A Major Nutritional Effect on Intestinal Stem Cells and Tumors

我们解剖了西式纯净饮食(NWD1)对肠道和结肠肿瘤发生的影响。 NWD1总结了小鼠的关键营养素摄入量，每种营养素的流行病学水平都与患心脏病的风险较高有关。 在西方发达国家，肠癌的发病率很高。我们证实，NWD1可以放大和加速肠道 许多小鼠遗传模型中的肿瘤，无论遗传病因和侵袭性由单一或 多基因驱动因素。此外，当长期喂食野生型小鼠时，我们和其他人建立了NWD1 会导致零星的肠道和结肠肿瘤。在这些散发性肿瘤发展之前至少一年，有 组织病理学正常粘膜中上皮细胞和潜在的分子通路的成熟改变。 这导致了研究NWD1是如何改变肠道干细胞的细胞和分子生物学的， 我们报告了一项令人惊讶的发现：NWD1损害了肠道中Lgr5hi细胞的干细胞功能 动态平衡和肿瘤发生，但这是通过动员Bmi1+细胞作为干细胞来弥补的 这些功能。此外，遗传和饮食实验发现，维生素D暴露和维生素D受体减少 信号在这些效应中很重要，这与克里弗斯报告的干细胞表达特征强烈一致 提示VDR信号与Lgr5hi干细胞功能有关。此外，我们注意到小鼠在肠道上的实验 干细胞使用的饮食中的维生素D水平远远高于美国人群的水平。这 提出了关于哪些干细胞以及如何在体内平衡和肿瘤发生中发挥作用的重要问题。 自前一次审查(提交供发表)以来的大量新数据确立了饮食的深远影响 暴露：i)Lgr5hi和Bmi1+细胞的编程(RNAseq)；ii)这些细胞的DNA修复能力； 三)随后在这些隐窝细胞中积累的突变的范围、频谱和特征 隔室(FACS分离的单个细胞的测序)。目标1解决了饮食诱导的假设 Lgr5hi干细胞编程、功能和表观遗传学改变的变化可以异常长寿，并且 因此，干细胞可以作为“哨兵细胞”来监测饮食干预的调节能力 这些改变是潜在的风险标志和干预目标。这将提供基本数据 关于干细胞功能调节中的基因和途径的协调，标记的识别 相对风险，以及如何优化干预策略。最初提交的AIM2主要是 完成，并对目标2进行了修订，以调查改变的DNA修复在 条件性APC时Lgr5+和Bmi1+人群的不同饮食对肿瘤发生的影响 突变专门针对这些细胞群体中的每一个。AIM2还决定了饮食对 在出现的肿瘤中，APC和整个基因组中突变的性质。目标3确定了 对肿瘤组织病理学的饮食和肿瘤上皮细胞的编程 Lgr5+或Bmi1+干细胞室。RNAScope和一种新的转录成像方法 测定Lgr5+和Bmi1+干细胞在肠道内环境稳定和肿瘤发生中的相对贡献 不同的饮食条件。

We have dissected impact of a western style purified diet (NWD1) on intestinal and colonic tumorigenesis. NWD1 recapitulates mouse intake of key nutrients each at their levels epidemiologically linked to higher risk for intestinal cancer in western developed countries. We established that NWD1 amplifies and accelerates intestinal tumors in many mouse genetic models, regardless of genetic etiology and aggressiveness driven by single or multiple genetic drivers. Moreover, when fed to wild-type mice long-term, we and others established that NWD1 causes sporadic intestinal and colon tumors. At least 1 year before these sporadic tumors develop, there is altered maturation of epithelial cells and underlying molecular pathways in the histopathologically normal mucosa. This led to investigating how the cell and molecular biology of intestinal stem cells are altered by the NWD1, We reported the surprising discovery that NWD1 compromises stem cell functions of Lgr5hi cells in intestinal homeostasis and tumorigenesis, but that this is compensated for by mobilization of Bmi1+ cells as stem cells for these functions. Moreover, genetic and dietary experiments identify decreased vitamin D exposure and Vdr signaling as important in these effects, consistent with Clevers' reported stem cell expression signature strongly implicating Vdr signaling in Lgr5hi stem cell functions. Further, we noted that mouse experiments on intestinal stem cells have used diets establishing vitamin D levels well above the range of that for the US population. This raises significant questions regarding which and how stem cells function in homeostasis and tumorigenesis. Extensive new data since the prior review (submitted for publication) establish profound effects of dietary exposures: i) on programming of Lgr5hi and Bmi1+ cells (RNAseq); ii) on capacity for DNA repair in these cells; iii) on subsequent extent, spectrum and signature of mutations that accumulate in cells of these crypt compartments (sequencing of FACS isolated single cells). Aim 1 addresses the hypothesis that dietary induced changes in Lgr5hi stem cell programming, function, and epigenetic alterations can be exceptionally long lived, and therefore that stem cells can be used as “sentinel cells” to monitor the ability of dietary intervention to modulate these alterations as potential markers of risk and targets for intervention. This will provide fundamental data regarding coordination of genes and pathways in regulation of stem cell function, identification of markers of relative risk, and how intervention strategies can be optimized. Aim2 of the initial submission has been largely accomplished, and Aim 2 is revised to investigate the hypothesis that altered DNA repair is important in differential dietary effects on tumorigenesis from the Lgr5+ and Bmi1+ populations when conditional Apc mutations are targeted specifically to each of these cell populations. Aim2 also determines the impact of diet on the nature of mutations in Apc and throughout the genome in the tumors that arise. Aim 3 determines impact of diet on tumor histopathology and programming of tumor epithelial cells when tumors are initiated from either the Lgr5+ or Bmi1+ stem cell compartments. RNAscope and a novel method of transcriptional imaging then determine relative contribution of Lgr5+ and Bmi1+ stem cells to intestinal homeostasis and tumorigenesis under different dietary conditions.