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Table 2
Scale used for clinical evaluation
0
Healthy
1
Flaccid tail. The mouse is held by the base of the tail. If it is healthy the tail is
always straight, otherwise it hangs down
1.5
If the mouse stumbles at least twice on the grid cage (unsteady gait, or ataxia)
2
Ataxia and/or hind-limbs paresis, or slow righting reflex (see Note 6)
3
Paralysis of hind limb and/or paresis of forelimbs
4
Paraparesis of fore limb
5
Moribund or death
3.4
Induction of EAE
1. Hold the mice from the neck and the tail.
2. Inject the mice subcutaneously (s.c.), slowly, with 100
μ
l of
emulsion in each flank and 100
μ
l at the tail base (see
Note 3).
3. After immunization (4-5 h), inject the mice intravenously
(i.v., see Note 4) with 500 ng of pertussis toxin resuspended
in 100
l of PBS. After 48 h, inject the mice again with pertus-
sin toxin, i.v., 500 ng/100
μ
l each.
4. Administer EPO or CEPO i.p., 200
μ
l/mouse in saline solu-
tion, three times per week, starting at different times after
immunization depending on the schedule (preventive, thera-
peutic or late therapeutic, see above).
μ
3.5 Clinical
Evaluation
The onset of the disease is usually 10-18 days after immunization.
Mice need to be checked daily for body weight and clinical score
(see Note 5). They will usually lose 1-2 g of weight in the days
before the onset. For clinical evaluation a scale with five steps is
used (Table 2 ).
3.6 Ef fi cacy
of Preventive or
Therapeutic EPO or
CEPO in Chronic-
Progressive EAE
C57BL/6 mice immunized with MOG suffer chronic progressive
EAE with onset around day 10. EPO inhibits the clinical score of
the disease when administered according to a preventive schedule,
and also according to a therapeutic schedule, starting at disease
onset (Fig. 1 ( 21 )). Also with a late therapeutic schedule of treat-
ment some improvement in the status of the animals was obtained,
as detected by comparing the curves reporting the body weight,
although no significant difference was observed in the clinical
score; interestingly, a late therapeutic schedule of CEPO treatment
significantly improved also the clinical score ( 21 ).
 
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