REGULATION OF ATROPHY-INDUCED PROGENITOR CELLS IN THE GASTRIC CORPUS

胃体中萎缩诱导的祖细胞的调节

• 批准号: 8695339
• 负责人: Jason C Mills
• 金额: $ 33.05万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-11 至 2017-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8695339
• 关键词: Ablation; Acids; Adult; Affect; Age; Animals; Arthritis; Atrophic; Bioinformatics; Biological Assay; Blood; CD44 gene; Cancer Etiology; Cell Death; Cell Differentiation process; Cell Proliferation; Cell Surface Proteins; Cells; Cessation of life; Characteristics; Chronic; Data; Databases; Developing Countries; Disadvantaged; Disease; E-Cadherin; Epithelial; Epithelial Cells; Epithelium; Estrogens; Frequencies; Gastric Parietal Cells; Gastric ulcer; Gene Expression; Generations; Genes; Goals; Hematopoietic stem cells; Homeostasis; Hour; Human; Hyaluronan; Inflammation; Inflammatory; Injection of therapeutic agent; Injury; Intestines; Kinetics; Label; Learning; Ligands; Malignant Neoplasms; Mediator of activation protein; Metaplasia; Minority; Molecular; Molecular Genetics; Molecular Profiling; Mus; Organ; Pathway interactions; Patients; Pattern; Pharmaceutical Preparations; Pharmacologic Substance; Phosphorylation; Physiological; Population; Process; Pyloric antrum; Regulation; Relative (related person); Research; Risk; Role; STAT3 gene; Secondary to; Selective Estrogen Receptor Modulators; Signal Transduction; Site; Staging; Stem cells; Stomach; Stomach Diseases; System; Tamoxifen; Techniques; Time; Tissues; Toxic effect; Tweens; Undifferentiated; United States; Work; adult stem cell; cancer risk; cell behavior; cost; genetic pedigree; human disease; inhibitor/antagonist; innovation; killings; laser capture microdissection; malignant stomach neoplasm; progenitor; prospective; research study; response; response to injury; stem; stem cell differentiation; stem cell niche; stem cell population; transcription factor; transcriptome sequencing

DESCRIPTION (provided by applicant): Despite recent breakthroughs in identifying prospective stem cell markers (e.g., LGR5) in the intestines and the gastric antrum, relatively little is known about the stem cells of the gastric (body) corpus epithelium. Our overall goal is t better characterize the corpus stem cell under normal conditions and in response to injury. Preliminary data show that the selective estrogen receptor modulator tamoxifen causes ablation of nearly all parietal cells, the acid-secreting lineage, in the mouse stomach within 3 days. Within 6 hours, parietal cells begin to die (atrophy), and progenitor cells in the isthmal stem cel niche of corpus gastric units begin to expand in response. We further find that: 1) the cell surface protein CD44 is required for normal stem cell homeostasis, because proliferation is halved in Cd44-/- relative to wildtype mice; and 2) CD44 marks the expanding isthmal progenitor population. CD44 activates STAT3 and is induced by ERK signaling. We show here that ERK and STAT3 are activated in early expanding progenitors, and levels of the ERK-induced transcription factor EGR1 peak at 3 days, in concert with maximal progenitor expansion. We hypothesize that parietal cell death leads to signals that activate the ERK pathway in isthmal stem/progenitor cells, which causes increased CD44 expression and increased proliferation. In Aim 1, we will determine whether ERK signaling is required for normal stem cell homeostasis and atrophy-induced expansion using both ERK pharmacological inhibition and pedigrees of mice with deficient (Egr1-/-) and overactive (Nab2-/-) ERK. In Aim 2, we will determine whether CD44 is required or sufficient for atrophy-induced progenitor expansion. We will examine kinetics of stem cell expansion in Cd44-/- mice, and we will either activate CD44 in wildtype mice ¿tamoxifen by injection of the CD44 ligand Hyaluronan or block CD44- Hyaluronan interactions with the inhibitor PEP-1. We will determine if CD44 induces STAT3 and/or if it is required for progenitor expansion. In Aim 3, we will isolate vehicle- and tamoxifen-treated CD44-expressing epithelial progenitor cells by FACS and laser-capture microdissection and then use RNA-Seq to generate gene expression profiles of the normal and atrophy-expanded progenitor cells that will then be integrated bioinformatically with our existing database of global gene expression in stem and progenitor cells. We will also perform limiting dilution assays to determine the stem cell frequency within the epithelial CD44 population. In humans, chronic inflammation can cause parietal cell atrophy, which in turn changes stem cell proliferation and differentiation. Atrophy predispose patients to gastric cancer, but changes in stem cells secondary to inflammation/injury are also an important and understudied aspect of adult diseases in multiple other tissues (e.g., hematopoietic stem cell differentiation changes in arthritis). Thus, the experiments proposed may help us begin to understand stem cell response to injury in multiple tissues and diseases. Finally, our new finding that the key drug tamoxifen causes gastric toxicity warrants further study in humans.

描述（由申请人提供）：尽管最近在确定肠道和胃窦中潜在的干细胞标记物（例如LGR5）方面取得了突破，但对胃（体）体上皮干细胞的了解相对较少。我们的总体目标是更好地表征正常条件下和损伤反应下的体干细胞。初步数据显示，选择性雌激素受体调节剂他莫昔芬在3天内导致小鼠胃中几乎所有的壁细胞（分泌酸的细胞系）消融。在6小时内，壁细胞开始死亡（萎缩），而胃主体的峡部干细胞生态位中的祖细胞开始相应扩张。我们进一步发现：1)细胞表面蛋白CD44是正常干细胞稳态所必需的，因为相对于野生型小鼠，CD44 -/-的增殖减少了一半；2) CD44标志着峡部祖细胞群体的扩大。CD44激活STAT3并由ERK信号诱导。我们在这里表明，ERK和STAT3在早期扩张的祖细胞中被激活，ERK诱导的转录因子EGR1的水平在第3天达到峰值，与最大的祖细胞扩张一致。我们假设壁细胞死亡导致信号激活峡干/祖细胞中的ERK通路，从而导致CD44表达增加和增殖增加。在Aim 1中，我们将通过ERK药理学抑制和ERK缺陷（Egr1-/-）和过度活跃（Nab2-/-）小鼠的谱系来确定ERK信号是否需要正常干细胞稳态和萎缩诱导的扩增。在Aim 2中，我们将确定CD44对于萎缩诱导的祖细胞扩增是必需的还是充分的。我们将研究Cd44-/-小鼠干细胞扩增的动力学，我们将通过注射Cd44配体透明质酸来激活野生型小鼠的Cd44，或者阻断Cd44-透明质酸与抑制剂PEP-1的相互作用。我们将确定CD44是否诱导STAT3和/或它是否是祖细胞扩增所必需的。在Aim 3中，我们将通过FACS和激光捕获显微解剖分离载体和他莫昔芬处理的表达cd44的上皮祖细胞，然后使用RNA-Seq生成正常和萎缩扩增祖细胞的基因表达谱，然后将其与我们现有的干细胞和祖细胞全球基因表达数据库进行生物信息学整合。我们还将进行限制性稀释试验，以确定上皮细胞CD44群体中的干细胞频率。在人类中，慢性炎症可导致顶叶细胞萎缩，进而改变干细胞的增殖和分化。萎缩使患者易患胃癌，但继发于炎症/损伤的干细胞变化也是多种其他组织中成人疾病的一个重要且未被充分研究的方面（例如，关节炎中的造血干细胞分化变化）。因此，提出的实验可能有助于我们开始了解干细胞对多种组织和疾病损伤的反应。最后，我们的新发现，关键药物他莫昔芬引起胃毒性值得在人类进一步研究。