Role of Gap Junctions in the Regulation of Cell Migration

该项目采用了尖端技术，用于识别间隙连接和测量细胞迁移。 将研究几种类型的肾上腺细胞，其在间隙连接表达中表现出广泛的变化，这是因为细胞的初始表型或通过细胞内第二信使的实验刺激/抑制。 在这些研究中，将深入探讨改变的表面间隙连接对细胞迁移调节的影响。间隙连接在细胞迁移中起关键作用的假设将得到解决。 将在具有或不具有间隙连接的肾上腺细胞培养物中研究迁移（特定目的I）。 将用特异性cAMP拮抗剂、激动剂或稀释剂处理细胞，并评估其迁移特征。 将使用以下技术测定迁移：（1）用膜可渗透染料预标记的细胞的核位移或细胞前缘位移的时间推移成像;（2）计数从伤口边缘移动到裸露区域的细胞数量;和（3）计数在硅模板区室中迁移的细胞的数量，在微流体室中，或者通过博伊登室过滤器。 一个组合的延时视频显微镜和免疫细胞化学技术将被用来识别和阐明的作用，间隙连接动力学在细胞migration.Cellstructural的贡献/更广泛的ImpactThe实验室将继续作为一个资源的培训本科生和研究生，博士后研究员，访问教授从少数民族机构，并指导高中学生和（影子计划）教师。 过去的NSF研究资金提供了接触和培训的学生，博士后研究员，和教师在细胞生物学，内分泌学和信号转导领域。通过在国家和国际一级举办的研讨会和讲习班，已经并将继续向来自若干学科的科学界成员提供信息。高中生将继续开发研究项目，并出席城市和国家科学博览会。最近，两名本科生在全国会议上获得了第一名，其他人将被鼓励跟随他们的道路。

This project incorporates cutting-edge techniques for the identification of gap junctions and measurements of cell migration. Several types of adrenal cells will be studied that exhibit a wide variation in gap junction expression, either because of the initial phenotype of the cell or through experimental stimulation/inhibition of second messengers within the cell. In these studies, the influence of altered surface gap junctions on the regulation of cell migration will be explored in depth. The hypothesis that gap junctions play a pivotal role in cell migration will be addressed. Migration will be studied in cultures of adrenal cells that either have or do not have gap junctions (Specific Aim I). Cells will be treated with specific cAMP antagonists, agonist or diluent, and their migration characteristics will be assessed. Migration will be assayed using the following techniques: (1) time lapse imaging of the nuclear displacement or cell leading edge displacement of cells prelabeled with membrane permeable dyes; (2) counting the number of cells moving from a wound edge into a denuded area; and (3) counting the number of cells migrating in a silicon template compartmentalization chamber, in a microfluidic chamber, or through a Boyden chamber filter. A combination of time-lapse video microscopic and immunocytochemical techniques will be used to identify and elucidate the role of gap junction dynamics in cell migration.Infrastructural Contributions/Broader ImpactThe laboratory will continue to serve as a resource for the training of undergraduate and graduate students, postdoctoral fellows, visiting professors from minority institutions, and for mentoring high school students and (shadow program) teachers. Past NSF research funding has provided for the exposure and training of students, postdoctoral fellows, and faculty in areas of cell biology, endocrinology and signal transduction. Through seminars and workshops presented both nationally and internationally, information has been, and will continue to be, provided to members of the scientific community from a number of disciplines. High school students will continue to develop research projects and present at city and state science fairs. Recently, two undergraduate students won first place awards at national meetings, and others will be encouraged to follow in their path.