Stem cell niche and Wnt in liver injury and regeneration.

描述(申请人提供)：卵圆细胞是成年的肝脏前体细胞，在肝脏再生中发挥生理作用，具有诱人的治疗潜力。它们被认为生活在门户周围区域的一个小生境中。然而，损伤后激活椭圆形细胞的信号尚不清楚。Wnt家族蛋白在多种哺乳动物组织中对成体干细胞的维持起着重要作用。WNT信号也参与肝脏的发育和修复。有证据表明，Wnt信号在激活卵圆细胞的损伤反应中起着中心作用。组织修复中的一个主要悬而未决的问题是，损伤如何导致重建组织所需的信号被激活。在一条新的研究路线上，Nusse实验室已经确定了一种转录调控元件(称为CAIRE，即顺式作用伤害反应元件)，它控制Wnt在受伤时的表达。通过使用与这种增强子相关联的报告基因，已在各种组织中检测到了CAIRE的功能。在初步工作中，实验室已经表明，它位于肝脏的门静脉周围区域，对损伤有反应。假设CAIRE在肝脏损伤后经历激活，从而激活椭圆形细胞隔室。它通过驱动再生反应所需的局部Wnt的表达来诱导卵圆细胞的增殖。CAIRE上游存在一条特定的信号通路，控制CAIRE的活性和Wnt的表达。拟议的研究将解决这些假设。首先，我们将研究CAIRE在三种不同形式的肝损伤中的作用，这三种肝损伤表现出不同程度的卵圆细胞激活。凯尔记者预计会在激活卵圆细胞群的损伤中表达。接下来，将产生CAIRE有条件地激活Wnt抑制因子DKK的转基因小鼠，以确定由CAIRE驱动的局部Wnt信号是否是卵圆细胞再生反应所必需的。WNT抑制有望导致损伤后卵圆细胞再生减少。最后，将确定结合并激活CAIRE的转录因子。生物信息学将被用来识别与CAIRE结合的潜在转录因子。将对这些因素进行审查，以确定这些因素是否对凯尔具有直接约束力。根据这些结果，CAIRE内的转录因子结合位点将发生突变。转基因小鼠将被用来评估功能丧失。预计这些小鼠的卵圆细胞再生反应会降低。这些研究将提供Wnt表达上游激活剂的第一个证据。他们将增加Wnt信号在肝损伤和随后涉及卵圆细胞的再生反应中发挥关键作用的证据。最后，这些结果将使我们深入了解生态位在将损伤反应与组织再生联系起来的作用。 公共卫生相关性：在美国，肝硬变是发病率和死亡率的最重要原因之一。卵圆细胞在肝硬变的细胞替代治疗中具有巨大的潜力。更好地理解来自利基的信号如何调控卵圆细胞介导的再生将是实现基于细胞的肝病治疗潜力的重要一步。

DESCRIPTION (provided by applicant): Oval cells are adult liver progenitor cells that play a physiologic role in liver regeneration and have attractive therapeutic potential. They are believed to reside in a niche within the periportal region. However, the signals that activate oval cells after injury are not known. The Wnt family of proteins plays important roles in adult stem cell maintenance in a variety of mammalian tissues. Wnt signaling is also involved in liver development and repair. Evidence suggests that Wnt signaling is central to the injury response that activates oval cells. A main unanswered question in tissue repair is how injury leads to activation of the signals necessary for rebuilding the tissue. Pursuing a novel line of research, the Nusse lab has identified a transcriptional control element (called CAIRE, for Cis Acting Injury Responsive Element) that governs the expression of Wnt in response to injury. Through the use of reporter genes linked to this enhancer, CAIRE function has been examined in various tissues. In preliminary work, the lab has shown that it is in the periportal region of the liver and responds to injury. The hypothesis is that CAIRE undergoes activation in response to liver injury that activates the oval cell compartment. It induces oval cell proliferation by driving local Wnt expression, which is required for the regenerative response. A specific signaling pathway exists upstream of CAIRE that controls CAIRE activity and Wnt expression. The proposed studies will address these hypotheses. First, the role of CAIRE in three distinct forms of liver injury that exhibit different degrees of oval cell activation will be examined. CAIRE reporter expression is expected in injury that activates the oval cell population. Next, transgenic mice in which CAIRE conditionally activates the Wnt inhibitor Dkk will be generated to determine if local Wnt signaling driven by CAIRE is required for the oval cell regenerative response. Wnt inhibition is expected to result in decreased oval cell regeneration after injury. Lastly, transcription factors that bind to and activate CAIRE will be identified. Bioinformatics will be used to identify potential transcription factors that bind to CAIRE. These factors will be examined for direct binding to CAIRE. Based on these results, mutations will be made in transcription factor binding sites within CAIRE. Transgenic mice will be made to assess for loss-of- function. These mice are expected to have decreased oval cell regenerative response. These studies will provide the first evidence of an upstream activator of Wnt expression. They will add to the evidence for a key role for Wnt signaling in liver injury and the subsequent regenerative response involving oval cells. Lastly, the results will give insight into the role of the niche in linking the injury response to tissue regeneration. PUBLIC HEALTH RELEVANCE: Cirrhosis is one of the most significant causes of morbidity and mortality in the United States. Oval cells have tremendous potential in cell replacement therapy for cirrhosis. Improved understanding of how signals from the niche regulate oval cell mediated regeneration will be an important step towards realizing the potential of cell based therapy for liver disease.