

0.00

0.02

0.04

0.06

0.08

0.10

0.12

0.14

Cerebral cortex Hippocampus Thalamus

(Arb

itra

ry u

nit

s)

Control Sandhoff

* *

Cerebral Blood Flow (CBF)

(comparison of 4-month-old mice)

Summary

• ADC (water diffusion in the brain) reduces in Sandhoff mice

as compared to their age-matched controls

• T2 relaxation time decreases in Sandhoff mice with the

progression of disease

• T1 relaxation time is short in Sandhoff mice as compared to

their age-matched controls.

• No significant differences in the magnetization transfer

ratios (MTRs) between control and Sandhoff mice.

• Significant increase in the cerebral blood flow (CBF) in the

terminal stage Sandhoff mice as compared to controls.

• This increase in the CBF appears to progress with the

disease

Conclusions

• This is the first time MRI study of Sandhoff mouse brain, and only limited data is available from clinical studies.

• These findings may have applications for disease diagnosis and therapy monitoring by non-invasive MRI methods.

Acknowledgements

Members of 1st. Floor

Mr David Smith

Dr M. Jeyakumar

Dr David Neville

Dr Terry Butters

Prof Raymond Dwek

Dr Fran PlattProf Peter Styles

Members of ENG

Dr Niki Sibson

Dr Kerry Broom

Dr John Lowe

Dr AM Blamire

Dr Justin Smith

Mr Andre ThomasMr Phillip Bradford

(Members of BSU)

Experimental Neuroimaging Group (ENG), University Lab. of Physiology