Characterizing the Role of Pancreatic Progenitors in Regeneration

描述胰腺祖细胞在再生中的作用

• 批准号: 9922906
• 负责人: Michael J Parsons
• 金额: $ 37.5万
• 依托单位: UNIVERSITY OF CALIFORNIA-IRVINE
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-07-01 至 2022-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9922906
• 关键词: Ablation; Adult; Affect; Architecture; Automobile Driving; B cell differentiation; Behavior; Beta Cell; Binding; Biological; Biological Models; Biology; Cell Line; Cells; Cilia; Collection; Data; Data Set; Development; Developmental Biology; Diabetes Mellitus; Disease; Endocrine; Fishes; Genes; Genetic Transcription; Genome; Genomic approach; Genomics; Goals; Grant; Haploidy; Heterozygote; Homeostasis; Human; Human Cell Line; Insulin; Insulin-Dependent Diabetes Mellitus; Islets of Langerhans; Lead; Maps; Mediating; Mediator of activation protein; Methods; Modeling; Molecular; Morphology; Natural regeneration; Non-Insulin-Dependent Diabetes Mellitus; Organogenesis; Pancreas; Pancreatic Centroacinar Cell; Pathway interactions; Pharmacology; Phenotype; Play; Prevalence; Process; Production; Recovery; Regulation; Reporter; Resources; Role; Route; Source; TACSTD1 gene; Techniques; Testing; Therapeutic; Transcript; Transgenic Organisms; Treatment Efficacy; Vertebrates; Work; World Health Organization; Zebrafish; beta cell replacement; cell regeneration; cell type; chromatin immunoprecipitation; cost effective; deep sequencing; dosage; insight; interest; knock-down; mutant; notch protein; novel; pancreas development; progenitor; regenerative; stem cells; tool; transcription factor; transcriptome

Project Summary The World Health Organization estimates that the global prevalence of diabetes in adults is 9%. Both Type 1 and Type 2 diabetes involve the issue of reduced β-cell mass; subsequently a cure for diabetes must involve β-cell replacement. Ideally, a cure would involve inducing regeneration via β-cell neogenesis from endogenous pancreatic progenitors. For these reasons we are interested in explicating the process of β-cell neogenesis, i.e., how β cells are formed from progenitors in the pancreas. Unlike their mammalian counterparts, we have shown that zebrafish readily regenerate their β cells following cell-specific ablation. Our goal is to identify the mechanisms behind the zebrafish’s capacity for β-cell neogenesis. Such molecular pathways could then be pharmacologically exploited in humans to induce β-cell neogenesis. We have recently made two discoveries critical to understanding how zebrafish so easily recover following β- cell ablation. First, we identified the progenitor source for β-cell neogenesis—namely a cell type called the centroacinar cell (CAC); second, we discovered that diminished activity of the Sox9b transcription factor leads to significantly accelerated regeneration. From these insights we hypothesized 1) Sox9b acts cell autonomously to maintain progenitor potency in adult CACs; 2) Diminishing Sox9b activity alters the behavior of CACs or their progeny in regeneration; and 3) The identification of downstream genes of SOX9 will elucidate molecular mechanisms that regulate β-cell differentiation. By testing these hypotheses in three complementary yet independent aims, we expect to discover if the differences in regeneration between sox9b heterozygotes and wildtypes is due either to pre-existing differences in morphology or the behavior of the CACs during regeneration. Furthermore, we will use genomic approaches to identify the direct downstream transcriptional targets of Sox9 homologs because we expect these targets will be the mediators of the sox9b haploinsufficient phenotype. As part of our preliminary data we knocked down SOX9 levels in the human PANC-1 cell line, a surrogate for pancreatic progenitors, and identified affected transcript levels. We have also used chromatin immunoprecipitation and deep sequencing (ChIP-seq) to identify where SOX9 binds in the PANC-1 genome. Putting these results together has allowed us to find direct targets of SOX9 transcriptional activity in PANC-1 cells. We have identified interesting genes downstream of SOX9, such as EpCAM, and identified biological pathways that SOX9 controls, such as cilia function and Notch regulation. Greatly encouraged by our preliminary results we now aim to expand this analysis to find SOX9 targets during development and regeneration. By the end of this proposed work we expect a better understanding of SOX9 function and the discovery of potential therapeutic routes to alleviate β-cell paucity in humans.

项目摘要 世界卫生组织估计，成人糖尿病的全球患病率为9％。两者类型1 和2型糖尿病涉及减少β细胞质量的问题；随后治愈糖尿病必须涉及 β细胞替代。理想情况下，治疗将涉及通过内生的β细胞新作用诱导再生 胰腺祖细胞。由于这些原因，我们有兴趣明确说明β细胞新发生的过程 即，如何由胰腺中的祖细胞形成β细胞。与他们的哺乳动物对应物不同，我们有 表明斑马鱼在细胞特异性消融后很容易再生其β细胞。我们的目标是确定 斑马鱼的β细胞新生成能力背后的机制。这样的分子途径可以是 在人类中探索的药理学诱导β细胞新生成。 最近，我们做出了两个发现，对于理解斑马鱼如何在β-之后如何轻易恢复至关重要的发现至关重要。 细胞消融。首先，我们确定了β细胞新发生的祖细胞来源，即一种称为的细胞类型 质心细胞（CAC）;其次，我们发现Sox9b转录因子引导的活性减少了 显着加速再生。从这些见解中，我们假设1）Sox9b作用细胞 自主维持成人CAC中的祖细胞效力； 2）减少Sox9b活动改变了 CAC的行为或它们在再生方面的进步； 3）鉴定下游基因的 Sox9将阐明调节β细胞分化的分子机制。通过测试这些 三个完善但独立的目标的假设，我们希望发现是否有差异 Sox9b杂合子与野生型之间的再生要么是由于先前存在的差异 形态或CAC在再生过程中的行为。此外，我们将使用基因组方法 确定SOX9同源物的直接下游转录目标，因为我们希望这些目标将 成为SOX9B单倍型表型的介体。 作为我们初步数据的一部分，我们在人类Panc-1细胞系中击倒Sox9水平，替代 胰腺祖细胞，并确定了受影响的转录水平。我们还使用了染色质 免疫沉淀和深度测序（CHIP-SEQ）识别SOX9在PANC-1基因组中结合的位置。 将这些结果放在一起使我们能够找到Panc-1中SOX9转录活动的直接目标 细胞。我们已经确定了Sox9下游的有趣基因，例如Epcam，并确定了生物学 Sox9控制的途径，例如纤毛功能和Notch调节。极大地鼓励我们 初步结果我们现在旨在扩展此分析，以在开发过程中找到Sox9目标 再生。 在这项拟议的工作结束时，我们希望对Sox9功能有更好的了解和发现 减轻人类β细胞稀少的潜在治疗途径。