Browsing by Subject "Receptors, Notch"
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Item Identification and Characterization of Non Small Cell Lung Cancer Stem Cells(2011-02-01) Sullivan, James Patrick; Minna, John D.The discovery of rare tumor cells with stem cell features first in myeloproliferative disease and later in solid tumors has emerged as an important area in cancer research. Through these studies the cancer stem cell model has emerged, which postulates that many tumors are initiated and progressed by a population of self-renewing malignant stem cells, referred to as cancer stem cells. This new tumor growth paradigm suggests that tumor metastasis and recurrence may be driven by a residual population of highly aggressive cancer stem cells. Furthermore this model argues that complete cancer remission may only be achieved by eradicating the malignant stem cell population charged as the source of tumor cell renewal. Lung cancer is the most commonly lethal form of cancer in the world with about 90% of the nearly one million new cases succumbing to the disease. While progress is being made in understanding lung cancer pathogenesis and improving therapy, prognosis remains poor. One approach to improving outcome in lung cancer has been to therapeutically target a unique, phenotypically defined lung cancer stem cell population. However despite the relatively rapid pace of cancer stem cell research in solid tumors such as breast, brain and colon cancers, similar progress in lung cancer remains hampered in part due to an incomplete understanding of lung stem cell hierarchy and the complex heterogeneity of the disease. To address this challenge, putative lung cancer stem cells were prospectively isolated from patient lung tumors and lung tumor cell lines using methods that have been reported to enrich for other stem cell populations in other cancers. As a result, a subpopulation of cells with elevated aldehyde dehydrogenase (ALDH) activity within many NSCLCs was identified with properties indicative of a cancer stem cell population including enhanced tumorigenicity in xenograft models, clonogenicity in culture and the capacity for self-renewal. In support of this, analysis of 282 clinically annotated non small cell lung cancer samples found elevated ALDH1A1 expression, the protein that drives ALDH in lung cancer, was associated with poor patient prognosis. Finally, molecular characterization of isolated ALDH+ lung cancer cells revealed elevated expression of stem cell transcripts including Notch signaling transcripts, suggesting enhanced pathway activity. Suppression of Notch signaling through chemical inhibition or knockdown of the proto-oncogene NOTCH3 resulted in a significant reduction in clonogenic ALDH+ cells indicating the importance of Notch signaling in lung cancer stem cell homeostasis and as a potential target for lung cancer stem cell directed therapy.Item The Role of Notch1 in Adult Hippocampal Neurogenesis and Function(2009-09-04) Ables, Jessica Lynn; Eisch, Amelia J.Neurogenesis occurs throughout life in the hippocampal subgranular zone (SGZ) and is potently stimulated by exercise, but the underlying mechanisms are still poorly defined. Notch1 is a master regulator of developmental neurogenesis, yet its role in adult hippocampal neurogenesis is unclear. To test the hypothesis that cell-intrinsic Notch1 is critical to both basal and exercise-induced SGZ neurogenesis, we generated Nestin-creERT2/R26R-YFP/Notch1loxP/loxP (Notch1 iKO) mice to inducibly ablate Notch1 in Nestin-expressing stem and progenitor SGZ cells. The total number of YFP+ SGZ cells increased over time in wild type littermates, but not in Notch1 iKO mice. Morphological and phenotypic analyses revealed that fewer YFP+ DG neurons were generated over time in Notch1 iKO mice due to smaller pools of YFP+ stem-like and progenitor cells. Likewise, neural progenitors isolated from Notch1 iKO mice were incapable of forming new neurospheres with extended passaging. While non-running Notch1 iKO mice had fewer YFP+ SGZ cells relative to wild type littermates, Notch1 iKO mice given 30 days access to a running wheel had equal number of YFP+ SGZ cells relative to controls, suggesting that running rescued total YFP+ SGZ cell number independent of Notch1. However, running did not rescue YFP+ stem-like cell number in Notch1 iKO mice, suggesting that the putative stem-like SGZ cells make little contribution to adult hippocampal neurogenesis in these conditions. From these data, we conclude that Notch1 in Nestin+ stem and progenitor cells is critical to maintain basal adult hippocampal neurogenesis, but is not critical for exercise-induced neurogenesis. Neurogenesis has also been implicating in depression and behavioral response to antidepressants. To determine if reduced neurogenesis contributed to depression- or anxiety-related behavior, we assessed several measures of depression and anxiety in Notch1 iKO mice. We found that Notch1 iKO mice did not differ from WT mice in their behavior, suggesting that reduced neurogenesis is not associated with mood disturbances.Item Spatial-Temporal Regulation of the Atonal Homolog 1 Gene(2007-12-17) Adams, Chris Aries; Johnson, Jane E.Controlled spatio-temporal expression of atonal homolog 1 is necessary for the correct development of the cerebellar granule cells, the dorsal interneuron 1 population, gut goblet cells and the cochlear and vestibular hair cells of mice. The purpose of these studies was to determine how atonal homolog 1 is regulated by analyzing the activity of atonal homolog 1 regulatory regions. This was accomplished by deleting different conserved regions in a bacterial artificial chromosome (318GFPBAC) containing ~180 kb of sequence 5' and 3' of atonal homolog 1 and assaying for green fluorescent protein expression in transgenic mice. Transgenic embryos were analyzed at embryonic day 10.5 in the neural tube, metencephalon and rhombencephalon and at embryonic day 16.5 in merkel cells, inner ear cells, vestibular cells and the cerebellum all tissues where endogenous atonal homolog 1 is expressed. Auto-regulation from an enhancer region 3' of the atonal homolog 1 gene (enhancer AB) was previously shown to be sufficient for atonal homolog 1-specific expression. In this paper, I show that enhancer AB is required for expression from the 318GFPBAC indicating that there are no other auto-regulatory elements for atonal homolog 1 expression in the 200 kb tested. Further, I have located an evolutionarily conserved region that has not previously been tested 12 kb 3' of the atonal homolog 1 coding region (enhancer C). When enhancer C is deleted from the 318GFPBAC, green fluorescent protein expression is not affected at embryonic day 10.5 or embryonic day 16.5. Also, enhancer C cannot drive green fluorescent protein expression by itself at embryonic day 10.5 or embryonic day 16.5. Thus no role for this conserved sequence around atonal homolog 1 was detected in these studies. To date, the AB enhancer is the only sequence shown in vivo to function in atonal homolog 1 regulation being both necessary and sufficient to direct expression in an atonal homolog 1 pattern.