An Optimized Mouse Model for Transient Ischemic Attack

Pedrono E, Durukan A, Strbian D, Marinkovic I, Shekhar S, Pitkonen M, Abo-Ramadan U, Tatlisumak T.

J Neuropathol Exp Neurol. 2010 Feb; 69(2): 188-95.

學生 : 黃怡靜

專討指導教授 : 鄭伯智老師 林宏榮老師

Introduction

http://marphilback.blogspot.com/2009/05/tia-mini-strokes-and-me.html

A transient ischemic attack (TIA) is a brief episode of neurologic dysfunction caused by focal brain or retinal ischemia, with clinical symptoms typically lasting less than one hour, and without evidence of acute infarction.

N Engl J Med 2002; 347: 1713-

16.

A transient ischemic attack (TIA) is a sudden focal neurologic deficit of cerebrovascular originlasting less than 24 hours and resolving without any residual symptoms or signs.

“MRI-negative”

Stroke 2008; 39: 3110-3115.

TIA Stroke (1/3)

Histopathology

Therapy

Animal model

Intraluminal suture MCAo (well-controlled reperfusion) 1. Ensures cerebral arterial occlusion and

reperfusion (LDF) 2. No permanent neurological deficit at 24 hrs3. No lesion on MRI (DWI, T2-WI) at 24 hrs

Materials and Methods

• Adult male NMRI mice, 22-44 g.• Suture occlusion model (MCAo). CBF - laser Doppler flowmetry (LDF). • MR imaging: 1) 4.7 T scanner, linear birdcage RF coil. 2) 7 coronal images, 1mm slice thickness. 3) DWI, T2-WI.• Neurological evaluation (6-point scale of sensorimotor skills):

Score

0 no deficit

1 failure to fully extend the left forepaw

2 circling to the left

3 decreased resistance to lateral push

4 no spontaneous walking with a depressed level of consciousness

5 dead

Journal of Cerebral Blood Flow & Metabolism (2004) 24, 668–

676.

Bregma

(LDF)

http://bmo.tnw.utwente.nl/vinay/principle_of_laser_doppler_flowm.htm0

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Doppler shift

• Histopathology: - hippocampus, caudoputamen, and frontoparietal cortex of

both hemispheres 1) H&E stain

2) TUNEL stain (* TUNEL positivity index)

Score

0 no change

1 scattered neuronal changes

2 selective neuronal necrosis (necrotic findings limited mainly

to specific neuron populations)

3 infarction (pan-necrosis characterized as the loss of affinity for

hematoxylin in all cell types)

Middle cerebral artery occlusion induced brain ischemic injury

Sham operation control group

(sham) (n=4)

Ischemia-Reperfusion group

(I/R) (n=6)

Physiology parameter Histology MRI assay

CBF, Tco, BW H&E stain TUNEL stain

20min 15min 10min 7.5min 5min 2.5min*12.5min

Reperfusion 24hrs

T2-WI DWI

Sensorimotor function

* 3 day (72 hrs) follow-up

1. Sham group

2. I/R group

0 min

Stabilized MCAoIschemia-reperfusion

Physiology parameters: CBF, Tco

24 hr * 72 hr

1. Sensorimotor function

2. MRI assay

3. Ischemic change

20 min, 15 min,

*12.5 min, 10 min, 7.5 min, 5 min, 2.5 min

• Statistical analysis: 1) Parametric data - means (± SD), unpaired t-test,

1-way analysis of variance.

2) Nonparametric data - medians, Mann-Whitney U test,

Kruskal-Wallis test.

3) Spearman correlation coefficient (r).

4) p < 0.05.

Results

Fig 1. Cortical cerebral blood flow (CBF) monitoring by laser Doppler flowmetry.

85% 83% 96%

Successful occlusion - decrease ≧ 75% CBF values from the baseline. Adequate reperfusion - recovery ≧ 50% CBF values from the baseline after suture withdrawal.

Fig 2. T2-weighted images at 24 hours after reperfusion.

optic chiasm level hippocampal level optic chiasm level hippocampal level

Neurological deficit Positive MRI

20 min 1 5 ‛

15 min 1 4 "

12.5 min 0 1 ª

10 min 0 0

7.5 min 0 0

5 min 0 0

2.5 min 0 0‛ : 1 small hippocampal infarct, 2 medium-sized subcortical infarcts, 2 large confluent infarcts" : 1 small cortical infarct, 3 medium-sized cortical and subcortical infarctsª : 1 small cortical infarct

3-day follow-up group (12.5min): no delayed change.

Fig 3. Postmortem assessments at 24 hours postreperfusion.

Ischemicchange

Duration of MCAo selective neuronal necrosis

pan-necrosis(r = 0.95)

lateral caudoputamen

Fig 4. Ischemic injury parameters. (A) Assessment with H&E.

** p = 0.003 versus control.

3-day follow-up group (12.5min): no delayed injury.

Selective neuronal necrosis (score 2) was a consistent feature in the ischemic frontoparietal cortex starting at 5 minutes of MCAo.

Fig 4. Ischemic injury parameters.

(B) Total TUNEL-positive cell counts of the right (ischemic) and left (contralateral) hemispheres. (C) Regional TUNEL counts.

** p = 0.003 versus control.

p < 0.001

3-day follow-up group (12.5min): no delayed injury.

Duration of MCAo

TUNEL positivity index

(r = 0.92)

Ann Neurol 1999;46:333-42.

Conclusion

MCAo of 10 minutes or less induced by the intraluminal suture

method is a reliable method of inducing a mouse model of TIA.

Animal species difference ?

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