Inhibition of Cdc42 activity extends lifespan and decreases circulating inflammatory cytokines in aged female C57BL/6 mice

Cdc42 is a small RhoGTPase regulating multiple functions in eukaryotic cells. The activity of Cdc42 is significantly elevated in several tissues of aged mice, while the Cdc42 gain-of-activity mouse model presents with a premature aging-like phenotype and with decreased lifespan. These data suggest a causal connection between elevated activity of Cdc42, aging, and reduced lifespan. Here, we demonstrate that systemic treatment of aged (75-week-old) female C57BL/6 mice with a Cdc42 activity-specific inhibitor (CASIN) for 4 consecutive days significantly extends average and maximum lifespan. Moreover, aged CASIN-treated animals displayed a youthful level of the aging-associated cytokines IL-1β, IL-1α, and INFγ in serum and a significantly younger epigenetic clock as based on DNA methylation levels in blood cells. Overall, our data show that systemic administration of CASIN to reduce Cdc42 activity in aged mice extends murine lifespan

The Spindle Assembly Checkpoint Is Required for Hematopoietic Progenitor Cell Engraftment

Summary: The spindle assembly checkpoint plays a pivotal role in preventing aneuploidy and transformation. Many studies demonstrate impairment of this checkpoint in cancer cells. While leukemia is frequently driven by transformed hematopoietic stem and progenitor cells (HSPCs), the biology of the spindle assembly checkpoint in such primary cells is not very well understood. Here, we reveal that the checkpoint is fully functional in murine progenitor cells and, to a lesser extent, in hematopoietic stem cells. We show that HSPCs arrest at prometaphase and induce p53-dependent apoptosis upon prolonged treatment with anti-mitotic drugs. Moreover, the checkpoint can be chemically and genetically abrogated, leading to premature exit from mitosis, subsequent enforced G1 arrest, and enhanced levels of chromosomal damage. We finally demonstrate that, upon checkpoint abrogation in HSPCs, hematopoiesis is impaired, manifested by loss of differentiation potential and engraftment ability, indicating a critical role of this checkpoint in HSPCs and hematopoiesis. : In this report, Andreas Brown and colleagues demonstrate that cycling hematopoietic stem and progenitor cells activate the spindle assembly checkpoint as a response to anti-mitotic stress. The authors further reveal that inhibition of the checkpoint causes impairment of progenitor function, whereas it appears to be less crucial for hematopoietic stem cells. Keywords: mitotic checkpoint, spindle assembly checkpoint, hematopoietic stem cells, hematopoietic progenitor cells, genome stability, hematopoiesis, SAC, reversine, micronucle

Increased hematopoietic stem cell mobilization in aged mice

Hematopoietic stem and progenitor cells (HSPCs) are located in the bone marrow in close association with a highly organized 3-dimensional structure formed by stroma cells, referred to as the niche. Mobilization of HSPCs from bone marrow to peripheral blood in response to granulocyte colony-stimulating factor (G-CSF) requires de-adhesion of HSPCs from the niche. The influence of aging of HSPCs on cell-stroma interactions has not been determined in detail. Using a mouse model of G-CSF–induced mobilization, we demonstrated that the ability to mobilize hematopoietic stem cells is approximately 5-fold greater in aged mice. Competitive mobilization experiments confirmed that enhanced mobilization ability was intrinsic to the stem cell. Enhanced mobilization efficiency of primitive hematopoietic cells from aged mice correlated with reduced adhesion of hematopoietic progenitor cells to stroma and with elevated levels of GTP-bound Cdc42. These results might indicate that stroma–stem cell interactions are dynamic over a lifetime and result in physiologically relevant changes in the biology of primitive hematopoietic cells with age

Canonical Wnt Signaling Ameliorates Aging of Intestinal Stem Cells

Although intestinal homeostasis is maintained by intestinal stem cells (ISCs), regeneration is impaired upon aging. Here, we first uncover changes in intestinal architecture, cell number, and cell composition upon aging. Second, we identify a decline in the regenerative capacity of ISCs upon aging because of a decline in canonical Wnt signaling in ISCs. Changes in expression of Wnts are found in stem cells themselves and in their niche, including Paneth cells and mesenchyme. Third, reactivating canonical Wnt signaling enhances the function of both murine and human ISCs and, thus, ameliorates aging-associated phenotypes of ISCs in an organoid assay. Our data demonstrate a role for impaired Wnt signaling in physiological aging of ISCs and further identify potential therapeutic avenues to improve ISC regenerative potential upon aging

Aging alters the epigenetic asymmetry of HSC division.

Hematopoietic stem cells (HSCs) balance self-renewal and differentiation to maintain homeostasis. With aging, the frequency of polar HSCs decreases. Cell polarity in HSCs is controlled by the activity of the small RhoGTPase cell division control protein 42 (Cdc42). Here we demonstrate-using a comprehensive set of paired daughter cell analyses that include single-cell 3D confocal imaging, single-cell transplants, single-cell RNA-seq, and single-cell transposase-accessible chromatin sequencing (ATAC-seq)-that the outcome of HSC divisions is strongly linked to the polarity status before mitosis, which is in turn determined by the level of the activity Cdc42 in stem cells. Aged apolar HSCs undergo preferentially self-renewing symmetric divisions, resulting in daughter stem cells with reduced regenerative capacity and lymphoid potential, while young polar HSCs undergo preferentially asymmetric divisions. Mathematical modeling in combination with experimental data implies a mechanistic role of the asymmetric sorting of Cdc42 in determining the potential of daughter cells via epigenetic mechanisms. Therefore, molecules that control HSC polarity might serve as modulators of the mode of stem cell division regulating the potential of daughter cells

TNF-α induces leukemic clonal evolution ex vivo in Fanconi anemia group C murine stem cells

The molecular pathogenesis of the myeloid leukemias that frequently occur in patients with Fanconi anemia (FA) is not well defined. Hematopoietic stem cells bearing inactivating mutations of FA complementation group C (FANCC) are genetically unstable and hypersensitive to apoptotic cytokine cues including IFN-γ and TNF-α, but neoplastic stem cell clones that arise frequently in vivo are resistant to these cytokines. Reasoning that the combination of genetic instability and cytokine hypersensitivity might create an environment supporting the emergence of leukemic stem cells, we tested the leukemia-promoting effects of TNF-α in murine stem cells. TNF-α exposure initially profoundly inhibited the growth of Fancc–/– stem cells. However, longer-term exposure of these cells promoted the outgrowth of cytogenetically abnormal clones that, upon transplantation into congenic WT mice, led to acute myelogenous leukemia. TNF-α induced ROS-dependent genetic instability in Fancc–/– but not in WT cells. The leukemic clones were TNF-α resistant but retained their characteristic hypersensitivity to mitomycin C and exhibited high levels of chromosomal instability. Expression of FANCC cDNA in Fancc–/– stem cells protected them from TNF-α–induced clonal evolution. We conclude that TNF-α exposure creates an environment in which somatically mutated preleukemic stem cell clones are selected and from which unaltered TNF-α–hypersensitive Fancc–/– stem cells are purged

Retroviral vector insertion sites associated with dominant hematopoietic clones mark “stemness” pathways

Evidence from model organisms and clinical trials reveals that the random insertion of retrovirus-based vectors in the genome of long-term repopulating hematopoietic cells may increase self-renewal or initiate malignant transformation. Clonal dominance of nonmalignant cells is a particularly interesting phenotype as it may be caused by the dysregulation of genes that affect self-renewal and competitive fitness. We have accumulated 280 retrovirus vector insertion sites (RVISs) from murine long-term studies resulting in benign or malignant clonal dominance. RVISs (22.5%) are located in or near (up to 100 kb [kilobase]) to known proto-oncogenes, 49.6% in signaling genes, and 27.9% in other or unknown genes. The resulting insertional dominance database (IDDb) shows substantial overlaps with the transcriptome of hematopoietic stem/progenitor cells and the retrovirus-tagged cancer gene database (RTCGD). RVISs preferentially marked genes with high expression in hematopoietic stem/progenitor cells, and Gene Ontology revealed an overrepresentation of genes associated with cell-cycle control, apoptosis signaling, and transcriptional regulation, including major “stemness” pathways. The IDDb forms a powerful resource for the identification of genes that stimulate or transform hematopoietic stem/progenitor cells and is an important reference for vector biosafety studies in human gene therapy