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Molecular regulation underlying differentiation of dental pulp stem cells

牙髓干细胞分化的分子调控

基本信息

批准号：
8812561
负责人：
Shaomian Yao
金额：
$ 36.22万
依托单位：
LOUISIANA STATE UNIV A&M COL BATON ROUGE
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-09-01 至 2020-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8812561
关键词：
3&apos Untranslated Regions Adult Binding Bioinformatics Bone Marrow Stem Cell Transplantation Cell Separation Cell Therapy Cells Code Dental Dental Pulp Development Disease Down-Regulation Elements Ethical Issues Fluorescence Future Gene Expression Gene Proteins Genes Global Change Goals Heat shock proteins Immune response In Vitro Injury Late Effects Light Medicine Messenger RNA Methods MicroRNAs Molecular Natural regeneration Northern Blotting Organ Patients Play Polyadenylation Porifera Property Proteins Proteome Rattus Regenerative Medicine Regulation Regulator Genes Regulatory Element Research Response Elements Reverse Transcriptase Polymerase Chain Reaction Role Signal Transduction Small RNA Source Stem cells Teratoma Testing Therapeutic Time Tissue Engineering Tissues Tooth structure Transfection Translations Transplantation Two-Dimensional Gel Electrophoresis Western Blotting adult stem cell design embryonic stem cell expression vector gain of function human disease improved induced pluripotent stem cell leukemia loss of function novel novel strategies overexpression prevent public health relevance repaired stem stem cell differentiation stemness tissue regeneration tumorigenesis two-dimensional vector

项目摘要

DESCRIPTION (provided by applicant): Utilization of stem cells to repair diseased or damaged tissues/organs represents a novel approach for remedies in regenerative medicine. Although embryonic stem cells (ESC) and induced pluripotent stem cells (iPS) have been studied, there are major drawbacks and concerns in using ESC and iPS; for example, tumorigenesis and teratomas formation can occur after their transplantation. It is not clear if the obstacles pertaining to ESC and iPS can be overcome in order to use them for therapeutics; at least there is still long way to go. In contrast, adult stem cells (AdSC) are used for treatment of human diseases. One example is the transplantation of bone marrow stem cells to cure or improve certain disorders, such as leukemia. Such cell- based therapies usually require large numbers of stem cells. Because AdSC lose their stem cell properties (stemness) including differentiation capability when cultured in vitro, primary isolated AdSC can only be used for limited passages, and propagation of a large quantity of high-potential stem cells from primary isolation is difficult. Thus, large amounts of tissues are needed for stem cell isolation, and continuous isolation of AdSC is required for multiple treatments. However, tissues, e.g., dental tissues that can be used for AdSC isolation are often limited. Stem cells isolated from dental pulp of extracted and exfoliated teeth have been shown to be a good source of stem cells for regenerative medicine. However, like other AdSC, these dental pulp stem cells (DPSCs) also lose their differentiation potential when cultured in vitro, which hampers their applications. The long-term goal of this research is to elucidate the genes and molecules regulating differentiation capability in the DPSCs. Preliminary studies showed that heat shock protein B8 (HspB8) was downregulated in cultured DPSCs when the cells lost differentiation capability, and knockdown of HspB8 expression in the early passage DPSCs caused the cells to lose differentiation capability. Further studies suggested that microRNAs (miRNAs) are responsible for downregulation of HspB8 expression in DPSCs. The central hypothesis is that upregulated expression of certain miRNAs in late passage DPSCs downregulates HspB8 and other regulatory genes, and in turn causes DPSCs to lose differentiation capability during in vitro culture. The project Aims: (1) to identify the miRNAs whose expression is substantially increased in late passage DPSCs, designated as late-passage upregulated miRNAs (LPU-miRNAs); (2) to identify the LPU-miRNAs that downregulate HspB8 expression; i.e., HspB8-targeting miRNAs, and (3) to study the effects of LPU-miRNAs and HspB8-targeting miRNAs on DPSC differentiation and on regulating other potential genes. Accomplishment of the Aims would be significant in understanding the roles of miRNAs and HspB8 in regulating the loss of differentiation seen in the long-term cultured DPSCs and will shed light on other AdSC as well. This would facilitate the development of cutting- edge methods for maintaining the stem cell properties in culturing DPSCs and AdSC, such that a large quantity of stem cells with high-differentiation potential can be obtained from expansion of primary isolated stem cells.

描述(由申请人提供)：利用干细胞修复病变或受损的组织/器官是再生医学中一种新的治疗方法。虽然胚胎干细胞(ESC)和诱导多能干细胞(IPS)已经被研究，但使用ESC和iPS存在着主要的缺陷和担忧，例如，它们移植后可能会发生肿瘤和畸胎瘤的形成。目前还不清楚是否有关ESC和iPS的障碍是可以克服的，以便将它们用于治疗；至少还有很长的路要走。相比之下，成体干细胞(ADSC)用于治疗人类疾病。一个例子是骨髓干细胞移植，以治愈或改善某些疾病，如白血病。这种基于细胞的疗法通常需要大量的干细胞。由于ADSC在体外培养时失去了包括分化能力在内的干细胞特性(干性)，原代分离的ADSC只能用于有限的传代，从原代分离的高潜能干细胞很难大量繁殖。因此，干细胞分离需要大量的组织，多次治疗需要连续分离ADSC。然而，可用于ADSC分离的组织，例如牙齿组织，通常是有限的。从拔除和剥离的牙齿的牙髓中分离出的干细胞已被证明是再生医学的良好干细胞来源。然而，与其他牙髓干细胞一样，这些牙髓干细胞在体外培养时也会失去分化潜能，这阻碍了它们的应用。本研究的长期目标是阐明DPSCs中调控分化能力的基因和分子。初步研究表明，当培养的DPSCs失去分化能力时，热休克蛋白B8(HspB8)的表达下调，在传代早期的DPSCs中HspB8的表达下调导致细胞失去分化能力。进一步的研究表明，microRNAs(MiRNAs)负责下调DPSCs中HspB8的表达。中心假说是，某些miRNAs在晚期传代的DPSCs中表达上调，下调HspB8和其他调控基因，进而导致DPSCs在体外培养过程中失去分化能力。本项目的目标是：(1)鉴定在晚期传代DPSCs中表达显著上调的miRNAs，命名为LPU-miRNAs(LPU-miRNAs)；(2)鉴定下调HspB8表达的LPU-miRNAs，即HspB8靶向miRNAs；(3)研究LPU-miRNAs和HspB8靶向miRNAs对DPSC分化和调控其他潜在基因的影响。这些目标的实现将对理解miRNAs和HspB8在调节长期培养的DPSCs分化缺失中的作用具有重要意义，也将为其他ADSC提供借鉴。这将有助于开发在培养DPSCs和ADSC时保持干细胞特性的尖端方法，以便通过原代分离干细胞的扩增获得大量具有高分化潜力的干细胞。