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Oxidative stress has been implicated in the pathogenesis of many human diseases including Fanconi anemia (FA), a genetic disorder associated with BM failure and cancer. Here we show that major antioxidant defense genes are downregulated in FA patients, and that gene down-regulation is selectively associated with increased oxidative DNA damage in the promoters of the antioxidant defense genes. Assessment of promoter activity and DNA damage repair kinetics shows that increased initial damage, rather than a reduced repair rate, contributes to the augmented oxidative DNA damage. Mechanistically, FA proteins act in concert with the chromatin-remodeling factor BRG1 to protect the promoters of antioxidant defense genes from oxidative damage. Specifically, BRG1 binds to the promoters of the antioxidant defense genes at steady state. On challenge with oxidative stress, FA proteins are recruited to promoter DNA, which correlates with significant increase in the binding of BRG1 within promoter regions. In addition, oxidative stress-induced FANCD2 ubiquitination is required for the formation of a FA-BRG1-promoter complex. Taken IntroductionOxidative DNA damage is a major source of genomic instability. The most prevalent lesion generated by intracellular reactive oxygen species (ROS) is 8-hydroxydeoxy guanosine (8-oxodG). This lesion causes G:C to T:A transversion mutations and is considered highly mutagenic. 1 There is compelling evidence that 8-oxodG levels are elevated in various human cancers. 2,3 and in animal models of tumors. 4,5 ROS-induced DNA damage can also result in single-or double-strand breaks, which are lethal to the cell if not repaired. 6,7 Although there is a great deal known about DNA repair, we have a limited understanding of the involvement of specific repair pathways in protecting cellular DNA from oxidative damaging agents, particularly ROS. The major pathways involved in DNA repair include repair of single-base damage by the base excision repair (BER) pathway, repair of lesions that distort the DNA helix by the nucleotide excision repair (NER) pathway, and repair of DNA double-strand breaks by homologous recombination (HR) and nonhomologous end-joining (NHEJ) pathways. [8][9][10] Although the specificity and efficiency of each of these DNA repair pathways is critical to ensure genome stability, the complexity of ROS-induced oxidative DNA damage may require coordination between these different pathways.Cells have developed a battery of defense mechanisms to protect against damage induced by oxidative stress. Antioxidant defense enzymes, including superoxide dismutases, catalase, glutathione peroxidases and peroxiredoxins, as well as nonenzymatic scavengers such as glutathione and carotenoids can directly eliminate ROS. 11 Other cellular enzymes can repair DNA damage induced by ROS. 12 Moreover, ROS can influence the selective activation of oxidative stress-responsive transcription factors.Indeed, the first line of defense against oxidative damage is the induction of stress-response genes, many of whi...

Salidroside is a phenylpropanoid glycoside isolated from the medicinal plant Rhodiola rosea, which has potent antioxidant properties. Here we show that salidroside prevented the loss of hematopoietic stem cells (HSCs) in mice under oxidative stress. Quiescent HSCs were recruited into cell cycling on in vivo challenge with oxidative stress, which was blocked by salidroside. Surprisingly, salidroside does not prevent the production of reactive oxygen species but reduces hydrogen peroxide-induced DNAstrand breaks in bone marrow cells enriched for HSCs. We tested whether salidroside enhances oxidative DNA damage repair in mice deficient for 5 DNA repair pathways known to be involved in oxidative DNA damage repair; we found that salidroside activated poly(ADPribose)polymerase-1 (PARP-1), a component of the base excision repair pathway, in mouse bone marrow HSCs as well as primary fibroblasts and human lymphoblasts. PARP-1 activation by salidroside protects quiescent HSCs from oxidative stress-induced cycling in native animals and self-renewal defect in transplanted recipients, which was abrogated by genetic ablation or pharmacologic inhibition of PARP-1. Together, these findings suggest that activation of PARP-1 by salidroside could affect the homeostasis and function of HSCs and contribute to the antioxidant effects of salidroside. IntroductionHematopoietic stem cells (HSCs) are a rare population of pluripotent cells that can self-renew and differentiate into various types of cells of the blood lineage. 1 Under steady physiologic conditions, the most primitive HSCs are in a quiescent state and reside in the bone marrow (BM) niche where they preserve the capacity to self-renew and to continue to produce all types of blood cells throughout a prolonged life span without depleting the regenerative cell pool. 2,3 In response to stress or stimulation, the HSCs can move out of the BM niche, entering cell cycle and undergoing division. In addition, the cycling HSCs may return to the BM niche and regain their quiescent state. 4 Disruption of HSC quiescence prematurely exhausts the stem cell pool and causes hematologic failure under various stresses, such as oxidative stress, cell cycling, and aging. 5,6 Oxidative stress, defined as an imbalance between the production of reactive oxygen species (ROS) and antioxidant defense, is most evident in states of aging and diseases such as BM failure and cancer. 7,8 Even in a healthy state, HSCs are exposed to various ROS, which are routinely generated during metabolic or inflammatory process. 9 ROS induce a variety of responses in HSCs, including cellular proliferation and apoptosis. 10,11 ROS can also cause DNA damage and drive HSCs into cell division, which is essential for DNA repair processes. 12,13 Similar to stem cells from other tissues, HSCs have developed several mechanisms to prevent the damage induced by oxidative stress. Antioxidant enzymes, including superoxide dismutases, catalase, glutathione peroxidases, and peroxiredoxins, can directly eliminate ROS. 10 Other cellula...

Patients with Fanconi anemia (FA) have a high risk of developing acute myeloid leukemia (AML). In this study, we attempted to identify cell-surface markers for leukemia-initiating cells in FA-AML patients. We found that the IL-3 receptor-␣ (IL-3R␣) is a promising candidate as an leukemia-initiating cell-specific antigen for FA-AML. Whereas IL-3R␣ expression is undetectable on normal CD34 ؉ CD38 ؊ HSCs, it is overexpressed on CD34 ؉ CD38 ؊ cells from FA patients with AML. IntroductionAcute myeloid leukemia (AML), a heterogeneous group of hematologic malignancies characterized by an accumulation of clonal myeloid progenitor cells that do not differentiate normally, 1,2 comprises approximately 25% of childhood acute leukemias. 3 The treatment of AML remains a challenge, and most AML patients will die of their disease within 1-2 years of diagnosis. 4 Conventional chemotherapeutic agents have been successful to some degree in treating AML, but now appear to have reached their maximum potential. Even with high-dose chemotherapy, only 30%-40% of AML patients survive, which is due mainly to relapse of the disease. 5 Recently, novel therapeutic strategies for AML have focused on immune-based therapy through monoclonal antibodies that target and destroy leukemic blasts via specific cell receptors. 6,7 These therapies were designed with the aim of selectively killing malignant cells that express unique antigens while sparing normal cells.One of the recent advances in the AML field is the postulation that AML arises from a rare population of leukemic stem cells (LSCs). 8,9 Phenotypic and functional analyses show that LSCs reside in the CD34 ϩ CD38 Ϫ compartment, the primitive stem/ progenitor population that also contains normal HSCs. 10 Further studies demonstrated that both normal HSCs and LSCs share the properties of quiescence and self-renewal. [8][9][10] This relatively dormant property of LSCs may contribute to the pattern of remission and subsequent relapse that is typical of the response to cytotoxic chemotherapy in AML. Therefore, it is believed that although most AML blasts can be eradicated by cytotoxic therapy, LSCs may be resistant to killing by chemotherapeutic agents. Recent studies have suggested that several antigens, such as CD33, CD44, CD96, CD123, and CLL-1, are specifically expressed in AML LSCs but not in normal HSCs. [11][12][13][14][15][16][17] Because it is believed that LSCs are the most relevant target population for novel antileukemic therapy, these unique antigens present opportunities for selectively targeting AML LSCs.One of the best studied AML models is Fanconi anemia (FA), a genetic disorder associated with bone marrow failure, clonal proliferation of HSCs and progenitor cells, and progression to myelodysplastic syndrome (MDS) and AML. [18][19][20] FA is caused by a deficiency in any of the 14 genes that encode the FANCA, FANCB, FANCC, FANCD1/BRCA2, FANCD2, FANCE, FANCF, FANCG, FANCI, FANCJ/BRIP1, FANCL, FANCM, FANCN/PALB2, and FANCO/RAD51C proteins. [21][22][23][24] The biologic function ...

Hematopoietic stem cells (HSCs) can either self-renew or differentiate into various types of cells of the blood lineage. Signaling pathways that regulate this choice of self-renewal versus differentiation are currently under extensive investigation. Here we report that deregulation of Notch signaling skews HSC differentiation in mouse models of Fanconi anemia (FA), a genetic disorder associated with bone marrow failure and progression to leukemia and other cancers. In mice expressing a transgenic Notch reporter, deletion of the Fanca or Fancc gene enhances Notch signaling in multipotential progenitors (MPPs), which is correlated with decreased phenotypic long-term HSCs and increased formation of MPP1 progenitors. Furthermore, we found an inverse correlation between Notch signaling and self-renewal capacity in FA hematopoietic stem and progenitor cells. Significantly, FA deficiency in MPPs deregulates a complex network of genes in the Notch and canonical NF-κB pathways. Genetic ablation or pharmacologic inhibition of NF-κB reduces Notch signaling in FA MPPs to near wide-type level, and blocking either NF-κB or Notch signaling partially restores FA HSC quiescence and self-renewal capacity. Taken together, these results suggest a functional crosstalk between Notch signaling and NF-κB pathway in regulation of HSC differentiation.