Palatogenesis: engineering, pathways and pathologies

Cleft palate represents the second most common birth defect and carries substantial physiologic and social challenges for affected patients, as they often require multiple surgical interventions during their lifetime. A number of genes have been identified to be associated with the cleft palate phenotype, but etiology in the majority of cases remains elusive. In order to better understand cleft palate and both surgical and potential tissue engineering approaches for repair, we have performed an in-depth literature review into cleft palate development in humans and mice, as well as into molecular pathways underlying these pathologic developments. We summarize the multitude of pathways underlying cleft palate development, with the transforming growth factor beta superfamily being the most commonly studied. Furthermore, while the majority of cleft palate studies are performed using a mouse model, studies focusing on tissue engineering have also focused heavily on mouse models. A paucity of human randomized controlled studies exists for cleft palate repair, and so far, tissue engineering approaches are limited. In this review, we discuss the development of the palate, explain the basic science behind normal and pathologic palate development in humans as well as mouse models and elaborate on how these studies may lead to future advances in palatal tissue engineering and cleft palate treatments.     

Palatogenesis: morphogenetic and molecular mechanisms of secondary palate development

Mammalian palatogenesis is a highly regulated morphogenetic process during which the embryonic primary and secondary palatal shelves develop as outgrowths from the medial nasal and maxillary prominences, respectively, remodel and fuse to form the intact roof of the oral cavity. The complexity of control of palatogenesis is reflected by the common occurrence of cleft palate in humans. Although the embryology of the palate has long been studied, the past decade has brought substantial new knowledge of the genetic control of secondary palate development. Here, we review major advances in the understanding of the morphogenetic and molecular mechanisms controlling palatal shelf growth, elevation, adhesion and fusion, and palatal bone formation.     

Temporal and Spatial Expression of Hoxa-2 During Murine Palatogenesis

1. Mice homozygous for a targeted mutation of the Hoxa-2 gene are born with a bilateral cleft of the secondary palate associated with multiple head and cranial anomalies and these animals die within 24 hr of birth (Gendron-Maguire et al., 1993; Rijli et al., 1993; Mallo and Gridley, 1996). We have determined the spatial and temporal expression of the Hoxa-2 homeobox protein in the developing mouse palate at embryonic stages E12, E13, E13.5, E14, E14.5, and E15.2. Hoxa-2 is expressed in the mesenchyme and epithelial cells of the palate at E12, but is progressively restricted to the tips of the growing palatal shelves at E13.3. By the E13.5 stage of development, Hoxa-2 protein was found to be expressed throughout the palatal shelf. These observations correlate with palatal shelf orientation and Hoxa-2 protein may play a direct or indirect role in guiding the palatal shelves vertically along side the tongue, starting with the tips of the palatal shelves at E13, followed by the entire palatal shelf at E13.5.4. As development progresses to E14, the stage at which shelf elevation occurs, Hoxa-2 protein is downregulated in the palatal mesenchyme but remains in the medial edge epithelium. Expression of Hoxa-2 continues in the medial edge epithelium until the fusion of opposing palatal shelves.5. By the E15 stage of development, Hoxa-2 is downregulated in the palate and expression is localized in the nasal and oral epithelia.6. In an animal model of phenytoin-induced cleft palate, we report that Hoxa-2 mRNA and protein expression were significantly decreased, implicating a possible functional role of the Hoxa-2 gene in the development of phenytoin-induced cleft palate.7. A recent report by Barrow and Capecchi (1999), has illustrated the importance of tongue posture during palatal shelf closure in Hoxa-2 mutant mice. This along with our new findings of the expression of the Hoxa-2 protein during palatogenesis has shed some light on the putative role of this gene in palate development.     

Palatogenesis in hamster. II. Ultrastructural observations on the closure of palate

Formation of secondary palate in hamster was studied with electron microscopy. Prior to assuming horizontal position, the palatal shelves were covered by a two to three cell layer thick epithelium which was separated from the underlying mesenchyme by an intact basal lamina. Epithelial cells were attached to each other by desmosomes. Early hemidesmosomes could be identified as thickenings of the cytoplasmic membrane opposing the basal lamina. Epithelial cells, like other embryonic cells, contained only few organelles but were rich in polyribosomes. As the horizontal shelves approached each other towards the midline, lysosomes and tonofilaments appeared in the superficial and basal cells of the epithelia. Superficial cells showed degeneration and eventual lysis. Fusion of the opposing epithelia occurred between the deeper cells by means of newly formed desmosomes. The epithelial seam resulting from fusion of the epithelia was limited on each side by a continuous basal lamina. Its subsequent thining and eventual fragmentation resulted from the loss of cells by autophagy. There was no evidence of mesenchymal invasion of the epithelial seam. Mesenchymal macrophages appeared in the later stage of palatogenesis and were responsible for phagocytosis of cellular debris. Formation of the soft palate was basically similar to that of the secondary hard palate and occurred by fusion of the opposing shelves. Similarly, anterior closure of the palate occurred by fusion of the lower end of the nasal septum to the primary and secondary palates. Hyperplasia of the opposing epithelia, prior to their fusion, was often seen. It is suggested that formation of the palate occurs in predictable and coordinated fashion and that timely appearance of lysosomes causing lysis of intervening epithelia is of great significance in normal palatogenesis.