Pompe's disease - RePub - Erasmus Universiteit Rotterdam

Chapter 1
Fig. 3. Targeting vectors used for creating mouse modesl for Pompe’s disease.
20
TK
RI
3
H
TK
4
B B
RV
5 6 7 8 9 10 11
5 6 7 8 9 10 11
E
12 13
B
H RI
14 15 16
12 13 13 14
3 4 5 6 7 8 9 10 11 12 13 14 15 16
NEO
TK
9 10 11 12 13 15 16
B=BamHI; E=EagI; RV=EcoRV; H=HindIII; RI=EcoRI; =LoxP
NEO
NEO
RI
Mouse Gaa locus
exon 3-16
targeting vector
13 neo
/13 neo
targeting vector
6 neo
/6 neo
and D6/D6
targeting vector
D14 neo
/D14 neo
The (13 neo /13 neo ) mouse
Disruption of the GAA gene in exon 13 (13 neo /13 neo ) resulted in the absence of acid αglucosidase
RNA, and the mice were fully defi cient in acid α-glucosidase activity in all
investigated tissues. Lysosomal glycogen storage was seen as early as eight days after birth
in skeletal muscle and heart. Accumulation of lysosomal glycogen was also found in Schwanncells
of the peripheral nervous system, in motor neurons and in the brain, as in the infantile
form of Pompe’s disease (74, 80).
The pathological evaluation of this mouse model gave more insight in specifi c clinical features
of the disease in humans. For example, the extensive glycogen accumulation in smooth
muscle of the upper digestive tract and masseter muscle could explain the feeding diffi culties
reported in humans. Pathological changes in the smooth muscle cells of the blood vessels
could result in weakening of the blood vessel wall and the formation of an aneurysm with risk
of rupture as reported in the literature about adult Pompe’s disease (85-87, 122).
However, these (13 neo /13 neo ) mice do not show overt clinical symptoms until adulthood despite
their total defi ciency of acid α-glucosidase. Although, at 12 weeks of age the knockout mice
are less active than their wild-type littermates when tested in a running wheel (123).
However, the difference is not statistically signifi cant due to intra group variation. The mice
were backcrossed for ten generations into the FVB and C57Bl/6 background in an attempt to
decrease the variation. It is our impression that the mice with a C57Bl/6 background show
symptoms earlier than mice with the FVB background, but statistically reliablke information
is still missing (98).
The (6 neo /6 neo ), (∆6/∆6) and (∆14 neo /∆14 neo ) mice
Disruption of exon 6 (6 neo /6 neo ) was chosen by Raben et al. to create a mouse model of
infantile Pompe’s disease. But, as it was realised that the phenotype could be ‘embryonic
lethal’ additional approaches were taken to create a mouse model with milder symptoms.
Raben et al. predicted that the complete deletion of exon 6 (∆6/∆6) was potentially associated
with a relatively milder phenotype based on the occurrence of a similar mutation (splicing
out of exon 6 and inclusion of 7 amino acids) in a patient with juvenile Pompe’s disease, as
published by Adams et al. (124). Homozygous ∆6/∆6 mice would thus have a better prognosis
than the 6 neo /6 neo mice. The two LoxP sites fl anking exon 6 in the targeting construct were
used to achieve this goal. Exon 6 was deleted by crossing the 6 neo /6 neo mice with Cre gene
expressing transgenic mice of strain FVB. Notably, the 6 neo /6 neo mouse model was created in
the C57Bl/6 background so that the ∆6/∆6 KO mice had a mixed background of 129, C57Bl/6
and FVB.