Forced chondrocyte expression of sonic hedgehog impairs joint formation affecting proliferation and apoptosis.

Proliferation and apoptosis are two fundamental processes that occur during limb development, and in particular in joint formation. To study the role of hedgehog proteins in limbs, we have misexpressed Sonic Hedgehog specifically in chondrocytes. We found that the appendicular skeleton was severely misshapen while pelvic and shoulder girdles developed normally. In particular, we detected fusion of the elbow/knee joint, no definite carpal/tarsal, metacarpal/metatarsal bones and absence of distinct phalanges, fused in a continuous cartilaginous rod. Molecular markers of joints, such as Gdf5 and sFrp2 were absent at presumptive joint sites and Tenascin C, a molecule associated with joint formation and expressed in permanent cartilage, was expressed in a wider region in transgenic animals as compared to the wild type. The ratio of proliferating to non-proliferating chondrocytes was about two times higher in transgenic developing cartilage as compared to the wild type. Accordingly, the proapoptotic gene Bax was barely detectable in the growth plate of transgenic mice and Tunel assay showed the absence of apoptosis in presumptive joints at E15.5. Taken together, these results suggest that misexpression of Sonic Hedgehog causes apoptosis and proliferation defects leading to the lack of joint cavity and fusion of selected limb skeletal elements.