Growth/Differentiation Factors in Organogenesis

器官发生中的生长/分化因素

• 批准号: 6949783
• 负责人: ALAN PERANTONI
• 金额: --
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6949783
• 关键词: biological signal transduction; carcinogenesis; cell differentiation; cell growth regulation; cell line; chemokine receptor; fibroblast growth factor; growth factor; growth factor receptors; histogenesis; interleukin 1; kidney; kidney neoplasms; leukemia inhibitory factor; mesenchyme; microarray technology; receptor binding; renal tubule; tissue /cell culture; transforming growth factors

Morphogenesis in the differentiating metanephros is regulated by reciprocal interactions between ureteric bud epithelia and the metanephric mesenchyme. The ureteric bud invades the overlying metanephric mesenchyme and induces conversion of the mesenchyme into stromal and epithelial elements, which form the nephron. Concurrently, the mesenchyme stimulates the ureteric bud to grow, branch, and eventually form the collecting duct system of the adult kidney. The Differentiation and Neoplasia Section has focused on the elucidation of mechanisms of inductive signaling in metanephric development, seeking (1) the ligands responsible for nephronic differentiation, (2) other non-inductive regulatory factors of nephrogenesis, (3) the molecular targets of induction, and (4) alterations in inductive signaling during tumorigenesis. Previously, we reported the establishment of a cell line from the renal inductor, ureteric bud, and have now identified three families of secreted growth factors/cytokines that cooperate to induce epithelial conversion, tubule formation, and glomerulogenesis. These include the fibroblast growth factors (notably FGF2 and 9), gp130-binding proteins (leukemia inhibitory factor; LIF), and transforming growth factor-beta (Tgf-beta2, activin A and B, and gdf11). Individually, these factors induce differentiation, but at a rate significantly slower than in vivo, while in combination, they cooperate in accelerating tubule formation to a rate consistent with the in vivo process. In efforts to understand the basis for this cooperation and to define the signaling pathways responsible for induction, we have begun a systematic assessment of potential pathways that may be involved. As part of this effort, we have generated an inducible immortalized cell line of metanephric mesenchyme that can, under inductive conditions, differentiate to form epithelial-like structures consistent with tubule formation. We have characterized this line extensively now and found that it responds to the three families of inductive factors and expresses markers appropriate to nephronic epithelialization. This line is now being applied to the elucidation of signaling events that mediate or regulate differentiation of this progenitor. From microarray analysis, we have observed high levels of interleukin-1 expression by our ureteric bud cell line and gro-alpha expression in mesenchyme. Il-1 can stimulate gro-alpha in the mesenchyme, which in turn, binds to the chemokine receptors CXCR1 and 2. A block in signaling through this receptor dramatically inhibits development of the metanephros, suggesting that it plays a critical role in the growth and maintenance of metanephric tissues and that it may also function in a similar capacity during tumorigenesis. As an extension to these studies of normal development, we have also now initiated investigations into the status of inductive signaling pathways in renal tumors. Thus far, we have evaluated several cell lines derived from nephroblastomas and renal cell carcinomas. Remarkably, we are finding a consistent alteration in patterns of Stat phosphorylation, which mediates LIF signaling. We have established that these altered patterns exist in primary human tumor tissues from nephroblastomas but not in adjacent normal tissues. Furthermore, abrogation of altered Stat signaling inhibits tumorigenesis as measured by nephroblastoma cell growth in soft agar, suggesting that the altered patterns contribute to the neoplastic process. Finally, we previously observed the down regulation of a transcriptional coactivating factor with epithelial conversion, i.e., CITED1. We have now found that the protein interacts both physically and functionally with members of the COUP family of transcription factors. Furthermore, COUP apparently mediates an interaction between CITED1 and Smad4 to regulate signaling through this pathway. Since we have also observed an ability of CITED1 to inhibit Wnt/Tcf activation, it may represent one of the first examples of a factor capable of simultaneously regulating two critical pathways in nephrogenesis and is consistent with our hypothesis that CITED1 may function as a gatekeeper for metanephric blastemal cell differentiation.

后肾分化过程中的形态发生受输尿管芽上皮和后肾间质相互作用的调控。输尿管芽侵入上覆的后肾间质，诱导间质转化为间质和上皮成分，形成肾元。同时，间质刺激输尿管芽生长、分支，最终形成成人肾脏的集管系统。分化和瘤变部分侧重于阐明后肾发育中诱导信号的机制，寻求(1)负责肾分化的配体，(2)肾形成的其他非诱导调节因子，(3)诱导的分子靶点，(4)肿瘤发生过程中诱导信号的改变。先前，我们报道了从肾诱导剂输尿管芽中建立的细胞系，现在已经确定了三个分泌生长因子/细胞因子家族，它们共同诱导上皮转化、小管形成和肾小球形成。这些包括成纤维细胞生长因子（特别是FGF2和9），gp130结合蛋白（白血病抑制因子；LIF）和转化生长因子- β （tgf - β 2，激活素A和B， gdf11）。单独而言，这些因素诱导分化，但其速度明显低于体内，而结合起来，它们合作加速小管形成，其速度与体内过程一致。为了了解这种合作的基础，并确定负责诱导的信号通路，我们已经开始对可能涉及的潜在通路进行系统评估。作为这项工作的一部分，我们已经产生了一个可诱导的后肾间质永生化细胞系，它可以在诱导条件下分化成与小管形成一致的上皮样结构。我们现在已经广泛地描述了这条线，发现它对三种诱导因子家族有反应，并表达适合肾上皮化的标记物。这条线现在被应用于阐明介导或调节该祖细胞分化的信号事件。通过微阵列分析，我们观察到输尿管芽细胞系中白细胞介素-1的高水平表达和间质中gro- α的表达。Il-1可以刺激间质中的gro- α，而gro- α又与趋化因子受体CXCR1和2结合。通过该受体的信号传导阻滞会显著抑制后肾的发育，这表明它在后肾组织的生长和维持中起着关键作用，并且在肿瘤发生过程中也可能发挥类似的作用。作为这些正常发育研究的延伸，我们现在也开始了对肾肿瘤诱导信号通路状态的研究。到目前为止，我们已经评估了来自肾母细胞瘤和肾细胞癌的几种细胞系。值得注意的是，我们发现Stat磷酸化模式的一致改变，它介导LIF信号传导。我们已经确定这些改变的模式存在于肾母细胞瘤的原发人类肿瘤组织中，但不存在于邻近的正常组织中。此外，通过软琼脂中肾母细胞瘤细胞的生长测量，改变的Stat信号的消除抑制了肿瘤的发生，这表明改变的模式有助于肿瘤的形成过程。最后，我们之前观察到与上皮转化相关的转录共激活因子（即CITED1）的下调。我们现在已经发现该蛋白在物理和功能上与COUP家族的转录因子成员相互作用。此外，COUP显然介导了CITED1和Smad4之间的相互作用，通过该途径调节信号传导。由于我们也观察到CITED1抑制Wnt/Tcf激活的能力，它可能是能够同时调节肾形成中两个关键途径的因子的第一个例子之一，并且与我们的假设一致，即CITED1可能作为后肾囊胚细胞分化的看门人。