Mechanisms Involved in Mammary Morphogenesis

乳腺形态发生的机制

• 批准号: 6989354
• 负责人: Joan Siefert Brugge
• 金额: $ 14.87万
• 依托单位: WHITEHEAD INSTITUTE FOR BIOMEDICAL RES
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-03-16 至 2009-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6989354
• 关键词: BCL2 gene /protein; acinar cell; apoptosis; carcinogenesis; carcinoma; cell line; gene induction /repression; histogenesis; mammary epithelium; mammary gland; neoplasm /cancer genetics; neoplastic growth

The filling of a hollow lumen within an intact glandular structure is a hallmark of early tumorigenesis. However the mechanisms involved in the formation of a glandular lumen and its filling during the development of carcinoma-in-situ are poorly understood. We have developed an in vitro three-dimensional (3D) culture model to investigate alterations in the architecture and growth properties of glandular epithelial structures (acini) during early stages of mammary carcinogenesis. We have found that formation of the lumen involves selective death of cells in the center of the acini. Filling of the luminal space, as provoked by oncogenes, requires not only induction of constitutive proliferation but also anti-apoptotic signals that allow cells to survive in the luminal space. We have identified the proapoptotic Bcl-2 protein Bim as critical mediator of apoptosis during lumen formation and found that its expression is regulated by oncogenes that escape death in the luminal space. Bim was also found to be induced and critically involved in apoptosis induced by detachment of monolayer cells from matrix (a process referred to as anoikis) and several lines of evidence suggest that matrix deprivation may be involved in luminal cell death during morphogenesis. This proposal describes plans to elucidate the mechanisms that regulate cell death induced by Bim during both lumen formation and anoikis, identify other proteins that collaborate with Bim to mediate cell death, and determine the mechanisms whereby oncogenes allow cells to escape cell death induced by these apoptotic mediators. This study should provide important information relating to events associated with the development of carcinoma-in-situ tumors in humans. These investigations should provide important information relating to events associated with the development of carcinoma-in-situ tumors in humans. Studies to examine the regulation of Bim during anoikis, lumen formation and oncogenesis will involve investigations of the mechanism involved in Bim induction and activation in both models, analysis of how Bim is downregulated or inactivated by oncogenes, and elucidation of the basis for protection of mammary epithelial cells from Bim-induced death operating in acinar cells that are attached to matrix. Identification of other proapoptotic proteins will involve affinity isolation techniques as well as strategies that induce loss of function of other pro-apoptotic proteins. We will use retroviral genetic screens to identify additional proteins that protect cells from detachment induced death or luminal clearing in acini. Lastly, we will examine whether the processes that we define in the immortalized MCF-10A cells are operative in primary human cells.

完整腺体结构内空腔的填充是早期肿瘤发生的标志。然而，对原位癌发展过程中腺腔形成及其填充的机制知之甚少。我们开发了一种体外三维 (3D) 培养模型，用于研究乳腺早期阶段腺上皮结构（腺泡）的结构和生长特性的变化 致癌作用。我们发现管腔的形成涉及腺泡中心细胞的选择性死亡。由癌基因引起的管腔空间的填充不仅需要诱导组成型增殖，还需要允许细胞在管腔空间中存活的抗凋亡信号。我们已经确定促凋亡 Bcl-2 蛋白 Bim 是管腔形成过程中细胞凋亡的关键介质，并发现其表达受到癌基因的调节 在光明空间中逃脱死亡。还发现 Bim 被诱导并关键参与单层细胞从基质脱离（称为失巢凋亡的过程）诱导的细胞凋亡，并且多种证据表明基质剥夺可能与形态发生过程中的管腔细胞死亡有关。该提案描述了阐明在管腔形成和失巢凋亡过程中调节 Bim 诱导的细胞死亡的机制的计划，确定 与 Bim 合作介导细胞死亡的其他蛋白质，并确定癌基因允许细胞逃避这些凋亡介质诱导的细胞死亡的机制。这项研究应该提供与人类原位癌肿瘤发展相关的事件的重要信息。这些研究应该提供与人类原位癌肿瘤发展相关的事件的重要信息。 检验Bim在失巢、管腔形成和肿瘤发生过程中的调节的研究将涉及研究两种模型中Bim诱导和激活所涉及的机制，分析Bim如何被癌基因下调或失活，并阐明乳腺保护的基础来自 Bim 诱导死亡的上皮细胞在附着于基质的腺泡细胞中起作用。其他促凋亡蛋白的鉴定将涉及亲和分离技术以及诱导其他促凋亡蛋白功能丧失的策略。我们将使用逆转录病毒遗传筛选来识别其他蛋白质，这些蛋白质可以保护细胞免受分离诱导的死亡或腺泡腔内的清除。最后，我们将检查我们在永生化 MCF-10A 细胞中定义的过程是否在原代人类细胞中有效。