PDG Links Stem Cell Niche to Pancreatic Epithelial Renewal, Repair and Cancer

描述(申请人提供)：干细胞壁龛负责全身大多数上皮表面的终生更新。这些细胞的失调被认为是导致 在化生和癌症方面。到目前为止，人们对这些体细胞干细胞的位置或它们对胰腺上皮再生和癌症的贡献知之甚少。我们实验室最近发现了一种新的上皮干细胞隔室--胰腺导管腺体(PDG)。这些腺体独特地表达Shh和TFF2以及其他已知存在于祖细胞干细胞利基中的发育基因。作为对急性损伤的反应，这些腺体能够进行不对称分裂，从而产生分化的子代细胞，然后这些子代细胞迁移到主管道中重新填充。作为对慢性炎症的反应，这些细胞可以经历胃肠道化生和囊性肥大，在组织和分子上类似于侧枝IPMN，已知的PDAC的前体。对人IPMN的评估显示，PDG形成了这些大的乳头突起的增殖性基底隐窝段。这项建议的重点是表征PDG间隔在上皮再生和癌变中的作用。[SA1]这一目标中提出的实验将确定PDG作为负责上皮更新和再生的上皮祖干细胞利基的作用。我们建议使用命运映射，体内谱系标记策略，特别是标记PDG间隔，结合一个具有良好特征的急、慢性胰腺损伤模型，来确认这些间隔包含上皮祖细胞。在体外和体内，也提出了确定这些细胞是否具有多潜能的策略。我们的初步数据表明，这个隔室至少有两个干细胞群体，标记保留细胞(LRC)和瞬时扩增细胞(TA)。为了更好地描述这两个 干细胞群体我们提出了一个发育子目标，通过从最少的(10个细胞)样本中通过mRNA-Seq来评估转录组，以确定这些细胞的独特标记，以及了解在再生和炎症性化生过程中调节这一间隔室的关键途径。[SA2]探讨PDG在胰腺癌前病变、旁支IPMN和肿瘤形成中的作用机制。为了达到这一目的，我们建议在小鼠体内使用谱系标记和PDG细胞特异性癌基因激活策略来确定PDG是IPMN和侵袭性癌症的起源室。通过使用线粒体突变图谱策略，在人和小鼠标本中进一步深入了解前体病变形成的机制，表明IPMN是通过生态位继承、克隆转换和腺体分裂形成的。低、中、高级别发育不良IPMN的SMART-SEQ序列可能有助于我们了解参与IPMN发生、发展和侵袭的关键途径。了解这个隔室如何促进再生和癌症，将使我们对癌症预防、早期诊断和治疗有新的见解。

DESCRIPTION (provided by applicant): Stem cell niches are responsible for life-long renewal of most epithelial surfaces throughout the body. Dysregulation of these cells is believed to result in metaplasia and cancer. To date little is known about these somatic stem cell niches or their contribution to pancreatic epithelial regeneration and cancer. Our laboratory has recently discovered a novel epithelial stem cell compartment, Pancreatic Duct Glands (PDG). These glands uniquely express Shh and TFF2 as well as other developmental genes known to reside in progenitor stem cell niches. In response to acute injury these glands are capable of asymmetric division that gives rise to differentiated daughter cells which then migrate to repopulate the main ducts. In response to chronic inflammation these cells can undergo a GI metaplasia and cystic hypertrophy which histologically and molecularly resemble side-branch IPMN, known precursors of PDAC. Evaluation of human IPMN reveals that PDG form the proliferative basilar crypt segment of these large papillary projections. This proposal is focused on characterizing the role of the PDG compartment in epithelial regeneration and carcinogenesis. [SA1] The experiments proposed in this aim will determine the role of PDG as an epithelial progenitor stem cell niche responsible for epithelial renewal and regeneration. We propose to use a fate mapping, in vivo lineage tagging strategy specifically tagging the PDG compartment, in combination with a well-characterized model of acute and chronic pancreatic injury, to confirm that these compartments contain epithelial progenitor cells. In vitro and in viv strategies are also proposed to determine if these cells have multipotential capacity. Our preliminary data suggests that this compartment has at least two populations of stem cells, label retaining cells (LRC) and transient amplifying cells (TA). In order to better characterize this two stem cell population we propose a developmental sub-aim to evaluate the transcriptome by mRNA-Seq from minimal (10-cell) samples in order to identify unique markers for these cells, as well as understand key pathways that regulate this compartment during regeneration and inflammatory metaplasia. [SA2] To determine the mechanisms by which PDG contribute to the formation of pancreatic cancer precursor lesions, side-branch IPMN and cancer. To accomplish this aim we propose to use in an in vivo lineage tagging and PDG cell-specific oncogene activation strategy in mice to identify PDG as the compartment of origin for IPMN and invasive cancer. Further insight into the mechanisms underlying the formation of precursor lesions will be gained in human and mouse specimens by using a mitochondrial mutational mapping strategy to show that IPMN are formed by niche succession, monoclonal conversion and gland fission. mRNA SMART- Seq of low, moderate and high grade dysplastic IPMN may allow us to understand key pathways involved in initiation, progression, and invasion. Understanding how this compartment contributes to regeneration and cancer will give us new insights into cancer prevention, early diagnosis and treatment.