scholarly journals Acute and endothelial-specific Robo4 deletion affect hematopoietic stem cell trafficking independent of VCAM1

PLoS ONE ◽  
2021 ◽  
Vol 16 (8) ◽  
pp. e0255606
Author(s):  
Stephanie Smith-Berdan ◽  
Alyssa Bercasio ◽  
Leah Kramer ◽  
Bryan Petkus ◽  
Lindsay Hinck ◽  
...  
Keyword(s):  
Stem Cell ◽  
Endothelial Cell ◽  
Hematopoietic Stem Cell ◽  
Transmembrane Protein ◽  
Vascular Endothelial Cell ◽  
Surface Protein ◽  
Cell Trafficking ◽  
Wild Type ◽  
Normal Numbers ◽  
Hematopoietic Stem

Hematopoietic stem cell (HSC) trafficking is regulated by a number of complex mechanisms. Among them are the transmembrane protein Robo4 and the vascular cell adhesion molecule, VCAM1. Endothelial VCAM1 is a well-known regulator of hematopoietic cell trafficking, and our previous studies revealed that germline deletion of Robo4 led to impaired HSC trafficking, with an increase in vascular endothelial cell (VEC) numbers and downregulation of VCAM1 protein on sinusoidal VECs. Here, we utilized two Robo4 conditional deletion models in parallel with Robo4 germline knockout mice (R4KO) to evaluate the effects of acute and endothelial cell-specific Robo4 deletion on HSC trafficking. Strikingly similar to the R4KO, the acute deletion of Robo4 resulted in altered HSC distribution between the bone marrow and blood compartments, despite normal numbers of VECs and wild-type levels of VCAM1 cell surface protein on sinusoidal VECs. Additionally, consistent with the R4KO mice, acute loss of Robo4 in the host perturbed long-term engraftment of donor wild-type HSCs and improved HSC mobilization to the peripheral blood. These data demonstrate the significant role that endothelial Robo4 plays in directional HSC trafficking, independent of alterations in VEC numbers and VCAM1 expression.

scholarly journals Albumin Modifies Responses to Hematopoietic Stem Cell Mobilizing Agents in Mice

Cells ◽  
2019 ◽  
Vol 9 (1) ◽  
pp. 4 ◽  
Author(s):  
Eva Danner ◽  
Halvard Bonig ◽  
Eliza Wiercinska
Keyword(s):  
Stem Cell ◽  
Hematopoietic Stem Cell ◽  
Wild Type ◽  
Cxcr4 Antagonist ◽  
Hematopoietic Stem ◽  
Split Dose ◽  
Healthy Donors ◽  
Hematopoietic Stem Cell Mobilization ◽  
Csf Levels ◽  
Cell Mobilization

Albumin, the most abundant plasma protein, not only controls osmotic blood pressure, but also serves as a carrier for various small molecules, including pharmaceuticals. Its impact on pharmacological properties of many drugs has been extensively studied over decades. Here, we focus on its interaction with the following mobilizing agents: Granulocyte-colony stimulating factor (G-CSF) and AMD3100, where such analyses are lacking. These compounds are widely used for hematopoietic stem cell mobilization of healthy donors or patients. Using albumin-deficient (Alb−/−) mice, we studied the contribution of albumin to mobilization outcomes. Mobilization with the bicyclam CXCR4 antagonist AMD3100 was attenuated in Alb−/− mice compared to wild-type littermates. By contrast, mobilization with recombinant human G-CSF (rhG-CSF), administered twice daily over a five-day course, was significantly increased in Alb−/− mice. In terms of a mechanism, we show that rhG-CSF bioavailability in the bone marrow is significantly improved in Alb−/− mice, compared to wild-type (WT) littermates, where rhG-CSF levels dramatically drop within a few hours of the injection. These observations likely explain the favorable mobilization outcomes with split-dose versus single-dose administration of rhG-CSF to healthy donors.

A Novel MBT-Containing Protein, Hemp, Plays Essential Roles In Skeletal Formation and Hematopoietic Stem Cell Function.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 1597-1597
Author(s):  
Phyo Wai Htun ◽  
Keiyo Takubo ◽  
Hideaki Oda ◽  
Feng Ma ◽  
Kenjiro Kosaki ◽  
...  
Keyword(s):  
Stem Cell ◽  
Hematopoietic Stem Cell ◽  
Cell Function ◽  
Conflicts Of Interest ◽  
Conditional Knockout ◽  
Thoracic Vertebrae ◽  
Wild Type ◽  
Hematopoietic Stem ◽  
Epigenetic Regulator ◽  
Skeletal Formation

Abstract Abstract 1597 Hemp (hematopoietic expressed mammalian polycomb, also denoted as mbt-containing 1) gene was originally identified in the hematopoietic stem cell (HSC)-enriched fraction of the mouse fetal liver (FL). It encodes a protein containing a putative Cys2-Cys2 zinc-finger region, followed by four tandem malignant brain tumor (MBT) repeats, which is frequently observed in polycomb gene (PcG) proteins. The structural characteristics strongly suggest that Hemp functions as an epigenetic regulator, but its biological role remains unknown. To address this issue, we generated hemp-deficient (hemp–/–) mice. Hemp–/– mice died soon after birth. Although no abnormalities were detected in internal organs, skeletal analysis exhibited a variety of malformations. Severe deformities were observed in the thoracic cavity, strongly suggesting that hemp–/– mice died of respiratory failure. Interestingly, they showed malformations of cervical and thoracic vertebrae, which were different from typical homeotic transformations observed in PcG-deficient mice. These results suggest that Hemp governs downstream target genes in distinct manners from conventional PcG proteins. The hematopoietic analysis of hemp in the FL showed that hemp is preferentially expressed in CD150+LSK and CD150–LSK HSC fractions in the hematopoietic hierarchy. Hemp–/– FL contained a significantly reduced number of hematopoietic cells and produced fewer number of hematopoietic colonies as compared to hemp+/+ FL. The decreases correlated with reduced number of CD150+LSK HSCs in hemp–/– FL, which generated much fewer hematopoietic colonies in the HPP-CFC assay. In addition, the competitive repopulation assay exhibited that the hematopoietic reconstitution ability of hemp–/– FL CD150+LSK HSCs was significantly impaired. Moreover, microarray analysis revealed that expression levels of several genes, such as Prdm16, Sox4, and Erdr1 were altered in hemp–/– FL HSCs. Since hemp–/– mice died at neonate, the role of Hemp in adult hematopoiesis remains to be elucidated. To address this issue, we generated hemp conditional knockout (cKO) mice. Acquired deletion of Hemp in the hematopoietic tissues was successfully achieved by crossing hempflox/flox mice with MxCre mice and stimulating the compound mice with pIpC. Analysis of the hematopoietic tissues revealed that the cell numbers of Mac+Gr1– and Mac+Gr1+ fractions in the hemp cKO bone marrow (BM) were significantly increased and decreased, respectively, as compared to those of the wild-type BM. However, no apparent differences have so far been observed between hemp cKO and wild-type littermates in functional analyses, such as colony forming activity and competitive repopulation ability of the BM cells. Here, we report that a novel MBT-containing protein, Hemp, plays essential roles in skeletal formation and HSC function during embryogenesis and also contributes to myeloid differentiation in adult hematopoiesis. Since Hemp likely functions as an epigenetic regulator, further studies will be required to clarify whether and what methylated lysine residues Hemp interacts with through the MBT repeats, what kind of genes are direct targets of Hemp, and how Hemp exerts its biological activity. Disclosures: No relevant conflicts of interest to declare.

The F-Box Protein NIPA Regulates the Hematopoietic Stem Cell Pool

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 2330-2330
Author(s):  
Stefanie Kreutmair ◽  
Anna Lena Illert ◽  
Rouzanna Istvanffy ◽  
Melanie Sickinger ◽  
Christina Eckl ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
Hematopoietic Stem Cell ◽  
Bone Marrow Cells ◽  
Wild Type ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Cell Pool ◽  
F Box Protein ◽  
Stem Cell Pool

Abstract Abstract 2330 Hematopoietic stem cells (HSCs) are characterized by their ability to self-renewal and multilineage differentiation. Since mostly HSCs exist in a quiescent state re-entry into cell cycle is essential for their regeneration and differentiation and the expression of numerous cell cycle regulators must be tightly controlled. We previously characterized NIPA (Nuclear Interaction Partner of ALK) as a F-Box protein that defines an oscillating ubiquitin E3 ligase targeting nuclear cyclin B1 in interphase thus contributing to the timing of mitotic entry. To examine the function of NIPA on vivo, we generated NIPA deficient animals, which are viable but sterile due to a defect in recombination and testis stem cell maintenance. To further characterize the role of NIPA in stem cell maintenance and self-renewal we investigated hematopoiesis in NIPA deficient animals. Peripheral blood counts taken at different ages revealed no apparent difference between NIPA knockout and wild type mice in numbers and differentiation. In contrast, looking at the hematopoietic stem cell pool, FACS analyses of bone marrow showed significantly decreased numbers of Lin-Sca1+cKit+ (LSK) cells in NIPA deficient animals, where LSKs were reduced to 40% of wild type littermates (p=0,0171). This effect was only apparent in older animals, where physiologically higher LSK numbers have to compensate for the exhaustion of the stem cell pool. Additionally, older NIPA deficient mice have only half the amount of multi myeloid progenitors (MMPs) in contrast to wild type animals. To examine efficient activation of stem cells to self-renew in response to myeloid depression, we treated young and old mice with the cytotoxic drug (5-FU) four days before bone marrow harvest. As expected, 5-FU activated hematopoietic progenitors in wild type animals, whereas NIPA deficient progenitors failed to compensate to 5-FU depression, e.g. LSKs of NIPA knockout mice were reduced to 50% of wild type levels (p<0.001), CD150+CD34+ Nipa deficient cells to 20% of wild type levels (p<0.0001). Interestingly, these effects were seen in all NIPA deficient animals independent of age, allowing us to trigger the self-renewal phenotype by activating the hematopoietic stem cell pool. Using competitive bone marrow transplantation assays, CD45.2 positive NIPA deficient or NIPA wild type bone marrow cells were mixed with CD45.1 positive wild type bone marrow cells and transplanted into lethally irradiated CD45.2 positive recipient mice. Thirty days after transplantation, FACS analysis of peripheral blood and bone marrow showed reduced numbers of NIPA knockout cells in comparison to NIPA wild type bone marrow recipient mice. This result was even more severe with aging of transplanted mice, where NIPA deficient cells were reduced to less than 10% of the level of wild type cells in bone marrow of sacrificed mice 6 months after transplantation, pointing to a profound defect in repopulation capacity of NIPA deficient HSCs. Taken together our results demonstrate a unique and critical role of NIPA in regulating the primitive hematopoietic compartment as a regulator of self-renewal, cycle capacity and HSC expansion. Disclosures: No relevant conflicts of interest to declare.

Interruption of IFNγR Signaling Results in Altered T Cell Trafficking in Vivo and Abrogation of GvHD While Maintaining a Robust Gvl Response

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 455-455
Author(s):  
Jaebok Choi ◽  
Edward Dela Ziga ◽  
Julie Ritchey ◽  
Lynne Collins ◽  
Julie Prior ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Hematopoietic Stem Cell ◽  
Cell Trafficking ◽  
Hematopoietic Stem ◽  
T Cell Trafficking ◽  
Target Organs ◽  
Donor T Cells

Abstract Abstract 455 Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is the only curative treatment for patients with relapsed/refractory leukemia, and marrow failure states such as myelodysplasia and aplastic anemia. However, allo-HSCT is complicated by allogeneic donor T cell-mediated graft-versus-host disease (GvHD) which can be life-threatening especially in recipients of unrelated or HLA-mismatched hematopoietic stem cell products. These same alloreactive donor T cells also mediate a beneficial graft-versus-leukemia (GvL) effect. Thus, the clinical goal in allo-HSCT is to minimize GvHD while maintaining GvL. Recent studies have suggested that this might be achieved by infusing regulatory T cells (Tregs) which in some preclinical models suppress GvHD-causing alloreactive donor T cells but have only limited effects on GvL-promoting alloreactive donor T cells. Unfortunately, Tregs exist in low frequency in the peripheral blood, are costly to purify and expand, and after expansion are difficult to isolate due to the lack of cell surface markers, all of which prevent their routine use in the clinic. Thus, alternative therapeutic approaches that do not require Tregs are needed. We have found that interferon gamma receptor deficient (IFNγR−/−) allogeneic donor T cells induce significantly less GvHD in both a MHC fully-mismatched (B6 (H-2b) → Balb/c (H-2d)) and a minor-mismatched (B6 (H-2b) → B6×129(H-2b)) allo-HSCT models compared to WT T cells. In addition, IFNγR−/− donor T cells maintain a beneficial GvL effect, which has been examined in both systemic leukemia and solid tumor models using luciferase-expressing A20 cells derived from Balb/c. We find that IFNγR−/− T cells migrate primarily to the spleen while WT T cells to GI tract and peripheral lymph nodes (LNs) using bioluminescence imaging (BLI), suggesting that altered T cell trafficking of IFNγR−/− T cells to GvHD target organs might be the major reason for the reduced GvHD. We further demonstrate that the IFNγR-mediated signaling in alloreactive donor T cells is required for expression of CXCR3 which has been implicated in trafficking of T cells to areas of inflammation and target organs, commonly known to be the sites of GvHD. Indeed, CXCR3−/− T cells recapitulate the reduced GvHD potential of IFNγR−/− T cells. In addition, forced overexpression of CXCR3 in IFNγR−/− T cells via retroviral transduction partially rescues the GvHD defect observed in IFNγR−/− T cells. We next examine if inhibition of IFNγR signaling using a small molecule inhibitor can recapitulate the anti-GVHD effects seen in IFNγR−/− T cells. We find that INCB018424, an inhibitor of JAK1/JAK2 which are the mediators of IFNγR signaling, blocks CXCR3 expression in vitro. Most importantly, in vivo administration of INCB018424 after allo-HSCT alters T cell trafficking and significantly reduces GvHD. Thus, the IFNγR signaling pathway represents a promising therapeutic target for future efforts to mitigate GvHD while maintaining GvL after allo-HSCT. Moreover, this pathway can be exploited in other diseases besides GvHD such as those from organ transplantation, chronic inflammatory diseases and autoimmune diseases. Disclosures: DiPersio: genzyme: Honoraria.

MiR-29a is Essential in Leukemic Transformation and Maintaining Hematopoietic Stem Cell Self-Renewal

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 4792-4792
Author(s):  
Wenhuo Hu ◽  
James Dooley ◽  
Luisa Cimmino ◽  
Adrian Liston ◽  
Christopher Y. Park
Keyword(s):  
Gene Expression ◽  
Stem Cell ◽  
Hematopoietic Stem Cell ◽  
Protein Translation ◽  
Epigenetic Mark ◽  
Mouse Bone Marrow ◽  
Wild Type ◽  
Leukemic Transformation ◽  
Self Renewal ◽  
Hematopoietic Stem

Abstract MicroRNAs are small non-coding RNAs that interfere with gene expression by degrading messenger RNAs (mRNAs) or blocking protein translation. Expression profiling studies has identified miRNAs that regulate normal and malignant hematopoietic stem cell function. Our previous studies showed that ectopic expression of miR-29a in mouse bone marrow cells induced a myeloproliferative disorder that progressed to acute myeloid leukemia (AML). Over-expression of miR-29b in AML cell lines has been reported to induce apoptosis by negatively regulating Dnmt3a. We recently found that miR-29a positively regulates hematopoietic stem cell (HSC) self-renewal and proliferation using a knockout mouse model of miR-29ab. miR-29a null mice contained significantly lower HSC numbers and miR-29a null HSCs exhibited markedly decreased reconstitution ability in both competitive and non-competitive transplantation assays. To investigate the mechanism of miR-29a action, we performed transcriptomal profiling of miR-29a null HSCs and found that miR-29a null HSCs exhibit a gene expression pattern more similar to wild-type committed progenitors than wild-type HSCs. We identified Dnmt3a as one dysregulated miR-29a target as showing increased expression in miR-29a null HSCs, and haplodeficiency of Dnmt3a partly restores miR-29a deficient HSC function. In order to test the requirement for miR-29a in myeloid leukemogenesis, we transduced miR-29a deficient Lin-c-Kit+Sca-1+ (LSK) cells with the oncogenic MLL-AF9 fusion gene, and found that the development of AML from these cells was markedly delayed. We found that Meis1, Ccna2, Hoxa5 and Hoxa9 transcripts were significantly downregulated in miR-29a null LSK cells compared to WT LSK cells, but they were similarly induced in MLL-AF9 transformed c-Kit+Mac-1+ cells. To investigate whether the epigenetic dysregulation resulting from miR-29a deletion may underlie this transformation-resistant phenotype, we examined the distribution of the active epigenetic mark, H3K79me2, in c-Kit+Mac-1+ miR-29a null cells using a ChIP-Seq assay. After analyzing H3K39me2 peaks using model-based analysis of ChIP-Seq, we identified 4281 and 3649 genes associated with this active epigenetic mark using a duplicated ChIP-Seq analysis, with an overlap of 3164 genes (66.39%). Using public available ChIP-Seq data, we compared our results with the genes associated with the H3K79me2 mark in normal immature LSK cells (9282 genes), granulocyte-macrophage progenitors (GMPs, 8556 genes), and MLL-AF9 transformed GMP cells (L-GMP, 8578 genes), and found 4234, 4111, 4046, and 4766 genes were also identified have an active H3K79me2 mark in MLL-AF9 transformed miR-29a null cells. These data indicate that miR-29a loss inactivates a large group of genes activated by the MLL-AF9 oncogene. We also found that 379 genes were associated with H3K79me2 peaks in both normal LSK and MLL-AF9 transformed miR-29a null c-Kit+Mac-1+ cells, but were absent of this epigenetic marker in L-GMP, suggesting that these genes confer self-renewal and proliferation capacities to normal HSCs. In addition, suppression of these genes are important in leukemic transformation by MLL-AF9, and finally the reactivation of these genes in miR-29a null cells compromises the leukemogenesis ability of MLL-AF9. Interestingly, out of these 379 genes, we were able to identify 18 genes that were potential miR-29a targets including Akt3, Map4k4, Dnmt3a, et al. This suggests the direct and indirect effects from miR-29a in regulating its target gene networks at transcriptional and post-transcriptional levels. Our studies found miR-29a is essential in maintaining HSC function and loss of miR-29a abrogate the leukemogenesis capacity of MLL-AF9. Disclosures No relevant conflicts of interest to declare.

Endothelial cell-selective adhesion molecule is a novel human hematopoietic stem cell marker associated with a subset of human leukemias

2015 ◽  
Vol 43 (9) ◽  
pp. S69
Author(s):  
Tomohiko Ishibashi ◽  
Takafumi Yokota ◽  
Hirokazu Tanaka ◽  
Michiko Ichii ◽  
Takao Sudo ◽  
...  
Keyword(s):  
Stem Cell ◽  
Endothelial Cell ◽  
Hematopoietic Stem Cell ◽  
Adhesion Molecule ◽  
Stem Cell Marker ◽  
Cell Marker ◽  
Hematopoietic Stem

scholarly journals Hemogenic Endothelium-like Changes Were Derived to an Adult Human Dermal Fibroblast By Electrical Stimulation

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 5129-5129
Author(s):  
Takuji Matsuo ◽  
Ryosuke Shirasaki ◽  
Oka Yoko ◽  
Tadashi Yamamoto ◽  
Jun Ooi ◽  
...  
Keyword(s):  
Stem Cell ◽  
Electrical Stimulation ◽  
Endothelial Cell ◽  
Cell Fate ◽  
Dermal Fibroblast ◽  
Vascular Endothelial Cell ◽  
Human Dermal Fibroblast ◽  
Vascular Endothelial ◽  
Hematopoietic Stem ◽  
Morphologic Changes

Abstract Background and Aims: We previously reported that when an adult human dermal fibroblast (HDF) was cultured with interleukin (IL)-1-beta (b) and erythropoietin (EPO), hematopoiesis-related molecules expressed. And, lymphatic duct-neogenesis genes also expressed, and vascular endothelial growth factor (VEGF)-A, and -C were produced. When anti-human VEGF-C antibody (Ab) was added to the cultures, hematopoiesis-related genes expressed; however, morphologic changes were not observed (54th ASH, 18th EHA). Reported findings on nuclear transfer reveal that electrical stimulation induces nuclear fusion and changes cell-fate. We observed the effect of electrical stimulation to IL-1-b-stimulated HDF. Materials and Methods: HDF was cultured with IL-1-b, EPO, VEGF-A, and anti-human VEGF-C Ab for 14 days. Then, cells were suspended in electrical stimulation-buffer (0.25 M d-sorbitol, 0.1 mM Ca-acetate, 0.5 mM Mg-acetate, 1 mg/mL fatty-acid-free BSA, 0.5 mM HEPES), and incubated on ice for 10 minutes. Cells were stimulated at 110V 20mA (0.2 cm electrode, gap), and were further cultured with k/o DMEM containing 20% KSR and SCF, IL-6, FL, IGF-2, and VEGF-A. Morphological changes and expressions of vascular endothelial cell-related genes, and hematopoiesis-related ones were observed. Results: When HDF was stimulated electrically, a few cells showed vascular endothelial cell-morphology after two days. When cells were further cultured in a hematopoietic stem cell-culturing condition, a part of endothelial cell-morphology changed to that of non-adherent hematopoietic cells or blastic colonies, in which increased expression-levels of SCL, CD41, GATA-2, CD34, and CD45 were accepted with significant statistical difference. Discussion: Recent reports reveal that a kind of vascular endothelial cells, called hemogenic endothelium, can convert to a hematopoietic cell, and works an important role in hematopoietic stem cell-generation. We hypothesized that when cells expressed a kind of specific transcripts, some stimulation triggerred to convert morphologic changes and cell-fate as are observed in nuclear fusion. And, our observations indicated that when IL-1-b-stimulated HDF, expressing hematopoiesis-related transcripts, were stimulated electrically, cellular morphology changed. Currently, we are precisely analyzing electrically stimulated HDF’s biological characteristics. Disclosures No relevant conflicts of interest to declare.

scholarly journals The Effects of Galactic Cosmic Radiation Exposure on Hematopoietic Stem Cell Dysfunction and Oncogenesis

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 5297-5297
Author(s):  
Rutulkumar Patel ◽  
Scott Welford ◽  
Stanton L. Gerson
Keyword(s):  
Stem Cell ◽  
Mismatch Repair ◽  
Hematopoietic Stem Cell ◽  
Cosmic Radiation ◽  
Whole Body ◽  
Wild Type ◽  
Radiation Quality ◽  
Hematopoietic Stem ◽  
Dna Mismatch

Abstract Natural sources of radiation in space include galactic cosmic rays (GCR), solar energetic particles (SPE) and trapped energetic particles in a planetary magnetic field. These different sources of space radiation consist of protons of various energies, particle nuclei of high energy and charge (HZE) and neutrons of different energies. These sources are difficult to shield because of their high energies and dense ionization patterns, thus posing significant health risks to astronauts on long term inter-planetary missions. Efforts to protect astronauts from harmful cosmic radiation require a deeper understanding of the effects of GCR on human health. In particular, very little is known about the effects of GCR exposure on the hematopoietic stem cell (HSC) population and whether disruptions in genetic stability in HSCs could result in the development of hematopoietic malignancies in astronauts on deep space missions. The average age of shuttle crew has risen above 46 years, and our work and others have shown that HSCs display diminished function with age. Recent data from our group has demonstrated that middle-aged individuals show frequent defects in DNA mismatch repair (MMR) in HSCs. MMR corrects DNA mismatches generated by DNA polymerase during replication which prevents mutations from becoming permanent in dividing cells. Thus, MMR plays a crucial role in the DNA damage response pathway to prevent short-term mutagenesis and long-term tumorigenesis. Several human MMR proteins have been identified as MutS and MutL homologues consisting of MSH2 and MLH1 heterodimers that functions in DNA mismatch/damage recognition, endonuclease activity and termination of mismatch-provoked excision. Our group has shown that humans accumulate microsatellite instability (MSI) with acquired loss of MLH1 protein in hematopoietic stem and progenitor cells as a function of age. Therefore, we employed a DNA mismatch repair deficient mouse model (MLH1+/- and MLH1-/-) to study the effects of different radiation sources including 56Fe, 28Si, 4He, 1H and ᵞ-rays on HSCs to examine HSCs of potential astronaut population under GCR conditions. The complete blood count (CBC) data after 5 months and 9 months of whole body irradiation with different ions showed a slight dose-dependent decrease in all blood counts but absence of any significant difference in CBC of MLH1+/+ vs MLH1+/- mice. In addition, CFU and competitive repopulation data demonstrated a radiation quality effects on HSC function, but not an MLH1 effect. These results demonstrate that hematopoietic stem cell function is normal and that a MLH1 defect does not differentiate progenitor and mature effector cells following HZE radiation. To study long term effects of different ions on the potential for disease progression in a MLH1 dependent manner, we performed whole body irradiation with 56Fe, 28Si, 1H and ᵞ-rays on MLH1+/+ and MLH1+/- mice and followed them up to 18 months post exposure. We observed that MLH1+/- mice show dramatic increases in lymphomagenesis 10-12 months after 56Fe irradiation compared to wild type mice, with greater than 60 % of MLH1+/- mice developed lymphomas at doses 10 cGy and 100 cGy compared to less than 10 % of wild type. For comparison, roughly 10 % of MLH1+/- mice developed lymphomas when mice were treated with whole body sparsely ionizing ᵞ-rays at 100 cGy compared to none of the control. Thus the date show that MMR defects in HSCs lead to sensitization to radiation induced hematopoietic malignancy and that radiation quality effects exacerbate the sensitivity. The findings could have profound effects on astronaut screening, as well as lead to important questions regarding safety of ion therapy and development of second malignancies for cancer patients who remain on Earth. Disclosures No relevant conflicts of interest to declare.

Activity of a Heptad of Transcription Factors Is Associated with Stem Cell Programs and Clinical Outcome in Acute Myeloid Leukaemia

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 3525-3525
Author(s):  
Eva Diffner ◽  
Dominik Beck ◽  
Emma Gudgin ◽  
Julie Thoms ◽  
Kathy Knezevic ◽  
...  
Keyword(s):  
Risk Factor ◽  
Stem Cell ◽  
Transcription Factors ◽  
Clinical Outcome ◽  
Hematopoietic Stem Cell ◽  
Conflicts Of Interest ◽  
Leukemic Cells ◽  
Wild Type ◽  
Hematopoietic Stem ◽  
Acute Myeloid

Abstract Abstract 3525 Leukaemic transformation is driven by aberrant transcriptional programs often in combination with abnormal proliferative signalling. These programs operate in normal hematopoiesis where they are involved in hematopoietic stem cell (HSC) proliferation and maintenance. ERG is a component of normal and leukemic stem cell signatures and high ERG expression has been proposed as a risk factor for poor prognosis in acute myeloid leukemia (AML). However, mechanisms that underlie ERG expression in AML and how its expression relates to leukemic stemness are unknown. We report that ERG expression in AML is associated with activity of the ERG+85 stem cell enhancer (SCE) and a heptad of transcription factors that combinatorially regulate genes in normal HSCs. Gene expression signatures derived from ERG+85 stem cell enhancer (Fig A) and heptad activity (Fig B) predict clinical outcome in a cytogenetically normal cohort of AML (CN-AML) patients when ERG expression alone fails. The heptad signature is an independent risk factor for poor overall and event-free survival (Fig C). There were no long-term survivors amongst patients with a heptad signature, FLT3 mutations and wild-type NPM1 (Fig D) pointing to a hitherto unappreciated link between aberrant signaling and transcriptional mediators of hematopoietic stem cell identity. In two independent cohorts, the heptad signature was as closely associated with wild-type NPM1 AML as the HOX signature was with mutant NPM1 AML (Fig E–F) suggestive of a collective role for these transcription factors in mediating the leukemic signature in the former. Taken together, these results show that key transcriptional regulators cooperate in establishing stem cell signatures in leukemic cells and that the underlying spectrum of somatic mutations contributes to the development of these signatures and modulate their influence on clinical outcome. Disclosures: No relevant conflicts of interest to declare.

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