Remodeling of Bone Marrow Hematopoietic Stem Cell Niches Promotes Myeloid Cell Expansion during Premature or Physiological Aging

Hematopoietic stem cells (HSCs) residing in the bone marrow (BM) accumulate during aging but are functionally impaired. However, the role of HSC-intrinsic and -extrinsic aging mechanisms remains debated. Megakaryocytes promote quiescence of neighboring HSCs. Nonetheless, whether megakaryocyte-HSC interactions change during pathological/natural aging is unclear. Premature aging in Hutchinson-Gilford progeria syndrome recapitulates physiological aging features, but whether these arise from altered stem or niche cells is unknown. Here, we show that the BM microenvironment promotes myelopoiesis in premature/physiological aging. During physiological aging, HSC-supporting niches decrease near bone but expand further from bone. Increased BM noradrenergic innervation promotes β2-adrenergic-receptor(AR)-interleukin-6-dependent megakaryopoiesis. Reduced β3-AR-Nos1 activity correlates with decreased endosteal niches and megakaryocyte apposition to sinusoids. However, chronic treatment of progeroid mice with β3-AR agonist decreases premature myeloid and HSC expansion and restores the proximal association of HSCs to megakaryocytes. Therefore, normal/premature aging of BM niches promotes myeloid expansion and can be improved by targeting the microenvironment.Y.-H.O. received fellowships from Alborada Scholar-ship (University of Cambridge), Trinity-Henry Barlow Scholarship (Universityof Cambridge), and R.O.C. Government Scholarship to Study Abroad (GSSA). A.G.G. received fellowships from the Ramon Areces Foundationand the LaCaixa Foundation. C.K. was supported by Marie Curie Career Inte-gration (H2020-MSCA-IF-2015-70841). S.M.-F. was supported by Red TerCel (ISCIII-Spanish Cell Therapy Network). V.A. is supported by grants from theSpanish Ministerio de Economıa,Industria y Competitividad (MEIC) with co-funding from the Fondo Europeo de Desarrollo Regional (FEDER, ‘‘Una manerade hacer Europa’’) (SAF2016-79490-R), the Instituto de Salud Carlos III (AC16/00091 and AC17/00067), the Fundacio Marato TV3 (122/C/2015), and the Progeria Research Foundation (Established Investigator Award 2014–52). TheCNIC is supported by the Instituto de Salud Carlos III (ISCIII), the Ministerio de Ciencia, Innovacion y Universidades (MCIU), and the Pro CNIC Foundation,and is a Severo Ochoa Center of Excellence (SEV-2015-0505). This work wassupported by core support grants from the Wellcome Trust and the MRC to theCambridge Stem Cell Institute, MEIC (SAF-2011-30308), Ramon y Cajal Program Grant (RYC-2009-04703), ConSEPOC-Comunidad de Madrid (S2010/BMD-2542), National Health Service Blood and Transplant (United Kingdom), European Union’s Horizon 2020 research (ERC-2014-CoG-64765 and MarieCurie Career Integration grant FP7-PEOPLE-2011-RG-294096), and a Programme Foundation Award from Cancer Research UK to S.M.-F., who wasalso supported in part by an International Early Career Scientist grant fromthe Howard Hughes Medical Institute.S

Remodeling of Bone Marrow Hematopoietic Stem Cell Niches Promotes Myeloid Cell Expansion during Premature or Physiological Aging.

Hematopoietic stem cells (HSCs) residing in the bone marrow (BM) accumulate during aging but are functionally impaired. However, the role of HSC-intrinsic and -extrinsic aging mechanisms remains debated. Megakaryocytes promote quiescence of neighboring HSCs. Nonetheless, whether megakaryocyte-HSC interactions change during pathological/natural aging is unclear. Premature aging in Hutchinson-Gilford progeria syndrome recapitulates physiological aging features, but whether these arise from altered stem or niche cells is unknown. Here, we show that the BM microenvironment promotes myelopoiesis in premature/physiological aging. During physiological aging, HSC-supporting niches decrease near bone but expand further from bone. Increased BM noradrenergic innervation promotes β2-adrenergic-receptor(AR)-interleukin-6-dependent megakaryopoiesis. Reduced β3-AR-Nos1 activity correlates with decreased endosteal niches and megakaryocyte apposition to sinusoids. However, chronic treatment of progeroid mice with β3-AR agonist decreases premature myeloid and HSC expansion and restores the proximal association of HSCs to megakaryocytes. Therefore, normal/premature aging of BM niches promotes myeloid expansion and can be improved by targeting the microenvironment.We thank A.R. Green for advice and support; M. García-Fernández, C. Fielding, C. Kapeni, X. Langa, and other current and former members of the S.M.-F group for help and discussions; A. Barettino and A. Macías (CNIC), D. Pask, T. Hamilton, the Central Biomedical Services and Cambridge NIHR BRC Cell Phenotyping Hub for technical assistance; H. Jolin and A.N.J. McZenzie (MRC Laboratory of Molecular Biology, Cambridge, UK) for help with milliplex analyses. Y.-H.O. received fellowships from Alborada Scholarship (University of Cambridge), Trinity-Henry Barlow Scholarship (University of Cambridge) and R.O.C. Government Scholarship to Study Abroad (GSSA) A.G.G. received fellowships from Ramón Areces and LaCaixa Foundations. C.K. was supported by Marie Curie Career Integration grant H2020-MSCA-IF-2015-70841. S.M.F., by Red TerCel (ISCIII-Spanish Cell Therapy Network). VA is supported by grants from the Spanish Ministerio de Economía, Industria y Competitividad (MEIC) with cofunding from the Fondo Europeo de Desarrollo Regional (FEDER, “Una manera de hacer Europa”) (SAF2016-79490-R), the Instituto de Salud Carlos III (AC16/00091), the Fundació Marató TV3 (122/C/2015), and the Progeria Research Foundation (Established Investigator Award 2014–52). The CNIC is supported by the Instituto de Salud Carlos III (ISCIII), the Ministerio de Ciencia, Innovación y Universidades (MCNU) and the Pro CNIC Foundation, and is a Severo Ochoa Center of Excellence (SEV-2015-0505). This work was supported by core support grants from the Wellcome Trust and the MRC to the Cambridge Stem Cell Institute, the Spanish Ministry of Economy and Competitiveness (SAF-2011-30308), Ramón y Cajal Program grant RYC-2009-04703, ConSEPOC-Comunidad de Madrid S2010/BMD-2542, National 427 Health Service Blood and Transplant (United Kingdom), European Union’s Horizon 428 2020 research (ERC-2014-CoG-64765 and Marie Curie Career Integration grant FP7- 429 PEOPLE-2011-RG-294096) and a Programme Foundation Award from Cancer Research 430 UK to S.M.-F., who was also supported in part by an International Early Career Scientist 431 grant of the Howard Hughes Medical Institute

Titre Environnement de la moelle osseuse et domiciliation des cellules souches hématopoïétiques

International audienceLa moelle osseuse est un tissu complexe peuplé par divers types cellulaires. Elle est le siège de l'hématopoïèse, processus selon lequel les cellules souches hématopoïétiques vont mener à la formation des cellules hautement spécialisées du sang. Comme pour tous les tissus des organismes multicellulaires, les CSH sont très rares mais en renouvellement constant ce qui leur permet une production des cellules hématopoïétiques tout au long de la vie de l'individu. Cette longévité est régulée par des facteurs moléculaires intrinsèques dont les facteurs de transcription et les régulateurs épigénétiques. Cependant, ces propriétés intrinsèques sont largement modulées par différents facteurs issus de leur environnement immédiat et produits à la fois par les cellules hématopoïétiques et non hématopoïétiques. En effet, en plus d'être le siège de l'hématopoïèse, la moelle osseuse abrite une multitude de populations cellulaires très diverses de type mésenchymateux, endothélial, myofibroblastique et nerveux formant des territoires où les CSH sont enclavées. Dans ces territoires, les molécules d'adhérence, les cytokines et les chimiokines déterminent la localisation spatiale des CSH qui se répartissent au niveau de structures anatomiques spécialisées appelés « niches ». Certaines de ces niches hébergent préférentiellement des CSH activées tandis que d'autres maintiennent le pool de CSH dans un état de quiescence. Au travers d'interactions cellules-cellules ou cellules-protéines de la matrice extracellulaire, les propriétés d'autorenouvellement et de différentiation peuvent ainsi perdurer tout au long de la vie de l'individu

Titre Environnement de la moelle osseuse et domiciliation des cellules souches hématopoïétiques

International audienceLa moelle osseuse est un tissu complexe peuplé par divers types cellulaires. Elle est le siège de l'hématopoïèse, processus selon lequel les cellules souches hématopoïétiques vont mener à la formation des cellules hautement spécialisées du sang. Comme pour tous les tissus des organismes multicellulaires, les CSH sont très rares mais en renouvellement constant ce qui leur permet une production des cellules hématopoïétiques tout au long de la vie de l'individu. Cette longévité est régulée par des facteurs moléculaires intrinsèques dont les facteurs de transcription et les régulateurs épigénétiques. Cependant, ces propriétés intrinsèques sont largement modulées par différents facteurs issus de leur environnement immédiat et produits à la fois par les cellules hématopoïétiques et non hématopoïétiques. En effet, en plus d'être le siège de l'hématopoïèse, la moelle osseuse abrite une multitude de populations cellulaires très diverses de type mésenchymateux, endothélial, myofibroblastique et nerveux formant des territoires où les CSH sont enclavées. Dans ces territoires, les molécules d'adhérence, les cytokines et les chimiokines déterminent la localisation spatiale des CSH qui se répartissent au niveau de structures anatomiques spécialisées appelés « niches ». Certaines de ces niches hébergent préférentiellement des CSH activées tandis que d'autres maintiennent le pool de CSH dans un état de quiescence. Au travers d'interactions cellules-cellules ou cellules-protéines de la matrice extracellulaire, les propriétés d'autorenouvellement et de différentiation peuvent ainsi perdurer tout au long de la vie de l'individu

CABLES1 Deficiency Impairs Quiescence and Stress Responses of Hematopoietic Stem Cells in Intrinsic and Extrinsic Manners

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CXCL12/CXCR4 pathway is activated by oncogenic JAK2 in a PI3K-dependent manner

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In vitro and in vivo efficacy of an anti-CD203c conjugated antibody (AGS-16C3F) in mouse models of advanced systemic mastocytosis

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CXCR4/CXCL12 axis counteracts hematopoietic stem cell exhaustion through selective protection against oxidative stress

International audienceHematopoietic stem cells (HSCs) undergo self-renewal to maintain hematopoietic homeostasis for lifetime, which is regulated by the bone marrow (BM) microenvironment. The chemokine receptor CXCR4 and its ligand CXCL12 are critical factors supporting quiescence and BM retention of HSCs. Here, we report an unknown function of CXCR4/CXCL12 axis in the protection of HSCs against oxidative stress. Disruption of CXCR4 receptor in mice leads to increased endogenous production of reactive oxygen species (ROS), resulting in p38 MAPK activation, increased DNA double-strand breaks and apoptosis leading to marked reduction in HSC repopulating potential. Increased ROS levels are directly responsible for exhaustion of the HSC pool and are not linked to loss of quiescence of CXCR4-deficient HSCs. Furthermore, we report that CXCL12 has a direct rescue effect on oxidative stress-induced HSC damage at the mitochondrial level. These data highlight the importance of CXCR4/CXCL12 axis in the regulation of lifespan of HSCs by limiting ROS generation and genotoxic stress. Reactive oxygen species (ROS) are produced during oxidative respiration or through exogenous environmental stresses, such as ionizing radiations or genotoxic treatments. Physiological concentrations of ROS play a role in signal transduction 1,2 , but at high concentrations they can oxidize cell constituents leading to protein carbonyla-tion, lipid peroxidation and DNA damage that activate multiple apoptosis pathways 3. By far, the most important source for ROS is mitochondria 4,5 and their endogenous production as by-products of aerobic respiration is thought to be the cause of most oxidative damages observed in mammals and particularly during aging 6,7. To avoid accumulation of oxidative stress, cells have evolved mechanisms to fine-tune ROS levels. They involve distinct groups of specialized proteins such as superoxide dismutase (SOD), catalase and glutathione peroxidase. Reduced glutathione (GSH), which also exists in the cell in its oxidized form (GSSG), is considered as the most abundant molecule among endogenous antioxidants. Alteration in its redox status serves as an indicator of oxida-tive stress when antioxidant defense mechanisms are not completely efficient and is a common feature of ageing and many pathological situations including AIDS, neurodegenerative diseases and cancer. Hematopoietic stem cells (HSCs) are defined as cells capable of both self-renewal and differentiation into any of the hematopoietic cell lineages, properties that allow hematopoietic reconstitution 8,9. Long-term maintenance of HSCs is precisely regulated by the equilibrium between proliferation and quiescence to maintain their number

Dual role of EZH2 in megakaryocyte differentiation

International audienceAbstract EZH2, the enzymatic component of PRC2, has been identified as a key factor in hematopoiesis. EZH2 loss-of-function mutations have been found in myeloproliferative neoplasms, particularly in myelofibrosis, but the precise function of EZH2 in megakaryopoiesis is not fully delineated. Here, we show that EZH2 inhibition by small molecules and short hairpin RNA induces megakaryocyte (MK) commitment by accelerating lineage marker acquisition without change in proliferation. Later in differentiation, EZH2 inhibition blocks proliferation and polyploidization and decreases proplatelet formation. EZH2 inhibitors similarly reduce MK polyploidization and proplatelet formation in vitro and platelet levels in vivo in a JAK2V617F background. In transcriptome profiling, the defect in proplatelet formation was associated with an aberrant actin cytoskeleton regulation pathway, whereas polyploidization was associated with an inhibition of expression of genes involved in DNA replication and repair and an upregulation of cyclin-dependent kinase inhibitors, particularly CDKN1A and CDKN2D. The knockdown of CDKN1A and to a lesser extent CDKN2D could partially rescue the percentage of polyploid MKs. Moreover, H3K27me3 and EZH2 chromatin immunoprecipitation assays revealed that CDKN1A is a direct EZH2 target and CDKN2D expression is not directly regulated by EZH2, suggesting that EZH2 controls MK polyploidization directly through CDKN1A and indirectly through CDKN2D