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1.
Blood ; 136(4): 441-454, 2020 07 23.
Artículo en Inglés | MEDLINE | ID: mdl-32369572

RESUMEN

Chemotherapy and irradiation cause DNA damage to hematopoietic stem cells (HSCs), leading to HSC depletion and dysfunction and the risk of malignant transformation over time. Extrinsic regulation of HSC DNA repair is not well understood, and therapies to augment HSC DNA repair following myelosuppression remain undeveloped. We report that epidermal growth factor receptor (EGFR) regulates DNA repair in HSCs following irradiation via activation of the DNA-dependent protein kinase-catalytic subunit (DNA-PKcs) and nonhomologous end joining (NHEJ). We show that hematopoietic regeneration in vivo following total body irradiation is dependent upon EGFR-mediated repair of DNA damage via activation of DNA-PKcs. Conditional deletion of EGFR in hematopoietic stem and progenitor cells (HSPCs) significantly decreased DNA-PKcs activity following irradiation, causing increased HSC DNA damage and depressed HSC recovery over time. Systemic administration of epidermal growth factor (EGF) promoted HSC DNA repair and rapid hematologic recovery in chemotherapy-treated mice and had no effect on acute myeloid leukemia growth in vivo. Further, EGF treatment drove the recovery of human HSCs capable of multilineage in vivo repopulation following radiation injury. Whole-genome sequencing analysis revealed no increase in coding region mutations in HSPCs from EGF-treated mice, but increased intergenic copy number variant mutations were detected. These studies demonstrate that EGF promotes HSC DNA repair and hematopoietic regeneration in vivo via augmentation of NHEJ. EGF has therapeutic potential to promote human hematopoietic regeneration, and further studies are warranted to assess long-term hematopoietic effects.


Asunto(s)
Reparación del ADN por Unión de Extremidades , Receptores ErbB/metabolismo , Hematopoyesis/fisiología , Células Madre Hematopoyéticas/metabolismo , Regeneración , Animales , Proteínas de Unión al Calcio/genética , Proteínas de Unión al Calcio/metabolismo , Daño del ADN , Proteína Quinasa Activada por ADN/genética , Proteína Quinasa Activada por ADN/metabolismo , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Factor de Crecimiento Epidérmico/genética , Factor de Crecimiento Epidérmico/metabolismo , Receptores ErbB/genética , Células Madre Hematopoyéticas/citología , Humanos , Ratones
2.
Nat Commun ; 10(1): 3667, 2019 08 14.
Artículo en Inglés | MEDLINE | ID: mdl-31413255

RESUMEN

Receptor type protein tyrosine phosphatase-sigma (PTPσ) is primarily expressed by adult neurons and regulates neural regeneration. We recently discovered that PTPσ is also expressed by hematopoietic stem cells (HSCs). Here, we describe small molecule inhibitors of PTPσ that promote HSC regeneration in vivo. Systemic administration of the PTPσ inhibitor, DJ001, or its analog, to irradiated mice promotes HSC regeneration, accelerates hematologic recovery, and improves survival. Similarly, DJ001 administration accelerates hematologic recovery in mice treated with 5-fluorouracil chemotherapy. DJ001 displays high specificity for PTPσ and antagonizes PTPσ via unique non-competitive, allosteric binding. Mechanistically, DJ001 suppresses radiation-induced HSC apoptosis via activation of the RhoGTPase, RAC1, and induction of BCL-XL. Furthermore, treatment of irradiated human HSCs with DJ001 promotes the regeneration of human HSCs capable of multilineage in vivo repopulation. These studies demonstrate the therapeutic potential of selective, small-molecule PTPσ inhibitors for human hematopoietic regeneration.


Asunto(s)
Apoptosis/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Células Madre Hematopoyéticas/efectos de los fármacos , Proteínas Tirosina Fosfatasas Clase 2 Similares a Receptores/antagonistas & inhibidores , Regeneración/efectos de los fármacos , Regulación Alostérica , Animales , Antimetabolitos Antineoplásicos/farmacología , Apoptosis/efectos de la radiación , Fluorouracilo/farmacología , Células Madre Hematopoyéticas/efectos de la radiación , Humanos , Ratones , Radiación , Regeneración/efectos de la radiación , Proteína bcl-X/efectos de los fármacos , Proteína bcl-X/metabolismo , Proteína de Unión al GTP rac1/efectos de los fármacos , Proteína de Unión al GTP rac1/metabolismo , Proteínas de Unión al GTP rho/efectos de los fármacos , Proteínas de Unión al GTP rho/metabolismo
3.
Nat Med ; 23(1): 91-99, 2017 01.
Artículo en Inglés | MEDLINE | ID: mdl-27918563

RESUMEN

The role of osteolineage cells in regulating hematopoietic stem cell (HSC) regeneration following myelosuppression is not well understood. Here we show that deletion of the pro-apoptotic genes Bak and Bax in osterix (Osx, also known as Sp7 transcription factor 7)-expressing cells in mice promotes HSC regeneration and hematopoietic radioprotection following total body irradiation. These mice showed increased bone marrow (BM) levels of the protein dickkopf-1 (Dkk1), which was produced in Osx-expressing BM cells. Treatment of irradiated HSCs with Dkk1 in vitro increased the recovery of both long-term repopulating HSCs and progenitor cells, and systemic administration of Dkk1 to irradiated mice increased hematopoietic recovery and improved survival. Conversely, inducible deletion of one allele of Dkk1 in Osx-expressing cells in adult mice inhibited the recovery of BM stem and progenitor cells and of complete blood counts following irradiation. Dkk1 promoted hematopoietic regeneration via both direct effects on HSCs, in which treatment with Dkk1 decreased the levels of mitochondrial reactive oxygen species and suppressed senescence, and indirect effects on BM endothelial cells, in which treatment with Dkk1 induced epidermal growth factor (EGF) secretion. Accordingly, blockade of the EGF receptor partially abrogated Dkk1-mediated hematopoietic recovery. These data identify Dkk1 as a regulator of hematopoietic regeneration and demonstrate paracrine cross-talk between BM osteolineage cells and endothelial cells in regulating hematopoietic reconstitution following injury.


Asunto(s)
Células de la Médula Ósea/metabolismo , Autorrenovación de las Células , Células Madre Hematopoyéticas/metabolismo , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Osteoblastos/metabolismo , Regeneración , Factores de Transcripción/metabolismo , Irradiación Corporal Total , Animales , Médula Ósea/metabolismo , Citocinas/metabolismo , Células Endoteliales/metabolismo , Ensayo de Inmunoadsorción Enzimática , Factor de Crecimiento Epidérmico/metabolismo , Receptores ErbB/antagonistas & inhibidores , Citometría de Flujo , Perfilación de la Expresión Génica , Células Madre Hematopoyéticas/efectos de los fármacos , Células Madre Hematopoyéticas/efectos de la radiación , Péptidos y Proteínas de Señalización Intercelular/farmacología , Ratones , Mitocondrias/metabolismo , Traumatismos Experimentales por Radiación , Especies Reactivas de Oxígeno , Factor de Transcripción Sp7 , Proteína Destructora del Antagonista Homólogo bcl-2/genética , Proteína X Asociada a bcl-2/genética
4.
Cell Rep ; 17(6): 1584-1594, 2016 11 01.
Artículo en Inglés | MEDLINE | ID: mdl-27806297

RESUMEN

Imprinted genes are differentially expressed by adult stem cells, but their functions in regulating adult stem cell fate are incompletely understood. Here we show that growth factor receptor-bound protein 10 (Grb10), an imprinted gene, regulates hematopoietic stem cell (HSC) self-renewal and regeneration. Deletion of the maternal allele of Grb10 in mice (Grb10m/+ mice) substantially increased HSC long-term repopulating capacity, as compared to that of Grb10+/+ mice. After total body irradiation (TBI), Grb10m/+ mice demonstrated accelerated HSC regeneration and hematopoietic reconstitution, as compared to Grb10+/+ mice. Grb10-deficient HSCs displayed increased proliferation after competitive transplantation or TBI, commensurate with upregulation of CDK4 and Cyclin E. Furthermore, the enhanced HSC regeneration observed in Grb10-deficient mice was dependent on activation of the Akt/mTORC1 pathway. This study reveals a function for the imprinted gene Grb10 in regulating HSC self-renewal and regeneration and suggests that the inhibition of Grb10 can promote hematopoietic regeneration in vivo.


Asunto(s)
Autorrenovación de las Células/genética , Proteína Adaptadora GRB10/deficiencia , Eliminación de Gen , Impresión Genómica , Células Madre Hematopoyéticas/citología , Células Madre Hematopoyéticas/metabolismo , Regeneración , Animales , Células de la Médula Ósea/citología , Proliferación Celular , Proteína Adaptadora GRB10/metabolismo , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Ratones Endogámicos C57BL , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transducción de Señal , Irradiación Corporal Total
5.
J Clin Invest ; 125(1): 177-82, 2015 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-25415437

RESUMEN

Hematopoietic stem cell (HSC) function is regulated by activation of receptor tyrosine kinases (RTKs). Receptor protein tyrosine phosphatases (PTPs) counterbalance RTK signaling; however, the functions of receptor PTPs in HSCs remain incompletely understood. We found that a receptor PTP, PTPσ, was substantially overexpressed in mouse and human HSCs compared with more mature hematopoietic cells. Competitive transplantation of bone marrow cells from PTPσ-deficient mice revealed that the loss of PTPσ substantially increased long-term HSC-repopulating capacity compared with BM cells from control mice. While HSCs from PTPσ-deficient mice had no apparent alterations in cell-cycle status, apoptosis, or homing capacity, these HSCs exhibited increased levels of activated RAC1, a RhoGTPase that regulates HSC engraftment capacity. shRNA-mediated silencing of PTPσ also increased activated RAC1 levels in wild-type HSCs. Functionally, PTPσ-deficient BM cells displayed increased cobblestone area-forming cell (CAFC) capacity and augmented transendothelial migration capacity, which was abrogated by RAC inhibition. Specific selection of human cord blood CD34⁺CD38⁻CD45RA⁻lin⁻ PTPσ⁻ cells substantially increased the repopulating capacity of human HSCs compared with CD34⁺CD38⁻CD45RA⁻lin⁻ cells and CD34⁺CD38⁻CD45RA⁻lin⁻PTPσ⁺ cells. Our results demonstrate that PTPσ regulates HSC functional capacity via RAC1 inhibition and suggest that selecting for PTPσ-negative human HSCs may be an effective strategy for enriching human HSCs for transplantation.


Asunto(s)
Células Madre Hematopoyéticas/enzimología , Proteínas Tirosina Fosfatasas Clase 2 Similares a Receptores/fisiología , Animales , Células Cultivadas , Trasplante de Células Madre Hematopoyéticas , Humanos , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Endogámicos NOD , Neuropéptidos/metabolismo , Migración Transendotelial y Transepitelial , Proteína de Unión al GTP rac1/metabolismo
6.
J Clin Invest ; 124(11): 4753-8, 2014 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-25250571

RESUMEN

Hematopoietic stem cells (HSCs) are highly susceptible to ionizing radiation-mediated death via induction of ROS, DNA double-strand breaks, and apoptotic pathways. The development of therapeutics capable of mitigating ionizing radiation-induced hematopoietic toxicity could benefit both victims of acute radiation sickness and patients undergoing hematopoietic cell transplantation. Unfortunately, therapies capable of accelerating hematopoietic reconstitution following lethal radiation exposure have remained elusive. Here, we found that systemic administration of pleiotrophin (PTN), a protein that is secreted by BM-derived endothelial cells, substantially increased the survival of mice following radiation exposure and after myeloablative BM transplantation. In both models, PTN increased survival by accelerating the recovery of BM hematopoietic stem and progenitor cells in vivo. PTN treatment promoted HSC regeneration via activation of the RAS pathway in mice that expressed protein tyrosine phosphatase receptor-zeta (PTPRZ), whereas PTN treatment did not induce RAS signaling in PTPRZ-deficient mice, suggesting that PTN-mediated activation of RAS was dependent upon signaling through PTPRZ. PTN strongly inhibited HSC cycling following irradiation, whereas RAS inhibition abrogated PTN-mediated induction of HSC quiescence, blocked PTN-mediated recovery of hematopoietic stem and progenitor cells, and abolished PTN-mediated survival of irradiated mice. These studies demonstrate the therapeutic potential of PTN to improve survival after myeloablation and suggest that PTN-mediated hematopoietic regeneration occurs in a RAS-dependent manner.


Asunto(s)
Proteínas Portadoras/fisiología , Citocinas/fisiología , Hematopoyesis , Proteínas ras/metabolismo , Animales , Trasplante de Médula Ósea , Células Cultivadas , Ratones , Traumatismos Experimentales por Radiación/terapia , Regeneración , Transducción de Señal
7.
Nat Med ; 19(3): 295-304, 2013 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-23377280

RESUMEN

The mechanisms that regulate hematopoietic stem cell (HSC) regeneration after myelosuppressive injury are not well understood. We identified epidermal growth factor (EGF) to be highly enriched in the bone marrow serum of mice bearing deletion of Bak and Bax in TIE2-expressing cells in Tie2Cre; Bak1(-/-); Bax(flox/-) mice. These mice showed radioprotection of the HSC pool and 100% survival after a lethal dose of total-body irradiation (TBI). Bone marrow HSCs from wild-type mice expressed functional EGF receptor (EGFR), and systemic administration of EGF promoted the recovery of the HSC pool in vivo and improved the survival of mice after TBI. Conversely, administration of erlotinib, an EGFR antagonist, decreased both HSC regeneration and the survival of mice after TBI. Mice with EGFR deficiency in VAV-expressing hematopoietic cells also had delayed recovery of bone marrow stem and progenitor cells after TBI. Mechanistically, EGF reduced radiation-induced apoptosis of HSCs and mediated this effect through repression of the proapoptotic protein PUMA. Our findings show that EGFR signaling regulates HSC regeneration after myelosuppressive injury.


Asunto(s)
Factor de Crecimiento Epidérmico/metabolismo , Factor de Crecimiento Epidérmico/farmacología , Receptores ErbB/metabolismo , Hematopoyesis , Células Madre Hematopoyéticas/efectos de la radiación , Traumatismos Experimentales por Radiación/tratamiento farmacológico , Regeneración , Animales , Apoptosis/efectos de la radiación , Proteínas Reguladoras de la Apoptosis/biosíntesis , Médula Ósea/efectos de la radiación , Células de la Médula Ósea/efectos de la radiación , Células Cultivadas , Receptores ErbB/antagonistas & inhibidores , Clorhidrato de Erlotinib , Femenino , Células Madre Hematopoyéticas/fisiología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Inhibidores de Proteínas Quinasas/farmacología , Quinazolinas/farmacología , Transducción de Señal/efectos de la radiación , Proteínas Supresoras de Tumor/biosíntesis , Irradiación Corporal Total , Proteína Destructora del Antagonista Homólogo bcl-2/genética , Proteína X Asociada a bcl-2/genética
8.
Cell Rep ; 2(4): 964-75, 2012 Oct 25.
Artículo en Inglés | MEDLINE | ID: mdl-23084748

RESUMEN

The mechanisms through which the bone marrow (BM) microenvironment regulates hematopoietic stem cell (HSC) fate remain incompletely understood. We examined the role of the heparin-binding growth factor pleiotrophin (PTN) in regulating HSC function in the niche. PTN(-/-) mice displayed significantly decreased BM HSC content and impaired hematopoietic regeneration following myelosuppression. Conversely, mice lacking protein tyrosine phosphatase receptor zeta, which is inactivated by PTN, displayed significantly increased BM HSC content. Transplant studies revealed that PTN action was not HSC autonomous, but rather was mediated by the BM microenvironment. Interestingly, PTN was differentially expressed and secreted by BM sinusoidal endothelial cells within the vascular niche. Furthermore, systemic administration of anti-PTN antibody in mice substantially impaired both the homing of hematopoietic progenitor cells to the niche and the retention of BM HSCs in the niche. PTN is a secreted component of the BM vascular niche that regulates HSC self-renewal and retention in vivo.


Asunto(s)
Células de la Médula Ósea/citología , Proteínas Portadoras/metabolismo , Citocinas/metabolismo , Células Madre Hematopoyéticas/citología , Animales , Anticuerpos/inmunología , Proteínas Portadoras/genética , Células Cultivadas , Quimiocina CXCL12/metabolismo , Citocinas/deficiencia , Citocinas/genética , Trasplante de Células Madre Hematopoyéticas , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Proteínas Tirosina Fosfatasas Clase 5 Similares a Receptores/metabolismo , Nicho de Células Madre
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