Gene Expression Profiling of Adult Olfactory Stem and Progenitor Cells

描述(申请人提供)：嗅觉上皮(OE)在损伤后从解剖和功能上恢复到接近正常的能力，这种能力贯穿整个生命，在神经系统中是独一无二的。然而，我们对嗅茎及其下游祖细胞的分子表型几乎一无所知，这种无知严重阻碍了我们分离它们、了解它们的调节或以可控的方式使用它们的能力。我们正在提出探索性和发展性研究，旨在解决我们知识中的巨大差距。我们将分离出三种特殊类型的球状基底细胞(GBCs)，在其功能不同的群体中，发现了一种广泛的多能干细胞或干细胞样细胞：多潜能的GBCS、形成支持细胞的GBCS和转运性放大的GBCS。不同种类的GBCs将由它们在甲基溴损伤小鼠OE后重新出现的时间以及它们在包括Hes1和Mash1在内的一组碱性螺旋-环-螺旋(BHLH)转录因子中的表达来定义。表达bHLHTF的细胞将由FACS根据其表面标记将其定义为GBCs，并且GFP的表达与每个单独的TF平行(来自转基因或内源性基因座)。对于每个TF定义的群体，我们将进行Affymetrix微阵列分析，旨在描述其基因表达模式。此外，在正常OE中，有丝分裂静止和BrdU标记的GBCs是一个潜在的干细胞群体。我们也将用流式细胞仪分离它们，并对它们进行基因图谱分析。作为这些研究的结果，我们将对静止和激活的嗅觉干细胞及其下游前体细胞有一个全面的了解，并将准备开始对功能调节途径进行假说驱动的评估。OE的干细胞在成年人类中很容易获得，而且有一种天然的生成神经元的倾向。此外，嗅觉定位的胚胎细胞与成体干细胞在分子和功能上都很相似，可以形成嗅神经的特化胶质细胞，这可能会促进脊髓损伤后的功能恢复。在这里收集的信息可能最终使我们能够使用这些成体干细胞来治疗神经系统损伤和退行性疾病。

DESCRIPTION (provided by applicant): The capacity of the olfactory epithelium (OE) to recover both anatomically and functionally after injury to near-normal, a capacity that extends throughout life, is unique in the nervous system. However, we know next to nothing about the molecular phenotype of the olfactory stems nor their downstream progenitors, an ignorance that severely hampers our ability to isolate them, understand their regulation, or use them in a controllable manner. We are proposing exploratory and developmental studies designed to address that vast gap in our knowledge. We will isolate three specific types of globose basal cells (GBCs), among whose functionally heterogeneous population, a broadly pluripotent, stem or stem-like cell is found: multipotent GBCs, sustentacular cell- forming GBCs, and transit-amplifying GBCs. The different kinds of GBCs will be defined by the timing of their re-emergence after methyl bromide lesion of the mouse OE and by their expression of one among a group of basic Helix-Loop-Helix (bHLH) transcription factors that will include Hes1 and Mash1. The bHLH TF-expressing cells will be isolated by FACS on the basis of markers on their surface that define them as GBCs and the expression of GFP in parallel with each individual TF (from either a transgen or the endogenous gene locus). For each TF-defined population we will carry out AffyMetrix microarray analysis designed to profile its pattern of gene expression. In addition, mitotically quiescent and BrdU label-retaining GBCs in the normal OE are a potential stem cell population. We will isolate them by FACS as well and subject them to gene profiling. As a consequence of these studies, we will have a comprehensive understanding of resting and activated olfactory stem cells and their downstream progenitors and will be poised to begin hypothesis-driven assessments of functional regulatory pathways. The stem cells of the OE are both easily accessible in adult humans and have a natural propensity to make neurons. Furthermore, the embryonic cells of the olfactory placode, which the adult stem cells resemble in both molecular and functional terms, give rise to the specialized glia of the olfactory nerve, which may promote functional recovery after spinal cord injury. The information gathered here may allow us eventually to use these adult stem cells in treating nervous system damage and degenerative disease.