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Fig. 4. Impaired calvarial osteogenesis in En1 mutant mice.
(A) Whole-mount in situ hybridization analysis of calvarial
Opn expression, showing that En1-/- embryos
exhibit a delay in the commencement of osteogenesis (E14.5), followed by
deficient osteoblast differentiation (arrows, E16.5-E18.5). (B) Section
ISH analysis demonstrating the absence of Osx expression in the
mesenchyme of En1-/- calvariae at E13.5. At E16.5,
Osx is expressed by wild-type osteoblasts of the frontal bone, but is
reduced in En1-/- osteoblasts (arrowheads). (C)
Analysis of osteopontin (Opn) and osteocalcin (Ocn) on
sagittal sections of wild-type and En1-/- calvariae at P1.
Although Opn normalizes to wild-type levels in
En1-/- calvariae postnatally, it is aberrantly expressed
by terminally differentiated osteocytes of the mutant frontal bones (inset,
arrowheads). By comparison, normal Ocn expression in ectoperiosteal
osteoblasts is nearly abolished in En1-/- calvariae.
(D) Northern blot analysis of Opn, Bsp, Ocn and Osx
expression by calvarial osteoblasts over a 21-day culture period. p,
parietal bone; f, frontal bone. Scale bars: 1.5 mm in A; 0.1 mm in B;
0.2 mm in C.
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