Cleft palate and cleft lip are among the most common congenital birth defects in humans occurring once in every 500-700 live births. Orofacial clefts are disfiguring, can affect respiration, speech, and feeding, and may require surgeries to repair. The two main types of orofacial clefts are cleft lip with or without cleft palate (CL/P) and cleft palate (CP) alone. Development of the secondary palate is a complex event and abnormalities during secondary palate development can lead to cleft palate. Many proteins, including transcription factors, growth factors and their receptors, and tissue re-modelling enzymes are involved in the coordinated process of driving secondary palate development. Adhesion of palatal shelves could be the result of many cell adhesion molecules expressed on the cell surfaces, and loss of their expression could subsequently lead to clefting. The secreted growth factor transforming growth factor β3 (TGFβ3) is essential for successful formation of the secondary palate; it plays a crucial role in regulating epithelial-mesenchymal interactions leading to appropriate growth and fusion of palatal shelves. Previous studies have shown that TGFβ3-deficient mice have cleft secondary palate due defects in fusion of the palatal shelves. The gene encoding interferon regulatory factor 6 (IRF6) has also been found to be down-regulated in the medial edge epithelia of Tgfb3-/-.In the current research, I have investigated the expression patterns of a series of cell adhesion molecules in wild-type and Tgfb3-/- mice to determine whether they were down-regulated in the medial edge epithelia of the mutant mice. In complementary studies, I have investigated whether the cleft palate phenotype observed in Tgfb3-/- mice could be rescued by re-expression of Irf6. Using an in vitro palatal organ culture system, I showed that addition of IRF6 adenovirus restored palatal fusion in Tgfb3-/- mice, midline epithelial seam degeneration being observed in treated cultures. These data were complemented using an in vivo strategy in which I generated a transgenic mouse in which Irf6 was expressed under the control of the Keratin 14 promoter to up-regulate Irf6 expression in the palatal epithelia. The results of these experiments indicated that that the cleft palate phenotype observed in Tgfb3-/- embryos could be rescued by the K14Irf6Flag transgene. Together, these results indicate that IRF6 lies downstream of TGFβ3 in the molecular cascade driving fusion of the palatal shelves. In our preliminary microarray analysis of Tgfb3 -/-mice treated with exogenous TGFB3 to determine whether any potential candidate genes involved in craniofacial development were upregulated or down regulated, no candidate genes were found to have any profound changes in expression between wild type and Tgfb3-/-. However genes involved in extracellular matrix proteins, growth factors, and transcription factors were highlighted in the data obtained.
|Date of Award||1 Aug 2011|
- The University of Manchester
|Supervisor||Michael Dixon (Supervisor)|