The role of Activin A signaling in esophageal cell invasion

激活素A信号在食管细胞侵袭中的作用

基本信息

批准号：
9096042
负责人：
CLAUDIA D ANDL
金额：
$ 30.14万
依托单位：
UNIVERSITY OF CENTRAL FLORIDA
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-07-01 至 2018-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9096042
关键词：
Activins Address Affect Autocrine Communication Biological Biological Assay Cells Chronic Data Development Disease Dominant-Negative Mutation E-Cadherin E-Cadherin Staining Method Embryonic Development Epithelial Epithelial Cells Epithelial-Stromal Communication Epithelium Esophageal Esophageal Squamous Cell Esophagus Fibroblasts Fibrous capsule of kidney Gastrointestinal Diseases Goals Growth Homeostasis In Vitro Investigation Knowledge Maintenance Malignant Squamous Cell Neoplasm Mediating Mesenchymal Mission Modeling Oncogenic Paracrine Communication Pathogenesis Play Public Health Regulation Reporting Research Role Signal Pathway Signal Transduction Signaling Molecule System Testing Tissue Recombination Transforming Growth Factor beta Transforming Growth Factors Tumor Tissue Xenograft Model Xenograft procedure activin A autocrine base in vivo in vivo Model innovation insight keratinocyte member mouse model neoplastic cell novel novel strategies overexpression paracrine receptor response small hairpin RNA tumorigenesis

项目摘要

DESCRIPTION (provided by applicant): The transforming growth factor ß (TGFß) superfamily of signaling molecules fulfills essential functions in development and disease. Activins are members of this superfamily. Activin A (Act A) and TGFß overlap in their intracellular signaling cascades and function and have multiple common downstream targets. We have previously demonstrated that coordinated loss of TGFß receptor II and E-cadherin results in cell invasion. These genetically manipulated esophageal keratinocytes show activation of the common downstream target Smad2 despite the absence of functional TGFß receptor II. These data indicate that Act A may be responsible for Smad2 activation in the absence of canonical TGFß signaling in esophageal keratinocytes. The interaction of the epithelium with the surrounding microenvironment is essential for maintaining epithelial homeostasis. A knowledge gap exists as no mechanism for the dual function of Act A in the maintenance and disruption of homeostasis similar to TGFß has been described yet. Our long-term goal is to produce mechanistic insights into the signaling pathways mediating epithelial-mesenchymal crosstalk. Our central hypothesis for this study is that Act A signaling is a major regulator of epithelial-mesenchymal transition (EMT) and epithelial-mesenchymal crosstalk in the esophagus. Our preliminary data show the increased secretion of Act A in organotypic cultures and increased cell invasion of esophageal keratinocytes in Boyden chamber assays after Act A stimulation. Therefore, we propose to utilize state-of-the-art approaches and novel mouse models to demonstrate the role of Act A in squamous homeostasis in vitro and in vivo. These data will advance our understanding of TGFß superfamily signaling in the esophagus. To test our hypothesis we pursue the following specific aims: 1) To determine how Act A signaling results in EMT and esophageal cell invasion. 1a) To define autocrine Act A signaling in esophageal epithelial cells and 1b) To analyze the functional consequences of ActRIIB loss. 2) To determine how paracrine Act A signaling induces fibroblast activation. 3) To identify the role of Act A in esophageal homeostasis in vivo. In the first aim, an established shRNA approach to suppress Act A and ActRIIB, as well as overexpression thereof will be used to show the causal effects of Activin signaling on EMT and cell invasion. We have shown the successful use of organotypic cultures to demonstrate the disruption of epithelial homeostasis by altered epithelial-mesenchymal crosstalk. We will utilize this system to identify how paracrine Act A secretion induces fibroblast activation in the second aim. In specific aim 3, we make use of mouse models with targeted loss of E-cadherin and TGFß receptor in the esophagus, which we have successfully developed. The approach is innovative, because it mimics the microenvironment by incorporating stromal and epithelial components in the analysis. The proposed research is significant, because it is expected to identify the role of a novel player, Activin A, in esophageal homeostasis.

描述（由适用提供）：信号分子的转化生长因子ß（TGFß）超家族符合发育和疾病中的基本功能。激活素是该超家族的成员。激活素A（ACT A）和TGFß在其细胞内信号级联和功能中重叠，并具有多个常见的下游靶标。我们以前已经证明，TGFß受体II和E-钙粘蛋白的协调损失会导致细胞侵袭。这些一般操纵的食管角质形成细胞显示了不存在功能性TGFß受体II的常见下游靶标SMAD2目的地的激活。这些数据表明，在食管角质形成细胞中没有规范的TGFß信号传导的情况下，ACT A可能导致SMAD2激活。上皮与周围微环境的相互作用对于维持上皮稳态至关重要。知识差距是尚未描述了ACT A双重功能在维持和破坏与TGFß类似的稳态中的双重功能的机制。我们的长期目标是对介导上皮 - 间质串扰的信号通路产生机理见解。我们在这项研究的中心假设是，ACT A信号传导是食管中上皮 - 间质转变（EMT）和上皮 - 间质串扰的主要调节剂。我们的初步数据表明，在有机培养物中，ACT A的分泌增加，并增加了A刺激后，在Boyden室内刺激中食管角质形成细胞的细胞侵袭。因此，我们建议利用最先进的方法和新型的小鼠模型来证明A ac a在体外和体内的鳞状稳态中的作用。这些数据将促进我们对食管中TGFß超家族信号的理解。为了检验我们的假设，我们将以下特定目的提取：1）确定ACT A的信号如何导致EMT和食管细胞侵袭。 1A）在食管上皮细胞中定义自分泌作用A信号传导和1B）分析Actriib损失的功能后果。 2）确定旁分泌如何在成纤维细胞激活中作用A信号传导。 3）确定A法在体内的食管稳态中的作用。在第一个目的中，将使用已建立的SHRNA抑制A和ACTRIIB的方法，以及其过表达，将用于显示激活素信号传导对EMT和细胞侵袭的因果作用。我们已经证明了有机培养物的成功使用，以通过改变上皮 - 间质串扰来证明上皮稳态的破坏。我们将利用该系统来确定旁分泌作用A分泌如何在第二个目标中诱导成纤维细胞激活。在特定的目标3中，我们利用了在食管中靶向损失E-钙粘着蛋白和TGFß受体的靶向损失的小鼠模型，我们已经成功地开发了这些模型。该方法具有创新性，因为它通过将基质和上皮成分掺入分析中模仿了微环境。拟议的研究很重要，因为它有望确定新型玩家Activin A在食管稳态中的作用。