This project will investigate the hypothesis that the growth factor TRANCE directly regulates both cartilage maturation and bone growth in the appendicular and craniofacial skeleton using osteopetrotic rats and mice as model systems. TRANCE is required for bone resorption, but recent reports and the preliminary data presented herein, make it probable that it also regulates bone growth and chondrocyte maturation in the growth plates of (endochondral) long bones, suture development and growth in the (intramembranous) craniofacial skeleton, an collagen gene switching in both sites. Because of the unique growth plate chondrodystrophy present in the toothless osteopetrotic rat and the TRANCE knock-out mouse, and the presence of the TRANCE receptor, RANK, in the region of the growth plate most affected by the TRANCE knockout mutation, the following are hypothesized: TRANCE directly regulates chondrocyte progression through the proliferative and hypertrophic stages, that the toothless rat is in fact a naturally-occurring TRANCE loss-of-function mutation, and that TRANCE signaling is independent of the Ihh/PTHrP cascade of growth regulation.
Three specific aims are presented to test these hypotheses.
In Specific Aim 1, normal and toothless rat TRANCE will be cloned and sequenced, the protein level measured, and receptor-binding activity tested to rule in or out a mutation affecting TRANCE function in the toothless rat.
In Specific Aim 2, TRANCE binding to chondrocytes in culture and the resultant signal transduction effects (phosphorylation/activity of IkB, PKB, c-src) will be tested, along with its ability to affect chondrocyte growth and apoptosis in metatarsal culture. Expression of known growth plate regulators and collagens in long bones and collagen types in the skull will be measured to assess the extent to which the mutations disrupt them.
In Specific Aim 3, in vivo studies will determine cell proliferation and cell death in the growth plates of both mutations to understand the cellular basis for the tissue distortions. TRANCE may find uses in cartilage culture systems. Epiphyseal chondrodysplasias or other defects in bone growth (limb or facial deformities) and metabolism may be impacted if new potential points for pharmacological or gene therapy interventions are discovered.
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