ANALYSIS OF PRIMARY PALATE FORMATION

这项研究计划的长期目标是确定 发展机制负责形成，扩大和 面部原基的结合。 目前的具体目标代表了 持续分析上皮间充质细胞 细胞通讯和细胞增殖的相互作用和模式。 先前的研究表明，上皮细胞的存在对人类的健康有着深远的影响。 对间充质细胞活力和增殖的影响。 将荧光示踪剂显微注射到外植体的细胞中 重组的组织揭示了缝隙连接通讯的模式， 下层间充质。 其他研究，采用免疫荧光 间隙连接蛋白（27 kDa和43 kDa）抗体定位 在特定的地点表现出独特的分布模式， 作为鼻基板（27 kDa），和相关的分布模式 细胞增殖（43 kDa）。 根据这些发现， 以下将进行。具体目标#1：通信功能不全的细胞 将在器官培养系统中制备和使用群体， 测试假设：a）上皮和 间充质通过间隙连接传递B）间隙连接的阻断 上皮-间充质界面的通讯将影响 邻近的有交通能力的间充质的增殖。 具体目标#2：免疫细胞化学，使用针对43 kDa的抗体 间隙连接蛋白，将用于测试假设a）， 43 kDa蛋白质存在于面部原基中，并分布在面部原基中。 区域特异性方式B）其分布与先前相关 确定面部原基中细胞增殖的速率 分布是一致的（或不一致的）， 27 kDa间隙连接蛋白。 具体目标#3：假设 27 kDa基因的表达受发育调控， a）确定所述基因的表达是否 B）确定是否表达 该基因在鼻基板形成过程中受到时间调节c） 确定所述基因的表达是否与所述疾病相关。 鼻基板发育过程中的特殊形态发生事件。 上皮-间质分离和重组，器官培养， 显微注射，电穿孔，流式细胞术，免疫细胞化学， 将使用原位杂交和放射自显影。

The long range goal of this research program is to identify the developmental mechanisms responsible for the formation, enlargement and union of the facial primordia. The present specific aims represent continuing analysis of the relationship between epithelial-mesenchymal interaction and patterns of cell communication and cell proliferation. Prior studies indicated that the presence of epithelium had a profound effect on the viability and proliferation of subjacent mesenchyme. Microinjection of fluorescent tracers into cells of explants of recombined tissues revealed patterns of gap junctional communication in subjacent mesenchyme. Other studies, employing immunofluorescent localization with antibodies to gap junction proteins (27kDa and 43kDa) demonstrated unique patterns of distribution in specific locations, such as the nasal placode (27kDa), and patterns of distribution associated with cell proliferation (43kDa). In light of these findings, the following will be done. Specific Aim #1: Communication-incompetent cell populations will be prepared and employed in organ culture systems to test the hypotheses: a) developmental signals between epithelium and mesenchyme are transmitted by gap junctions b) blockage of gap junctional communication at the epithelial-mesenchymal interface will affect proliferation of the adjacent communication-competent mesenchyme. Specific Aim #2: Immunocytochemistry, employing antibodies to the 43kDa gap junction protein, will be used to test the hypotheses a) that the 43kDa protein is present in the facial primordia and is distributed in a region-specific manner b) its distribution is correlated with previously determined rates of cell proliferation in the facial primordia c) the distribution is concordant (or discordant) with the distribution of the 27kDa gap junction protein. Specific Aim #3: The hypothesis that expression of the gene for the 27kDa is developmentally regulated will be tested by a) determining whether expression of the gene is region-specific in the nasal placode b) determining whether expression of the gene is temporally regulated during nasal placode formation c) determining whether expression of the gene is associated with a particular morphogenetic event during nasal placode development. Epithelial-mesenchymal separation and recombination, organ culture, microinjection, electroporation, flow cytometry, immunocytochemistry, in situ hybridization, and autoradiography will be utilized.