UBC Theses and Dissertations
Analysis of WNT5A function in development and disease using the chicken model Hosseini-Farahabadi, Sara
Mouse and human genetic data suggests that Wnt5a is required for jaw development but the specific role in facial skeletogenesis and morphogenesis is unknown. The aim of this thesis is to study functions of WNT5A during mandibular development in chicken embryos. We initially determined that WNT5A is expressed in developing Meckel's cartilage but in mature cartilage expression was decreased to background. This pattern suggested that WNT5A is regulating chondrogenesis so to determine whether initiation, differentiation or maintenance of matrix was affected I used primary cultures of mandibular mesenchyme. I found that Wnt5a conditioned media allowed normal initiation and differentiation of cartilage but the matrix was subsequently lost. Collagen II and aggrecan, two matrix markers, were decreased in treated cultures. Degradation of matrix was due to the induction of metalloproteinases, MMP1, MMP13, and ADAMTS5 and was rescued by an MMP antagonist. The effects of Wnt5a on cartilage were mainly due to stimulation of the non-canonical JNK/PCP pathway as opposed to antagonism of the canonical Wnt pathway. To increase the clinical relevance of my work I studied the functional consequences of two human WNT5A mutations (C182R and C83S) causing human Robinow syndrome. Retroviruses containing mutant and wild-type versions of WNT5A caused shortening of beaks and limbs; however, the phenotypes were more frequent and severe with mutations. Mechanisms responsible for micrognathia were assessed. Decreased cell proliferation and impaired chondrocyte organization and intercalation were seen with all constructs. The effects of mutant proteins on the migration of mesenchymal cells were tested in organ cultures of the mandible. The C83S and to a lesser extent C182R forms of WNT5A inhibited the normal migration of dye-labeled mesenchymal cells. The lack of cell migration was similar to that reported in Wnt5a null mice and therefore suggested that the WNT5A mutations are causing a loss-of-function. We conclude that WNT5A is required during early chondrogenesis to block canonical signaling thereby allowing cartilage to form. In addition, WNT5A is required for cells to migrate within the mandible and perhaps to form the elongated shape of the lower jaw. Finally WNT5A in conditions of excess has detrimental effects on cartilage integrity.
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