Molecular control of secondary palate development

Compared with the embryonic development of other organs, development of the secondary palate is seemingly simple. However, each step of palatogenesis, from initiation until completion, is subject to a tight molecular control that is governed by epithelial-mesenchymal interactions. The importance of a rigorous molecular regulation of palatogenesis is reflected when loss of function of a single protein generates cleft palate, a frequent malformation with a complex etiology. Genetic studies in humans and targeted mutations in mice have identified numerous factors that play key roles during palatogenesis. This review highlights the current understanding of the molecular and cellular mechanisms involved in normal and abnormal palate development with special respect to recent advances derived from studies of mouse models.     

Muscle development in vitro following X irradiation

Myogenic cells obtained from 12-day-old embryonic chicken hind limb and breast muscle were exposed to 5000 rads of X irradiation. Although 10% of the initial cell dissociates were killed by irradiation, the remaining cells were comparable to controls in plating efficiency and light microscopic morphology. Moreover, there was no increase or loss of cells for at least 72 hr in vitro when plated at a density of 2 × 10⁶ cells/60-mm plate. It was found that muscle cell fusion after irradiation proceeded at the same rate and to the same relative extent as in control cultures. Myotubes developed normally; cross-striations were prominent by 5 to 7 days of culture and the cells maintained a well-differentiated state for periods of at least 3 weeks in vitro. In control cultures continuously labeled with 1 μCi/ml of [³H]TdR, 75% of the nuclei within myotubes were heavily labeled by 118 hr; less than 15% of the nuclei within syncytia of irradiated cultures were labeled. Quantitative microphotometry of Feulgen-stained cultures demonstrated that all nuclei within control and irradiated myotubes contained the 2C complement of DNA. Similar experiments conducted with cells released from limbs and breasts of 10-day-old embryos revealed lower absolute levels of cytoplasmic fusion in both control and irradiated samples, however, there was slightly more cell death after exposure to X rays in 10-day-old than 12-day-old material. Nevertheless, considerable cell fusion occurred in irradiated limb and breast cell cultures, consistent with the conclusion that the commitment to myogenesis of prefusion myoblasts is extremely stable even in the face of massive ionizing radiation and that neither cell division nor replication of DNA is an obligatory prerequisite for the in vitro fusion and subsequent differentiation of skeletal muscle obtained from 10- and 12-day-old chick embryos.     

In vitro development of the hamster and chick secondary palate

A series of experiments were undertaken to compare the in vitro behaviour of the medial edge epithelium (MEE) of hamster, in which palatal shelves normally fuse, and chick, in which they do not fuse. Homotypic pairs of hamster and chick embryo palatal processes, single palatal processes, and heterotypic palatal shelves of both animals were grown in vitro. The results indicated that contact between palatal shelves may not be crucial for MEE differentiation in mammals. The ability to acquire pre-fusion characteristics may be present in mammalian palatal tissue from their early development and may be expressed by cessation of DNA synthesis in the MEE, elevation of cAMP, and MEE cell death. Isolated chick palatal shelf cultured under identical conditions did not express these mammalian pre-fusion characteristics. When MEE of hamster and chick palatal shelves were placed in contact with one another, the intervening epithelia underwent cytolysis. This could be due to either the destruction of chick MEE by lysosomal enzymes liberated from adjacent degenerating hamster MEE cells, or by induction of cell death in chick MEE by hamster mesenchyme. Heterotypic palatal tissue combinations also suggest that release of lysosomal enzymes in the hamster MEE, which leads to its dissolution, may be the terminal event in epithelial differentiation prior to the establishment of mesenchymal continuity. It is suggested that an inverse relationship exists between DNA synthesis and cAMP levels during palatogenesis: when palate closes (as in mammals) the MEE is eliminated by increasing cAMP levels, whereas when palate remains open (as in birds) low level of cAMP preserve the integrity of MEE by supporting DNA synthesis.     

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.     

Effect of continuous irradiation on the thrombopoietic system in mice

In this paper the changes of the megakaryocyte number of the spleen and those of thrombocyte number of the peripheral blood were studied during continuous irradiation with a dose rate of 9.57 to 957 mGy/day. Beginning with a dose rate of 95.7 mGy/day the thrombocyte number of the blood and the megakaryocyte number of the spleen of irradiated animals decreased significantly. Whereas the thrombocyte number remained permanently decreased, the cell number of the megakaryocyte type is increased temporarily to a clearly higher level as compared with the controls on the 100th day of irradiation approximately. This is especially true for the middle dose rate. During this time of irradiation nucleolar hyperchromatosis as well as pycnosis were observed in many megakaryocytes.     

Identification and profiles of microRNAs in different development stages of miniature pig secondary palate

Cleft palate is one of the most frequent craniofacial malformation birth defects. Miniature pigs (Sus scrofa) are a valuable alternative large animal model to explore human palate development. Presently, the microRNA (miRNA) expression profiles in miniature pigs during palatogenesis from embryonic day (E) 30 to 50 were identified. A total of 2044 known miRNAs and 192 novel miRNAs were identified. The functional characteristics of their potential target genes were identified using Gene Ontology function and Kyoto Encyclopedia of Genes and Genomes pathway analysis. MiRNAs displayed diverse expression levels among the different stages. Using Short Time-series Expression Miner software to investigate the expression patterns of miRNAs from E30–50, all miRNAs were clustered into 20 profiles. The profiles showing miRNAs expression decreased (profile 0)/increased (profile 19) from E30–50 were the main patterns during palatogenesis. Hub genes of four significant modules were identified by weighted correlation network analysis, including ssc-miR-98, ssc-miR-27a_R + 1, and ssc-miR-150, etc. which might be novel potential targets for regulating palate development. The data are expected to improve the understanding of palate development and the etiology of cleft palate in further studies.     

Vertical development of the secondary palate in hamster embryos following exposure to 6-mercaptopurine

Cellular aspects of vertical development of the secondary palate were examined in control and 6-mercaptopurine (6MP)-treated hamster embryos. Cross-sectional area of the palatal shelf was measured and the numbers of both epithelial and mesenchymal cells counted. Also, in 6MP-treated palates the damaged mesenchymal cells, characterized by the presence of dense bodies, were counted. DNA synthesis in both control and treated fetuses was measured by 3H-thymidine incorporation. The results indicated that both the shelf area and cell numbers increased with age in control and 6MP-treated palates. However, in controls the mesenchymal cell density and DNA synthesis showed two peaks that were absent following 6MP treatment. Unlike controls, in treated embryos the damage to mesenchymal cells became increasingly pronounced between days 10:00 and 10:12 but subsided by day 11:00 of gestation. It is suggested that a major force in the development of the initial primordia and early vertical development of the palatal shelf may be provided by a spurt of DNA synthesis in the mesenchymal cells resulting in their increased number. After 6MP treatment, depression of DNA synthesis and consequent reduction in the mesenchymal cell number and density followed by cell damage lead to retardation in the vertical development of the palatal shelves.     

The genesis of Thy-1-lymphomas in NFS mice exposed to X irradiation

The kinetics of the appearance of potentially leukemic cells (PoLCs) for radiation-induced lymphoma in NFS mice was investigated by the opposite sex (male----female) transplantation assay. The origin of the cells of the lymphomas that developed in the host was decided by sex chromosome markers. The bone marrow and the spleen cells collected from mice 30 days after fractionated irradiation (1.7 Gy X 4) gave rise, upon transfer to 4-Gy-irradiated hosts, to tumors of either donor or host origin. Most tumors of donor origin were thymine-1-negative (Thy-1-) and surface immunoglobulin negative and classified as nonthymic lymphoma, while the tumors of host origin were mainly Thy-1-positive thymic lymphoma. In contrast, neither the bone marrow nor the thymus contained any PoLCs for thymic lymphoma 30 days after split-dose irradiation. These results indicate that PoLCs for Thy-1-lymphoma were induced in the bone marrow and spleens of NFS mice by the split-dose regimen which developed exclusively T-cell lymphomas in the absence of cell grafting.