Aging and Reprogramming of Somatic Cells to Pluripotency

体细胞的衰老和重编程至多能性

• 批准号: 7907777
• 负责人: PETER J HORNSBY
• 金额: $ 15.07万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-15 至 2012-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7907777
• 关键词: Abnormal Cell; Adult; Age; Aging; Aging-Related Process; Animals; Back; Basic Science; Biopsy; Callithrix; Cell Aging; Cell Lineage; Cell Therapy; Cell surface; Cells; Characteristics; Chromatin; Coculture Techniques; Defect; Development; Disease; Elderly; Environment; Exhibits; Fibroblasts; Future; Genes; Germ Lines; Gold; Haplorhini; Health; Health Status; Housing; Human; Immunodeficient Mouse; Immunohistochemistry; In Vitro; Individual; Literature; Living Donors; Longevity; Malignant - descriptor; Mesenchymal; Modeling; Monkeys; Morphology; Mus; Nature; Newborn Infant; Organ; Parkinson Disease; Patients; Predisposition; Primates; Property; Skin; Somatic Cell; Subfamily lentivirinae; System; Telomerase; Teratoma; Testing; Therapeutic Effect; Tissues; Transplantation; Work; age effect; age related; c-myc Genes; dopaminergic neuron; embryonic stem cell; frailty; in vivo; induced pluripotent stem cell; insight; neuron development; nonhuman primate; novel; pluripotency; public health relevance; transcription factor

DESCRIPTION (provided by applicant): Aging alters the properties of somatic cells, resulting in declines in tissue and organ function. Reprogramming of somatic cells to induced pluripotent stem cells (iPS cells) provides the opportunity to test whether aging at the cell level is reversible and whether the changes in cell properties that are caused by aging can be erased and the cell returned to a completely pluripotent state. A novel aspect of this proposal is that our reprogramming studies will be performed using cells from a small nonhuman primate, the marmoset. Marmosets have the shortest life span of any anthropoid primate; therefore, they can more efficiently be used to compare reprogramming in early- versus late-life donors. In a future development of the work proposed here, reprogrammed/re-differentiated cells can be transplanted back into the same individual animal from which the somatic cells were derived, thus providing a "gold standard" test of the ability of reprogrammed/redifferentiated cells to function normally in an in vivo environment. Specific Aim 1: We hypothesize that aging will impair reprogramming by transcription factors, so that cells derived from older donors will exhibit a greater tendency to be misprogrammed and less ability to be correctly reprogrammed to a true pluripotent state. This will be tested by the retroviral introduction of reprogramming genes (Oct4, Sox2, Klf4, with or without c-Myc) into skin fibroblasts from marmosets of a range of ages, newborn to very old (~13 years). We will examine reprogramming in resultant cell colonies; defects in reprogramming may be observed as an aging-related increase in misprogramming, producing cells that lack pluripotency and which may have acquired malignant properties instead. If this is correct, future work will aim to elucidate those aging processes that interfere with reprogramming. However, we also anticipate that even at very old age some cells will be correctly reprogrammed to a pluripotent state. Specific Aim 2: We hypothesize that cells that have been correctly reprogrammed to pluripotency, even from old donors, will be able to properly redifferentiate along defined lineages when exposed to appropriate in vitro environments. We propose to induce them to differentiate to dopaminergic neurons. Proper differentiation will be assessed by development of neuronal morphology and expression of dopaminergic neuron-specific markers. If defects in redifferentiation or abnormal cell properties emerge as characteristics of iPS cells formed from old donors, further studies will focus on the nature of these aging-related changes that interfere with redifferentiation. These studies provide the necessary basic science that supports future patient-specific cell therapy. In the marmoset model, we can determine whether cells can be derived from skin biopsies, subjected to reprogramming/redifferentiation, and then safely transplanted back into the donor to exert a desired therapeutic effect, including therapies of late-life diseases such as Parkinson's disease. PUBLIC HEALTH RELEVANCE: Project Narrative Aging causes many changes in cells that eventually result in declines in body function, frailty, and increased susceptibility to diseases. The possibility that these cellular aging changes are reversible has been raised by recent studies that show that it is possible to take skin cells from adults and reprogram them to an embryonic stem cell-like state. In this proposal we will test the effects of aging on the ability to reprogram skin cells from marmoset monkeys, in which there is the potential of retransplanting embryonic stem cell-like cells back into the donor animal to test a therapeutic effect.

描述(申请人提供)：衰老会改变体细胞的特性，导致组织和器官功能的下降。将体细胞重新编程为诱导多能干细胞(iPS细胞)提供了测试细胞水平的衰老是否可逆以及老化引起的细胞特性变化是否可以消除并使细胞恢复到完全多能状态的机会。这项提议的一个新方面是，我们的重新编程研究将使用一种小型非人灵长类动物--绒猴的细胞进行。在所有类人灵长类动物中，绒猴的寿命最短；因此，它们可以更有效地用于比较早期捐赠者和晚期捐赠者的重编程。在这里提出的工作的未来发展中，重新编程/重新分化的细胞可以被移植回衍生体细胞的同一个体动物中，从而提供了重新编程/重新分化的细胞在体内环境中正常发挥作用的能力的“金标准”测试。具体目标1：我们假设衰老将损害转录因子的重编程，因此来自老年捐赠者的细胞将表现出更大的错误编程倾向，并且更不能正确地重编程为真正的多能性状态。这将通过逆转录病毒将重编程基因(Oct4、Sox2、Klf4，带有或不带有c-Myc)导入不同年龄的绒猴的皮肤成纤维细胞来进行测试，这些绒猴从新生儿到非常年长的(~13岁)。我们将研究结果细胞克隆中的重新编程；重新编程中的缺陷可能被观察到与年龄相关的错误编程增加，产生缺乏多能性的细胞，而这些细胞可能获得了恶性特性。如果这是正确的，未来的工作将致力于阐明那些干扰重新编程的衰老过程。然而，我们也预计，即使在非常年老的时候，一些细胞也会被正确地重新编程为多能状态。具体目标2：我们假设，被正确地重新编程为多能性的细胞，即使是来自老捐赠者的细胞，当暴露在适当的体外环境中时，也能够沿着确定的谱系正确地重新分化。我们建议诱导它们分化为多巴胺能神经元。适当的分化将通过神经元形态的发展和多巴胺能神经元特异性标志物的表达来评估。如果再分化缺陷或异常细胞特性作为由老年供者形成的iPS细胞的特征出现，进一步的研究将集中在这些干扰再分化的衰老相关变化的本质上。这些研究提供了必要的基础科学，支持未来针对患者的细胞治疗。在绒猴模型中，我们可以确定是否可以从皮肤活检组织中提取细胞，经过重新编程/重新分化，然后安全地移植回捐赠者体内，以发挥预期的治疗效果，包括治疗帕金森氏症等晚期疾病。公共卫生相关性：项目叙事衰老导致细胞发生许多变化，最终导致身体功能下降、虚弱和疾病易感性增加。最近的研究提出了这些细胞老化变化是可逆的可能性，这些研究表明，从成年人身上提取皮肤细胞并将其重新编程为胚胎干细胞状态是可能的。在这项提案中，我们将测试衰老对猴皮肤细胞重新编程能力的影响，在这种能力中，有可能将类似胚胎干细胞的细胞重新移植回捐赠者动物体内，以测试治疗效果。