RNA sequencing of slow-and fast-cycling epidermal stem cells vs hair follicle stem cells in young vs old mice
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ABSTRACT: This study was performed to compare transcriptomic changes in the heterogeneous mouse skin epidermal stem cells and hair follicle stem cells (HFSC) populations during chronological aging. Slow-cycling stem cells (label retaining cells, LRCs), fast-cycling stem cells (non-label retaining cells, nLRCs) and hair follicle stem cells express unique gene signatures in young age (2 months old) and have independent stem cell identities. The changes in aging stem cells lineage identities have been a topic of discussion and here we examined if distinct stem cells cycling speed affects their aging process by comparing the transcriptomes of slow-and fast-cycling epidermal stem cells. Our data indicates the loss of unique stem cell identities in aging slow or fast-cycling epidermal stem cells or HFSC at 2 years of age with intermediary effects seen at 1.5 year old.
Project description:This study was performed to compare the transcriptome of epidermal stem cells in young (2 months old) and middle aged (1 year old) fibulin 7 wild type (WT) compared to fibulin 7 knockout (KO) mice. Fbln7 gene was an ECM gene significantly upregulated in aging fast-cycling stem cells and this RNA-sequencing was done to investigate the role of Fbln7 in aging epidermal stem cells. Data indicates differences in gene expressions occuring between WT and KO at 1 year old but not at 2 months old, suggesting an age dependent role of fibulin 7. At 1 year old, other matrix-related and remodelling genes were affected, together with inflammatory responses of stem cells and lineage fate abberations, as in the case of 2 years old fast-cycling epidermal stem cells.
Project description:The extracellular matrix (ECM) plays a crucial role in building the extracellular environment and translating extracellular information into biochemical signals that sustain organ functions. Fibulin-5 is a multifunctional ECM protein essential for the formation of elastic fibers and the regulation of cellular functions through integrin binding. Fibulin-5 expression decreases with aging in human skin; however, its functional significance remains unknown. To address the roles of fibulin-5 in regulating epidermal stem cells during skin aging, Fbln5 knockout mice were examined for changes in their cellular and molecular phenotypes. Loss of Fbln5 in mice results in early impairments of epidermal stem cell properties, similar to the chronological aging of the skin. Fibulin-5 deficiency results in the suppression of integrins and other cell junctional molecules, leading to the inactivation of YAP signaling in epidermal stem cells. The reduced YAP signal is associated with the down-regulation of the fast-cycling epidermal stem cell marker, SLC1A3, in human skin and primary keratinocytes. These findings underscore the important role of fibulin-5 in governing the balance of epidermal stem cell populations during skin aging via crosstalk between the extracellular environment and intracellular signaling.
Project description:Reprogramming somatic cells to induced pluripotency by Yamanaka factors is usually slow and inefficient, and is thought to be a stochastic process. We identified a privileged somatic cell state, from which acquisition of pluripotency could occur in a non-stochastic manner. Subsets of murine hematopoietic progenitors are privileged, whose progeny cells predominantly adopt the pluripotent fate with activation of endogenous Oct4 locus after 4-5 divisions in reprogramming conditions. Privileged cells display an ultrafast cell cycle of ~8 hours. In fibroblasts, a subpopulation cycling at a similar ultrafast speed is observed after 6 days of factor expression, and is increased by p53-knockdown. This ultrafast-cycling population accounts for >99% of the bulk reprogramming activity in wildtype or p53-knockdown fibroblasts. We compared the transcriptomes of the fast cycling cells with those of slower hematopoietic progenitors, bulk fibroblasts and established iPS cells. 3-5 replicates for each of the six cell types were included: 4 replicates for established iPS cells, 4 replicates for bulk mouse embryonic fibroblasts (MEF), 4 replicates for fast cycling MEF, 4 replicates for slow cycling MEF, 5 replicates for fast cycling granulocyte monocyte progenitors (GMP) and 3 replicates for slow cycling GMP.
Project description:Human epidermal stem cells transit from a slow cycling to an actively proliferating state to contribute to homeostasis. Both stem cell states differ in their cell cycle profiles but must remain guarded from differentiation and senescence. Here we show that Cbx4, a Polycomb Repressive Complex-1 (PRC1)-associated protein, maintains human epidermal stem cells slow-cycling and undifferentiated, while protecting them from senescence. Interestingly, abrogating the polycomb activity of Cbx4 impairs its anti-senescent function without affecting stem cell differentiation, indicating that differentiation and senescence are independent processes in human epidermis. Conversely, Cbx4 inhibits stem cell activation and differentiation through its SUMO ligase activity. Global transcriptome and chromatin occupancy analyses indicate that Cbx4 regulates modulators of epidermal homeostasis and represses factors, such as Ezh2, Dnmt1, and Bmi1, to prevent the activate stem cell state. Our results suggest that distinct Polycomb complexes balance epidermal stem cell dormancy and activation, while continually preventing senescence and differentiation.
Project description:Human epidermal stem cells transit from a slow cycling to an actively proliferating state to contribute to homeostasis. Both stem cell states differ in their cell cycle profiles but must remain guarded from differentiation and senescence. Here we show that Cbx4, a Polycomb Repressive Complex-1 (PRC1)-associated protein, maintains human epidermal stem cells slow-cycling and undifferentiated, while protecting them from senescence. Interestingly, abrogating the polycomb activity of Cbx4 impairs its anti-senescent function without affecting stem cell differentiation, indicating that differentiation and senescence are independent processes in human epidermis. Conversely, Cbx4 inhibits stem cell activation and differentiation through its SUMO ligase activity. Global transcriptome and chromatin occupancy analyses indicate that Cbx4 regulates modulators of epidermal homeostasis and represses factors, such as Ezh2, Dnmt1, and Bmi1, to prevent the activate stem cell state. Our results suggest that distinct Polycomb complexes balance epidermal stem cell dormancy and activation, while continually preventing senescence and differentiation.
Project description:We identified slow-cycling cells (SCCs) in Ewing sarcoma using a label retention assay with CFSE. We labeled cells of SK-ES-1, an Ewing sarcoma cell line, with CFSE. After 5 days culture, we isolated cells retaining strong fluorescence (upper, ~10%) as SCCs and other cells (lower, ~90%) as non-slow-cycling cells (non-SCCs) using FACS AriaTM Ⅲ cell sorter.
Project description:Hair loss is one of the typical aging phenotypes in mammals, yet the underlying mechanism(s) is unclear. Here we report that hair follicle stem cell (HFSC) aging causes the stepwise miniaturization of hair follicles and eventual hair loss both in wild-type mice and in humans. In vivo fate analysis of HFSCs revealed that the DNA damage response in HFSCs causes proteolysis of Type XVII Collagen (COL17A1/BP180) to trigger “HFSC aging”, characterized by their loss of stemness signature and epidermal commitment. Those aged HFSCs are cyclically eliminated from the skin through their terminal epidermal differentiation, thereby causing hair follicle miniaturization. That process can be recapitulated by Col17a1 deficiency and prevented by forced maintenance of COL17A1 in HFSCs, demonstrating that stem cell homeostasis is the keystone against ultimate execution of the tissue/organ aging program.