Fig. 8.

Defective expression of angiogenic factors in Ctgf mutant growth plates. Growth plates of neonatal radii are shown in all panels. (A) PECAM immunostaining demonstrates the existence of blood vessels in the intertrabecular spaces of the metaphysis, but the fine network of capillaries seen at the ossification zone (broken line) in wild-type animals is not as extensive in Ctgf mutants. Scale bar: 100 μm. (B) MMP9 immunostaining is reduced in neonatal mutants. (C) Mmp9 RNA levels are reduced in neonatal mutants. The triangular bars above the photograph of the gel represent increasing numbers of amplification cycles (see Materials and Methods). Data are from three pairs on Ctgf mutants and wild-type littermates. Expression of Mmp9 was normalized to Gapdh in each reaction. *P<0.05. (D) TRAP-positive cells (arrows) are present in normal levels in the bone marrow of Ctgf mutants, but in reduced levels at the cartilage-bone junction, indicating a defect in recruitment of osteoclasts/chondroclasts to the hypertrophic region. MMP13 protein is present at diminished levels in hypertrophic chondrocytes in Ctgf mutants. H, hypertrophic zone; M, metaphysis.