Hematopoietic stem cells (HSCs) undergo self-renewing cell divisions and maintain blood production for their lifetime. Appropriate control of HSC self-renewal is crucial for the maintenance of ...hematopoietic homeostasis. Here we show that activation of p38 MAPK in response to increasing levels of reactive oxygen species (ROS) limits the lifespan of HSCs in vivo. In Atm−/− mice, elevation of ROS levels induces HSC-specific phosphorylation of p38 MAPK accompanied by a defect in the maintenance of HSC quiescence. Inhibition of p38 MAPK rescued ROS-induced defects in HSC repopulating capacity and in the maintenance of HSC quiescence, indicating that the ROS-p38 MAPK pathway contributes to exhaustion of the stem cell population. Furthermore, prolonged treatment with an antioxidant or an inhibitor of p38 MAPK extended the lifespan of HSCs from wild-type mice in serial transplantation experiments. These data show that inactivation of p38 MAPK protects HSCs against loss of self-renewal capacity. Our characterization of molecular mechanisms that limit HSC lifespan may lead to beneficial therapies for human disease.
Cardiovascular disease (CVD), the most common morbidity resulting from atherosclerosis, remains a frequent cause of death. Efforts to develop effective therapeutic strategies have focused on vascular ...inflammation as a critical pathology driving atherosclerosis progression. Nonetheless, molecular mechanisms underlying this activity remain unclear. Here, we ask whether angiopoietin-like protein 2 (Angptl2), a proinflammatory protein, contributes to vascular inflammation that promotes atherosclerosis progression.
Histological analysis revealed abundant Angptl2 expression in endothelial cells and macrophages infiltrating atheromatous plaques in patients with cardiovascular disease. Angptl2 knockout in apolipoprotein E-deficient mice (ApoE(-/-)/Angptl2(-/-)) attenuated atherosclerosis progression by decreasing the number of macrophages infiltrating atheromatous plaques, reducing vascular inflammation. Bone marrow transplantation experiments showed that Angptl2 deficiency in endothelial cells attenuated atherosclerosis development. Conversely, ApoE(-/-) mice crossed with transgenic mice expressing Angptl2 driven by the Tie2 promoter (ApoE(-/-)/Tie2-Angptl2 Tg), which drives Angptl2 expression in endothelial cells but not monocytes/macrophages, showed accelerated plaque formation and vascular inflammation because of increased numbers of infiltrated macrophages in atheromatous plaques. Tie2-Angptl2 Tg mice alone did not develop plaques but exhibited endothelium-dependent vasodilatory dysfunction, likely because of decreased production of endothelial cell-derived nitric oxide. Conversely, Angptl2(-/-) mice exhibited less severe endothelial dysfunction than did wild-type mice when fed a high-fat diet. In vitro, Angptl2 activated proinflammatory nuclear factor-κB signaling in endothelial cells and increased monocyte/macrophage chemotaxis.
Endothelial cell-derived Angptl2 accelerates vascular inflammation by activating proinflammatory signaling in endothelial cells and increasing macrophage infiltration, leading to endothelial dysfunction and atherosclerosis progression.
Hematopoietic stem cells (HSCs) are maintained in an undifferentiated quiescent state within a bone marrow niche. Here we show that Foxo3a, a forkhead transcription factor that acts downstream of the ...PTEN/PI3K/Akt pathway, is critical for HSC self-renewal. We generated gene-targeted Foxo3a−/− mice and showed that, although the proliferation and differentiation of Foxo3a−/− hematopoietic progenitors were normal, the number of colony-forming cells present in long-term cocultures of Foxo3a−/− bone marrow cells and stromal cells was reduced. The ability of Foxo3a−/− HSCs to support long-term reconstitution of hematopoiesis in a competitive transplantation assay was also impaired. Foxo3a−/− HSCs also showed increased phosphorylation of p38MAPK, an elevation of ROS, defective maintenance of quiescence, and heightened sensitivity to cell-cycle-specific myelotoxic injury. Finally, HSC frequencies were significantly decreased in aged Foxo3a−/− mice compared to the littermate controls. Our results demonstrate that Foxo3a plays a pivotal role in maintaining the HSC pool.
Cell–cell and cell–extracellular matrix interactions between hematopoietic stem cells (HSCs) and their niches are critical for the maintenance of stem cell properties. Here, it is demonstrated that a ...cell adhesion molecule, N‐cadherin, is expressed in hematopoietic stem/progenitor cells (HSPCs) and plays a critical role in the regulation of HSPC engraftment. Furthermore, overexpression of N‐cadherin in HSCs promoted quiescence and preserved HSC activity during serial bone marrow (BM) transplantation (BMT). Inhibition of N‐cadherin by the transduction of N‐cadherin short hairpin (sh) RNA (shN‐cad) reduced the lodgment of donor HSCs to the endosteal surface, resulting in a significant reduction in long‐term engraftment. shN‐cad‐transduced cells were maintained in the spleen for six months after BMT, indicating that N‐cadherin expression in HSCs is specifically required in the BM. These findings suggest that N‐cadherin‐mediated cell adhesion is functionally essential for the regulation of HSPC activities in the BM niche.
Repeated cell divisions and aging impair stem cell function. However, the mechanisms by which this occurs are not fully understood. Here we show that protection of telomeres 1A (Pot1a), a component ...of the Shelterin complex that protects telomeres, improves haematopoietic stem cell (HSC) activity during aging. Pot1a is highly expressed in young HSCs, but declines with age. In mouse HSCs, Pot1a knockdown increases DNA damage response (DDR) and inhibits self-renewal. Conversely, Pot1a overexpression or treatment with POT1a protein prevents DDR, maintained self-renewal activity and rejuvenated aged HSCs upon ex vivo culture. Moreover, treatment of HSCs with exogenous Pot1a inhibits the production of reactive oxygen species, suggesting a non-telomeric role for Pot1a in HSC maintenance. Consistent with these results, treatment with exogenous human POT1 protein maintains human HSC activity in culture. Collectively, these results show that Pot1a/POT1 sustains HSC activity and can be used to expand HSC numbers ex vivo.Repeated cell divisions induce DNA damage in haematopoietic stem cells (HSC) and telomeres are sensitive to this damage. Here, the authors show in murine HSCs that the telomere binding protein POT1a inhibited the production of reactive oxygen species, and rejuvenated aged HSCs.
The 'ataxia telangiectasia mutated' (Atm) gene maintains genomic stability by activating a key cell-cycle checkpoint in response to DNA damage, telomeric instability or oxidative stress. Mutational ...inactivation of the gene causes an autosomal recessive disorder, ataxia-telangiectasia, characterized by immunodeficiency, progressive cerebellar ataxia, oculocutaneous telangiectasia, defective spermatogenesis, premature ageing and a high incidence of lymphoma. Here we show that ATM has an essential function in the reconstitutive capacity of haematopoietic stem cells (HSCs) but is not as important for the proliferation or differentiation of progenitors, in a telomere-independent manner. Atm-/- mice older than 24 weeks showed progressive bone marrow failure resulting from a defect in HSC function that was associated with elevated reactive oxygen species. Treatment with anti-oxidative agents restored the reconstitutive capacity of Atm-/- HSCs, resulting in the prevention of bone marrow failure. Activation of the p16INK4a-retinoblastoma (Rb) gene product pathway in response to elevated reactive oxygen species led to the failure of Atm-/- HSCs. These results show that the self-renewal capacity of HSCs depends on ATM-mediated inhibition of oxidative stress.
During postnatal life, the bone marrow (BM) supports both self-renewal and differentiation of hematopoietic stem cells (HSCs) in specialized microenvironments termed stem cell niches. Cell-cell and ...cell-extracellular matrix interactions between HSCs and their niches are critical for the maintenance of HSC properties. Here, we analyzed the function of N-cadherin in the regulation of the proliferation and long-term repopulation activity of hematopoietic stem/progenitor cells (HSPCs) by the transduction of N-cadherin shRNA. Inhibition of N-cadherin expression accelerated cell division in vitro and reduced the lodgment of donor HSPCs to the endosteal surface, resulting in a significant reduction in long-term engraftment. Cotransduction of N-cadherin shRNA and a mutant N-cadherin that introduced the silent mutations to shRNA target sequences rescued the accelerated cell division and reconstitution phenotypes. In addition, the requirement of N-cadherin for HSPC engraftment appears to be niche specific, as shN-cad–transduced lineage−Sca-1+c-Kit+ cells successfully engrafted in spleen, which lacks an osteoblastic niche. These findings suggest that N-cad–mediated cell adhesion is functionally required for the establishment of hematopoiesis in the BM niche after BM transplantation.
Prostaglandin E2 (PGE2) regulates hematopoietic stem/progenitor cell (HSPC) activity. However, the receptor(s) responsible for PGE2 signaling remains unclear. Here, we identified EP4 as a receptor ...activated by PGE2 to regulate HSPCs. Knockdown of Ep4 in HSPCs reduced long-term reconstitution capacity, whereas an EP4-selective agonist induced phosphorylation of GSK3β and β-catenin and enhanced long-term reconstitution capacity. Next, we analyzed the niche-mediated effect of PGE2 in HSPC regulation. Bone marrow mesenchymal progenitor cells (MPCs) expressed EP receptors, and stimulation of MPCs with PGE2 significantly increased their ability to support HSPC colony formation. Among the EP receptor agonists, only an EP4 agonist facilitated the formation of HSPC colonies after the coculture with MPCs. PGE2 up-regulated the expression of cytokine-, cell adhesion-, extracellular matrix-, and protease-related genes in MPCs. We also examined the function of PGE2/EP4 signaling in the recovery of the HSPCs after myelosuppression. The administration of PGE2 or an EP4 agonist facilitated the recovery of HSPCs from 5-fluorouracil (5-FU)–induced myelosuppression, indicating a role for PGE2/EP4 signaling in this process. Altogether, these data suggest that EP4 is a key receptor for PGE2-mediated direct and indirect regulation of HSPCs.
•PGE2 signaling positively regulates hematopoietic stem cells both directly and via activation of a nonhematopoietic cell population.•EP4 is a major receptor for the PGE2-mediated regulation of hematopoietic stem and progenitor cells.
In order to maintain the homeostasis of the hematopoietic system, hematopoietic stem cells (HSCs) need to be maintained while slowly dividing over their lifetime. However, repeated cell divisions ...lead to the gradual accumulation of DNA damage and ultimately impair HSC function. Since telomeres are particularly fragile when subjected to replication stress, cells have several defense machinery to protect telomeres. Moreover, HSCs must protect their genome against possible DNA damage, while maintaining telomere length. A group of proteins called the shelterin complex are deeply involved in this two-way role, and it is highly resistant to the replication stress to which HSCs are subjected. Most shelterin-deficient experimental models suffer acute cytotoxicity and severe phenotypes, as each shelterin component is essential for telomere protection. The Tin2 point mutant mice show a dyskeratosis congenita (DC) like phenotype, and the Tpp1 deletion impairs the hematopoietic system. POT1/Pot1a is highly expressed in HSCs and contributes to the maintenance of the HSC pool during in vitro culture. Here, we discuss the role of shelterin molecules in HSC regulation and review current understanding of how these are regulated in the maintenance of the HSC pool and the development of hematological disorders.
Telomerase reverse transcriptase (TERT) contributes to the prevention of aging by a largely unknown mechanism that is unrelated to telomere lengthening. The current study used ataxia-telangiectasia ...mutated (ATM) and TERT doubly deficient mice to evaluate the contributions of 2 aging-regulating molecules, TERT and ATM, to the aging process. ATM and TERT doubly deficient mice demonstrated increased progression of aging and had shorter lifespans than ATM-null mice, while TERT alone was insufficient to affect lifespan. ATM-TERT doubly null mice show in vivo senescence, especially in hematopoietic tissues, that was dependent on p16INK4a and p19ARF, but not on p21. As their HSCs show decreased stem cell activities, accelerated aging seen in these mice has been attributed to impaired stem cell function. TERT-deficient HSCs are characterized by reactive oxygen species (ROS) fragility, which has been suggested to cause stem cell impairment during aging, and apoptotic HSCs are markedly increased in these mice. p38MAPK activation was indicated to be partially involved in ROS-induced apoptosis in TERT-null HSCs, and BCL-2 is suggested to provide a part of the protective mechanisms of HSCs by TERT. The current study demonstrates that TERT mitigates aging by protecting HSCs under stressful conditions through telomere length-independent mechanisms.
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