Research Article: Rapamycin increases oxidative stress response gene expression in adult stem cells

Date Published: April 18, 2012

Publisher: Impact Journals LLC

Author(s): Amber E. Kofman, Margeaux R. McGraw, Christopher J. Payne.

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Abstract

Balancing quiescence with proliferation is of paramount importance for adult stem cells in order to avoid hyperproliferation and cell depletion. In some models, stem cell exhaustion may be reversed with the drug rapamycin, which was shown can suppress cellular senescence in vitro and extend lifespan in animals. We hypothesized that rapamycin increases the expression of oxidative stress response genes in adult stem cells, and that these gene activities diminish with age. To test our hypothesis, we exposed mice to rapamycin and then examined the transcriptome of their spermatogonial stem cells (SSCs). Gene expression microarray analysis revealed that numerous oxidative stress response genes were upregulated upon rapamycin treatment, including superoxide dismutase 1, glutathione reductase, and delta-aminolevulinate dehydratase. When we examined the expression of these genes in 55-week-old wild type SSCs, their levels were significantly reduced compared to 3-week-old SSCs, suggesting that their downregulation is coincident with the aging process in adult stem cells. We conclude that rapamycin-induced stimulation of oxidative stress response genes may promote cellular longevity in SSCs, while a decline in gene expression in aged stem cells could reflect the SSCs’ diminished potential to alleviate oxidative stress, a hallmark of aging.

Partial Text

Cell senescence may contribute to adult stem cell exhaustion, compromising the maintenance of cell lineages within the body [1]. Recent evidence suggests that the cumulative exposure to reactive oxygen species (ROS) and DNA damage can lead to the decline of adult stem cells both in population and in regenerative capacity. For example, hematopoietic stem cells (HSCs) from mice lacking forkhead box O (FOXO) family transcription factors exhibit higher levels of ROS, accompanied by short-term hyperproliferation that is then followed by increased apoptosis that depletes the HSC pool [2, 3]. Epithelial stem cells in the epidermis (ESCs) that are engineered to constitutively transduce wingless-related MMTV integration site (WNT) signals in mice rapidly divide in the short term, but then undergo cell senescence and disappear from the ESC compartment [4]. Neural stem cells (NSCs), meanwhile, decline in number and function within the subventricular zone of lateral ventricles in the aging mouse brain due to genomic instability and upregulated cyclin-dependent kinase inhibitor 2a (Cdkn2a; p16Ink4a), which activates DNA-damage response pathways that induce apoptosis or senescence [5, 6]. Spermatogonial stem cells (SSCs) exhibit a loss of regenerative ability during aging in vivo and in vitro, with the downregulation of several genes important for self-renewal [7-9, 10].

The maintenance of adult stem cells, including SSCs, is critical to ensure the continuous production of differentiated cells within that lineage as organisms age. Mouse SSC self-renewal is promoted through GDNF signaling and mTORC1 antagonism by the PLZF-mediated activation of Redd1 [12]. Chronic exposure of mouse testes to rapamycin expands the SSC pool in vivo and increases Gfra1 and Ret expression [12]. The present study demonstrated that along with Gfra1 and Ret, additional SSC self-renewal genes (Lin28b, Nanos 2, Foxo1) and oxidative stress response genes (Alad, Sod1, Gsr) are upregulated in rapamycin-exposed SSCs. LIN28B suppresses microRNA biogenesis through interactions with the let-7 precursor, and is enriched in undifferentiated germ cells within the testis [32]. The functional role of LIN28B in SSCs is not yet clear, but the protein exhibits a striking temporal co-expression in germ cells with PLZF, suggesting a possible regulatory association with this transcription factor (unpublished observations). NANOS2 is an RNA-binding protein that acts downstream of GFRA1 to promote SSC self-renewal, and is required for stem cell maintenance [33, 34]. FOXO1, a transcription factor, regulates the expression of Ret and other genes in SSCs and is required for their homeostasis [35]. Collectively, these findings identify a transcriptional network that is enhanced when mTORC1 is inhibited by rapamycin.

 

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