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order to initiate folding.
51
Three additional families were
identified with mutations in
PPIB
. The patients from the
first two families had a diagnosis consistent with OI II.
The infants died shortly after birth and collagen isolated
from fibroblasts had delayed electrophoretic mobility,
indicating collagen overmodification.52
52
The patient from
the third family was milder in presentation, with a diag-
nosis of OI III
/
IV. The proband had poorly modeled long
bones that eventually required intramedullary rodding.
52
Collagen isolated from mesenchymal stem cells (MSCs)
had a delay in electrophoretic mobility.
52
The variation
observed in patients with documented mutations in
PPIB
suggests that there is a range of alleles, from hypo-
morphic alleles that result in mild phenotypes to com-
plete loss-of-function alleles that result in a more severe
phenotype.
glomeruli showing abnormal PAS staining, mesangial
cell hyperplasia and collagen fibril deposition.
16
The skin
showed hyper-laxity and was thin and disorganized at
the histological examination.
16
This work suggests that
the pulmonary deficits observed in patients with docu-
mented mutations in
CRTAP
could be due to primary
lung defects in addition to the secondary defects caused
by scoliosis and rib fractures.
The
Lepre1
KO mice were similar in presentation to the
Crtap
KO mice. The null mice were significantly smaller,
developed kyphoscoliosis, had low bone density, shorten-
ing of the long bone segments with rhizomelia present,
bone defects as shown by X-ray and uCT, and a decrease
in overall body fat.
53
The analysis of tendon collagen
showed a large increase in the number of small diameter
collagen fibrils with heterogeneity in overall fibril con-
tour.
53
As shown by the absence of staining in the pari-
etal bone of the skull, the
Lepre1
KO mice have a delay in
ossification.53
53
Analysis of the skin showed less densely
packed dermis with fewer collagen fibrils, with a con-
comitant decrease in skin thickness.
53
Chrondrocytes in
the hypertrophic zone were less organized, which could
account for the shortening of the long bone segments.
53
Biochemically, the
Lepre1
KO mice had collagen overmod-
ification, as observed by delayed gel electrophoresis.
53
There was a complete absence of prolyl 3-hydroxylation
but a slight increase in the percentage of residues that
were 4-hydroxylated.
53
In addition, the increase in lysyl
hydroxylation and glucose galactosyl-hydroxylysine sug-
gests that there is a slight increase in the melting tempera-
ture of type I collagen.
53
Along with the overmodification,
the secretion rate was delayed in the
Lepre1
KO mice.
53
Ppib
mutant mice displayed many similar abnormali-
ties when compared with the
Lepre1
and
Crtap
KO mice,
including decreased body size and weight, pronounced
kyphosis, reduced BMD and collagen overmodification
together with a complete absence of prolyl 3-hydroxyl-
ation at Proline 986 of type I and II collagen.
12
However,
unlike the
Crtap
and
Lepre1
mice, the
Ppib
KO mice did
not present with rhizomelia, suggesting an absence of
chondrodysplasia.
12
Skin findings were similar to that
described in the
Crtap
KO mice, with a decrease in pic-
rosirius red staining suggesting reduced presence of col-
lagen.
12
As a result, the skin is more elastic and requires
less force to break.
12
MOUSE PHENOTYPES WITH MUTATIONS
IN
CR
TAP
,
LEPRE1
AND
PPIB
G
ENES
The generation of
Crtap
,
Lepre1
and
Ppib
KO mice has
been described and the study of their phenotype has
helped to understand some of the underlying molecular
defects in recessive OI.
In the initial paper by Morello et al.,
Crtap
-null mice
were found to have chondro-osseus dysplasia with rhi-
zomelia, kyphosis and severe osteopenia (
Figure 14.2
).
11
In addition, analysis for the prolyl 3-hydroxylation site
of fibrillar collagen demonstrated a complete absence
of Proline 986 hydroxylation. With the observation of
postnatal growth delay and the shortening of the first
segment of the limbs, the growth plate was analyzed
and found to have slightly disorganized growth col-
umns of proliferating chondrocytes with focal areas of
cell dropout.
11
Analysis of the skeleton using radiogra-
phy and histomorphometry showed severe osteopenia
in
Crtap
KO mice.
11
Osteoblasts
in vivo
produced osteoid
at a slower rate and the osteoid mineralized quickly.
11
Ultrastructural analysis of the mutant collagen fibrils
using electron microscopy demonstrated a thicker diam-
eter when compared to WT (
Figure 14.2
).
11
The collagen
was also overmodified based on electrophoretic mobil-
ity.
11
The initial description found that
Crtap
-null mice
had primary defects of the bone and cartilage but
Crtap
is also expressed in the lung parenchyma, in the vis-
ceral and parietal epithelial cells of the glomerulus, in
the peritubular capillaries of the kidney and in dermal
fibroblasts.11
11
Crtap−/−
mice were found to have primary
defects of the lung, skin and kidney which were consis-
tent with a generalized connective tissue abnormality.
Work by Baldridge et al. demonstrated that the lungs
had a diffuse increase in alveolar airway space accom-
panied by thinning of the alveolar walls.
16
In the kid-
ney, there was focal glomerular sclerosis with several
PUTATIVE ROLE OF COLLAGEN
PROLYL 3-HYDROXYLATION AND
PATHOGENETIC MECHANISMS OF
M
UTATIONS IN THESE GEN
ES
Collagen prolyl 3-hydroxylation is an ancient modi-
fication and 3Hyp residues are present in collagens
from very primitive organisms such as Porifera that
