Protective effects of HemoHIM on immune and hematopoietic systems against γ-irradiation.

PubMed

Park, Hae-Ran; Jo, Sung-Kee; Jung, Uhee; Yee, Sung-Tae; Kim, Sung-Ho

2014-02-01

We examined the effect of HemoHIM on the protective efficacy of hematopoietic stem cells and on the recovery of immune cells against sublethal doses of ionizing radiation. Two-month-old mice were exposed to γ-rays at a dose of 8, 6.5, or 5 Gy for a30-day survival study, endogenous spleen colony formation, or other experiments, respectively. HemoHIM was injected intraperitoneally before and after irradiation. Our results showed that HemoHIM significantly decreased the mortality of sublethally irradiated mice. The HemoHIM administration decreased the apoptosis of bone marrow cells in irradiated mice. On the other hand, HemoHIM increased the formation of endogenous spleen colony in irradiated mice. In irradiated mice, the recovery of total leukocytes in the peripheral blood and lymphocytes in the spleen were enhanced significantly by HemoHIM. Moreover, the function of B cells, T cells, and NK cells regenerated in irradiated mice were significantly improved by the administration of HemoHIM. HemoHIM showed an ideal radioprotector for protecting hematopoietic stem cells and for accelerating the recovery of immune cells. We propose HemoHIM as a beneficial supplement drug during radiotherapy to alleviate adverse radiation-induced effects for cancer patients. Copyright © 2013 John Wiley & Sons, Ltd.

Whole-Body Proton Irradiation Causes Long-Term Damage to Hematopoietic Stem Cells in Mice

PubMed Central

Chang, Jianhui; Feng, Wei; Wang, Yingying; Luo, Yi; Allen, Antiño R.; Koturbash, Igor; Turner, Jennifer; Stewart, Blair; Raber, Jacob; Hauer-Jensen, Martin; Zhou, Daohong; Shao, Lijian

2016-01-01

Space flight poses certain health risks to astronauts, including exposure to space radiation, with protons accounting for more than 80% of deep-space radiation. Proton radiation is also now being used with increasing frequency in the clinical setting to treat cancer. For these reasons, there is an urgent need to better understand the biological effects of proton radiation on the body. Such improved understanding could also lead to more accurate assessment of the potential health risks of proton radiation, as well as the development of improved strategies to prevent and mitigate its adverse effects. Previous studies have shown that exposure to low doses of protons is detrimental to mature leukocyte populations in peripheral blood, however, the underlying mechanisms are not known. Some of these detriments may be attributable to damage to hematopoietic stem cells (HSCs) that have the ability to self-renew, proliferate and differentiate into different lineages of blood cells through hematopoietic progenitor cells (HPCs). The goal of this study was to investigate the long-term effects of low-dose proton irradiation on HSCs. We exposed C57BL/6J mice to 1.0 Gy whole-body proton irradiation (150 MeV) and then studied the effects of proton radiation on HSCs and HPCs in the bone marrow (BM) 22 weeks after the exposure. The results showed that mice exposed to 1.0 Gy whole-body proton irradiation had a significant and persistent reduction of BM HSCs compared to unirradiated controls. In contrast, no significant changes were observed in BM HPCs after proton irradiation. Furthermore, irradiated HSCs and their progeny exhibited a significant impairment in clonogenic function, as revealed by the cobblestone area-forming cell (CAFC) and colony-forming cell assays, respectively. These long-term effects of proton irradiation on HSCs may be attributable to the induction of chronic oxidative stress in HSCs, because HSCs from irradiated mice exhibited a significant increase in NADPH

Lentiviral-mediated genetic correction of hematopoietic and mesenchymal progenitor cells from Fanconi anemia patients.

PubMed

Jacome, Ariana; Navarro, Susana; Río, Paula; Yañez, Rosa M; González-Murillo, Africa; Lozano, M Luz; Lamana, Maria Luisa; Sevilla, Julian; Olive, Teresa; Diaz-Heredia, Cristina; Badell, Isabel; Estella, Jesus; Madero, Luis; Guenechea, Guillermo; Casado, José; Segovia, Jose C; Bueren, Juan A

2009-06-01

Previous clinical trials based on the genetic correction of purified CD34(+) cells with gamma-retroviral vectors have demonstrated clinical efficacy in different monogenic diseases, including X-linked severe combined immunodeficiency, adenosine deaminase deficient severe combined immunodeficiency and chronic granulomatous disease. Similar protocols, however, failed to engraft Fanconi anemia (FA) patients with genetically corrected cells. In this study, we first aimed to correlate the hematological status of 27 FA patients with CD34(+) cell values determined in their bone marrow (BM). Strikingly, no correlation between these parameters was observed, although good correlations were obtained when numbers of colony-forming cells (CFCs) were considered. Based on these results, and because purified FA CD34(+) cells might have suboptimal repopulating properties, we investigated the possibility of genetically correcting unselected BM samples from FA patients. Our data show that the lentiviral transduction of unselected FA BM cells mediates an efficient phenotypic correction of hematopoietic progenitor cells and also of CD34(-) mesenchymal stromal cells (MSCs), with a reported role in hematopoietic engraftment. Our results suggest that gene therapy protocols appropriate for the treatment of different monogenic diseases may not be adequate for stem cell diseases like FA. We propose a new approach for the gene therapy of FA based on the rapid transduction of unselected hematopoietic grafts with lentiviral vectors (LVs).

Lentiviral-mediated Genetic Correction of Hematopoietic and Mesenchymal Progenitor Cells From Fanconi Anemia Patients

PubMed Central

Jacome, Ariana; Navarro, Susana; Río, Paula; Yañez, Rosa M; González-Murillo, Africa; Luz Lozano, M; Lamana, Maria Luisa; Sevilla, Julian; Olive, Teresa; Diaz-Heredia, Cristina; Badell, Isabel; Estella, Jesus; Madero, Luis; Guenechea, Guillermo; Casado, José; Segovia, Jose C; Bueren, Juan A

2009-01-01

Previous clinical trials based on the genetic correction of purified CD34+ cells with γ-retroviral vectors have demonstrated clinical efficacy in different monogenic diseases, including X-linked severe combined immunodeficiency, adenosine deaminase deficient severe combined immunodeficiency and chronic granulomatous disease. Similar protocols, however, failed to engraft Fanconi anemia (FA) patients with genetically corrected cells. In this study, we first aimed to correlate the hematological status of 27 FA patients with CD34+ cell values determined in their bone marrow (BM). Strikingly, no correlation between these parameters was observed, although good correlations were obtained when numbers of colony-forming cells (CFCs) were considered. Based on these results, and because purified FA CD34+ cells might have suboptimal repopulating properties, we investigated the possibility of genetically correcting unselected BM samples from FA patients. Our data show that the lentiviral transduction of unselected FA BM cells mediates an efficient phenotypic correction of hematopoietic progenitor cells and also of CD34− mesenchymal stromal cells (MSCs), with a reported role in hematopoietic engraftment. Our results suggest that gene therapy protocols appropriate for the treatment of different monogenic diseases may not be adequate for stem cell diseases like FA. We propose a new approach for the gene therapy of FA based on the rapid transduction of unselected hematopoietic grafts with lentiviral vectors (LVs). PMID:19277017

Hematopoietic Stem Cells as a Novel Source of Dental Tissue Cells.

PubMed

Wilson, Katie R; Kang, In-Hong; Baliga, Uday; Xiong, Ying; Chatterjee, Shilpak; Moore, Emily; Parthiban, Beneta; Thyagarajan, Krishnamurthy; Borke, James L; Mehrotra, Shikhar; Kirkwood, Keith L; LaRue, Amanda C; Ogawa, Makio; Mehrotra, Meenal

2018-05-23

While earlier studies have suggested that cells positive for hematopoietic markers can be found in dental tissues, it has yet to be confirmed. To conclusively demonstrate this, we utilized a unique transgenic model in which all hematopoietic cells are green fluorescent protein + (GFP + ). Pulp, periodontal ligament (PDL) and alveolar bone (AvB) cell culture analysis demonstrated numerous GFP + cells, which were also CD45 + (indicating hematopoietic origin) and co-expressed markers of cellular populations in pulp (dentin matrix protein-1, dentin sialophosphoprotein, alpha smooth muscle actin [ASMA], osteocalcin), in PDL (periostin, ASMA, vimentin, osteocalcin) and in AvB (Runx-2, bone sialoprotein, alkaline phosphatase, osteocalcin). Transplantation of clonal population derived from a single GFP + hematopoietic stem cell (HSC), into lethally irradiated recipient mice, demonstrated numerous GFP + cells within dental tissues of recipient mice, which also stained for markers of cell populations in pulp, PDL and AvB (used above), indicating that transplanted HSCs can differentiate into cells in dental tissues. These hematopoietic-derived cells deposited collagen and can differentiate in osteogenic media, indicating that they are functional. Thus, our studies demonstrate, for the first time, that cells in pulp, PDL and AvB can have a hematopoietic origin, thereby opening new avenues of therapy for dental diseases and injuries.

Isolation, Characterization, and Transplantation of Bone Marrow-Derived Cell Components with Hematopoietic Stem Cell Niche Properties

PubMed Central

Ahmadbeigi, Naser; Vasei, Mohammad; Gheisari, Yousof; Mortazavi, Yousef; Azadmanesh, Kayhan; Omidkhoda, Azadeh; Janzamin, Ehsan; Nardi, Nance Beyer

2013-01-01

Although the unique role of hematopoietic stem cell (HSC) niche in hematopoiesis has long been recognized, unsuccessful isolation of intact niche units limited their in vitro study, manipulation, and therapeutic application. Here, we isolated cell complexes based on size fractionation from mouse bone marrow (BM), characterized the derived cells, and transplanted them to irradiated mice. These cell complexes were the origin of both BM mesenchymal stem cells and various hematopoietic lineages when kept in appropriate culture conditions. They also had the potential of recruiting circulating HSC. Intraperitoneal transplantation of these structures into irradiated mice not only showed long-lasting hematopoietic multilineage reconstitution, but also could recover the stromal cells of BM. In conclusion, this study for the first time provides evidences on the feasibility and efficacy of transplantation of HSC in association with their native specialized microenvironment. As the molecular cross-talk between HSC and niche is crucial for their proper function, the proposed method could be considered as a novel hematopoietic transplantation strategy. PMID:23879861

Ineffective vaccination against solid tumors can be enhanced by hematopoietic cell transplantation.

PubMed

Filatenkov, Alexander; Müller, Antonia M S; Tseng, William Wei-Lin; Dejbakhsh-Jones, Sussan; Winer, Daniel; Luong, Richard; Shizuru, Judith A; Engleman, Edgar G; Strober, Samuel

2009-12-01

Vaccination with tumor Ags has not been an effective treatment for solid tumors. The goal of the current study was to determine whether a combination of vaccination and hematopoietic cell transplantation (HCT) can effectively treat primary, disseminated, or metastatic CT26 and MC38 murine colon tumors. Vaccination of tumor-bearing mice with irradiated tumor cells and CpG adjuvant failed to alter progressive tumor growth. However, mice bearing primary, disseminated lung, or metastatic liver tumors were uniformly cured after administration of total body irradiation, followed by the transplantation of hematopoietic progenitor cells and T cells from syngeneic, but not allogeneic vaccinated donors. Requirements for effective treatment of tumors included irradiation of hosts, vaccination of donors with both tumor cells and CpG, transfer of both CD4(+) and CD8(+) T cells along with progenitor cells, and ability of donor cells to produce IFN-gamma. Irradiation markedly increased the infiltration of donor T cells into the tumors, and the combined irradiation and HCT altered the balance of tumor-infiltrating cells to favor CD8(+) effector memory T cells as compared with CD4(+)CD25(+)FoxP3(+) T regulatory cells. The combination of vaccination and autologous hematopoietic cell transplantation was also effective in treating tumors. In conclusion, these findings show that otherwise ineffective vaccination to solid nonhematologic tumors can be dramatically enhanced by HCT.

Mitigating the effects of Xuebijing injection on hematopoietic cell injury induced by total body irradiation with γ rays by decreasing reactive oxygen species levels.

PubMed

Li, Deguan; Lu, Lu; Zhang, Junling; Wang, Xiaochun; Xing, Yonghua; Wu, Hongying; Yang, Xiangdong; Shi, Zhexin; Zhao, Mingfeng; Fan, Saijun; Meng, Aimin

2014-06-12

Hematopoietic injury is the most common side effect of radiotherapy. However, the methods available for the mitigating of radiation injury remain limited. Xuebijing injection (XBJ) is a traditional Chinese medicine used to treat sepsis in the clinic. In this study, we investigated the effects of XBJ on the survival rate in mice with hematopoietic injury induced by γ ray ionizing radiation (IR). Mice were intraperitoneally injected with XBJ daily for seven days after total body irradiation (TBI). Our results showed that XBJ (0.4 mL/kg) significantly increased 30-day survival rates in mice exposed to 7.5 Gy TBI. This effect may be attributable to improved preservation of white blood cells (WBCs) and hematopoietic cells, given that bone marrow (BM) cells from XBJ-treated mice produced more granulocyte-macrophage colony forming units (CFU-GM) than that in the 2 Gy/TBI group. XBJ also decreased the levels of reactive oxygen species (ROS) by increasing glutathione (GSH) and superoxide dismutase (SOD) levels in serum and attenuated the increased BM cell apoptosis caused by 2 Gy/TBI. In conclusion, these findings suggest that XBJ enhances the survival rate of irradiated mice and attenuates the effects of radiation on hematopoietic injury by decreasing ROS production in BM cells, indicating that XBJ may be a promising therapeutic candidate for reducing hematopoietic radiation injury.

Hematopoietic tissue repair under chronic low daily dose irradiation

NASA Astrophysics Data System (ADS)

Seed, T. M.

The capacity of the hematopoietic system to repair constantly accruing cellular damage under chronic, low daily dose gamma irradiation is essential for the maintenance of a functional hematopoietic system, and, in turn, long term survival. In certain individuals, however, such continuous cycles of damage and repair provide an essential inductive environment for selected types of hematopathologies, e.g., myeloid leukemia (ML). In our laboratory we have been studying temporal and causal relationships between hematopoietic capacity, associated repair functions, and propensities for hematologic disease in canines under variable levels of chronic radiation stress (0.3-26.3 cGy d^-1). Results indicate that the maximum exposure rate tolerated by the hematopoietic system is highly individual-specific (three major responding subgroups identified) and is based largely on the degree to which repair capacity, and, in turn, hematopoietic restoration, is augmented under chronic exposure. In low-tolerance individuals (prone to aplastic anemia, subgroup 1), the failure to augment basic repair functions seemingly results in a progressive accumulation of genetic and cellular damage within vital progenitorial marrow compartments (particularly marked within erythroid compartments) that results in loss of reproductive capacity and ultimately in collapse of the hematopoietic system. The high-tolerance individuals (radioaccommodated and either prone- or not prone to ML, subgroup 2 & 3) appear to minimize the accumulating damage effect of daily exposures by extending repair functions, which preserves reproductive integrity and fosters regenerative hematopoietic responses. As the strength of the regenerative response manifests the extent of repair augmentation, the relatively strong response of high-tolerance individuals progressing to patent ML suggests an insufficiency of repair quality rather than repair quantity. The kinetics of these repair-mediated, regenerative hematopoietic

Whole body proton irradiation causes acute damage to bone marrow hematopoietic progenitor and stem cells in mice.

PubMed

Chang, Jianhui; Wang, Yingying; Pathak, Rupak; Sridharan, Vijayalakshmi; Jones, Tamako; Mao, Xiao Wen; Nelson, Gregory; Boerma, Marjan; Hauer-Jensen, Martin; Zhou, Daohong; Shao, Lijian

2017-12-01

Exposure to proton irradiation during missions in deep space can lead to bone marrow injury. The acute effects of proton irradiation on hematopoietic stem and progenitor cells remain undefined and thus were investigated. We exposed male C57BL/6 mice to 0.5 and 1.0 Gy proton total body irradiation (proton-TBI, 150 MeV) and examined changes in peripheral blood cells and bone marrow (BM) progenitors and LSK cells 2 weeks after exposure. 1.0 Gy proton-TBI significantly reduced the numbers of peripheral blood cells compared to 0.5 Gy proton-TBI and unirradiated animals, while the numbers of peripheral blood cell counts were comparable between 0.5 Gy proton-TBI and unirradiated mice. The frequencies and numbers of LSK cells and CMPs in BM of 0.5 and 1.0 Gy irradiated mice were decreased in comparison to those of normal controls. LSK cells and CMPs and their progeny exhibited a radiation-induced impairment in clonogenic function. Exposure to 1.0 Gy increased cellular apoptosis but not the production of reactive oxygen species (ROS) in CMPs two weeks after irradiation. LSK cells from irradiated mice exhibited an increase in ROS production and apoptosis. Exposure to proton-TBI can induce acute damage to BM progenitors and LSK cells.

Endothelial transplantation rejuvenates aged hematopoietic stem cell function

PubMed Central

Poulos, Michael G.; Gutkin, Michael C.; Llanos, Pierre; Gilleran, Katherine; Rabbany, Sina Y.; Butler, Jason M.

2017-01-01

Age-related changes in the hematopoietic compartment are primarily attributed to cell-intrinsic alterations in hematopoietic stem cells (HSCs); however, the contribution of the aged microenvironment has not been adequately evaluated. Understanding the role of the bone marrow (BM) microenvironment in supporting HSC function may prove to be beneficial in treating age-related functional hematopoietic decline. Here, we determined that aging of endothelial cells (ECs), a critical component of the BM microenvironment, was sufficient to drive hematopoietic aging phenotypes in young HSCs. We used an ex vivo hematopoietic stem and progenitor cell/EC (HSPC/EC) coculture system as well as in vivo EC infusions following myelosuppressive injury in mice to demonstrate that aged ECs impair the repopulating activity of young HSCs and impart a myeloid bias. Conversely, young ECs restored the repopulating capacity of aged HSCs but were unable to reverse the intrinsic myeloid bias. Infusion of young, HSC-supportive BM ECs enhanced hematopoietic recovery following myelosuppressive injury and restored endogenous HSC function in aged mice. Coinfusion of young ECs augmented aged HSC engraftment and enhanced overall survival in lethally irradiated mice by mitigating damage to the BM vascular microenvironment. These data lay the groundwork for the exploration of EC therapies that can serve as adjuvant modalities to enhance HSC engraftment and accelerate hematopoietic recovery in the elderly population following myelosuppressive regimens. PMID:29035282

Sodium Caseinate (CasNa) Induces Mobilization of Hematopoietic Stem Cells in a BALB/c Mouse Model

PubMed Central

Santiago-Osorio, Edelmiro; Ledesma-Martínez, Edgar; Aguiñiga-Sánchez, Itzen; Poblano-Pérez, Ignacio; Weiss-Steider, Benny; Montesinos-Montesinos, Juan José; de Lourdes Mora-García, María

2015-01-01

Background Hematopoietic stem cells transplantation has high clinical potential against a wide variety of hematologic, metabolic, and autoimmune diseases and solid tumors. Clinically, hematopoietic stem cells derived from peripheral blood are currently used more than those obtained from sources such as bone marrow. However, mobilizing agents used in the clinic tend to fail in high rates, making the number of mobilized cells insufficient for transplantation. We investigated whether sodium caseinate induces functional mobilization of hematopoietic stem cells into peripheral blood of Balb/c mice. Material/Methods Using a mouse model, we administrated sodium caseinate or Plerixafor, a commercial mobilizing agent, and analyzed counts of hematopoietic stem cells in peripheral blood, and then cells were transplanted into lethally irradiated mice to restore hematopoiesis. All assays were performed at least twice. Results We found that sodium caseinate increases the number of mononuclear cells in peripheral blood with the immunophenotype of hematopoietic stem cells (0.2 to 0.5% LSK cells), allowing them to form colonies of various cell lineages in semisolid medium (p<0.05). This effect is similar to that of Plerixafor, and cells transplanted into lethally irradiated mice can restore hematopoiesis at higher percentages than mononuclear cells mobilized by Plerixafor (40% vs. 20%, respectively). Further, a secondary transplant rescued a separate group of irradiated mice from death, proving definitive evidence of hematopoietic reconstitution after hematopoietic stem cells transplantation. Data are presented as mean ± standard deviation. To determine significant differences between the data, one-way ANOVA and the Tukey test were used. Conclusions Collectively these results show the utility of sodium caseinate as a mobilizer of hematopoietic stem cells and its potential clinical application in transplantation settings. PMID:26409928

Sodium Caseinate (CasNa) Induces Mobilization of Hematopoietic Stem Cells in a BALB/c Mouse Model.

PubMed

Santiago-Osorio, Edelmiro; Ledesma-Martínez, Edgar; Aguiñiga-Sánchez, Itzen; Poblano-Pérez, Ignacio; Weiss-Steider, Benny; Montesinos-Montesinos, Juan José; Mora-García, María de Lourdes

2015-09-25

BACKGROUND Hematopoietic stem cells transplantation has high clinical potential against a wide variety of hematologic, metabolic, and autoimmune diseases and solid tumors. Clinically, hematopoietic stem cells derived from peripheral blood are currently used more than those obtained from sources such as bone marrow. However, mobilizing agents used in the clinic tend to fail in high rates, making the number of mobilized cells insufficient for transplantation. We investigated whether sodium caseinate induces functional mobilization of hematopoietic stem cells into peripheral blood of Balb/c mice. MATERIAL AND METHODS Using a mouse model, we administrated sodium caseinate or Plerixafor, a commercial mobilizing agent, and analyzed counts of hematopoietic stem cells in peripheral blood, and then cells were transplanted into lethally irradiated mice to restore hematopoiesis. All assays were performed at least twice. RESULTS We found that sodium caseinate increases the number of mononuclear cells in peripheral blood with the immunophenotype of hematopoietic stem cells (0.2 to 0.5% LSK cells), allowing them to form colonies of various cell lineages in semisolid medium (p<0.05). This effect is similar to that of Plerixafor, and cells transplanted into lethally irradiated mice can restore hematopoiesis at higher percentages than mononuclear cells mobilized by Plerixafor (40% vs. 20%, respectively). Further, a secondary transplant rescued a separate group of irradiated mice from death, proving definitive evidence of hematopoietic reconstitution after hematopoietic stem cells transplantation. Data are presented as mean ± standard deviation. To determine significant differences between the data, one-way ANOVA and the Tukey test were used. CONCLUSIONS Collectively these results show the utility of sodium caseinate as a mobilizer of hematopoietic stem cells and its potential clinical application in transplantation settings.

Long-term hematopoietic stem cell damage in a murine model of the hematopoietic syndrome of the acute radiation syndrome.

PubMed

Chua, Hui Lin; Plett, P Artur; Sampson, Carol H; Joshi, Mandar; Tabbey, Rebeka; Katz, Barry P; MacVittie, Thomas J; Orschell, Christie M

2012-10-01

Residual bone marrow damage (RBMD) persists for years following exposure to radiation and is believed to be due to decreased self-renewal potential of radiation-damaged hematopoietic stem cells (HSC). Current literature has examined primarily sublethal doses of radiation and time points within a few months of exposure. In this study, the authors examined RBMD in mice surviving lethal doses of total body ionizing irradiation (TBI) in a murine model of the Hematopoietic Syndrome of the Acute Radiation Syndrome (H-ARS). Survivors were analyzed at various time points up to 19 mo post-TBI for hematopoietic function. The competitive bone marrow (BM) repopulating potential of 150 purified c-Kit+ Sca-1+ lineage- CD150+ cells (KSLCD150+) remained severely deficient throughout the study compared to KSLCD150+ cells from non-TBI age-matched controls. The minimal engraftment from these TBI HSCs is predominantly myeloid, with minimal production of lymphocytes both in vitro and in vivo. All classes of blood cells as well as BM cellularity were significantly decreased in TBI mice, especially at later time points as mice aged. Primitive BM hematopoietic cells (KSLCD150+) displayed significantly increased cell cycling in TBI mice at all time points, which may be a physiological attempt to maintain HSC numbers in the post-irradiation state. Taken together, these data suggest that the increased cycling among primitive hematopoietic cells in survivors of lethal radiation may contribute to long-term HSC exhaustion and subsequent RBMD, exacerbated by the added insult of aging at later time points.

Soluble factor(s) from bone marrow cells can rescue lethally irradiated mice by protecting endogenous hematopoietic stem cells.

PubMed

Zhao, Yi; Zhan, Yuxia; Burke, Kathleen A; Anderson, W French

2005-04-01

Ionizing radiation-induced myeloablation can be rescued via bone marrow transplantation (BMT) or administration of cytokines if given within 2 hours after radiation exposure. There is no evidence for the existence of soluble factors that can rescue an animal after a lethal dose of radiation when administered several hours postradiation. We established a system that could test the possibility for the existence of soluble factors that could be used more than 2 hours postirradiation to rescue animals. Animals with an implanted TheraCyte immunoisolation device (TID) received lethal-dose radiation and then normal bone marrow Lin- cells were loaded into the device (thereby preventing direct interaction between donor and recipient cells). Animal survival was evaluated and stem cell activity was tested with secondary bone marrow transplantation and flow cytometry analysis. Donor cell gene expression of five antiapoptotic cytokines was examined. Bone marrow Lin- cells rescued lethally irradiated animals via soluble factor(s). Bone marrow cells from the rescued animals can rescue and repopulate secondary lethally irradiated animals. Within the first 6 hours post-lethal-dose radiation, there is no significant change of gene expression of the known radioprotective factors TPO, SCF, IL-3, Flt-3 ligand, and SDF-1. Hematopoietic stem cells can be protected in lethally irradiated animals by soluble factors produced by bone marrow Lin- cells.

Evaluation of hematopoietic potential generated by transplantation of muscle-derived stem cells in mice.

PubMed

Farace, Francoise; Prestoz, Laetitita; Badaoui, Sabrina; Guillier, Martine; Haond, Celine; Opolon, Paule; Thomas, Jean-Leon; Zalc, Bernard; Vainchenker, William; Turhan, Ali G

2004-02-01

Muscle tissue of adult mice has been shown to contain stem cells with hematopoietic repopulation ability in vivo. To determine the functional characteristics of stem cells giving rise to this hematopoietic activity, we have performed hematopoietic reconstitution experiments by the use of muscle versus marrow transplantation in lethally irradiated mice and followed the fate of transplanted cells by Y-chimerism using PCR and fluorescence in situ hybridization (FISH) analysis. We report here that transplantation of murine muscle generate a major hematopoietic chimerism at the level of CFU-C, CFU-S, and terminally-differentiated cells in three generations of lethally irradiated mice followed up to 1 year after transplantation. This potential is totally abolished when muscle grafts were performed by the use of muscle from previously irradiated mice. As compared to marrow transplantation, muscle transplants were able to generate similar potencies to give rise to myeloid, T, B, and natural killer (NK) cells. Interestingly, marrow stem cells that have been generated in primary and then in secondary recipients were able to contribute efficiently to myofibers in the muscle tissue of tertiary recipients. Altogether, our data demonstrate that muscle-derived stem cells present a major hematopoietic repopulating ability with evidence of self-replication in vivo. They are radiation-sensitive and similar to marrow-derived stem cells in terms of their ability to generate multilineage hematopoiesis. Finally, our data demonstrate that muscle-derived hematopoietic stem cells do not lose their ability to contribute to myofiber generation after at least two rounds of serial transplantation, suggesting a potential that is probably equivalent to that generated by marrow transplantation.

Decrease in hematopoietic stem cell domains as a delayed effect of x-irradiation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Maloney, M.A.; Lamela, R.A.; Patt, H.M.

Although the hematopoietic integrity of locally X-irradiated sites can be restored for a time even after fairly large doses, a secondary aplasia often occurs some months later. To gain further insight into this delayed effect within the framework of the stem cell regulatory domain hypothesis, we characterized the growth kinetics of spleen colony forming units (CFU-S) in WBB6FI-+/+ bone marrow transplanted into WBB6FI-W/WV mice in which one leg had been exposed to 10-30 Gy of X rays 4-5 months previously. Compared to unirradiated contralateral marrow, fewer CFU-S either reached the previously irradiated marrow or were seeded into sites that couldmore » support growth. The initial exponential growth of effectively seeded CFU-S was unchanged, but growth deceleration (inflection point) occurred at a lower level of CFU-S in marrow previously irradiated with 20-30 Gy. This change in the inflection point indicates a radiation dose-dependent decrease consistent with the decrease in bone marrow cellularity. The decrease in effective stem cell domains after 20 Gy was calculated to be about 35%. We interpret these results to reflect the highly localized nature of delayed radiation damage to the marrow microenvironment.« less

Mismatch repair deficient hematopoietic stem cells are preleukemic stem cells

PubMed Central

Gerson, Stanton L.

2017-01-01

Whereas transformation events in hematopoietic malignancies may occur at different developmental stages, the initial mutation originates in hematopoietic stem cells (HSCs), creating a preleukemic stem cell (PLSC). Subsequent mutations at either stem cell or progenitor cell levels transform the PLSC into lymphoma/leukemia initiating cells (LIC). Thymic lymphomas have been thought to develop from developing thymocytes. T cell progenitors are generated from HSCs in the bone marrow (BM), but maturation and proliferation of T cells as well as T-lymphomagenesis depends on both regulatory mechanisms and microenvironment within the thymus. We studied PLSC linked to thymic lymphomas. In this study, we use MSH2-/- mice as a model to investigate the existence of PLSC and the evolution of PLSC to LIC. Following BM transplantation, we found that MSH2-/- BM cells from young mice are able to fully reconstitute multiple hematopoietic lineages of lethally irradiated wild-type recipients. However, all recipients developed thymic lymphomas within three and four months post transplantation. Transplantation of different fractions of BM cells or thymocytes from young health MSH2-/- mice showed that an HSC enriched fraction always reconstituted hematopoiesis followed by lymphoma development. In addition, lymphomas did not occur in thymectomized recipients of MSH2-/- BM. These results suggest that HSCs with DNA repair defects such as MSH2-/- are PLSCs because they retain hematopoietic function, but also carry an obligate lymphomagenic potential within their T-cell progeny that is dependent on the thymic microenvironment. PMID:28767666

Melanoma induced immunosuppression is mediated by hematopoietic dysregulation.

PubMed

Kamran, Neha; Li, Youping; Sierra, Maria; Alghamri, Mahmoud S; Kadiyala, Padma; Appelman, Henry D; Edwards, Marta; Lowenstein, Pedro R; Castro, Maria G

2018-01-01

Tumors are associated with expansion of immunosuppressive cells such as tumor associated macrophages (TAMs), regulatory T cells (Tregs) and myeloid derived suppressor cells (MDSCs). These cells promote tumor growth, angiogenesis, metastasis and immune escape. Cancer patients frequently present symptoms such as anemia, leukocytosis and/or cytopenia; associated with poor prognosis. To uncover tumor-mediated hematopoietic abnormalities and identify novel targets that can be harnessed to improve tumor-specific immune responses, we investigated the hematopoietic stem and progenitor cell compartment in melanoma bearing mice. We show that melanoma growth results in expansion of myeloid lineages such as MDSCs, macrophages and DCs along with a reduction in mature RBCs and platelets. Mature B lymphocytes in the blood and BM of melanoma mice were also reduced. Mice bearing melanoma showed extramedullary hematopoiesis in the spleen. Increased expansion of myeloid lineages occurred directly at the level of stem and progenitor cells. The reduction in mature B lymphocytes resulted from a block at the Pro-B cell stage in the bone marrow. Addition of recombinant IL-3 to bone marrow cells resulted in the expansion of committed myeloid progenitors including common myeloid precursors, granulocyte-monocyte precursors and megakaryocyte-erythrocyte precursors. In vivo , IL-3 receptor stimulation in melanoma bearing mice using an IL-3 antibody also resulted in a robust expansion of committed myeloid progenitors and hematopoietic stem cells. Collectively our findings demonstrate that tumor growth plays a pivotal role in reprogramming the host immune system by impacting hematopoiesis directly at the level of stem cell compartment.

Repurposing Treprostinil for Enhancing Hematopoietic Progenitor Cell Transplantation

PubMed Central

Kazemi, Zahra; Bergmayr, Christian; Prchal-Murphy, Michaela; Javaheri, Tahereh; Themanns, Madeleine; Pham, Ha T. T.; Strohmaier, Wolfgang; Sexl, Veronika; Zebedin-Brandl, Eva

2016-01-01

Activation of Gs-coupled receptors enhances engraftment of hematopoietic stem and progenitor cells (HSPCs). We tested the hypothesis that treprostinil, a prostacyclin analog approved for the treatment of pulmonary hypertension, can be repurposed to improve hematopoietic stem cell transplantation. Murine and human HSPCs were isolated from bone marrow and umbilical cord blood, respectively. Prostanoid receptor agonists and the combination thereof with forskolin were tested for their capacity to stimulate [3H]cAMP accumulation in HSPCs. Three independent approaches were employed to verify the ability of agonist-activated HSPCs to reconstitute the bone marrow in lethally irradiated recipient mice. The underlying mechanism was explored in cellular migration assays and by blocking C-X-C motif chemokine receptor 4 (CXCR4). Among several prostanoid agonists tested in combination with forskolin, treprostinil was most efficacious in raising intracellular cAMP levels in murine and human HPSCs. Injection of murine and human HSPCs, which had been pretreated with treprostinil and forskolin, enhanced survival of lethally irradiated recipient mice. Survival was further improved if recipient mice were subcutaneously administered treprostinil (0.15 mg kg−1 8 h−1) for 10 days. This regimen also reduced the number of HSPCs required to rescue lethally irradiated mice. Enhanced survival of recipient mice was causally related to treprostinil-enhanced CXCR4-dependent migration of HSPCs. Treprostinil stimulates the engraftment of human and murine hematopoietic stem cells without impairing their capacity for self-renewal. The investigated dose range corresponds to the dose approved for human use. Hence, these findings may be readily translated into a clinical application. PMID:26989084

Purified hematopoietic stem cell engraftment of rare niches corrects severe lymphoid deficiencies without host conditioning

PubMed Central

Bhattacharya, Deepta; Rossi, Derrick J.; Bryder, David; Weissman, Irving L.

2006-01-01

In the absence of irradiation or other cytoreductive conditioning, endogenous hematopoietic stem cells (HSCs) are thought to fill the unique niches within the bone marrow that allow maintenance of full hematopoietic potential and thus prevent productive engraftment of transplanted donor HSCs. By transplantation of purified exogenous HSCs into unconditioned congenic histocompatible strains of mice, we show that ∼0.1–1.0% of these HSC niches are available for engraftment at any given point and find no evidence that endogenous HSCs can be displaced from the niches they occupy. We demonstrate that productive engraftment of HSCs within these empty niches is inhibited by host CD4+ T cells that recognize very subtle minor histocompatibility differences. Strikingly, transplantation of purified HSCs into a panel of severe combined immunodeficient (SCID) mice leads to a rapid and complete rescue of lymphoid deficiencies through engraftment of these very rare niches and expansion of donor lymphoid progenitors. We further demonstrate that transient antibody-mediated depletion of CD4+ T cells allows short-term HSC engraftment and regeneration of B cells in a mouse model of B(-) non-SCID. These experiments provide a general mechanism by which transplanted HSCs can correct hematopoietic deficiencies without any host conditioning or with only highly specific and transient lymphoablation. PMID:16380511

Early Hematopoietic Zinc Finger Protein Prevents Tumor Cell Recognition by Natural Killer Cells1

PubMed Central

La Rocca, Rosanna; Fulciniti, Mariateresa; Lakshmikanth, Tadepally; Mesuraca, Maria; Ali, Talib Hassan; Mazzei, Valerio; Amodio, Nicola; Catalano, Lucio; Rotoli, Bruno; Ouerfelli, Ouathek; Grieco, Michele; Gulletta, Elio; Bond, Heather M.; Morrone, Giovanni; Ferrone, Soldano; Carbone, Ennio

2009-01-01

Early hematopoietic zinc finger/zinc finger protein 521 (EHZF/ZNF521) is a novel zinc finger protein expressed in hematopoietic stem and progenitor cells and is down-regulated during their differentiation. Its transcript is also abundant in some hematopoietic malignancies. Analysis of the changes in the antigenic profile of cells transfected with EHZF cDNA revealed up-regulation of HLA class I cell surface expression. This phenotypic change was associated with an increased level of HLA class I H chain, in absence of detectable changes in the expression of other Ag-processing machinery components. Enhanced resistance of target cells to NK cell-mediated cytotoxicity was induced by enforced expression of EHZF in the cervical carcinoma cell line HeLa and in the B lymphoblastoid cell line IM9. Preincubation of transfected cells with HLA class I Ag-specific mAb restored target cell susceptibility to NK cell-mediated lysis, indicating a specific role for HLA class I Ag up-regulation in the NK resistance induced by EHZF. A potential clinical significance of these findings is further suggested by the inverse correlation between EHZF and MHC class I expression levels, and autologous NK susceptibility of freshly explanted multiple myeloma cells. PMID:19342626

Thrombopoietin protects hematopoietic stem cells from retrotransposon-mediated damage by promoting an antiviral response.

PubMed

Barbieri, Daniela; Elvira-Matelot, Emilie; Pelinski, Yanis; Genève, Laetitia; de Laval, Bérengère; Yogarajah, Gayathri; Pecquet, Christian; Constantinescu, Stefan N; Porteu, Françoise

2018-05-07

Maintenance of genomic integrity is crucial for the preservation of hematopoietic stem cell (HSC) potential. Retrotransposons, spreading in the genome through an RNA intermediate, have been associated with loss of self-renewal, aging, and DNA damage. However, their role in HSCs has not been addressed. Here, we show that mouse HSCs express various retroelements (REs), including long interspersed element-1 (L1) recent family members that further increase upon irradiation. Using mice expressing an engineered human L1 retrotransposition reporter cassette and reverse transcription inhibitors, we demonstrate that L1 retransposition occurs in vivo and is involved in irradiation-induced persistent γH2AX foci and HSC loss of function. Thus, RE represents an important intrinsic HSC threat. Furthermore, we show that RE activity is restrained by thrombopoietin, a critical HSC maintenance factor, through its ability to promote a potent interferon-like, antiviral gene response in HSCs. This uncovers a novel mechanism allowing HSCs to minimize irradiation-induced injury and reinforces the links between DNA damage, REs, and antiviral immunity. © 2018 Barbieri et al.

Hematopoietic stem and progenitor cells regulate the regeneration of their niche by secreting Angiopoietin-1

PubMed Central

Zhou, Bo O; Ding, Lei; Morrison, Sean J

2015-01-01

Hematopoietic stem cells (HSCs) are maintained by a perivascular niche in bone marrow but it is unclear whether the niche is reciprocally regulated by HSCs. Here, we systematically assessed the expression and function of Angiopoietin-1 (Angpt1) in bone marrow. Angpt1 was not expressed by osteoblasts. Angpt1 was most highly expressed by HSCs, and at lower levels by c-kit+ hematopoietic progenitors, megakaryocytes, and Leptin Receptor+ (LepR+) stromal cells. Global conditional deletion of Angpt1, or deletion from osteoblasts, LepR+ cells, Nes-cre-expressing cells, megakaryocytes, endothelial cells or hematopoietic cells in normal mice did not affect hematopoiesis, HSC maintenance, or HSC quiescence. Deletion of Angpt1 from hematopoietic cells and LepR+ cells had little effect on vasculature or HSC frequency under steady-state conditions but accelerated vascular and hematopoietic recovery after irradiation while increasing vascular leakiness. Hematopoietic stem/progenitor cells and LepR+ stromal cells regulate niche regeneration by secreting Angpt1, reducing vascular leakiness but slowing niche recovery. DOI: http://dx.doi.org/10.7554/eLife.05521.001 PMID:25821987

Prostaglandin E2 increases hematopoietic stem cell survival and accelerates hematopoietic recovery after radiation injury

PubMed Central

Porter, Rebecca L.; Georger, Mary; Bromberg, Olga; McGrath, Kathleen E.; Frisch, Benjamin J.; Becker, Michael W.; Calvi, Laura M.

2013-01-01

Hematopoietic stem and progenitor cells (HSPCs), which continuously maintain all mature blood cells, are regulated within the marrow microenvironment. We previously reported that pharmacologic treatment of naïve mice with prostaglandin E2 (PGE2) expands HSPCs. However, the cellular mechanisms mediating this expansion remain unknown. Here we demonstrate that PGE2 treatment in naïve mice inhibits apoptosis of HSPCs without changing their proliferation rate. In a murine model of sub-lethal total body irradiation (TBI), in which HSPCs are rapidly lost, treatment with a long-acting PGE2 analogue (dmPGE2) reversed the apoptotic program initiated by TBI. dmPGE2 treatment in vivo decreased the loss of functional HSPCs following radiation injury, as demonstrated both phenotypically and by their increased reconstitution capacity. The antiapoptotic effect of dmPGE2 on HSPCs did not impair their ability to differentiate in vivo, resulting instead in improved hematopoietic recovery after TBI. dmPGE2 also increased microenvironmental cyclooxygenase-2 expression and expanded the α-SMA+ subset of marrow macrophages, thus enhancing the bone marrow microenvironmental response to TBI. Therefore, in vivo treatment with PGE2 analogues may be particularly beneficial to HSPCs in the setting of injury by targeting them both directly and also through their niche. The current data provide rationale for in vivo manipulation of the HSPC pool as a strategy to improve recovery after myelosuppression. PMID:23169593

Effects of Developmental Activation of the Aryl Hydrocarbon Receptor by 2,3,7,8-Tetrachlorodibenzo-p-dioxin on Long-term Self-renewal of Murine Hematopoietic Stem Cells.

PubMed

Laiosa, Michael D; Tate, Everett R; Ahrenhoerster, Lori S; Chen, Yuhong; Wang, Demin

2016-07-01

Human epidemiological and animal studies suggest that developmental exposure to contaminants that activate the aryl hydrocarbon receptor (AHR) lead to suppression of immune system function throughout life. The persistence of immune deficiency throughout life suggests that the cellular target of AHR activation is a fetal hematopoietic progenitor or stem cell. The aim of this study was to identify the effects of transplacental exposure to an AHR agonist on long-term self-renewal of fetal hematopoietic stem cells. Pregnant C57BL/6 or AHR+/- mice were exposed to the AHR agonist, 2,3,7,8-tetra-​chlorodibenzo-p-dioxin (TCDD). On day 14 of gestation, hematopoietic progenitors from wild-type or AHR-deficient fetuses were placed into in vitro T-lymphocyte differentiation cultures to identify the effects of transplacental TCDD on AHR activation in the fetus. We next analyzed the fetal hematopoietic progenitor cells for changes in reactive oxygen species (ROS). Finally, hematopoietic progenitors from fetuses exposed transplacentally to TCDD were mixed 1:1 with cells from congenic controls and used to reconstitute lethally irradiated recipients for analysis of long-term self-renewal potential. Our findings suggested that the effects of TCDD on the developing hematopoietic system were mediated by direct AHR activation in the fetus. Furthermore, developmental AHR activation by TCDD increased ROS in the fetal hematopoietic stem cells, and the elevated ROS was associated with a reduced capacity of the TCDD-exposed fetal cells to compete with control cells in a mixed competitive irradiation/reconstitution assay. Our findings indicate that AHR activation by TCDD in the fetus during pregnancy leads to impairment of long-term self-renewal of hematopoietic stem cells. Laiosa MD, Tate ER, Ahrenhoerster LS, Chen Y, Wang D. 2016. Effects of developmental activation of the aryl hydrocarbon receptor by 2,3,7,8-tetrachlorodibenzo-p-dioxin on long-term self-renewal of murine hematopoietic

Hematopoietic cell differentiation from embryonic and induced pluripotent stem cells

PubMed Central

2013-01-01

Pluripotent stem cells, both embryonic stem cells and induced pluripotent stem cells, are undifferentiated cells that can self-renew and potentially differentiate into all hematopoietic lineages, such as hematopoietic stem cells (HSCs), hematopoietic progenitor cells and mature hematopoietic cells in the presence of a suitable culture system. Establishment of pluripotent stem cells provides a comprehensive model to study early hematopoietic development and has emerged as a powerful research tool to explore regenerative medicine. Nowadays, HSC transplantation and hematopoietic cell transfusion have successfully cured some patients, especially in malignant hematological diseases. Owing to a shortage of donors and a limited number of the cells, hematopoietic cell induction from pluripotent stem cells has been regarded as an alternative source of HSCs and mature hematopoietic cells for intended therapeutic purposes. Pluripotent stem cells are therefore extensively utilized to facilitate better understanding in hematopoietic development by recapitulating embryonic development in vivo, in which efficient strategies can be easily designed and deployed for the generation of hematopoietic lineages in vitro. We hereby review the current progress of hematopoietic cell induction from embryonic stem/induced pluripotent stem cells. PMID:23796405

Mitophagy in hematopoietic stem cells

PubMed Central

Joshi, Aashish; Kundu, Mondira

2013-01-01

Hematopoietic stem cells (HSCs) are inherently quiescent and self-renewing, yet can differentiate and commit to multiple blood cell types. Intracellular mitochondrial content is dynamic, and there is an increase in mitochondrial content during differentiation and lineage commitment in HSCs. HSCs reside in a hypoxic niche within the bone marrow and rely heavily on glycolysis, while differentiated and committed progenitors rely on oxidative phosphorylation. Increased oxidative phosphorylation during differentiation and commitment is not only due to increased mitochondrial content but also due to changes in mitochondrial cytosolic distribution and efficiency. These changes in the intracellular mitochondrial landscape contribute signals toward regulating differentiation and commitment. Thus, a functional relationship exists between the mitochondria in HSCs and the state of the HSCs (i.e., stemness vs. differentiated). This review focuses on how autophagy-mediated mitochondrial clearance (i.e., mitophagy) may affect HSC mitochondrial content, thereby influencing the fate of HSCs and maintenance of hematopoietic homeostasis. PMID:24135495

Hematopoietic Stem Cell Regeneration Enhanced by Ectopic Expression of ROS-detoxifying Enzymes in Transplant Mice

PubMed Central

Miao, Weimin; XuFeng, Richard; Park, Moo-Rim; Gu, Haihui; Hu, Linping; Kang, Jin Wook; Ma, Shihui; Liang, Paulina H; Li, Yanxin; Cheng, Haizi; Yu, Hui; Epperly, Michael; Greenberger, Joel; Cheng, Tao

2013-01-01

High levels of reactive oxygen species (ROS) can exhaust hematopoietic stem cells (HSCs). Thus, maintaining a low state of redox in HSCs by modulating ROS-detoxifying enzymes may augment the regeneration potential of HSCs. Our results show that basal expression of manganese superoxide dismutase (MnSOD) and catalase were at low levels in long-term and short-term repopulating HSCs, and administration of a MnSOD plasmid and lipofectin complex (MnSOD-PL) conferred radiation protection on irradiated recipient mice. To assess the intrinsic role of elevated MnSOD or catalase in HSCs and hematopoietic progenitor cells, the MnSOD or catalase gene was overexpressed in mouse hematopoietic cells via retroviral transduction. The impact of MnSOD and catalase on hematopoietic progenitor cells was mild, as measured by colony-forming units (CFUs). However, overexpressed catalase had a significant beneficial effect on long-term engraftment of transplanted HSCs, and this effect was further enhanced after an insult of low-dose γ-irradiation in the transplant mice. In contrast, overexpressed MnSOD exhibited an insignificant effect on long-term engraftment of transplanted HSCs, but had a significant beneficial effect after an insult of sublethal irradiation. Taken together, these results demonstrate that HSC function can be enhanced by ectopic expression of ROS-detoxifying enzymes, especially after radiation exposure in vivo. PMID:23295952

Selection of genetically modified hematopoietic cells in vitro and in vivo using alkylating agent lysomustine.

PubMed

Rozov, F N; Grinenko, T S; Levit, G L; Krasnov, V P; Belyavsky, A V

2010-09-15

Efficient gene transfer into hematopoietic stem cells is vital for the success of gene therapy of hematopoietic and immune system disorders. An in vivo selection system based on a mutant form of the O(6)-methylguanine-DNA-methyltransferase gene (MGMTm) is considered one of the more promising strategies for expansion of hematopoietic cells transduced with viral vectors. Here we demonstrate that MGMTm-expressing cells can be efficiently selected using lysomustine, a nitrosourea derivative of lysine. K562 and murine bone marrow cells expressing MGMTm are protected from the cytotoxic action of lysomustine in vitro. We also show in a murine model that MGMTm-transduced hematopoietic cells can be expanded in vivo on transplantation into sublethally irradiated recipients followed by lysomustine treatment. These results indicate that lysomustine can be used as a potent novel chemoselection drug applicable for gene therapy of hematopoietic and immune system disorders. 2010 Elsevier Inc. All rights reserved.

Adenosine signaling promotes hematopoietic stem and progenitor cell emergence

PubMed Central

Jing, Lili; Tamplin, Owen J.; Chen, Michael J.; Deng, Qing; Patterson, Shenia; Kim, Peter G.; Durand, Ellen M.; McNeil, Ashley; Green, Julie M.; Matsuura, Shinobu; Ablain, Julien; Brandt, Margot K.; Schlaeger, Thorsten M.; Huttenlocher, Anna; Daley, George Q.; Ravid, Katya

2015-01-01

Hematopoietic stem cells (HSCs) emerge from aortic endothelium via the endothelial-to-hematopoietic transition (EHT). The molecular mechanisms that initiate and regulate EHT remain poorly understood. Here, we show that adenosine signaling regulates hematopoietic stem and progenitor cell (HSPC) development in zebrafish embryos. The adenosine receptor A2b is expressed in the vascular endothelium before HSPC emergence. Elevated adenosine levels increased runx1+/cmyb+ HSPCs in the dorsal aorta, whereas blocking the adenosine pathway decreased HSPCs. Knockdown of A2b adenosine receptor disrupted scl+ hemogenic vascular endothelium and the subsequent EHT process. A2b adenosine receptor activation induced CXCL8 via cAMP–protein kinase A (PKA) and mediated hematopoiesis. We further show that adenosine increased multipotent progenitors in a mouse embryonic stem cell colony-forming assay and in embryonic day 10.5 aorta-gonad-mesonephros explants. Our results demonstrate that adenosine signaling plays an evolutionary conserved role in the first steps of HSPC formation in vertebrates. PMID:25870200

The transcriptional landscape of hematopoietic stem cell ontogeny

PubMed Central

McKinney-Freeman, Shannon; Cahan, Patrick; Li, Hu; Lacadie, Scott A.; Huang, Hsuan-Ting; Curran, Matthew; Loewer, Sabine; Naveiras, Olaia; Kathrein, Katie L.; Konantz, Martina; Langdon, Erin M.; Lengerke, Claudia; Zon, Leonard I.; Collins, James J.; Daley, George Q.

2012-01-01

Transcriptome analysis of adult hematopoietic stem cells (HSC) and their progeny has revealed mechanisms of blood differentiation and leukemogenesis, but a similar analysis of HSC development is lacking. Here, we acquired the transcriptomes of developing HSC purified from >2500 murine embryos and adult mice. We found that embryonic hematopoietic elements clustered into three distinct transcriptional states characteristic of the definitive yolk sac, HSCs undergoing specification, and definitive HSCs. We applied a network biology-based analysis to reconstruct the gene regulatory networks of sequential stages of HSC development and functionally validated candidate transcriptional regulators of HSC ontogeny by morpholino-mediated knock-down in zebrafish embryos. Moreover, we found that HSCs from in vitro differentiated embryonic stem cells closely resemble definitive HSC, yet lack a Notch-signaling signature, likely accounting for their defective lymphopoiesis. Our analysis and web resource (http://hsc.hms.harvard.edu) will enhance efforts to identify regulators of HSC ontogeny and facilitate the engineering of hematopoietic specification. PMID:23122293

Effect of tocopherol-monoglucoside (TMG), a water-soluble glycosylated derivate of vitamin E, on hematopoietic recovery in irradiated mice.

PubMed

Cherdyntseva, Nadezda; Shishkina, Anna; Butorin, Ivan; Murase, Hironobu; Gervas, Polina; Kagiya, Tsutomu V

2005-03-01

A preparation of alpha-tocopherol monoglucoside (TMG) administered i.p. at a dose of 600 mg/kg immediately after whole body gamma irradiation was examined for its radioprotective efficacy towards bone marrow and peripheral blood nucleated cells. When mice received X-rays at a dose of 5,6 Gy, a marked decrease in bone marrow karyocytes and a reduction of peripheral leukocytes within the early post-irradiated period were observed. However these changes were attenuated in TMG-treated mice. Significant protection of blood lymphocytes was found for the TMG group of mice. The return to normal value of the reduced blood leukocyte count starting from the 8th day was more rapid in TMG-treated mice than in untreated irradiated mice. TMG administration was found to enhance hematopoietic recovery, as measured by the exceeded nucleated bone marrow cell count due to elevated amount of both lymphoid and granulocytic elements in the TMG-group, in comparison with that of both control irradiated and non-irradiated animals. These findings indicate that the radioprotective effect of TMG is apparently realized through its influence on hematopoietic system.

Identification of a population of cells with hematopoietic stem cell properties in mouse aorta-gonad-mesonephros cultures

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nobuhisa, Ikuo; Ohtsu, Naoki; Okada, Seiji

The aorta-gonad-mesonephros (AGM) region is a primary source of definitive hematopoietic cells in the midgestation mouse embryo. In cultures of dispersed AGM regions, adherent cells containing endothelial cells are observed first, and then non-adherent hematopoietic cells are produced. Here we report on the characterization of hematopoietic cells that emerge in the AGM culture. Based on the expression profiles of CD45 and c-Kit, we defined three cell populations: CD45{sup low} c-Kit{sup +} cells that had the ability to form hematopoietic cell colonies in methylcellulose media and in co-cultures with stromal cells; CD45{sup low} c-Kit{sup -} cells that showed a granulocyte morphology;more » CD45{sup high} c-Kit{sup low/-} that exhibited a macrophage morphology. In co-cultures of OP9 stromal cells and freshly prepared AGM cultures, CD45{sup low} c-Kit{sup +} cells from the AGM culture had the abilities to reproduce CD45{sup low} c-Kit{sup +} cells and differentiate into CD45{sup low} c-Kit{sup -} and CD45{sup high} c-Kit{sup low/-} cells, whereas CD45{sup low} c-Kit{sup -} and CD45{sup high} c-Kit{sup low/-} did not produce CD45{sup low} c-Kit{sup +} cells. Furthermore, CD45{sup low} c-Kit{sup +} cells displayed a long-term repopulating activity in adult hematopoietic tissue when transplanted into the liver of irradiated newborn mice. These results indicate that CD45{sup low} c-Kit{sup +} cells from the AGM culture have the potential to reconstitute multi-lineage hematopoietic cells.« less

The biology of NK cells and their receptors affects clinical outcomes after hematopoietic cell transplantation (HCT).

PubMed

Foley, Bree; Felices, Martin; Cichocki, Frank; Cooley, Sarah; Verneris, Michael R; Miller, Jeffrey S

2014-03-01

Natural killer (NK) cells were first identified for their capacity to reject bone marrow allografts in lethally irradiated mice without prior sensitization. Subsequently, human NK cells were detected and defined by their non-major histocompatibility complex (MHC)-restricted cytotoxicity toward transformed or virally infected target cells. Karre et al. later proposed 'the missing self hypothesis' to explain the mechanism by which self-tolerant cells could kill targets that had lost self MHC class I. Subsequently, the receptors that recognize MHC class I to mediate tolerance in the host were identified on NK cells. These class I-recognizing receptors contribute to the acquisition of function by a dynamic process known as NK cell education or licensing. In the past, NK cells were assumed to be short lived, but more recently NK cells have been shown to mediate immunologic memory to secondary exposures to cytomegalovirus infection. Because of their ability to lyse tumors with aberrant MHC class I expression and to produce cytokines and chemokines upon activation, NK cells may be primed by many stimuli, including viruses and inflammation, to contribute to a graft-versus-tumor effect. In addition, interactions with other immune cells support the therapeutic potential of NK cells to eradicate tumor and to enhance outcomes after hematopoietic cell transplantation. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Memory T cells: A helpful guard for allogeneic hematopoietic stem cell transplantation without causing graft-versus-host disease.

PubMed

Huang, Wei; Chao, Nelson J

2017-12-01

Graft-versus-host disease (GVHD) is a major complication of allogeneic hematopoietic stem cell transplantation (AHSCT) and the major cause of nonrelapse morbidity and mortality of AHSCT. In AHSCT, donor T cells facilitate hematopoietic stem cell (HSC) engraftment, contribute to anti-infection immunity, and mediate graft-versus-leukemia (GVL) responses. However, activated alloreactive T cells also attack recipient cells in vital organs, leading to GVHD. Different T-cell subsets, including naïve T (T N ) cells, memory T (T M ) cells, and regulatory T (T reg ) cells mediate different forms of GVHD and GVL; T N cells mediate severe GVHD, whereas T M cells do not cause GVHD, but preserve T-cell function including GVL. In addition, metabolic reprogramming controls T-cell differentiation and activation in these disease states. This minireview focuses on the role and the related mechanisms of T M cells in AHSCT, and the potential manipulation of T cells in AHSCT. Copyright © 2017 King Faisal Specialist Hospital & Research Centre. Published by Elsevier B.V. All rights reserved.

Generating autologous hematopoietic cells from human-induced pluripotent stem cells through ectopic expression of transcription factors.

PubMed

Hwang, Yongsung; Broxmeyer, Hal E; Lee, Man Ryul

2017-07-01

Hematopoietic cell transplantation (HCT) is a successful treatment modality for patients with malignant and nonmalignant disorders, usually when no other treatment option is available. The cells supporting long-term reconstitution after HCT are the hematopoietic stem cells (HSCs), which can be limited in numbers. Moreover, finding an appropriate human leukocyte antigen-matched donor can be problematic. If HSCs can be stably produced in large numbers from autologous or allogeneic cell sources, it would benefit HCT. Induced pluripotent stem cells (iPSCs) established from patients' own somatic cells can be differentiated into hematopoietic cells in vitro. This review will highlight recent methods for regulating human (h) iPSC production of HSCs and more mature blood cells. Advancements in transcription factor-mediated regulation of the developmental stages of in-vivo hematopoietic lineage commitment have begun to provide an understanding of the molecular mechanism of hematopoiesis. Such studies involve not only directed differentiation in which transcription factors, specifically expressed in hematopoietic lineage-specific cells, are overexpressed in iPSCs, but also direct conversion in which transcription factors are introduced into patient-derived somatic cells which are dedifferentiated to hematopoietic cells. As iPSCs derived from patients suffering from genetically mutated diseases would express the same mutated genetic information, CRISPR-Cas9 gene editing has been utilized to differentiate genetically corrected iPSCs into normal hematopoietic cells. IPSCs provide a model for molecular understanding of disease, and also may function as a cell population for therapy. Efficient differentiation of patient-specific iPSCs into HSCs and progenitor cells is a potential means to overcome limitations of such cells for HCT, as well as for providing in-vitro drug screening templates as tissue-on-a-chip models.

Exonuclease 1 is a critical mediator of survival during DNA double strand break repair in nonquiescent hematopoietic stem and progenitor cells.

PubMed

Desai, Amar; Qing, Yulan; Gerson, Stanton L

2014-02-01

Hematopoietic stem cell (HSC) populations require DNA repair pathways to maintain their long-term survival and reconstitution capabilities, but mediators of these processes are still being elucidated. Exonuclease 1 (Exo1) participates in homologous recombination (HR) and Exo1 loss results in impaired 5' HR end resection. We use cultured Exo1(mut) fibroblasts and bone marrow to demonstrate that loss of Exo1 function results in defective HR in cycling cells. Conversely, in Exo1(mut) mice HR is not required for maintenance of quiescent HSCs at steady state, confirming the steady state HSC reliance on nonhomologous end joining (NHEJ). Exo1(mut) mice sustained serial repopulation, displayed no defect in competitive repopulation or niche occupancy, and exhibited no increased sensitivity to whole body ionizing radiation. However, when Exo1(mut) HSCs were pushed into cell cycle in vivo with 5-fluorouracil or poly IC, the hematopoietic population became hypersensitive to IR, resulting in HSC defects and animal death. We propose Exo1-mediated HR is dispensable for stem cell function in quiescent HSC, whereas it is essential to HSC response to DNA damage processing after cell cycle entry, and its loss is not compensated by intact NHEJ. In HSCs, the maintenance of stem cell function after DNA damage is dependent on the DNA repair capacity, segregated by active versus quiescent points in cell cycle. © AlphaMed Press.

Inflamm-Aging of Hematopoiesis, Hematopoietic Stem Cells, and the Bone Marrow Microenvironment

PubMed Central

Kovtonyuk, Larisa V.; Fritsch, Kristin; Feng, Xiaomin; Manz, Markus G.; Takizawa, Hitoshi

2016-01-01

All hematopoietic and immune cells are continuously generated by hematopoietic stem cells (HSCs) and hematopoietic progenitor cells (HPCs) through highly organized process of stepwise lineage commitment. In the steady state, HSCs are mostly quiescent, while HPCs are actively proliferating and contributing to daily hematopoiesis. In response to hematopoietic challenges, e.g., life-threatening blood loss, infection, and inflammation, HSCs can be activated to proliferate and engage in blood formation. The HSC activation induced by hematopoietic demand is mediated by direct or indirect sensing mechanisms involving pattern recognition receptors or cytokine/chemokine receptors. In contrast to the hematopoietic challenges with obvious clinical symptoms, how the aging process, which involves low-grade chronic inflammation, impacts hematopoiesis remains undefined. Herein, we summarize recent findings pertaining to functional alternations of hematopoiesis, HSCs, and the bone marrow (BM) microenvironment during the processes of aging and inflammation and highlight some common cellular and molecular changes during the processes that influence hematopoiesis and its cells of origin, HSCs and HPCs, as well as the BM microenvironment. We also discuss how age-dependent alterations of the immune system lead to subclinical inflammatory states and how inflammatory signaling might be involved in hematopoietic aging. Our aim is to present evidence supporting the concept of “Inflamm-Aging,” or inflammation-associated aging of hematopoiesis. PMID:27895645

Tracking single hematopoietic stem cells in vivo using high-throughput sequencing in conjunction with viral genetic barcoding

PubMed Central

Lu, Rong; Neff, Norma F.; Quake, Stephen R.; Weissman, Irving L.

2011-01-01

Disentangling cellular heterogeneity is a challenge in many fields, particularly in the stem cell and cancer biology fields. Here, we demonstrate how to combine viral genetic barcoding with high-throughput sequencing to track single cells in a heterogeneous population. We use this technique to track the in vivo differentiation of unitary hematopoietic stem cells (HSCs). The results are consistent with single cell transplantation studies, but require two orders of magnitude fewer mice. In addition to its high throughput, the high sensitivity of the technique allows for a direct examination of the clonality of sparse cell populations such as HSCs. We show how these capabilities offer a clonal perspective of the HSC differentiation process. In particular, our data suggests that HSCs do not equally contribute to blood cells after irradiation-mediated transplantation, and that two distinct HSC differentiation patterns co-exist in the same recipient mouse post irradiation. This technique can be applied to any viral accessible cell type for both in vitro and in vivo processes. PMID:21964413

Hematopoietic Stem Cells in Neural-crest Derived Bone Marrow.

PubMed

Jiang, Nan; Chen, Mo; Yang, Guodong; Xiang, Lusai; He, Ling; Hei, Thomas K; Chotkowski, Gregory; Tarnow, Dennis P; Finkel, Myron; Ding, Lei; Zhou, Yanheng; Mao, Jeremy J

2016-12-21

Hematopoietic stem cells (HSCs) in the endosteum of mesoderm-derived appendicular bones have been extensively studied. Neural crest-derived bones differ from appendicular bones in developmental origin, mode of bone formation and pathological bone resorption. Whether neural crest-derived bones harbor HSCs is elusive. Here, we discovered HSC-like cells in postnatal murine mandible, and benchmarked them with donor-matched, mesoderm-derived femur/tibia HSCs, including clonogenic assay and long-term culture. Mandibular CD34 negative, LSK cells proliferated similarly to appendicular HSCs, and differentiated into all hematopoietic lineages. Mandibular HSCs showed a consistent deficiency in lymphoid differentiation, including significantly fewer CD229 + fractions, PreProB, ProB, PreB and B220 + slgM cells. Remarkably, mandibular HSCs reconstituted irradiated hematopoietic bone marrow in vivo, just as appendicular HSCs. Genomic profiling of osteoblasts from mandibular and femur/tibia bone marrow revealed deficiencies in several HSC niche regulators among mandibular osteoblasts including Cxcl12. Neural crest derived bone harbors HSCs that function similarly to appendicular HSCs but are deficient in the lymphoid lineage. Thus, lymphoid deficiency of mandibular HSCs may be accounted by putative niche regulating genes. HSCs in craniofacial bones have functional implications in homeostasis, osteoclastogenesis, immune functions, tumor metastasis and infections such as osteonecrosis of the jaw.

Helping Yourself by Offering Help: Mediators of Expressive Helping in Survivors of Hematopoietic Stem Cell Transplant.

PubMed

Williamson, Timothy J; Stanton, Annette L; Austin, Jane E; Valdimarsdottir, Heiddis B; Wu, Lisa M; Krull, Jennifer L; Rini, Christine M

2017-10-01

A randomized experiment by Rini et al. (Health Psychol. 33(12):1541-1551, 2014) demonstrated that expressive helping, which involves three expressive writing sessions regarding hematopoietic stem cell transplant, followed by one writing session directed toward helping other stem cell transplant recipients, reduced psychological distress and bothersome physical symptoms among stem cell transplant recipients with elevated survivorship problems, relative to a neutral writing control condition. The current study evaluated whether word use reflective of emotional expression, cognitive processing, and change in perspective mediates the effects of expressive helping. The essays of 67 stem cell transplant recipients with high survivorship problems were analyzed with Linguistic Inquiry and Word Count. Multiple mediation modeling was used to test the hypothesized mechanisms of expressive helping on distress and bothersome physical symptoms. Relative to the control condition, expressive helping produced significant reductions in psychological distress and marginal reductions in physical symptom bother in the analyzed subset of participants from the parent study. Results indicated that positive emotion word use significantly mediated effects of expressive helping on reduced distress, but only for participants who used average (compared to above or below average) rates of negative emotion words. Cognitive processing and change in perspective did not significantly mediate benefits of expressive helping. Expressive helping carried its positive effects on distress through participants' higher expression of positive emotions when coupled with moderate rates of negative emotions. Findings highlight the benefit of expressing both positive and negative emotions in stressful situations.

Hematologic changes after total body irradiation and autologous transplantation of hematopoietic peripheral blood progenitor cells in dogs with lymphoma.

PubMed

Escobar, C; Grindem, C; Neel, J A; Suter, S E

2012-03-01

Dogs with and without lymphoma have undergone hematopoietic cell transplantation in a research setting for decades. North Carolina State University is currently treating dogs with B- and T-cell lymphoma in a clinical setting with autologous peripheral blood progenitor cell transplants, using peripheral blood CD34+ progenitor cells harvested using an apheresis machine. Complete blood counts were performed daily for 15 to 19 days posttransplantation to monitor peripheral blood cell nadirs and subsequent CD34+ cell engraftment. This study documents the hematologic toxicities of total body irradiation in 10 dogs and the subsequent recovery of the affected cell lines after peripheral blood progenitor cell transplant, indicating successful CD34+ engraftment. All peripheral blood cell lines, excluding red blood cells, experienced grade 4 toxicities. All dogs had ≥ 500 neutrophils/μl by day 12, while thrombocytopenia persisted for many weeks. All dogs were clinically normal at discharge.

Cord blood-derived CD34+ hematopoietic cells with low mitochondrial mass are enriched in hematopoietic repopulating stem cell function.

PubMed

Romero-Moya, Damia; Bueno, Clara; Montes, Rosa; Navarro-Montero, Oscar; Iborra, Francisco J; López, Luis Carlos; Martin, Miguel; Menendez, Pablo

2013-07-01

The homeostasis of the hematopoietic stem/progenitor cell pool relies on a fine-tuned balance between self-renewal, differentiation and proliferation. Recent studies have proposed that mitochondria regulate these processes. Although recent work has contributed to understanding the role of mitochondria during stem cell differentiation, it remains unclear whether the mitochondrial content/function affects human hematopoietic stem versus progenitor function. We found that mitochondrial mass correlates strongly with mitochondrial membrane potential in CD34(+) hematopoietic stem/progenitor cells. We, therefore, sorted cord blood CD34(+) cells on the basis of their mitochondrial mass and analyzed the in vitro homeostasis and clonogenic potential as well as the in vivo repopulating potential of CD34(+) cells with high (CD34(+) Mito(High)) versus low (CD34(+) Mito(Low)) mitochondrial mass. The CD34(+) Mito(Low) fraction contained 6-fold more CD34(+)CD38(-) primitive cells and was enriched in hematopoietic stem cell function, as demonstrated by its significantly greater hematopoietic reconstitution potential in immuno-deficient mice. In contrast, the CD34(+) Mito(High) fraction was more enriched in hematopoietic progenitor function with higher in vitro clonogenic capacity. In vitro differentiation of CD34(+) Mito(Low) cells was significantly delayed as compared to that of CD34(+) Mito(High) cells. The eventual complete differentiation of CD34(+) Mito(Low) cells, which coincided with a robust expansion of the CD34(-) differentiated progeny, was accompanied by mitochondrial adaptation, as shown by significant increases in ATP production and expression of the mitochondrial genes ND1 and COX2. In conclusion, cord blood CD34(+) cells with low levels of mitochondrial mass are enriched in hematopoietic repopulating stem cell function whereas high levels of mitochondrial mass identify hematopoietic progenitors. A mitochondrial response underlies hematopoietic stem/progenitor cell

Low-dose radiation (LDR) induces hematopoietic hormesis: LDR-induced mobilization of hematopoietic progenitor cells into peripheral blood circulation.

PubMed

Li, Wei; Wang, Guanjun; Cui, Jiuwei; Xue, Lu; Cai, Lu

2004-11-01

The aim of this study was to investigate the stimulating effect of low-dose radiation (LDR) on bone marrow hematopoietic progenitor cell (HPC) proliferation and peripheral blood mobilization. Mice were exposed to 25- to 100-mGy x-rays. Bone marrow and peripheral blood HPCs (BFU-E, CFU-GM, and c-kit+ cells) were measured, and GM-CSF, G-CSF, and IL-3 protein and mRNA expression were detected using ELISA, slot blot hybridization, and Northern blot methods. To functionally evaluate LDR-stimulated and -mobilized HPCs, repopulation of peripheral blood cells in lethally irradiated recipients after transplantation of LDR-treated donor HPCs was examined by WBC counts, animal survival, and colony-forming units in the recipient spleens (CFUs-S). 75-mGy x-rays induced a maximal stimulation for bone marrow HPC proliferation (CFU-GM and BFU-E formation) 48 hours postirradiation, along with a significant increase in HPC mobilization into peripheral blood 48 to 72 hours postradiation, as shown by increases in CFU-GM formation and proportion of c-kit+ cells in the peripheral mononuclear cells. 75-mGy x-rays also maximally induced increases in G-CSF and GM-CSF mRNA expression in splenocytes and levels of serum GM-CSF. To define the critical role of these hematopoietic-stimulating factors in HPC peripheral mobilization, direct administration of G-CSF at a dose of 300 microg/kg/day or 150 microg/kg/day was applied and found to significantly stimulate GM-CFU formation and increase c-kit+ cells in the peripheral mononuclear cells. More importantly, 75-mGy x-rays plus 150 microg/kg/day G-CSF (LDR/150-G-CSF) produced a similar effect to that of 300 microg/kg/day G-CSF alone. Furthermore, the capability of LDR-mobilized donor HPCs to repopulate blood cells was confirmed in lethally irradiated recipient mice by counting peripheral WBC and CFUs-S. These results suggest that LDR induces hematopoietic hormesis, as demonstrated by HPC proliferation and peripheral mobilization, providing a

CCND1-CDK4-mediated cell cycle progression provides a competitive advantage for human hematopoietic stem cells in vivo.

PubMed

Mende, Nicole; Kuchen, Erika E; Lesche, Mathias; Grinenko, Tatyana; Kokkaliaris, Konstantinos D; Hanenberg, Helmut; Lindemann, Dirk; Dahl, Andreas; Platz, Alexander; Höfer, Thomas; Calegari, Federico; Waskow, Claudia

2015-07-27

Maintenance of stem cell properties is associated with reduced proliferation. However, in mouse hematopoietic stem cells (HSCs), loss of quiescence results in a wide range of phenotypes, ranging from functional failure to extensive self-renewal. It remains unknown whether the function of human HSCs is controlled by the kinetics of cell cycle progression. Using human HSCs and human progenitor cells (HSPCs), we report here that elevated levels of CCND1-CDK4 complexes promoted the transit from G0 to G1 and shortened the G1 cell cycle phase, resulting in protection from differentiation-inducing signals in vitro and increasing human leukocyte engraftment in vivo. Further, CCND1-CDK4 overexpression conferred a competitive advantage without impacting HSPC numbers. In contrast, accelerated cell cycle progression mediated by elevated levels of CCNE1-CDK2 led to the loss of functional HSPCs in vivo. Collectively, these data suggest that the transition kinetics through the early cell cycle phases are key regulators of human HSPC function and important for lifelong hematopoiesis. © 2015 Mende et al.

Cytohesin 1 regulates homing and engraftment of human hematopoietic stem and progenitor cells.

PubMed

Rak, Justyna; Foster, Katie; Potrzebowska, Katarzyna; Talkhoncheh, Mehrnaz Safaee; Miharada, Natsumi; Komorowska, Karolina; Torngren, Therese; Kvist, Anders; Borg, Åke; Svensson, Lena; Bonnet, Dominique; Larsson, Jonas

2017-02-23

Adhesion is a key component of hematopoietic stem cell regulation mediating homing and retention to the niche in the bone marrow. Here, using an RNA interference screen, we identify cytohesin 1 (CYTH1) as a critical mediator of adhesive properties in primary human cord blood-derived hematopoietic stem and progenitor cells (HSPCs). Knockdown of CYTH1 disrupted adhesion of HSPCs to primary human mesenchymal stroma cells. Attachment to fibronectin and ICAM1, 2 integrin ligands, was severely impaired, and CYTH1-deficient cells showed a reduced integrin β1 activation response, suggesting that CYTH1 mediates integrin-dependent functions. Transplantation of CYTH1-knockdown cells to immunodeficient mice resulted in significantly lower long-term engraftment levels, associated with a reduced capacity of the transplanted cells to home to the bone marrow. Intravital microscopy showed that CYTH1 deficiency profoundly affects HSPC mobility and localization within the marrow space and thereby impairs proper lodgment into the niche. Thus, CYTH1 is a novel major regulator of adhesion and engraftment in human HSPCs through mechanisms that, at least in part, involve the activation of integrins. © 2017 by The American Society of Hematology.

Contribution of endothelial progenitors and proangiogenic hematopoietic cells to vascularization of tumor and ischemic tissue

PubMed Central

Kopp, Hans-Georg; Ramos, Carlos A.; Rafii, Shahin

2010-01-01

Purpose of review During the last several years, a substantial amount of evidence from animal as well as human studies has advanced our knowledge of how bone marrow derived cells contribute to neoangiogenesis. In the light of recent findings, we may have to redefine our thinking of endothelial cells as well as of perivascular mural cells. Recent findings Inflammatory hematopoietic cells, such as macrophages, have been shown to promote neoangiogenesis during tumor growth and wound healing. Dendritic cells, B lymphocytes, monocytes, and other immune cells have also been found to be recruited to neoangiogenic niches and to support neovessel formation. These findings have led to the concept that subsets of hematopoietic cells comprise proangiogenic cells that drive adult revascularization processes. While evidence of the importance of endothelial progenitor cells in adult vasculogenesis increased further, the role of these comobilized hematopoietic cells has been intensely studied in the last few years. Summary Angiogenic factors promote mobilization of vascular endothelial growth factor receptor 1-positive hematopoietic cells through matrix metalloproteinase-9 mediated release of soluble kit-ligand and recruit these proangiogenic cells to areas of hypoxia, where perivascular mural cells present stromal-derived factor 1 (CXCL-12) as an important retention signal. The same factors are possibly involved in mobilization of vascular endothelial growth factor receptor 2-positive endothelial precursors that may participate in neovessel formation. The complete characterization of mechanisms, mediators and signaling pathways involved in these processes will provide novel targets for both anti and proangiogenic therapeutic strategies. PMID:16567962

Epigenetic and Epitranscriptomic Factors Make a Mark on Hematopoietic Stem Cell Development.

PubMed

Kasper, Dionna M; Nicoli, Stefania

2018-03-01

Blood specification is a highly dynamic process, whereby committed hemogenic endothelial cells (ECs) progressively transdifferentiate into multipotent, self-renewing hematopoietic stem cells (HSCs). Massive changes in gene expression must occur to switch cell identity, however the factors that mediate such an effect were a mystery until recently. This review summarizes the higher-order mechanisms involved in endothelial to hematopoietic reprogramming identified thus far. Accumulating evidence from mouse and zebrafish studies reveal that numerous chromatin-modifying (epigenetic) and RNA-modifying (epitranscriptomic) factors are required for the formation of HSCs from hemogenic endothelium. These genes function throughout the endothelial-hematopoietic transition, suggesting a dynamic interplay between 'epi'-machineries. Epigenetic and epitranscriptomic regulation are key mechanisms for reshaping global EC gene expression patterns to those that support HSC production. Future studies that capture modification dynamics should bring us closer to a complete understanding of how HSCs transition from hemogenic endothelium at the molecular level.

MicroRNAs enriched in hematopoietic stem cells differentially regulate long-term hematopoietic output.

PubMed

O'Connell, Ryan M; Chaudhuri, Aadel A; Rao, Dinesh S; Gibson, William S J; Balazs, Alejandro B; Baltimore, David

2010-08-10

The production of blood cells depends on a rare hematopoietic stem-cell (HSC) population, but the molecular mechanisms underlying HSC biology remain incompletely understood. Here, we identify a subset of microRNAs (miRNAs) that is enriched in HSCs compared with other bone-marrow cells. An in vivo gain-of-function screen found that three of these miRNAs conferred a competitive advantage to engrafting hematopoietic cells, whereas other HSC miRNAs attenuated production of blood cells. Overexpression of the most advantageous miRNA, miR-125b, caused a dose-dependent myeloproliferative disorder that progressed to a lethal myeloid leukemia in mice and also enhanced hematopoietic engraftment in human immune system mice. Our study identifies an evolutionarily conserved subset of miRNAs that is expressed in HSCs and functions to modulate hematopoietic output.

Development of hematopoietic stem and progenitor cells from human pluripotent stem cells.

PubMed

Chen, Tong; Wang, Fen; Wu, Mengyao; Wang, Zack Z

2015-07-01

Human pluripotent stem cells (hPSCs), including human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs), provide a new cell source for regenerative medicine, disease modeling, drug discovery, and preclinical toxicity screening. Understanding of the onset and the sequential process of hematopoietic cells from differentiated hPSCs will enable the achievement of personalized medicine and provide an in vitro platform for studying of human hematopoietic development and disease. During embryogenesis, hemogenic endothelial cells, a specified subset of endothelial cells in embryonic endothelium, are the primary source of multipotent hematopoietic stem cells. In this review, we discuss current status in the generation of multipotent hematopoietic stem and progenitor cells from hPSCs via hemogenic endothelial cells. We also review the achievements in direct reprogramming from non-hematopoietic cells to hematopoietic stem and progenitor cells. Further characterization of hematopoietic differentiation in hPSCs will improve our understanding of blood development and expedite the development of hPSC-derived blood products for therapeutic purpose. © 2015 Wiley Periodicals, Inc.

Are hematopoietic stem cells involved in hepatocarcinogenesis?

PubMed

Facciorusso, Antonio; Antonino, Matteo; Del Prete, Valentina; Neve, Viviana; Scavo, Maria Principia; Barone, Michele

2014-08-01

THE LIVER HAS THREE CELL LINEAGES ABLE TO PROLIFERATE AFTER A HEPATIC INJURY: the mature hepatocyte, the ductular "bipolar" progenitor cell termed "oval cell" and the putative periductular stem cell. Hepatocytes can only produce other hepatocytes whereas ductular progenitor cells are considerate bipolar since they can give rise to biliary cells or hepatocytes. Periductular stem cells are rare in the liver, have a very long proliferation potential and may be multipotent, being this aspect still under investigation. They originate in the bone marrow since their progeny express genetic markers of donor hematopoietic cells after bone marrow transplantation. Since the liver is the hematopoietic organ of the fetus, it is possible that hematopoietic stem cells may reside in the liver of the adult. This assumption is proved by the finding that oval cells express hematopoietic markers like CD34, CD45, CD 109, Thy-1, c-kit, and others, which are also expressed by bone marrow-derived hematopoietic stem cells (BMSCs). Few and discordant studies have evaluated the role of BMSC in hepatocarcinogenesis so far and further studies in vitro and in vivo are warranted in order to definitively clarify such an issue.

Pleiotrophin regulates the expansion and regeneration of hematopoietic stem cells

PubMed Central

Himburg, Heather A; Muramoto, Garrett G; Daher, Pamela; Meadows, Sarah K; Russell, J Lauren; Doan, Phuong; Chi, Jen-Tsan; Salter, Alice B; Lento, William E; Reya, Tannishtha; Chao, Nelson; Chute, John P

2013-01-01

Hematopoietic stem cell (HSC) self-renewal is regulated by both intrinsic and extrinsic signals. Although some of the pathways that regulate HSC self-renewal have been uncovered, it remains largely unknown whether these pathways can be triggered by deliverable growth factors to induce HSC growth or regeneration. Here we show that pleiotrophin, a neurite outgrowth factor with no known function in hematopoiesis, efficiently promotes HSC expansion in vitro and HSC regeneration in vivo. Treatment of mouse bone marrow HSCs with pleiotrophin caused a marked increase in long-term repopulating HSC counts in culture, as measured in competitive repopulating assays. Treatment of human cord blood CD34+CDCD38−Lin− cells with pleiotrophin also substantially increased severe combined immunodeficient (SCID)-repopulating cell counts in culture, compared to input and cytokine-treated cultures. Systemic administration of pleiotrophin to irradiated mice caused a pronounced expansion of bone marrow stem and progenitor cells in vivo, indicating that pleiotrophin is a regenerative growth factor for HSCs. Mechanistically, pleiotrophin activated phosphoinositide 3-kinase (PI3K) signaling in HSCs; antagonism of PI3K or Notch signaling inhibited pleiotrophin-mediated expansion of HSCs in culture. We identify the secreted growth factor pleiotrophin as a new regulator of both HSC expansion and regeneration PMID:20305662

Pleiotrophin regulates the expansion and regeneration of hematopoietic stem cells.

PubMed

Himburg, Heather A; Muramoto, Garrett G; Daher, Pamela; Meadows, Sarah K; Russell, J Lauren; Doan, Phuong; Chi, Jen-Tsan; Salter, Alice B; Lento, William E; Reya, Tannishtha; Chao, Nelson J; Chute, John P

2010-04-01

Hematopoietic stem cell (HSC) self-renewal is regulated by both intrinsic and extrinsic signals. Although some of the pathways that regulate HSC self-renewal have been uncovered, it remains largely unknown whether these pathways can be triggered by deliverable growth factors to induce HSC growth or regeneration. Here we show that pleiotrophin, a neurite outgrowth factor with no known function in hematopoiesis, efficiently promotes HSC expansion in vitro and HSC regeneration in vivo. Treatment of mouse bone marrow HSCs with pleiotrophin caused a marked increase in long-term repopulating HSC numbers in culture, as measured in competitive repopulating assays. Treatment of human cord blood CD34(+)CDCD38(-)Lin(-) cells with pleiotrophin also substantially increased severe combined immunodeficient (SCID)-repopulating cell counts in culture, compared to input and cytokine-treated cultures. Systemic administration of pleiotrophin to irradiated mice caused a pronounced expansion of bone marrow stem and progenitor cells in vivo, indicating that pleiotrophin is a regenerative growth factor for HSCs. Mechanistically, pleiotrophin activated phosphoinositide 3-kinase (PI3K) signaling in HSCs; antagonism of PI3K or Notch signaling inhibited pleiotrophin-mediated expansion of HSCs in culture. We identify the secreted growth factor pleiotrophin as a new regulator of both HSC expansion and regeneration.

Are hematopoietic stem cells involved in hepatocarcinogenesis?

PubMed Central

Antonino, Matteo; Del Prete, Valentina; Neve, Viviana; Scavo, Maria Principia; Barone, Michele

2014-01-01

The liver has three cell lineages able to proliferate after a hepatic injury: the mature hepatocyte, the ductular “bipolar” progenitor cell termed “oval cell” and the putative periductular stem cell. Hepatocytes can only produce other hepatocytes whereas ductular progenitor cells are considerate bipolar since they can give rise to biliary cells or hepatocytes. Periductular stem cells are rare in the liver, have a very long proliferation potential and may be multipotent, being this aspect still under investigation. They originate in the bone marrow since their progeny express genetic markers of donor hematopoietic cells after bone marrow transplantation. Since the liver is the hematopoietic organ of the fetus, it is possible that hematopoietic stem cells may reside in the liver of the adult. This assumption is proved by the finding that oval cells express hematopoietic markers like CD34, CD45, CD 109, Thy-1, c-kit, and others, which are also expressed by bone marrow-derived hematopoietic stem cells (BMSCs). Few and discordant studies have evaluated the role of BMSC in hepatocarcinogenesis so far and further studies in vitro and in vivo are warranted in order to definitively clarify such an issue. PMID:25202697

Lipoprotein lipase regulates hematopoietic stem progenitor cell maintenance through DHA supply.

PubMed

Liu, Chao; Han, Tianxu; Stachura, David L; Wang, Huawei; Vaisman, Boris L; Kim, Jungsu; Klemke, Richard L; Remaley, Alan T; Rana, Tariq M; Traver, David; Miller, Yury I

2018-04-03

Lipoprotein lipase (LPL) mediates hydrolysis of triglycerides (TGs) to supply free fatty acids (FFAs) to tissues. Here, we show that LPL activity is also required for hematopoietic stem progenitor cell (HSPC) maintenance. Knockout of Lpl or its obligatory cofactor Apoc2 results in significantly reduced HSPC expansion during definitive hematopoiesis in zebrafish. A human APOC2 mimetic peptide or the human very low-density lipoprotein, which carries APOC2, rescues the phenotype in apoc2 but not in lpl mutant zebrafish. Creating parabiotic apoc2 and lpl mutant zebrafish rescues the hematopoietic defect in both. Docosahexaenoic acid (DHA) is identified as an important factor in HSPC expansion. FFA-DHA, but not TG-DHA, rescues the HSPC defects in apoc2 and lpl mutant zebrafish. Reduced blood cell counts are also observed in Apoc2 mutant mice at the time of weaning. These results indicate that LPL-mediated release of the essential fatty acid DHA regulates HSPC expansion and definitive hematopoiesis.

Desensitization for solid organ and hematopoietic stem cell transplantation

PubMed Central

Zachary, Andrea A; Leffell, Mary S

2014-01-01

Desensitization protocols are being used worldwide to enable kidney transplantation across immunologic barriers, i.e. antibody to donor HLA or ABO antigens, which were once thought to be absolute contraindications to transplantation. Desensitization protocols are also being applied to permit transplantation of HLA mismatched hematopoietic stem cells to patients with antibody to donor HLA, to enhance the opportunity for transplantation of non-renal organs, and to treat antibody-mediated rejection. Although desensitization for organ transplantation carries an increased risk of antibody-mediated rejection, ultimately these transplants extend and enhance the quality of life for solid organ recipients, and desensitization that permits transplantation of hematopoietic stem cells is life saving for patients with limited donor options. Complex patient factors and variability in treatment protocols have made it difficult to identify, precisely, the mechanisms underlying the downregulation of donor-specific antibodies. The mechanisms underlying desensitization may differ among the various protocols in use, although there are likely to be some common features. However, it is likely that desensitization achieves a sort of immune detente by first reducing the immunologic barrier and then by creating an environment in which an autoregulatory process restricts the immune response to the allograft. PMID:24517434

Inductive potential of recombinant human granulocyte colony-stimulating factor to mature neutrophils from x-irradiated human peripheral blood hematopoietic progenitor cells.

PubMed

Katsumori, Takeo; Yoshino, Hironori; Hayashi, Masako; Takahashi, Kenji; Kashiwakura, Ikuo

2009-11-01

Recombinant human granulocyte colony-stimulating factor (rhG-CSF) has been used for treatment of neutropenia. Filgrastim, Nartograstim, and Lenograstim are clinically available in Japan. However, the differences in potential benefit for radiation-induced disorder between these types of rhG-CSFs remain unknown. Therefore, the effects of three different types of rhG-CSFs on granulocyte progenitor cells and expansion of neutrophils from nonirradiated or 2 Gy X-irradiated human CD34+ hematopoietic progenitor cells were examined. For analysis of granulocyte colony-forming units (CFU-G) and a surviving fraction of CFU-G, nonirradiated or X-irradiated CD34+ cells were cultured in methylcellulose containing rhG-CSF. These cells were cultured in serum-free medium supplemented with rhG-CSF, and the expansion and characteristics of neutrophils were analyzed. All three types of rhG-CSFs increased the number of CFU-G in a dose-dependent manner; however, Lenograstim is superior to others because of CFU-G-derived colony formation at relatively low doses. The surviving fraction of CFU-G was independent of the types of rhG-CSFs. Expansion of neutrophils by rhG-CSF was largely attenuated by X-irradiation, though no significant difference in neutrophil number was observed between the three types of rhG-CSFs under both nonirradiation and X-irradiation conditions. In terms of functional characteristics of neutrophils, Lenograstim-induced neutrophils produced high levels of reactive oxygen species compared to Filgrastim, when rhG-CSF was applied to nonirradiated CD34(+) cells. In conclusion, different types of rhG-CSFs lead to different effects when rhG-CSF is applied to nonirradiated CD34+ cells, though Filgrastim, Nartograstim, and Lenograstim show equal effects on X-irradiated CD34+ cells.

The Sequence of Cyclophosphamide and Myeloablative Total Body Irradiation in Hematopoietic Cell Transplantation for Patients with Acute Leukemia.

PubMed

Holter-Chakrabarty, Jennifer L; Pierson, Namali; Zhang, Mei-Jie; Zhu, Xiaochun; Akpek, Görgün; Aljurf, Mahmoud D; Artz, Andrew S; Baron, Frédéric; Bredeson, Christopher N; Dvorak, Christopher C; Epstein, Robert B; Lazarus, Hillard M; Olsson, Richard F; Selby, George B; Williams, Kirsten M; Cooke, Kenneth R; Pasquini, Marcelo C; McCarthy, Philip L

2015-07-01

Limited clinical data are available to assess whether the sequencing of cyclophosphamide (Cy) and total body irradiation (TBI) changes outcomes. We evaluated the sequence in 1769 (CyTBI, n = 948; TBICy, n = 821) recipients of related or unrelated hematopoietic cell transplantation who received TBI (1200 to 1500 cGY) for acute leukemia from 2003 to 2010. The 2 cohorts were comparable for median age, performance score, type of leukemia, first complete remission, Philadelphia chromosome-positive acute lymphoblastic leukemia, HLA-matched siblings, stem cell source, antithymocyte globulin use, TBI dose, and type of graft-versus-host disease (GVHD) prophylaxis. The sequence of TBI did not significantly affect transplantation-related mortality (24% versus 23% at 3 years, P = .67; relative risk, 1.01; P = .91), leukemia relapse (27% versus 29% at 3 years, P = .34; relative risk, .89, P = .18), leukemia-free survival (49% versus 48% at 3 years, P = .27; relative risk, .93; P = .29), chronic GVHD (45% versus 47% at 1 year, P = .39; relative risk, .9; P = .11), or overall survival (53% versus 52% at 3 years, P = .62; relative risk, .96; P = .57) for CyTBI and TBICy, respectively. Corresponding cumulative incidences of sinusoidal obstruction syndrome were 4% and 6% at 100 days (P = .08), respectively. This study demonstrates that the sequence of Cy and TBI does not impact transplantation outcomes and complications in patients with acute leukemia undergoing hematopoietic cell transplantation with myeloablative conditioning. Copyright © 2015 American Society for Blood and Marrow Transplantation. Published by Elsevier Inc. All rights reserved.

Autologous peripheral blood hematopoietic cell transplantation in dogs with T-cell lymphoma.

PubMed

Warry, E E; Willcox, J L; Suter, S E

2014-01-01

Peripheral blood hematopoietic cell transplantation (PBHCT) is a feasible treatment option for dogs with B-cell lymphoma. To examine apheresis and PBHCT outcomes in dogs diagnosed with T-cell lymphoma (TCL). Fifteen client-owned dogs diagnosed with high-grade TCL. After high-dose cyclophosphamide and rhG-colony-stimulating (rhG-CSF) factor treatment, peripheral blood mononuclear cells were collected using cell separators. The harvested cells then were infused after varying doses of total body irradiation (TBI). Postirradiation adverse effects were managed symptomatically and dogs were discharged upon evidence of hematopoietic engraftment. More than 2 × 10(6) CD34+ cells/kg were harvested from 15/15 dogs. Thirteen of 15 (87%) dogs engrafted appropriately, whereas 2 (13%) of the dogs died in the hospital. One dog developed cutaneous B-cell lymphoma 120 days post-PBHCT. The median disease-free interval and overall survival (OS) of the 13 dogs transplanted in first remission from the time of PBHCT were 184 and 240 days, respectively. Stage and substage of disease at diagnosis had no effect on OS. Two of 13 (15%) dogs were alive 741 and 772 days post-PBHCT. PBHCT may be considered as a treatment option for dogs with TCL. Copyright © 2014 by the American College of Veterinary Internal Medicine.

Inhibiting glycogen synthase kinase-3 mitigates the hematopoietic acute radiation syndrome in mice.

PubMed

Lee, Chang-Lung; Lento, William E; Castle, Katherine D; Chao, Nelson J; Kirsch, David G

2014-05-01

Exposure to a nuclear accident or radiological attack can cause death from acute radiation syndrome (ARS), which results from radiation injury to vital organs such as the hematopoietic system. However, the U.S. Food and Drug Administration (FDA) has not approved any medical countermeasures for this specific purpose. With growing concern over nuclear terrorism, there is an urgent need to develop small molecule deliverables that mitigate mortality from ARS. One emerging modulator of hematopoietic stem/progenitor cell (HSPC) activity is glycogen synthase kinase-3 (GSK-3). The inhibition of GSK-3 has been shown to augment hematopoietic repopulation in mouse models of bone marrow transplantation. In this study, we performed an in vitro screen using irradiated bone marrow mononuclear cells (BM-MNCs) to test the effects of four GSK-3 inhibitors: CHIR99021; 6-Bromoindirubin-3'-oxime (BIO); SB415286; and SB216763. This screen showed that SB216763 significantly increased the frequency of c-Kit(+) Lin(-) Sca1(+) (KLS) cells and hematopoietic colony-forming cells in irradiated BM-MNCs. Importantly, administration of a single dose of SB216763 to C57BL/6J mice by subcutaneous injection 24 h after total-body irradiation significantly improved hematopoietic recovery and mitigated hematopoietic ARS. Collectively, our results demonstrate that the GSK-3 inhibitor SB216763 is an effective medical countermeasure against acute radiation injury of the hematopoietic system.

Biology and flow cytometry of proangiogenic hematopoietic progenitors cells.

PubMed

Rose, Jonathan A; Erzurum, Serpil; Asosingh, Kewal

2015-01-01

During development, hematopoiesis and neovascularization are closely linked to each other via a common bipotent stem cell called the hemangioblast that gives rise to both hematopoietic cells and endothelial cells. In postnatal life, this functional connection between the vasculature and hematopoiesis is maintained by a subset of hematopoietic progenitor cells endowed with the capacity to differentiate into potent proangiogenic cells. These proangiogenic hematopoietic progenitors comprise a specific subset of bone marrow (BM)-derived cells that homes to sites of neovascularization and possess potent paracrine angiogenic activity. There is emerging evidence that this subpopulation of hematopoietic progenitors plays a critical role in vascular health and disease. Their angiogenic activity is distinct from putative "endothelial progenitor cells" that become structural cells of the endothelium by differentiation into endothelial cells. Proangiogenic hematopoietic progenitor cell research requires multidisciplinary expertise in flow cytometry, hematology, and vascular biology. This review provides a comprehensive overview of proangiogenic hematopoietic progenitor cell biology and flow cytometric methods to detect these cells in the peripheral blood circulation and BM. © 2014 International Society for Advancement of Cytometry.

l-Arginine is a Radioprotector for Hematopoietic Progenitor Cells

PubMed Central

Pearce, Linda L.; Zheng, Xichen; Martinez-Bosch, Sandra; Kerr, Patrick P.; Khlangwiset, Pornsri; Epperly, Michael W.; Fink, Mitchell P.; Greenberger, Joel S.; Peterson, Jim

2012-01-01

l-Arginine is shown to protect hematopoietic progenitor (32D cl 3) cells from death due to exposure to γ radiation (137Cs). Some of the other intermediates in the urea cycle, namely ornithine and citrulline, plus urea itself, were not found to have any significant impact on cell survival after irradiation. Intriguingly, supplementation of irradiated cells with l-arginine results in decreased production of peroxynitrite, suggesting that suppression of superoxide generation by nitric oxide synthase in one or more microenvironments is an important factor in the observed radioprotection. The absence of any radioprotective effect of l-arginine in cells at 3% oxygen also confirms the involvement of one or more oxygen-derived species. Knockdown experiments with nitric oxide synthase (NOS) siRNAs in cells and NOS knockout animals confirm that the observed radioprotection is associated with nNOS (NOS-1). l-Arginine also ameliorates the transient inhibition of the electron-transport chain complex I that occurs within 30 min of completing the dose (10 Gy) and that appears to be a functional marker for postirradiation mitochondrial oxidant production. PMID:22175298

Inhibition of T Cell Protein Tyrosine Phosphatase Enhances Interleukin-18-Dependent Hematopoietic Stem Cell Expansion

PubMed Central

Bourdeau, Annie; Trop, Sébastien; Doody, Karen M; Dumont, Daniel J; Tremblayef, Michel L

2013-01-01

The clinical application of hematopoietic progenitor cell-based therapies for the treatment of hematological diseases is hindered by current protocols, which are cumbersome and have limited efficacy to augment the progenitor cell pool. We report that inhibition of T-cell protein tyrosine phosphatase (TC-PTP), an enzyme involved in the regulation of cytokine signaling, through gene knockout results in a ninefold increase in the number of hematopoietic progenitors in murine bone marrow (BM). This effect could be reproduced using a short (48 hours) treatment with a pharmacological inhibitor of TC-PTP in murine BM, as well as in human BM, peripheral blood, and cord blood. We also demonstrate that the ex vivo use of TC-PTP inhibitor only provides a temporary effect on stem cells and did not alter their capacity to reconstitute all hematopoietic components in vivo. We establish that one of the mechanisms whereby inhibition of TC-PTP mediates its effects involves the interleukin-18 (IL-18) signaling pathway, leading to increased production of IL-12 and interferon-gamma by progenitor cells. Together, our results reveal a previously unrecognized role for IL-18 in contributing to the augmentation of the stem cell pool and provide a novel and simple method to rapidly expand progenitor cells from a variety of sources using a pharmacological compound. Stem Cells 2013;31:293–304 PMID:23135963

Hematopoietic stem cell-specific GFP-expressing transgenic mice generated by genetic excision of a pan-hematopoietic reporter gene.

PubMed

Perez-Cunningham, Jessica; Boyer, Scott W; Landon, Mark; Forsberg, E Camilla

2016-08-01

Selective labeling of specific cell types by expression of green fluorescent protein (GFP) within the hematopoietic system would have great utility in identifying, localizing, and tracking different cell populations in flow cytometry, microscopy, lineage tracing, and transplantation assays. In this report, we describe the generation and characterization of a new transgenic mouse line with specific GFP labeling of all nucleated hematopoietic cells and platelets. This new "Vav-GFP" mouse line labels the vast majority of hematopoietic cells with GFP during both embryonic development and adulthood, with particularly high expression in hematopoietic stem and progenitor cells (HSPCs). With the exception of transient labeling of fetal endothelial cells, GFP expression is highly selective for hematopoietic cells and persists in donor-derived progeny after transplantation of HSPCs. Finally, we also demonstrate that the loxP-flanked reporter allows for specific GFP labeling of different hematopoietic cell subsets when crossed to various Cre reporter lines. By crossing Vav-GFP mice to Flk2-Cre mice, we obtained robust and highly selective GFP expression in hematopoietic stem cells (HSCs). These data describe a new mouse model capable of directing GFP labeling exclusively of hematopoietic cells or exclusively of HSCs. Copyright © 2016 ISEH - International Society for Experimental Hematology. Published by Elsevier Inc. All rights reserved.

CCND1–CDK4–mediated cell cycle progression provides a competitive advantage for human hematopoietic stem cells in vivo

PubMed Central

Mende, Nicole; Kuchen, Erika E.; Lesche, Mathias; Grinenko, Tatyana; Kokkaliaris, Konstantinos D.; Hanenberg, Helmut; Lindemann, Dirk; Dahl, Andreas; Platz, Alexander; Höfer, Thomas; Calegari, Federico

2015-01-01

Maintenance of stem cell properties is associated with reduced proliferation. However, in mouse hematopoietic stem cells (HSCs), loss of quiescence results in a wide range of phenotypes, ranging from functional failure to extensive self-renewal. It remains unknown whether the function of human HSCs is controlled by the kinetics of cell cycle progression. Using human HSCs and human progenitor cells (HSPCs), we report here that elevated levels of CCND1–CDK4 complexes promoted the transit from G0 to G1 and shortened the G1 cell cycle phase, resulting in protection from differentiation-inducing signals in vitro and increasing human leukocyte engraftment in vivo. Further, CCND1–CDK4 overexpression conferred a competitive advantage without impacting HSPC numbers. In contrast, accelerated cell cycle progression mediated by elevated levels of CCNE1–CDK2 led to the loss of functional HSPCs in vivo. Collectively, these data suggest that the transition kinetics through the early cell cycle phases are key regulators of human HSPC function and important for lifelong hematopoiesis. PMID:26150472

Reduced hematopoietic stem cell frequency predicts outcome in acute myeloid leukemia.

PubMed

Wang, Wenwen; Stiehl, Thomas; Raffel, Simon; Hoang, Van T; Hoffmann, Isabel; Poisa-Beiro, Laura; Saeed, Borhan R; Blume, Rachel; Manta, Linda; Eckstein, Volker; Bochtler, Tilmann; Wuchter, Patrick; Essers, Marieke; Jauch, Anna; Trumpp, Andreas; Marciniak-Czochra, Anna; Ho, Anthony D; Lutz, Christoph

2017-09-01

In patients with acute myeloid leukemia and low percentages of aldehyde-dehydrogenase-positive cells, non-leukemic hematopoietic stem cells can be separated from leukemic cells. By relating hematopoietic stem cell frequencies to outcome we detected poor overall- and disease-free survival of patients with low hematopoietic stem cell frequencies. Serial analysis of matched diagnostic and follow-up samples further demonstrated that hematopoietic stem cells increased after chemotherapy in patients who achieved durable remissions. However, in patients who eventually relapsed, hematopoietic stem cell numbers decreased dramatically at the time of molecular relapse demonstrating that hematopoietic stem cell levels represent an indirect marker of minimal residual disease, which heralds leukemic relapse. Upon transplantation in immune-deficient mice cases with low percentages of hematopoietic stem cells of our cohort gave rise to leukemic or no engraftment, whereas cases with normal hematopoietic stem cell levels mostly resulted in multi-lineage engraftment. Based on our experimental data, we propose that leukemic stem cells have increased niche affinity in cases with low percentages of hematopoietic stem cells. To validate this hypothesis, we developed new mathematical models describing the dynamics of healthy and leukemic cells under different regulatory scenarios. These models suggest that the mechanism leading to decreases in hematopoietic stem cell frequencies before leukemic relapse must be based on expansion of leukemic stem cells with high niche affinity and the ability to dislodge hematopoietic stem cells. Thus, our data suggest that decreasing numbers of hematopoietic stem cells indicate leukemic stem cell persistence and the emergence of leukemic relapse. Copyright© 2017 Ferrata Storti Foundation.

Nitric oxide-mediated bystander signal transduction induced by heavy-ion microbeam irradiation

NASA Astrophysics Data System (ADS)

Tomita, Masanori; Matsumoto, Hideki; Funayama, Tomoo; Yokota, Yuichiro; Otsuka, Kensuke; Maeda, Munetoshi; Kobayashi, Yasuhiko

2015-07-01

In general, a radiation-induced bystander response is known to be a cellular response induced in non-irradiated cells after receiving bystander signaling factors released from directly irradiated cells within a cell population. Bystander responses induced by high-linear energy transfer (LET) heavy ions at low fluence are an important health problem for astronauts in space. Bystander responses are mediated via physical cell-cell contact, such as gap-junction intercellular communication (GJIC) and/or diffusive factors released into the medium in cell culture conditions. Nitric oxide (NO) is a well-known major initiator/mediator of intercellular signaling within culture medium during bystander responses. In this study, we investigated the NO-mediated bystander signal transduction induced by high-LET argon (Ar)-ion microbeam irradiation of normal human fibroblasts. Foci formation by DNA double-strand break repair proteins was induced in non-irradiated cells, which were co-cultured with those irradiated by high-LET Ar-ion microbeams in the same culture plate. Foci formation was suppressed significantly by pretreatment with an NO scavenger. Furthermore, NO-mediated reproductive cell death was also induced in bystander cells. Phosphorylation of NF-κB and Akt were induced during NO-mediated bystander signaling in the irradiated and bystander cells. However, the activation of these proteins depended on the incubation time after irradiation. The accumulation of cyclooxygenase-2 (COX-2), a downstream target of NO and NF-κB, was observed in the bystander cells 6 h after irradiation but not in the directly irradiated cells. Our findings suggest that Akt- and NF-κB-dependent signaling pathways involving COX-2 play important roles in NO-mediated high-LET heavy-ion-induced bystander responses. In addition, COX-2 may be used as a molecular marker of high-LET heavy-ion-induced bystander cells to distinguish them from directly irradiated cells, although this may depend on the time

Vitamin A-Retinoic Acid Signaling Regulates Hematopoietic Stem Cell Dormancy.

PubMed

Cabezas-Wallscheid, Nina; Buettner, Florian; Sommerkamp, Pia; Klimmeck, Daniel; Ladel, Luisa; Thalheimer, Frederic B; Pastor-Flores, Daniel; Roma, Leticia P; Renders, Simon; Zeisberger, Petra; Przybylla, Adriana; Schönberger, Katharina; Scognamiglio, Roberta; Altamura, Sandro; Florian, Carolina M; Fawaz, Malak; Vonficht, Dominik; Tesio, Melania; Collier, Paul; Pavlinic, Dinko; Geiger, Hartmut; Schroeder, Timm; Benes, Vladimir; Dick, Tobias P; Rieger, Michael A; Stegle, Oliver; Trumpp, Andreas

2017-05-18

Dormant hematopoietic stem cells (dHSCs) are atop the hematopoietic hierarchy. The molecular identity of dHSCs and the mechanisms regulating their maintenance or exit from dormancy remain uncertain. Here, we use single-cell RNA sequencing (RNA-seq) analysis to show that the transition from dormancy toward cell-cycle entry is a continuous developmental path associated with upregulation of biosynthetic processes rather than a stepwise progression. In addition, low Myc levels and high expression of a retinoic acid program are characteristic for dHSCs. To follow the behavior of dHSCs in situ, a Gprc5c-controlled reporter mouse was established. Treatment with all-trans retinoic acid antagonizes stress-induced activation of dHSCs by restricting protein translation and levels of reactive oxygen species (ROS) and Myc. Mice maintained on a vitamin A-free diet lose HSCs and show a disrupted re-entry into dormancy after exposure to inflammatory stress stimuli. Our results highlight the impact of dietary vitamin A on the regulation of cell-cycle-mediated stem cell plasticity. VIDEO ABSTRACT. Copyright © 2017. Published by Elsevier Inc.

Desensitization for solid organ and hematopoietic stem cell transplantation.

PubMed

Zachary, Andrea A; Leffell, Mary S

2014-03-01

Desensitization protocols are being used worldwide to enable kidney transplantation across immunologic barriers, i.e. antibody to donor HLA or ABO antigens, which were once thought to be absolute contraindications to transplantation. Desensitization protocols are also being applied to permit transplantation of HLA mismatched hematopoietic stem cells to patients with antibody to donor HLA, to enhance the opportunity for transplantation of non-renal organs, and to treat antibody-mediated rejection. Although desensitization for organ transplantation carries an increased risk of antibody-mediated rejection, ultimately these transplants extend and enhance the quality of life for solid organ recipients, and desensitization that permits transplantation of hematopoietic stem cells is life saving for patients with limited donor options. Complex patient factors and variability in treatment protocols have made it difficult to identify, precisely, the mechanisms underlying the downregulation of donor-specific antibodies. The mechanisms underlying desensitization may differ among the various protocols in use, although there are likely to be some common features. However, it is likely that desensitization achieves a sort of immune detente by first reducing the immunologic barrier and then by creating an environment in which an autoregulatory process restricts the immune response to the allograft. © 2014 The Authors. Immunological Reviews Published by John Wiley & Sons Ltd.

Regulation of long-term repopulating hematopoietic stem cells by EPCR/PAR1 signaling

PubMed Central

Gur-Cohen, Shiri; Kollet, Orit; Graf, Claudine; Esmon, Charles T.; Ruf, Wolfram; Lapidot, Tsvee

2016-01-01

The common developmental origin of endothelial and hematopoietic cells is manifested by coexpression of several cell surface receptors. Adult murine bone marrow (BM) long-term repopulating hematopoietic stem cells (LT-HSCs), endowed with the highest repopulation and self-renewal potential, express endothelial protein C receptor (EPCR), which is used as a marker to isolate them. EPCR/PAR1 signaling in endothelial cells has anticoagulant and anti-inflammatory roles, while thrombin/PAR1 signaling induces coagulation and inflammation. Recent studies define two new PAR1-mediated signaling cascades that regulate EPCR+ LT-HSC BM retention and egress. EPCR/PAR1 signaling facilitates LT-HSC BM repopulation, retention, survival, and chemotherapy resistance by restricting nitric oxide (NO) production, maintaining NOlow LT-HSC BM retention with increased VLA4 expression, affinity, and adhesion. Conversely, acute stress and clinical mobilization upregulate thrombin generation and activate different PAR1 signaling which overcomes BM EPCR+ LT-HSC retention, inducing their recruitment to the bloodstream. Thrombin/PAR1 signaling induces NO generation, TACE-mediated EPCR shedding, and upregulation of CXCR4 and PAR1, leading to CXCL12-mediated stem and progenitor cell mobilization. This review discusses new roles for factors traditionally viewed as coagulation related, which independently act in the BM to regulate PAR1 signaling in bone- and blood-forming progenitor cells, navigating their fate by controlling NO production. PMID:26928241

Impact of Abbreviated Filgrastim Schedule on Survival and Hematopoietic Recovery after Irradiation in Four Mouse Strains with Different Radiosensitivity

PubMed Central

Satyamitra, Merriline; Kumar, Vidya P.; Biswas, Shukla; Cary, Lynnette; Dickson, Leonora; Venkataraman, Srinivasan; Ghosh, Sanchita P.

2017-01-01

Filgrastim (Neupogen®, granulocyte-colony stimulating factor) is among the few countermeasures recommended for management of patients in the event of lethal total-body irradiation. Despite the plethora of studies using filgrastim as a radiation countermeasure, relatively little is known about the optimal dose schedule of filgrastim to mitigate radiation lethality. We evaluated the efficacy of filgrastim in improving 30-day survival of CD2F1 mice irradiated with a lethal dose (LD70/30) in the AFRRI cobalt-60 facility. We tested different schedules of 1, 3, 5,10 or 16 once-daily injections of filgrastim initiated one day after irradiation. Time optimization studies with filgrastim treatment were also performed, beginning 6–48 h postirradiation. Maximum survival was observed with 3 daily doses of 0.17 mg/kg filgrastim. Survival efficacy of the 3-day treatment was compared against the conventional 16-day filgrastim treatment after irradiation in four mouse strains with varying radiation sensitivities: C3H/HeN, C57BL/6, B6C3F1 and CD2F1. Blood indices, bone marrow histopathology and colony forming unit assays were also evaluated. Filgrastim significantly increased 30-day survival (P < 0.001) with a 3-day treatment compared to 16-day treatment. Filgrastim did not prevent cytopenia nadirs, but facilitated faster recovery of white blood cells, neutrophils, red blood cells, platelets, lymphocytes and hematocrits in all four strains. Accelerated hematopoietic recovery was also reflected in faster bone marrow reconstitution and significant increase in hematopoietic progenitors (P < 0.001) in all four mouse strains. These data indicate that prompt and abbreviated filgrastim treatment has potential benefit for triage in the event of a radiological incident for treating acute hematopoietic syndrome. PMID:28362168

Expression of Fas and Fas-ligand in donor hematopoietic stem and progenitor cells is dissociated from the sensitivity to apoptosis.

PubMed

Pearl-Yafe, Michal; Yolcu, Esma S; Stein, Jerry; Kaplan, Ofer; Shirwan, Haval; Yaniv, Isaac; Askenasy, Nadir

2007-10-01

The interaction between the Fas receptor and its cognate ligand (FasL) has been implicated in the mutual suppression of donor and host hematopoietic cells after transplantation. Following the observation of deficient early engraftment of Fas and FasL-defective donor cells and recipients, we determined the role of the Fas-FasL interaction. Donor cells were recovered after syngeneic (CD45.1-->CD45.2) transplants from various organs and assessed for expression of Fas/FasL in reference to lineage markers, carboxyfluorescein succinimidyl ester dilution, Sca-1 and c-kit expression. Naïve and bone marrow-homed cells were challenged for apoptosis ex vivo. The Fas receptor and ligand were markedly upregulated to 40% to 60% (p < 0.001 vs 5-10% in naïve cells) within 2 days after syngeneic transplantation, while residual host cells displayed modest and delayed upregulation of these molecules ( approximately 10%). All lin(-)Sca(+)c-kit(+) cells were Fas(+)FasL(+), including 95% of Sca-1(+) and 30% of c-kit(+) cells. Fas and FasL expression varied in donor cells that homed to bone marrow, spleen, liver and lung, and was induced by interaction with the stroma, irradiation, cell cycling, and differentiation. Bone marrow-homed donor cells challenged with supralethal doses of FasL were insensitive to apoptosis (3.2% +/- 1% vs 38% +/- 5% in naïve bone marrow cells), and engraftment was not affected by pretransplantation exposure of donor cells to an apoptotic challenge with FasL. There was no evidence of Fas-mediated suppression of donor and host cell activity after transplantation. Resistance to Fas-mediated apoptosis evolves as a functional characteristic of hematopoietic reconstituting stem and progenitor cells, providing them competitive engraftment advantage over committed progenitors.

Scorpion venom peptide SPVII promotes irradiated cells proliferation and increases the expression of the IL-3 receptor

PubMed Central

2013-01-01

Background The previous investigation demonstrated the radioprotective efficacy of peptides isolated from the venom of Buthus Martti Karsch. In this study, the effect of isolated scorpion venom peptide II (SVPII) on irradiated M-NFS-60 cells and mouse bone marrow mononuclear cells (BM-MNCs) was observed. The AlamarBlue cell viability assay, a colony-forming unit (CFU) assay, flow cytometry (FCM), immunofluorescence, and Western blotting were used to evaluate cell proliferation, cell cycle progression, and the expression of the IL-3 receptor (IL-3R) protein in non-irradiated and irradiated cells. Results Proliferation of irradiated M-NFS-60 cells was significantly accelerated by SPVII, and this effect was further enhanced by co-application of IL-3. Similarly, SPVII increased the number of BM-MNC CFUs and this proliferative effect was greater in the presence of SVPII plus IL-3. In addition, SPVII significantly altered cell cycle progression; SVPII enhanced the fraction of unirradiated M-NFS-60 cells in S phase and the fraction of irradiated M-NFS-60 cells arrested in G2/M. The expression of IL-3R protein by unirradiated M-NFS-60 cells was enhanced significantly by SVPII, and SVPII-induced IL-3R overexpression was 10-fold greater in irradiated M-NFS-60 cells. Conclusions These results indicated the hematopoietic growth factor (HGF)-like effects of SVPII on irradiated cells, possibly mediated by upregulation of IL-3R. PMID:23835458

Cadmium modulates hematopoietic stem and progenitor cells and skews toward myelopoiesis in mice

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhang, Yandong; Yu, Xinchun

The heavy metal cadmium (Cd) is known to modulate immunity and cause osteoporosis. However, how Cd influences on hematopoiesis remain largely unknown. Herein, we show that wild-type C57BL/6 (B6) mice exposed to Cd for 3 months had expanded bone marrow (BM) populations of long-term hematopoietic stem cells (LT-HSCs), common myeloid progenitors (CMPs) and granulocyte-macrophage progenitors (GMPs), while having reduced populations of multipotent progenitors (MPPs) and common lymphoid progenitors (CLPs). A competitive mixed BM transplantation assay indicates that BM from Cd-treated mice had impaired LT-HSC ability to differentiate into mature cells. In accordance with increased myeloid progenitors and decreased lymphoid progenitors,more » the BM and spleens of Cd-treated mice had more monocytes and/or neutrophils and fewer B cells and T cells. Cd impaired the ability of the non-hematopoietic system to support LT-HSCs, in that lethally irradiated Cd-treated recipients transplanted with normal BM cells had reduced LT-HSCs after the hematopoietic system was fully reconstituted. This is consistent with reduced osteoblasts, a known critical component for HSC niche, observed in Cd-treated mice. Conversely, lethally irradiated control recipients transplanted with BM cells from Cd-treated mice had normal LT-HSC reconstitution. Furthermore, both control mice and Cd-treated mice that received Alendronate, a clinical drug used for treating osteoporosis, had BM increases of LT-HSCs. Thus, the results suggest Cd increase of LT-HSCs is due to effects on HSCs and not on osteoblasts, although, Cd causes osteoblast reduction and impaired niche function for maintaining HSCs. Furthermore, Cd skews HSCs toward myelopoiesis. - Highlights: • Cd increases the number of LT-HSCs but impairs their development. • Cd-treated hosts have compromised ability to support LT-HSCs. • Cd promotes myelopoiesis at the expense of lymphopoiesis at the MPP level.« less

c-Myc-Induced Survivin Is Essential for Promoting the Notch-Dependent T Cell Differentiation from Hematopoietic Stem Cells

PubMed Central

Haque, Rizwanul; Song, Jianyong; Haque, Mohammad; Lei, Fengyang; Sandhu, Praneet; Ni, Bing; Zheng, Songguo; Fang, Deyu; Yang, Jin-Ming; Song, Jianxun

2017-01-01

Notch is indispensable for T cell lineage commitment, and is needed for thymocyte differentiation at early phases. During early stages of T cell development, active Notch prevents other lineage potentials including B cell lineage and myeloid cell (e.g., dendritic cell) lineage. Nevertheless, the precise intracellular signaling pathways by which Notch promotes T cell differentiation remain unclear. Here we report that the transcription factor c-Myc is a key mediator of the Notch signaling–regulated T cell differentiation. In a well-established in vitro differentiation model of T lymphocytes from hematopoietic stem cells, we showed that Notch1 and 4 directly promoted c-Myc expression; dominant-negative (DN) c-Myc inhibited early T cell differentiation. Moreover, the c-Myc expression activated by Notch signaling increased the expression of survivin, an inhibitor of apoptosis (IAP) protein. We further demonstrated that over-expression of c-Myc increased the abundance of survivin and the T cell differentiation thereof, whereas dn c-Myc reduced survivin levels and concomitantly retarded the differentiation. The c-Myc–dependent survivin induction is functionally germane, because Notch-dependent T cell differentiation was canceled by the depletion of survivin. These results identify both c-Myc and survivin as important mediators of the Notch signaling–regulated differentiation of T lymphocytes from hematopoietic stem cells. PMID:28272325

Hhex Regulates Hematopoietic Stem Cell Self-Renewal and Stress Hematopoiesis via Repression of Cdkn2a.

PubMed

Jackson, Jacob T; Shields, Benjamin J; Shi, Wei; Di Rago, Ladina; Metcalf, Donald; Nicola, Nicos A; McCormack, Matthew P

2017-08-01

The hematopoietically expressed homeobox transcription factor (Hhex) is important for the maturation of definitive hematopoietic progenitors and B-cells during development. We have recently shown that in adult hematopoiesis, Hhex is dispensable for maintenance of hematopoietic stem cells (HSCs) and myeloid lineages but essential for the commitment of common lymphoid progenitors (CLPs) to lymphoid lineages. Here, we show that during serial bone marrow transplantation, Hhex-deleted HSCs are progressively lost, revealing an intrinsic defect in HSC self-renewal. Moreover, Hhex-deleted mice show markedly impaired hematopoietic recovery following myeloablation, due to a failure of progenitor expansion. In vitro, Hhex-null blast colonies were incapable of replating, implying a specific requirement for Hhex in immature progenitors. Transcriptome analysis of Hhex-null Lin - Sca + Kit + cells showed that Hhex deletion leads to derepression of polycomb repressive complex 2 (PRC2) and PRC1 target genes, including the Cdkn2a locus encoding the tumor suppressors p16 Ink 4 a and p19 Arf . Indeed, loss of Cdkn2a restored the capacity of Hhex-null blast colonies to generate myeloid progenitors in vitro, as well as hematopoietic reconstitution following myeloablation in vivo. Thus, HSCs require Hhex to promote PRC2-mediated Cdkn2a repression to enable continued self-renewal and response to hematopoietic stress. Stem Cells 2017;35:1948-1957. © 2017 AlphaMed Press.

Latexin Inactivation Enhances Survival and Long-Term Engraftment of Hematopoietic Stem Cells and Expands the Entire Hematopoietic System in Mice.

PubMed

Liu, Yi; Zhang, Cuiping; Li, Zhenyu; Wang, Chi; Jia, Jianhang; Gao, Tianyan; Hildebrandt, Gerhard; Zhou, Daohong; Bondada, Subbarao; Ji, Peng; St Clair, Daret; Liu, Jinze; Zhan, Changguo; Geiger, Hartmut; Wang, Shuxia; Liang, Ying

2017-04-11

Natural genetic diversity offers an important yet largely untapped resource to decipher the molecular mechanisms regulating hematopoietic stem cell (HSC) function. Latexin (Lxn) is a negative stem cell regulatory gene identified on the basis of genetic diversity. By using an Lxn knockout mouse model, we found that Lxn inactivation in vivo led to the physiological expansion of the entire hematopoietic hierarchy. Loss of Lxn enhanced the competitive repopulation capacity and survival of HSCs in a cell-intrinsic manner. Gene profiling of Lxn-null HSCs showed altered expression of genes enriched in cell-matrix and cell-cell interactions. Thrombospondin 1 (Thbs1) was a potential downstream target with a dramatic downregulation in Lxn-null HSCs. Enforced expression of Thbs1 restored the Lxn inactivation-mediated HSC phenotypes. This study reveals that Lxn plays an important role in the maintenance of homeostatic hematopoiesis, and it may lead to development of safe and effective approaches to manipulate HSCs for clinical benefit. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Hematopoietic stem cell loss and hematopoietic failure in severe aplastic anemia is driven by macrophages and aberrant podoplanin expression.

PubMed

McCabe, Amanda; Smith, Julianne N P; Costello, Angelica; Maloney, Jackson; Katikaneni, Divya; MacNamara, Katherine C

2018-05-17

Severe aplastic anemia results from profound hematopoietic stem cell loss. T cells and interferon gamma have long been associated with severe aplastic anemia, yet the underlying mechanisms driving hematopoietic stem cell loss remain unknown. Using a mouse model of severe aplastic anemia, we demonstrate that interferon gamma-dependent hematopoietic stem cell loss required macrophages. Interferon gamma was necessary for bone marrow macrophage persistence, despite loss of other myeloid cells and hematopoietic stem cells. Depleting macrophages or abrogating interferon gamma signaling specifically in macrophages did not impair T cell activation or interferon gamma production in the bone marrow but rescued hematopoietic stem cells and reduced mortality. Thus, macrophages are not required for induction of interferon gamma in severe aplastic anemia and rather act as sensors of interferon gamma. Macrophage depletion rescued thrombocytopenia, increased bone marrow megakaryocytes, preserved platelet-primed stem cells, and increased the platelet-repopulating capacity of transplanted hematopoietic stem cells. In addition to the hematopoietic effects, severe aplastic anemia induced loss of non-hematopoietic stromal populations, including podoplanin-positive stromal cells. However, a subset of podoplanin-positive macrophages was increased during disease, and blockade of podoplanin in mice was sufficient to rescue disease. Our data further our understanding of disease pathogenesis demonstrating a novel role for macrophages as sensors of interferon gamma, thus illustrating an important role for the microenvironment in pathogenesis of severe aplastic anemia. Copyright © 2018, Ferrata Storti Foundation.

MHC class I D(k) expression in hematopoietic and nonhematopoietic cells confers natural killer cell resistance to murine cytomegalovirus.

PubMed

Xie, Xuefang; Stadnisky, Michael D; Coats, Ebony R; Ahmed Rahim, Mir Munir; Lundgren, Alyssa; Xu, Wenhao; Makrigiannis, Andrew P; Brown, Michael G

2010-05-11

NK cell-mediated murine cytomegalovirus (MCMV) resistance (Cmv(r)) is under H-2(k) control in MA/My mice, but the underlying gene(s) is unclear. Prior genetic analysis mapped Cmv(r) to the MHC class I (MHC-I) D(k) gene interval. Because NK cell receptors are licensed by and responsive to MHC class I molecules, D(k) itself is a candidate gene. A 10-kb genomic D(k) fragment was subcloned and microinjected into MCMV-susceptible (Cmv(s)) (MA/My.L-H2(b) x C57L)F(1) or (B6 x DBA/2)F(2) embryos. Transgenic founders, which are competent for D(k) expression and germline transgene transmission, were identified and further backcrossed to MA/My.L-H2(b) or C57L mice. Remarkably, D(k) expression delivered NK-mediated resistance in either genetic background. Further, NK cells with cognate inhibitory Ly49G receptors for self-MHC-I D(k) were licensed and critical in protection against MCMV infection. In radiation bone marrow chimeras, NK resistance was significantly diminished when MHC-I D(k) expression was restricted to only hematopoietic or nonhematopoietic cells. Thus, MHC-I D(k) is the H-2(k)-linked Cmv(r) locus; these findings suggest a role for NK cell interaction with D(k)-bearing hematopoietic and nonhematopoietic cells to shape NK-mediated virus immunity.

HOXB4 overexpression mediates very rapid stem cell regeneration and competitive hematopoietic repopulation.

PubMed

Antonchuk, J; Sauvageau, G; Humphries, R K

2001-09-01

Hox transcription factors have emerged as important regulators of hematopoiesis. In particular, we have shown that overexpression of HOXB4 in mouse bone marrow can greatly enhance the level of hematopoietic stem cell (HSC) regeneration achieved at late times (> 4 months) posttransplantation. The objective of this study was to resolve if HOXB4 increases the rate and/or duration of HSC regeneration, and also to see if this enhancement was associated with impaired production of end cells or would lead to competitive reconstitution of all compartments. Retroviral vectors were generated with the GFP reporter gene +/- HOXB4 to enable the isolation and direct tracking of transduced cells in culture or following transplantation. Stem cell recovery was measured by limit dilution assay for long-term competitive repopulating cells (CRU). HOXB4-overexpressing cells have enhanced growth in vitro, as demonstrated by their rapid dominance in mixed cultures and their shortened population doubling time. Furthermore, HOXB4-transduced cells have a marked competitive repopulating advantage in vivo in both primitive and mature compartments. CRU recovery in HOXB4 recipients was extremely rapid, reaching 25% of normal by 14 days posttransplant or some 80-fold greater than control transplant recipients, and attaining normal numbers by 12 weeks. Mice transplanted with even higher numbers of HOXB4-transduced CRU regenerated up to but not beyond the normal CRU levels. HOXB4 is a potent enhancer of primitive hematopoietic cell growth, likely by increasing self-renewal probability but without impairing homeostatic control of HSC population size or the rate of production and maintenance of mature end cells.

LRF-mediated Dll4 repression in erythroblasts is necessary for hematopoietic stem cell maintenance

PubMed Central

Lee, Sung-Uk; Maeda, Manami; Ishikawa, Yuichi; Li, Sierra Min; Wilson, Anne; Jubb, Adrian M.; Sakurai, Nagisa; Weng, Lihong; Fiorini, Emma; Radtke, Freddy; Yan, Minhong; MacDonald, H. Robson; Chen, Ching-Cheng

2013-01-01

Hematopoietic stem cells (HSCs) are the most primitive cells in the hematopoietic system and are under tight regulation for self-renewal and differentiation. Notch signals are essential for the emergence of definitive hematopoiesis in mouse embryos and are critical regulators of lymphoid lineage fate determination. However, it remains unclear how Notch regulates the balance between HSC self-renewal and differentiation in the adult bone marrow (BM). Here we report a novel mechanism that prevents HSCs from undergoing premature lymphoid differentiation in BM. Using a series of in vivo mouse models and functional HSC assays, we show that leukemia/lymphoma related factor (LRF) is necessary for HSC maintenance by functioning as an erythroid-specific repressor of Delta-like 4 (Dll4) expression. Lrf deletion in erythroblasts promoted up-regulation of Dll4 in erythroblasts, sensitizing HSCs to T-cell instructive signals in the BM. Our study reveals novel cross-talk between HSCs and erythroblasts, and sheds a new light on the regulatory mechanisms regulating the balance between HSC self-renewal and differentiation. PMID:23134786

Reduced Erg Dosage Impairs Survival of Hematopoietic Stem and Progenitor Cells.

PubMed

Xie, Ying; Koch, Mia Lee; Zhang, Xin; Hamblen, Melanie J; Godinho, Frank J; Fujiwara, Yuko; Xie, Huafeng; Klusmann, Jan-Henning; Orkin, Stuart H; Li, Zhe

2017-07-01

ERG, an ETS family transcription factor frequently overexpressed in human leukemia, has been implicated as a key regulator of hematopoietic stem cells. However, how ERG controls normal hematopoiesis, particularly at the stem and progenitor cell level, and how it contributes to leukemogenesis remain incompletely understood. Using homologous recombination, we generated an Erg knockdown allele (Erg kd ) in which Erg expression can be conditionally restored by Cre recombinase. Erg kd/kd animals die at E10.5-E11.5 due to defects in endothelial and hematopoietic cells, but can be completely rescued by Tie2-Cre-mediated restoration of Erg in these cells. In Erg kd/+ mice, ∼40% reduction in Erg dosage perturbs both fetal liver and bone marrow hematopoiesis by reducing the numbers of Lin - Sca-1 + c-Kit + (LSK) hematopoietic stem and progenitor cells (HSPCs) and megakaryocytic progenitors. By genetic mosaic analysis, we find that Erg-restored HSPCs outcompete Erg kd/+ HSPCs for contribution to adult hematopoiesis in vivo. This defect is in part due to increased apoptosis of HSPCs with reduced Erg dosage, a phenotype that becomes more drastic during 5-FU-induced stress hematopoiesis. Expression analysis reveals that reduced Erg expression leads to changes in expression of a subset of ERG target genes involved in regulating survival of HSPCs, including increased expression of a pro-apoptotic regulator Bcl2l11 (Bim) and reduced expression of Jun. Collectively, our data demonstrate that ERG controls survival of HSPCs, a property that may be used by leukemic cells. Stem Cells 2017;35:1773-1785. © 2017 AlphaMed Press.

Fanconi Anemia Mesenchymal Stromal Cells-Derived Glycerophospholipids Skew Hematopoietic Stem Cell Differentiation Through Toll-Like Receptor Signaling.

PubMed

Amarachintha, Surya; Sertorio, Mathieu; Wilson, Andrew; Li, Xiaoli; Pang, Qishen

2015-11-01

Fanconi anemia (FA) patients develop bone marrow (BM) failure or leukemia. One standard care for these devastating complications is hematopoietic stem cell transplantation. We identified a group of mesenchymal stromal cells (MSCs)-derived metabolites, glycerophospholipids, and their endogenous inhibitor, 5-(tetradecyloxy)-2-furoic acid (TOFA), as regulators of donor hematopoietic stem and progenitor cells. We provided two pieces of evidence that TOFA could improve hematopoiesis-supporting function of FA MSCs: (a) limiting-dilution cobblestone area-forming cell assay revealed that TOFA significantly increased cobblestone colonies in Fanca-/- or Fancd2-/- cocultures compared to untreated cocultures. (b) Competitive repopulating assay using output cells collected from cocultures showed that TOFA greatly alleviated the abnormal expansion of the donor myeloid (CD45.2+Gr1+Mac1+) compartment in both peripheral blood and BM of recipient mice transplanted with cells from Fanca-/- or Fancd2-/- cocultures. Furthermore, mechanistic studies identified Tlr4 signaling as the responsible pathway mediating the effect of glycerophospholipids. Thus, targeting glycerophospholipid biosynthesis in FA MSCs could be a therapeutic strategy to improve hematopoiesis and stem cell transplantation. © 2015 AlphaMed Press.

Brief Reports: Nfix Promotes Survival of Immature Hematopoietic Cells via Regulation of c-Mpl.

PubMed

Hall, Trent; Walker, Megan; Ganuza, Miguel; Holmfeldt, Per; Bordas, Marie; Kang, Guolian; Bi, Wenjian; Palmer, Lance E; Finkelstein, David; McKinney-Freeman, Shannon

2018-02-12

Hematopoietic stem and progenitor cells (HSPCs) are necessary for life-long blood production and replenishment of the hematopoietic system during stress. We recently reported that nuclear factor I/X (Nfix) promotes HSPC survival post-transplant. Here, we report that ectopic expression of Nfix in primary mouse HSPCs extends their ex vivo culture from about 20 to 40 days. HSPCs overexpressing Nfix display hypersensitivity to supportive cytokines and reduced apoptosis when subjected to cytokine deprivation relative to controls. Ectopic Nfix resulted in elevated levels of c-Mpl transcripts and cell surface protein on primary murine HSPCs as well as increased phosphorylation of STAT5, which is known to be activated down-stream of c-MPL. Blocking c-MPL signaling by removal of thrombopoietin or addition of a c-MPL neutralizing antibody negated the antiapoptotic effect of Nfix overexpression on cultured HSPCs. Furthermore, NFIX was capable of binding to and transcriptionally activating a proximal c-Mpl promoter fragment. In sum, these data suggest that NFIX-mediated upregulation of c-Mpl transcription can protect primitive hematopoietic cells from stress ex vivo. Stem Cells 2018. © AlphaMed Press 2018.

Pericarditis mediated by respiratory syncytial virus in a hematopoietic stem cell transplant patient.

PubMed

Rubach, M P; Pavlisko, E N; Perfect, J R

2013-08-01

We describe a case of pericarditis and large pericardial effusion in a 63-year-old African-American man undergoing autologous hematopoietic stem cell transplant for multiple myeloma. Pericardial tissue biopsy demonstrated fibrinous pericarditis, and immunohistochemistry stains were positive for respiratory syncytial virus. The patient improved with oral ribavirin and intravenous immune globulin infusions. © 2013 John Wiley & Sons A/S.

Deletion of the Imprinted Gene Grb10 Promotes Hematopoietic Stem Cell Self-Renewal and Regeneration.

PubMed

Yan, Xiao; Himburg, Heather A; Pohl, Katherine; Quarmyne, Mamle; Tran, Evelyn; Zhang, Yurun; Fang, Tiancheng; Kan, Jenny; Chao, Nelson J; Zhao, Liman; Doan, Phuong L; Chute, John P

2016-11-01

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 (Grb10 m/+ mice) substantially increased HSC long-term repopulating capacity, as compared to that of Grb10 +/+ mice. After total body irradiation (TBI), Grb10 m/+ 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. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

FGF7 supports hematopoietic stem and progenitor cells and niche-dependent myeloblastoma cells via autocrine action on bone marrow stromal cells in vitro

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ishino, Ruri; Minami, Kaori; Tanaka, Satowa

2013-10-11

Highlights: •FGF7 is downregulated in MED1-deficient mesenchymal cells. •FGF7 produced by mesenchymal stromal cells is a novel hematopoietic niche molecule. •FGF7 supports hematopoietic progenitor cells and niche-dependent leukemia cells. •FGF7 activates FGFR2IIIb of bone marrow stromal cells in an autocrine manner. •FGF7 indirectly acts on hematopoietic cells lacking FGFR2IIIb via stromal cells. -- Abstract: FGF1 and FGF2 support hematopoietic stem and progenitor cells (HSPCs) under stress conditions. In this study, we show that fibroblast growth factor (FGF7) may be a novel niche factor for HSPC support and leukemic growth. FGF7 expression was attenuated in mouse embryonic fibroblasts (MEFs) deficient formore » the MED1 subunit of the Mediator transcriptional coregulator complex. When normal mouse bone marrow (BM) cells were cocultured with Med1{sup +/+} MEFs or BM stromal cells in the presence of anti-FGF7 antibody, the growth of BM cells and the number of long-time culture-initiating cells (LTC-ICs) decreased significantly. Anti-FGF7 antibody also attenuated the proliferation and cobblestone formation of MB1 stromal cell-dependent myeloblastoma cells. The addition of recombinant FGF7 to the coculture of BM cells and Med1{sup −/−} MEFs increased BM cells and LTC-ICs. FGF7 and its cognate receptor, FGFR2IIIb, were undetectable in BM cells, but MEFs and BM stromal cells expressed both. FGF7 activated downstream targets of FGFR2IIIb in Med1{sup +/+} and Med1{sup −/−} MEFs and BM stromal cells. Taken together, we propose that FGF7 supports HSPCs and leukemia-initiating cells indirectly via FGFR2IIIb expressed on stromal cells.« less

Transformation of human mesenchymal cells and skin fibroblasts into hematopoietic cells.

PubMed

Harris, David M; Hazan-Haley, Inbal; Coombes, Kevin; Bueso-Ramos, Carlos; Liu, Jie; Liu, Zhiming; Li, Ping; Ravoori, Murali; Abruzzo, Lynne; Han, Lin; Singh, Sheela; Sun, Michael; Kundra, Vikas; Kurzrock, Razelle; Estrov, Zeev

2011-01-01

Patients with prolonged myelosuppression require frequent platelet and occasional granulocyte transfusions. Multi-donor transfusions induce alloimmunization, thereby increasing morbidity and mortality. Therefore, an autologous or HLA-matched allogeneic source of platelets and granulocytes is needed. To determine whether nonhematopoietic cells can be reprogrammed into hematopoietic cells, human mesenchymal stromal cells (MSCs) and skin fibroblasts were incubated with the demethylating agent 5-azacytidine (Aza) and the growth factors (GF) granulocyte-macrophage colony-stimulating factor and stem cell factor. This treatment transformed MSCs to round, non-adherent cells expressing T-, B-, myeloid-, or stem/progenitor-cell markers. The transformed cells engrafted as hematopoietic cells in bone marrow of immunodeficient mice. DNA methylation and mRNA array analysis suggested that Aza and GF treatment demethylated and activated HOXB genes. Indeed, transfection of MSCs or skin fibroblasts with HOXB4, HOXB5, and HOXB2 genes transformed them into hematopoietic cells. Further studies are needed to determine whether transformed MSCs or skin fibroblasts are suitable for therapy.

Transformation of Human Mesenchymal Cells and Skin Fibroblasts into Hematopoietic Cells

PubMed Central

Harris, David M.; Hazan-Haley, Inbal; Coombes, Kevin; Bueso-Ramos, Carlos; Liu, Jie; Liu, Zhiming; Li, Ping; Ravoori, Murali; Abruzzo, Lynne; Han, Lin; Singh, Sheela; Sun, Michael; Kundra, Vikas; Kurzrock, Razelle; Estrov, Zeev

2011-01-01

Patients with prolonged myelosuppression require frequent platelet and occasional granulocyte transfusions. Multi-donor transfusions induce alloimmunization, thereby increasing morbidity and mortality. Therefore, an autologous or HLA-matched allogeneic source of platelets and granulocytes is needed. To determine whether nonhematopoietic cells can be reprogrammed into hematopoietic cells, human mesenchymal stromal cells (MSCs) and skin fibroblasts were incubated with the demethylating agent 5-azacytidine (Aza) and the growth factors (GF) granulocyte-macrophage colony-stimulating factor and stem cell factor. This treatment transformed MSCs to round, non-adherent cells expressing T-, B-, myeloid-, or stem/progenitor-cell markers. The transformed cells engrafted as hematopoietic cells in bone marrow of immunodeficient mice. DNA methylation and mRNA array analysis suggested that Aza and GF treatment demethylated and activated HOXB genes. Indeed, transfection of MSCs or skin fibroblasts with HOXB4, HOXB5, and HOXB2 genes transformed them into hematopoietic cells. Further studies are needed to determine whether transformed MSCs or skin fibroblasts are suitable for therapy. PMID:21731684

Pluripotent stem cell models of Shwachman-Diamond syndrome reveal a common mechanism for pancreatic and hematopoietic dysfunction

PubMed Central

Tulpule, Asmin; Kelley, James M.; Lensch, M. William; McPherson, Jade; Park, In Hyun; Hartung, Odelya; Nakamura, Tomoka; Schlaeger, Thorsten M.; Shimamura, Akiko; Daley, George Q.

2013-01-01

Summary Shwachman-Diamond syndrome (SDS), a rare autosomal recessive disorder characterized by exocrine pancreatic insufficiency and hematopoietic dysfunction, is caused by mutations in the Shwachman-Bodian-Diamond syndrome (SBDS) gene. We created human pluripotent stem cell models of SDS by knock-down of SBDS in human embryonic stem cells (hESCs) and generation of induced pluripotent stem cell (iPSC) lines from two SDS patients. SBDS-deficient hESCs and iPSCs manifest deficits in exocrine pancreatic and hematopoietic differentiation in vitro, enhanced apoptosis and elevated protease levels in culture supernatants, which could be reversed by restoring SBDS protein expression through transgene rescue or by supplementing culture media with protease inhibitors. Protease-mediated auto-digestion provides a mechanistic link between the pancreatic and hematopoietic phenotypes in SDS, highlighting the utility of hESCs and iPSCs in obtaining novel insights into human disease. PMID:23602541

Neural Crossroads in the Hematopoietic Stem Cell Niche.

PubMed

Agarwala, Sobhika; Tamplin, Owen J

2018-05-29

The hematopoietic stem cell (HSC) niche supports steady-state hematopoiesis and responds to changing needs during stress and disease. The nervous system is an important regulator of the niche, and its influence is established early in development when stem cells are specified. Most research has focused on direct innervation of the niche, however recent findings show there are different modes of neural control, including globally by the central nervous system (CNS) and hormone release, locally by neural crest-derived mesenchymal stem cells, and intrinsically by hematopoietic cells that express neural receptors and neurotransmitters. Dysregulation between neural and hematopoietic systems can contribute to disease, however new therapeutic opportunities may be found among neuroregulator drugs repurposed to support hematopoiesis. Copyright © 2018 Elsevier Ltd. All rights reserved.

Hematopoietic Acute Radiation Syndrome (Bone marrow syndrome, Aplastic Anemia): Molecular Mechanisms of Radiation Toxicity.

NASA Astrophysics Data System (ADS)

Popov, Dmitri

Key Words: Aplastic Anemia (AA), Pluripotential Stem Cells (PSC) Introduction: Aplastic Anemia (AA) is a disorder of the pluripotential stem cells involve a decrease in the number of cells of myeloid, erythroid and megakaryotic lineage [Segel et al. 2000 ]. The etiology of AA include idiopathic cases and secondary aplastic anemia after exposure to drugs, toxins, chemicals, viral infections, lympho-proliferative diseases, radiation, genetic causes, myelodisplastic syndromes and hypoplastic anemias, thymomas, lymphomas. [Brodskyet al. 2005.,Modan et al. 1975., Szklo et al. 1975]. Hematopoietic Acute Radiation Syndrome (or Bone marrow syndrome, or Radiation-Acquired Aplastic Anemia) is the acute toxic syndrome which usually occurs with a dose of irradiation between 0.7 and 10 Gy (70- 1000 rads), depending on the species irradiated. [Waselenko et al., 2004]. The etiology of bone morrow damage from high-level radiation exposure results depends on the radiosensitivity of certain bone marrow cell lines. [Waselenko et al. 2004] Aplastic anemia after radiation exposure is a clinical syndrome that results from a marked disorder of bone marrow blood cell production. [Waselenko et al. 2004] Radiation hematotoxicity is mediated via genotoxic and other specific toxic mechanisms, leading to aplasia, cell apoptosis or necrosis, initiation via genetic mechanisms of clonal disorders, in cases such as the acute radiation-acquired form of AA. AA results from radiation injury to pluripotential and multipotential stem cells in the bone marrow. The clinical signs displayed in reticulocytopenia, anemia, granulocytopenia, monocytopenia, and thrombocytopenia. The number of marrow CD34+ cells (multipotential hematopoietic progenitors) and their derivative colony-forming unit{granulocyte-macrophage (CFU-GM) and burst forming unit {erythroid (BFU{E) are reduced markedly in patients with AA. [Guinan 2011, Brodski et al. 2005, Beutler et al.,2000] Cells expressing CD34 (CD34+ cell) are normally

Parasitic Infections in Hematopoietic Stem Cell Transplantation

PubMed Central

Jarque, Isidro; Salavert, Miguel; Pemán, Javier

2016-01-01

Parasitic infections are rarely documented in hematopoietic stem cell transplant recipients. However they may be responsible for fatal complications that are only diagnosed at autopsy. Increased awareness of the possibility of parasitic diseases both in autologous and allogeneic stem cell transplant patients is relevant not only for implementing preventive measures but also for performing an early diagnosis and starting appropriate therapy for these unrecognized but fatal infectious complications in hematopoietic transplant recipients. In this review, we will focus on parasitic diseases occurring in this population especially those with major clinical relevance including toxoplasmosis, American trypanosomiasis, leishmaniasis, malaria, and strongyloidiasis, among others, highlighting the diagnosis and management in hematopoietic transplant recipients. PMID:27413527

The biochemistry of hematopoietic stem cell development.

PubMed

Kaimakis, P; Crisan, M; Dzierzak, E

2013-02-01

The cornerstone of the adult hematopoietic system and clinical treatments for blood-related disease is the cohort of hematopoietic stem cells (HSC) that is harbored in the adult bone marrow microenvironment. Interestingly, this cohort of HSCs is generated only during a short window of developmental time. In mammalian embryos, hematopoietic progenitor and HSC generation occurs within several extra- and intraembryonic microenvironments, most notably from 'hemogenic' endothelial cells lining the major vasculature. HSCs are made through a remarkable transdifferentiation of endothelial cells to a hematopoietic fate that is long-lived and self-renewable. Recent studies are beginning to provide an understanding of the biochemical signaling pathways and transcription factors/complexes that promote their generation. The focus of this review is on the biochemistry behind the generation of these potent long-lived self-renewing stem cells of the blood system. Both the intrinsic (master transcription factors) and extrinsic regulators (morphogens and growth factors) that affect the generation, maintenance and expansion of HSCs in the embryo will be discussed. The generation of HSCs is a stepwise process involving many developmental signaling pathways, morphogens and cytokines. Pivotal hematopoietic transcription factors are required for their generation. Interestingly, whereas these factors are necessary for HSC generation, their expression in adult bone marrow HSCs is oftentimes not required. Thus, the biochemistry and molecular regulation of HSC development in the embryo are overlapping, but differ significantly from the regulation of HSCs in the adult. HSC numbers for clinical use are limiting, and despite much research into the molecular basis of HSC regulation in the adult bone marrow, no panel of growth factors, interleukins and/or morphogens has been found to sufficiently increase the number of these important stem cells. An understanding of the biochemistry of HSC

Selective inhibition of receptor activator of NF-κB ligand (RANKL) in hematopoietic cells improves outcome after experimental myocardial infarction.

PubMed

Slavic, Svetlana; Andrukhova, Olena; Ford, Kristopher; Handschuh, Stephan; Latic, Nejla; Reichart, Ursula; Sasgary, Soleman; Bergow, Claudia; Hofbauer, Lorenz C; Kostenuik, Paul J; Erben, Reinhold G

2018-05-08

The RANK (receptor activator of nuclear factor κB)/RANKL (RANK ligand)/OPG (osteoprotegerin) axis is activated after myocardial infarction (MI), but its pathophysiological role is not well understood. Here, we investigated how global and cell compartment-selective inhibition of RANKL affects cardiac function and remodeling after MI in mice. Global RANKL inhibition was achieved by treatment of human RANKL knock-in (huRANKL-KI) mice with the monoclonal antibody AMG161. huRANKL-KI mice express a chimeric RANKL protein wherein part of the RANKL molecule is humanized. AMG161 inhibits human and chimeric but not murine RANKL. To dissect the pathophysiological role of RANKL derived from hematopoietic and mesenchymal cells, we selectively exchanged the hematopoietic cell compartment by lethal irradiation and across-genotype bone marrow transplantation between wild-type and huRANKL-KI mice, exploiting the specificity of AMG161. After permanent coronary artery ligation, mice were injected with AMG161 or an isotype control antibody over 4 weeks post-MI. MI increased RANKL expression mainly in cardiomyocytes and scar-infiltrating cells 4 weeks after MI. Only inhibition of RANKL derived from hematopoietic cellular sources, but not global or mesenchymal RANKL inhibition, improved post-infarct survival and cardiac function. Mechanistically, hematopoietic RANKL inhibition reduced expression of the pro-inflammatory cytokine IL-1ß in the cardiac cellular infiltrate. In conclusion, inhibition of RANKL derived from hematopoietic cellular sources is beneficial to maintain post-ischemic cardiac function by reduction of pro-inflammatory cytokine production. Experimental myocardial infarction (MI) augments cardiac RANKL expression in mice. RANKL expression is increased in cardiomyocytes and scar-infiltrating cells after MI. Global or mesenchymal cell RANKL inhibition has no influence on cardiac function after MI. Inhibition of RANKL derived from hematopoietic cells improves heart

The Total Body Irradiation Schedule Affects Acute Leukemia Relapse After Matched T Cell–Depleted Hematopoietic Stem Cell Transplantation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Aristei, Cynthia, E-mail: cynthia.aristei@unipg.it; Carotti, Alessandra; Palazzari, Elisa

Purpose: We sought to determine whether the total body irradiation (TBI) schedule affected outcome in patients with acute leukemia in complete remission who received T cell–depleted allogeneic hematopoietic stem cell transplantation from HLA identical siblings. Methods and Materials: The study recruited 55 patients (median age, 48 years; age range, 20-66 years; 30 men and 25 women; 34 with acute myeloid leukemia and 21 with acute lymphoid leukemia). Hyperfractionated TBI (HTBI) (1.2 Gy thrice daily for 4 days [for a total dose of 14.4 Gy] from day −12 to day −9) was administered to 29 patients. Single-dose TBI (STBI) (8 Gy, at a median dose rate of 10.7 cGy/minmore » on day −9) was given to 26 patients. Results: All patients achieved primary, sustained engraftment with full donor-type chimerism. At 10 years, the overall cumulative incidence of transplant-related mortality was 11% (SE, ±0.1%). It was 7% (SE, ±0.2%) after HTBI and 15% (SE, ±0.5%) after STBI (P=.3). The overall cumulative incidence of relapse was 33% (SE, ±0.5). It was 13% (SE, ±0.5%) after HTBI and 46% (SE, ±1%) after STBI (P=.02). The overall probability of disease-free survival (DFS) was 59% (SE, ±7%). It was 67% (SE, ±0.84%) after HTBI and 37% (SE, ±1.4%) after STBI (P=.01). Multivariate analyses showed the TBI schedule was the only risk factor that significantly affected relapse and DFS (P=.01 and P=.03, respectively). Conclusions: In patients with acute leukemia, HTBI is more efficacious than STBI in eradicating minimal residual disease after HLA-matched T cell–depleted hematopoietic stem cell transplantation, thus affecting DFS.« less

Failure in activation of the canonical NF-κB pathway by human T-cell leukemia virus type 1 Tax in non-hematopoietic cell lines

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mizukoshi, Terumi; Komori, Hideyuki; Mizuguchi, Mariko

2013-09-01

Human T-cell leukemia virus type 1 (HTLV-1) Tax (Tax1) plays crucial roles in leukemogenesis in part through activation of NF-κB. In this study, we demonstrated that Tax1 activated an NF-κB binding (gpκB) site of the gp34/OX40 ligand gene in a cell type-dependent manner. Our examination showed that the gpκΒ site and authentic NF-κB (IgκB) site were activated by Tax1 in hematopoietic cell lines. Non-hematopoietic cell lines including hepatoma and fibroblast cell lines were not permissive to Tax1-mediated activation of the gpκB site, while the IgκB site was activated in those cells in association with binding of RelB. However RelA bindingmore » was not observed in the gpκB and IgκB sites. Our results suggest that HTLV-1 Tax1 fails to activate the canonical pathway of NF-κB in non-hematopoietic cell lines. Cell type-dependent activation of NF-κB by Tax1 could be associated with pathogenesis by HTLV-1 infection. - Highlights: • HTLV-1 Tax1 does not activate RelA of NF-κB in non-hematopoietic cell lines. • Tax1 activates the NF-κB non-canonical pathway in non-hematopoietic cell lines. • Tax1 does not induce RelA nuclear translocation in those cell lines, unlike TNFα. • The OX40L promoter κB site is activated by ectopic, but not endogenous, RelA.« less

Collection and use of circulating hematopoietic progenitor cells.

PubMed

Lee, J H; Klein, H G

1995-02-01

Although lymphocytes and monocytes are becoming increasingly important in transfusion therapy, peripheral stem cells have been responsible for the recent explosive interest in harvesting mononuclear cells from the peripheral circulation. Despite their low concentration in peripheral blood and the consequent difficulty in cell collection, circulating hematopoietic progenitor cells are collected and used almost routinely. These mononuclear cells, possessing the capacity for hematopoietic reconstitution and the potential for definitive therapy of a variety of disorders, have been the focus of recent intense interest in transfusion medicine.

Correction of glucocerebrosidase deficiency after retroviral-mediated gene transfer into hematopoietic progenitor cells from patients with Gaucher disease

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fink, J.K.; Correll, P.H.; Perry, L.K.

1990-03-01

Retroviral gene transfer has been used successfully to correct the glucocerebrosidase (GCase) deficiency in primary hematopoietic cells from patients with Gaucher disease. For this model of somatic gene therapy, the authors developed a high-titer, amphotropic retroviral vector designated NTG in which the human GCase gene was driven by the mutant polyoma virus enhancer/herpesvirus thymidine kinase gene (tk) promoter (Py{sup +}/Htk). NTG normalized GCase activity in transduced Gaucher fibroblasts and efficiently infected human monocytic and erythroleukemic cell lines. RNA blot-hybridization (Northern blot) analysis of these hemaptopoietic cell lines showed unexpectedly high-level expression from the Moloney murine leukemia virus long terminal repeatmore » (Mo-MLV LTR) and levels of Py{sup +}/Htk enhancer/promoter-initiated human GCase RNA that approximated endogenous GCase RNA levels. Furthermore, NTG efficiently infected human hematopoietic progenitor cells. Detection of the provirus in approximately one-third of NTG-infected progenitor colonies that had not been selected in G418-containing medium indicates that relative resistance to G418 underestimated the actual gene transfer efficiency. Northern blot analysis of NTG-infected, progenitor-derived cells showed expression from both the Mo-MLV LTR and the Py{sup +}/Htk enhancer/promoter. NTG-transduced hematopoietic progenitor cells from patients with Gaucher disease generated progeny in which GCase activity has been normalized.« less

Free iron catalyzes oxidative damage to hematopoietic cells/mesenchymal stem cells in vitro and suppresses hematopoiesis in iron overload patients.

PubMed

Lu, Wenyi; Zhao, Mingfeng; Rajbhandary, Sajin; Xie, Fang; Chai, Xiao; Mu, Juan; Meng, Juanxia; Liu, Yongjun; Jiang, Yan; Xu, Xinnv; Meng, Aimin

2013-09-01

Transfusional iron overload is of major concern in hematological disease. Iron-overload-related dyserythropoiesis and reactive oxygen species (ROS)-related damage to hematopoietic stem cell (HSC) function are major setbacks in treatment for such disorders. We therefore aim to investigate the effect of iron overload on hematopoiesis in the patients and explore the role of ROS in iron-induced oxidative damage in hematopoietic cells and microenvironment in vitro. The hematopoietic colony-forming capacity and ROS level of bone marrow cells were tested before and after iron chelation therapy. In vitro, we first established an iron overload model of bone marrow mononuclear cells (BMMNC) and umbilical cord-derived mesenchymal stem cells (UC-MSC). ROS level, cell cycle, and apoptosis were measured by FACS. Function of cells was individually studied by Colony-forming cell (CFC) assay and co-culture system. Finally, ROS-related signaling pathway was also detected by Western blot. After administering deferoxamine (DFO), reduced blood transfusion, increased neutrophil, increased platelet, and improved pancytopenia were observed in 76.9%, 46.2%, 26.9%, and 15.4% of the patients, respectively. Furthermore, the colony-forming capacity of BMMNC from iron overload patient was deficient, and ROS level was higher, which were partially recovered following iron chelation therapy. In vitro, exposure of BMMNC to ferric ammonium citrate (FAC) for 24 h decreased the ratio of CD34(+) cell from 0.91 ± 0.12% to 0.39 ± 0.07%. Excessive iron could also induce apoptosis, arrest cell cycle, and decrease function of BMMNC and UC-MSC, which was accompanied by increased ROS level and stimulated p38MAPK, p53 signaling pathway. More importantly, N-acetyl-L-cysteine (NAC) or DFO could partially attenuate cell injury and inhibit the signaling pathway induced by excessive iron. Our study shows that iron overload injures the hematopoiesis by damaging hematopoietic cell and hematopoietic

Single low-dose rHuIL-12 safely triggers multilineage hematopoietic and immune-mediated effects

PubMed Central

2014-01-01

Background Recombinant human interleukin 12 (rHuIL-12) regulates hematopoiesis and cell-mediated immunity. Based on these hematopoietic and immunomodulatory activities, a recombinant human IL-12 (rHuIL-12) is now under development to address the unmet need for a medical countermeasure against the hematopoietic syndrome of the acute radiation syndrome (HSARS) that occurs in individuals exposed to lethal radiation, and also to serve as adjuvant therapy that could provide dual hematopoietic and immunotherapeutic benefits in patients with cancer receiving chemotherapy. We sought to demonstrate in healthy subjects the safety of rHuIL-12 at single, low doses that are appropriate for use as a medical countermeasure for humans exposed to lethal radiation and as an immunomodulatory anti-cancer agent. Methods Two placebo-controlled, double-blinded studies assessed the safety, tolerability, pharmacokinetics and pharmacodynamics of rHuIL-12. The first-in-human (FIH) dose-escalation study randomized subjects to single subcutaneous injections of placebo or rHuIL-12 at 2, 5, 10, and 20 μg doses. Due to toxicity, dose was reduced to 15 μg and then to 12 μg. The phase 1b expansion study randomized subjects to the highest safe and well tolerated dose of 12 μg. Results Thirty-two subjects were enrolled in the FIH study: 4 active and 2 placebo at rHuIL-12 doses of 2, 5, 10, 12, and 15 μg; 1 active and 1 placebo at 20 μg. Sixty subjects were enrolled in the expansion study: 48 active and 12 placebo at 12 μg dose of rHuIL-12. In both studies, the most common adverse events (AEs) related to rHuIL-12 were headache, dizziness, and chills. No immunogenicity was observed. Elimination of rHuIL-12 was biphasic, suggesting significant distribution into extravascular spaces. rHuIL-12 triggered transient changes in neutrophils, platelets, reticulocytes, lymphocytes, natural killer cells, and CD34+ hematopoietic progenitor cells, and induced increases in interferon-γ and C-X-C motif

Distinct Brca1 Mutations Differentially Reduce Hematopoietic Stem Cell Function.

PubMed

Mgbemena, Victoria E; Signer, Robert A J; Wijayatunge, Ranjula; Laxson, Travis; Morrison, Sean J; Ross, Theodora S

2017-01-24

BRCA1 is a well-known DNA repair pathway component and a tissue-specific tumor suppressor. However, its role in hematopoiesis is uncertain. Here, we report that a cohort of patients heterozygous for BRCA1 mutations experienced more hematopoietic toxicity from chemotherapy than those with BRCA2 mutations. To test whether this reflects a requirement for BRCA1 in hematopoiesis, we generated mice with Brca1 mutations in hematopoietic cells. Mice homozygous for a null Brca1 mutation in the embryonic hematopoietic system (Vav1-iCre;Brca1 F22-24/F22-24 ) developed hematopoietic defects in early adulthood that included reduced hematopoietic stem cells (HSCs). Although mice homozygous for a huBRCA1 knockin allele (Brca1 BRCA1/BRCA1 ) were normal, mice with a mutant huBRCA1/5382insC allele and a null allele (Mx1-Cre;Brca1 F22-24/5382insC ) had severe hematopoietic defects marked by a complete loss of hematopoietic stem and progenitor cells. Our data show that Brca1 is necessary for HSC maintenance and normal hematopoiesis and that distinct mutations lead to different degrees of hematopoietic dysfunction. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Lentiviral gene therapy of murine hematopoietic stem cells ameliorates the Pompe disease phenotype.

PubMed

van Til, Niek P; Stok, Merel; Aerts Kaya, Fatima S F; de Waard, Monique C; Farahbakhshian, Elnaz; Visser, Trudi P; Kroos, Marian A; Jacobs, Edwin H; Willart, Monique A; van der Wegen, Pascal; Scholte, Bob J; Lambrecht, Bart N; Duncker, Dirk J; van der Ploeg, Ans T; Reuser, Arnold J J; Verstegen, Monique M; Wagemaker, Gerard

2010-07-01

Pompe disease (acid alpha-glucosidase deficiency) is a lysosomal glycogen storage disorder characterized in its most severe early-onset form by rapidly progressive muscle weakness and mortality within the first year of life due to cardiac and respiratory failure. Enzyme replacement therapy prolongs the life of affected infants and supports the condition of older children and adults but entails lifelong treatment and can be counteracted by immune responses to the recombinant enzyme. We have explored the potential of lentiviral vector-mediated expression of human acid alpha-glucosidase in hematopoietic stem cells (HSCs) in a Pompe mouse model. After mild conditioning, transplantation of genetically engineered HSCs resulted in stable chimerism of approximately 35% hematopoietic cells that overexpress acid alpha-glucosidase and in major clearance of glycogen in heart, diaphragm, spleen, and liver. Cardiac remodeling was reversed, and respiratory function, skeletal muscle strength, and motor performance improved. Overexpression of acid alpha-glucosidase did not affect overall hematopoietic cell function and led to immune tolerance as shown by challenge with the human recombinant protein. On the basis of the prominent and sustained therapeutic efficacy without adverse events in mice we conclude that ex vivo HSC gene therapy is a treatment option worthwhile to pursue.

Radiologic Differences between Bone Marrow Stromal and Hematopoietic Progenitor Cell Lines from Fanconi Anemia (Fancd2−/−) Mice

PubMed Central

Berhane, Hebist; Epperly, Michael W.; Goff, Julie; Kalash, Ronny; Cao, Shaonan; Franicola, Darcy; Zhang, Xichen; Shields, Donna; Houghton, Frank; Wang, Hong; Wipf, Peter; Parmar, Kalindi; Greenberger, Joel S.

2014-01-01

FancD2 plays a central role in the human Fanconi anemia DNA damage response (DDR) pathway. Fancd2−/− mice exhibit many features of human Fanconi anemia including cellular DNA repair defects. Whether the DNA repair defect in Fancd2−/− mice results in radiologic changes in all cell lineages is unknown. We measured stress of hematopoiesis in long-term marrow cultures and radiosensitivity in clonogenic survival curves, as well as comet tail intensity, total antioxidant stores and radiation-induced gene expression in hematopoietic progenitor compared to bone marrow stromal cell lines. We further evaluated radioprotection by a mitochondrial-targeted antioxidant GS-nitroxide, JP4-039. Hematopoiesis longevity in Fancd2−/− mouse long-term marrow cultures was diminished and bone marrow stromal cell lines were radiosensitive compared to Fancd2+/+ stromal cells (Fancd2−/− D0 = 1.4 ± 0.1 Gy, ñ = 5.0 ± 0.6 vs. Fancd2+/+ D0 = 1.6 ± 0.1 Gy, ñ = 6.7 ± 1.6), P = 0.0124 for D0 and P = 0.0023 for ñ, respectively). In contrast, Fancd2−/− IL-3-dependent hematopoietic progenitor cells were radioresistant (D0 = 1.71 ± 0.04 Gy and ñ = 5.07 ± 0.52) compared to Fancd2+/+ (D0 = 1.39 ± 0.09 Gy and ñ = 2.31 ± 0.85, P = 0.001 for D0). CFU-GM from freshly explanted Fancd2−/− marrow was also radioresistant. Consistent with radiosensitivity, irradiated Fancd2−/− stromal cells had higher DNA damage by comet tail intensity assay compared to Fancd2+/+ cells (P < 0.0001), slower DNA damage recovery, lower baseline total antioxidant capacity, enhanced radiation-induced depletion of antioxidants, and increased CDKN1A-p21 gene transcripts and protein. Consistent with radioresistance, Fancd2−/− IL-3-dependent hematopoietic cells had higher baseline and post irradiation total antioxidant capacity. While, there was no detectable alteration of radiation-induced cell cycle arrest with Fancd2−/− stromal cells, hematopoietic progenitor cells showed reduced G2/M

Metabolic syndrome in long-term survivors of childhood acute leukemia treated without hematopoietic stem cell transplantation: an L.E.A. study.

PubMed

Saultier, Paul; Auquier, Pascal; Bertrand, Yves; Vercasson, Camille; Oudin, Claire; Contet, Audrey; Plantaz, Dominique; Poirée, Marilyne; Ducassou, Stéphane; Kanold, Justyna; Tabone, Marie-Dominique; Dalle, Jean-Hugues; Lutz, Patrick; Gandemer, Virginie; Sirvent, Nicolas; Thouvenin, Sandrine; Berbis, Julie; Chambost, Hervé; Baruchel, André; Leverger, Guy; Michel, Gérard

2016-12-01

Cardiovascular conditions are serious long-term complications of childhood acute leukemia. However, few studies have investigated the risk of metabolic syndrome, a known predictor of cardiovascular disease, in patients treated without hematopoietic stem cell transplantation. We describe the overall and age-specific prevalence, and the risk factors for metabolic syndrome and its components in the L.E.A. (Leucémie de l'Enfant et de l'Adolescent) French cohort of childhood acute leukemia survivors treated without hematopoietic stem cell transplantation. The study included 650 adult patients (mean age at evaluation: 24.2 years; mean follow-up after leukemia diagnosis: 16.0 years). The prevalence of metabolic syndrome was 6.9% (95% CI 5.1-9.2). The age-specific cumulative prevalence at 20, 25, 30 and 35 years of age was 1.3%, 6.1%, 10.8% and 22.4%, respectively. The prevalence of decreased high-density lipoprotein cholesterol, increased triglycerides, increased fasting glucose, increased blood pressure and increased abdominal circumference was 26.8%, 11.7%, 5.8%, 36.7% and 16.7%, respectively. Risk factors significantly associated with metabolic syndrome in the multivariate analysis were male sex (OR 2.64; 95% CI 1.32-5.29), age at last evaluation (OR 1.10; 95% CI 1.04-1.17) and body mass index at diagnosis (OR 1.15; 95% CI 1.01-1.32). The cumulative steroid dose was not a significant risk factor. Irradiated and non-irradiated patients exhibited different patterns of metabolic abnormalities, with more frequent abdominal obesity in irradiated patients and more frequent hypertension in non-irradiated patients. Survivors of childhood acute leukemia are at risk of metabolic syndrome, even when treated without hematopoietic stem cell transplantation or central nervous system irradiation. A preventive approach with regular screening for cardiovascular risk factors is recommended. clinicaltrials.gov identifier:01756599. Copyright© Ferrata Storti Foundation.

[Analysis of thyroid lesions in childhood recipients after hematopoietic stem cell transplantation].

PubMed

Maeda, Naoko; Hamajima, Takashi; Yambe, Yuko; Sekimizu, Masahiro; Horibe, Keizo

2013-03-01

We performed a physical examination and ultrasonography of the thyroid gland in 24 patients who had received hematopoietic stem cell transplantation with a total-body irradiation (TBI)-containing regimen during childhood. When ultrasonography revealed thyroid nodules larger than 1 cm in diameter, fine-needle aspiration biopsies were performed. Of 5 patients with palpable masses and thyroid nodules larger than 1 cm, adenomatous goiter was diagnosed in 4 cases and thyroid cancer in 1. Of the remaining 19 patients in whom no palpable mass was detected in the physical examination, 5 had thyroid nodules (including 1 adenomatous goiter), 6 had cystic lesions, and 8 exhibited no abnormalities on ultrasonography. No significant differences in sex, age at transplantation, interval between transplantation and evaluation, primary disease, preconditioning regimen, status at transplantation, stem cell source, chronic graft-versus-host disease, hypogonadism, or hypothyroidism were observed between patients with and without nodules. Individuals who received hematopoietic stem cell transplantation with a TBI-containing regimen are at risk of secondary thyroid cancer due to radiotherapy and require regular clinical evaluations of the thyroid gland by palpation, and ultrasonography should be incorporated into these checkups.

Physiological Srsf2 P95H expression causes impaired hematopoietic stem cell functions and aberrant RNA splicing in mice.

PubMed

Kon, Ayana; Yamazaki, Satoshi; Nannya, Yasuhito; Kataoka, Keisuke; Ota, Yasunori; Nakagawa, Masahiro Marshall; Yoshida, Kenichi; Shiozawa, Yusuke; Morita, Maiko; Yoshizato, Tetsuichi; Sanada, Masashi; Nakayama, Manabu; Koseki, Haruhiko; Nakauchi, Hiromitsu; Ogawa, Seishi

2018-02-08

Splicing factor mutations are characteristic of myelodysplastic syndromes (MDS) and related myeloid neoplasms and implicated in their pathogenesis, but their roles in the development of MDS have not been fully elucidated. In the present study, we investigated the consequence of mutant Srsf2 expression using newly generated Vav1-Cre -mediated conditional knockin mice. Mice carrying a heterozygous Srsf2 P95H mutation showed significantly reduced numbers of hematopoietic stem and progenitor cells (HSPCs) and differentiation defects both in the steady-state condition and transplantation settings. Srsf2 -mutated hematopoietic stem cells (HSCs) showed impaired long-term reconstitution compared with control mice in competitive repopulation assays. Although the Srsf2 mutant mice did not develop MDS under the steady-state condition, when their stem cells were transplanted into lethally irradiated mice, the recipients developed anemia, leukopenia, and erythroid dysplasia, which suggests the role of replicative stress in the development of an MDS-like phenotype in Srsf2 -mutated mice. RNA sequencing of the Srsf2 -mutated HSPCs revealed a number of abnormal splicing events and differentially expressed genes, including several potential targets implicated in the pathogenesis of hematopoietic malignancies, such as Csf3r , Fyn , Gnas , Nsd1 , Hnrnpa2b1 , and Trp53bp1 Among the mutant Srsf2 -associated splicing events, most commonly observed were the enhanced inclusion and/or exclusion of cassette exons, which were caused by the altered consensus motifs for the recognition of exonic splicing enhancers. Our findings suggest that the mutant Srsf2 leads to a compromised HSC function by causing abnormal RNA splicing and expression, contributing to the deregulated hematopoiesis that recapitulates the MDS phenotypes, possibly as a result of additional genetic and/or environmental insults. © 2018 by The American Society of Hematology.

Isolation and clonal characterization of hematopoietic and liver stem cells.

PubMed

Nakauchi, Hiromitsu

2004-11-01

Prospective isolation of stem cells is essential to understanding the mechanisms that control their proliferation and differentiation. Using 9 monoclonal antibodies and fluorescence-activated cell sorting (FACS), we have succeeded in prospectively identifying hematopoietic stem cells (HSCs) in adult mouse bone marrow. Mouse HSCs were exclusively enriched in CD34 negative, c-Kit Sca-1 Lineage Marker (CD34 KSL) cells representing 0.004% of bone marrow (BM) mononuclear cells. When single CD34-KSL cells were transplanted individually into a lethally irradiated mouse, 25% of the recipient mice survived and showed long-term reconstitution of the BM, providing evidence for multipotency and a self-renewal capacity of HSCs. Using a similar approach, we also prospectively identified hepatic stem cells with multilineage differentiation potential and self-renewal capability in the c-Met CD49f c-Kit CD45 Ter119 fraction of cells isolated from day 13.5 fetal mouse liver. On cell transplantation, these cells differentiated into hepatocytes and cholangiocytes. As an alternative to the antibody based stem cell isolation, Hoechst33342 staining is useful. To understand the mechanism responsible for SP phenotype, we performed an expression cloning and identified bcrp-1/ABCG2 gene, a member of ATP binding-cassette (ABC) transporter family. Bcrp-1 is almost exclusively expressed in CD34 KSL cells among blood cells; however their expression in other tissue specific stem cells remains to be studied. With the use of FACS and monoclonal antibodies, hematopoietic and liver stem cells were prospectively isolated and characterized. HSCs could also be purified by Hoechst 33342 staining. By expression cloning, we identify bcrp-1/ABCG2 transporter as a molecule responsible for SP phenotype. Elucidation of the physiological role of bcrp-1/ABCG2 in HSCs may provide us with clues to understand the molecular mechanisms of stem cell self-renewal and differentiation.

Cutting the brakes on hematopoietic regeneration by blocking TGFβ to limit chemotherapy-induced myelosuppression

PubMed Central

Brenet, Fabienne; Scandura, Joseph M

2015-01-01

Hematopoietic stressors such as infection, bleeding, or toxic injury trigger a hematopoietic adaptation that sacrifices hematopoietic stem and progenitor cell (HSPC) quiescence to meet an urgent need for new blood cell production. Once the hematopoietic demands are adequately met, homeostasis must be restored. Transforming growth factor β (TGFβ) signaling is a central mediator mandating the return of HSPCs to quiescence after stress. Blockade of TGFβ signaling after hematopoietic stress delays the return of cycling HSPCs to quiescence and in so doing promotes hematopoietic stem cell (HSC) self-renewal and accelerates hematopoietic reconstitution. These findings open the door to new therapeutics that modulate the hematopoietic adaptation to stress. In this review, we will discuss the complex context-dependent activities of TGFβ in hematopoiesis and the potential benefits and limitations of using TGFβ pathway inhibitors to promote multilineage hematopoietic reconstitution after myelosuppressive chemotherapy. PMID:27308454

Dynamic equilibrium of heterogeneous and interconvertible multipotent hematopoietic cell subsets.

PubMed

Weston, Wendy; Zayas, Jennifer; Perez, Ruben; George, John; Jurecic, Roland

2014-06-06

Populations of hematopoietic stem cells and progenitors are quite heterogeneous and consist of multiple cell subsets with distinct phenotypic and functional characteristics. Some of these subsets also appear to be interconvertible and oscillate between functionally distinct states. The multipotent hematopoietic cell line EML has emerged as a unique model to study the heterogeneity and interconvertibility of multipotent hematopoietic cells. Here we describe extensive phenotypic and functional heterogeneity of EML cells which stems from the coexistence of multiple cell subsets. Each of these subsets is phenotypically and functionally heterogeneous, and displays distinct multilineage differentiation potential, cell cycle profile, proliferation kinetics, and expression pattern of HSC markers and some of the key lineage-associated transcription factors. Analysis of their maintenance revealed that on a population level all EML cell subsets exhibit cell-autonomous interconvertible properties, with the capacity to generate all other subsets and re-establish complete parental EML cell population. Moreover, all EML cell subsets generated during multiple cell generations maintain their distinct phenotypic and functional signatures and interconvertible properties. The model of EML cell line suggests that interconvertible multipotent hematopoietic cell subsets coexist in a homeostatically maintained dynamic equilibrium which is regulated by currently unknown cell-intrinsic mechanisms.

Fanconi anemia mesenchymal stromal cells-derived glycerophospholipids skew hematopoietic stem cell differentiation through Toll-like receptor signaling

PubMed Central

Amarachintha, Surya; Sertorio, Mathieu; Wilson, Andrew; Li, Xiaoli; Pang, Qishen

2015-01-01

Fanconi anemia (FA) patients develop bone marrow (BM) failure or leukemia. One standard care for these devastating complications is hematopoietic stem cell transplantation. We identified a group of mesenchymal stromal cells (MSCs)-derived metabolites, glycerophospholipids and their endogenous inhibitor, 5-(Tetradecyloxy)-2-furoic acid (TOFA), as regulators of donor hematopoietic stem and progenitor cells (HSPCs). We provided two pieces of evidence that TOFA could improve hematopoiesis-supporting function of FA MSCs: (1) limiting-dilution CAFC assay revealed that TOFA significantly increased cobblestone colonies in Fanca−/− or Fancd2−/− co-cultures compared to untreated co-cultures. (2) Competitive repopulating assay using output cells collected from co-cultures showed that TOFA greatly alleviated the abnormal expansion of the donor myeloid (CD45.2+Gr1+Mac1+) compartment in both peripheral blood and BM of recipient mice transplanted with cells from Fanca−/− or Fancd2−/− co-cultures. Further, mechanistic studies identified Tlr4 signaling as the responsible pathway mediating the effect of glycerophospholipids. Thus, targeting Glycerophospholipid biosynthesis in FA MSCs could be a therapeutic strategy to improve hematopoiesis and stem cell transplantation. PMID:26212365

The cell fate determinant Scribble is required for maintenance of hematopoietic stem cell function.

PubMed

Mohr, Juliane; Dash, Banaja P; Schnoeder, Tina M; Wolleschak, Denise; Herzog, Carolin; Tubio Santamaria, Nuria; Weinert, Sönke; Godavarthy, Sonika; Zanetti, Costanza; Naumann, Michael; Hartleben, Björn; Huber, Tobias B; Krause, Daniela S; Kähne, Thilo; Bullinger, Lars; Heidel, Florian H

2018-05-01

Cell fate determinants influence self-renewal potential of hematopoietic stem cells. Scribble and Llgl1 belong to the Scribble polarity complex and reveal tumor-suppressor function in drosophila. In hematopoietic cells, genetic inactivation of Llgl1 leads to expansion of the stem cell pool and increases self-renewal capacity without conferring malignant transformation. Here we show that genetic inactivation of its putative complex partner Scribble results in functional impairment of hematopoietic stem cells (HSC) over serial transplantation and during stress. Although loss of Scribble deregulates transcriptional downstream effectors involved in stem cell proliferation, cell signaling, and cell motility, these effectors do not overlap with transcriptional targets of Llgl1. Binding partner analysis of Scribble in hematopoietic cells using affinity purification followed by mass spectometry confirms its role in cell signaling and motility but not for binding to polarity modules described in drosophila. Finally, requirement of Scribble for self-renewal capacity also affects leukemia stem cell function. Thus, Scribble is a regulator of adult HSCs, essential for maintenance of HSCs during phases of cell stress.

Delta-Tocotrienol Protects Mouse and Human Hematopoietic Progenitors from Gamma-Irradiation Through Erk/mTOR Signaling

DTIC Science & Technology

2010-01-01

δ- tocotrienol protects mouse and human hematopoietic progenitors from γ-irradiation through Erk/mTOR signaling by Xiang Hong Li, Dadin Fu, Nabil H...print] Citation: Li XH, Fu D, Latif NH, Mullaney CP, Ney PH, Mog SR, Whitnall MH, Srinivasan V, and Xiao M. δ- tocotrienol protects mouse and human...2010 2. REPORT TYPE 3. DATES COVERED 00-00-2010 to 00-00-2010 4. TITLE AND SUBTITLE !- tocotrienol protects mouse and human hematopoietic

Mesenchymal stromal cell derived extracellular vesicles rescue radiation damage to murine marrow hematopoietic cells

PubMed Central

Wen, Sicheng; Dooner, Mark; Cheng, Yan; Papa, Elaine; Del Tatto, Michael; Pereira, Mandy; Deng, Yanhui; Goldberg, Laura; Aliotta, Jason; Chatterjee, Devasis; Stewart, Connor; Carpanetto, Andrea; Collino, Federica; Bruno, Stefania; Camussi, Giovanni; Quesenberry, Peter

2016-01-01

Mesenchymal stromal cells (MSC) have been shown to reverse radiation damage to marrow stem cells. We have evaluated the capacity of MSC-derived extracellular vesicles (MSC-EVs) to mitigate radiation injury to marrow stem cells at 4 hours to 7 days after irradiation. Significant restoration of marrow stem cell engraftment at 4, 24 and 168 hours post-irradiation by exposure to MSC-EVs was observed at 3 weeks to 9 months after transplant and further confirmed by secondary engraftment. Intravenous injection of MSC-EVs to 500cGy exposed mice led to partial recovery of peripheral blood counts and restoration of the engraftment of marrow. The murine hematopoietic cell line, FDC-P1 exposed to 500 cGy, showed reversal of growth inhibition, DNA damage and apoptosis on exposure to murine or human MSC-EVs. Both murine and human MSC-EVs reverse radiation damage to murine marrow cells and stimulate normal murine marrow stem cell/progenitors to proliferate. A preparation with both exosomes and microvesicles was found to be superior to either microvesicles or exosomes alone. Biologic activity was seen in freshly isolated vesicles and in vesicles stored for up to 6 months in 10% DMSO at −80°C. These studies indicate that MSC-EVs can reverse radiation damage to bone marrow stem cells. PMID:27150009

Lentiviral gene transduction of mouse and human hematopoietic stem cells.

PubMed

van Til, Niek P; Wagemaker, Gerard

2014-01-01

Lentiviral vectors can be used to genetically modify a broad range of cells. Hematopoietic stem cells (HSCs) are particularly suitable for lentiviral gene augmentation, because these cells can be enriched with relative ease from mouse bone marrow and human hematopoietic sources, and in principle require relatively limited cell numbers to completely reconstitute the hematopoietic system in vivo. Furthermore, lentiviral vectors are very efficient if pseudotyped with broad tropism envelope proteins. This chapter focuses on gene modification by the use of self-inactivating third-generation human immunodeficiency virus-derived lentiviral vectors for ex vivo HSC modification for both mouse and human application.

Lack of Phenotypical and Morphological Evidences of Endothelial to Hematopoietic Transition in the Murine Embryonic Head during Hematopoietic Stem Cell Emergence.

PubMed

Iizuka, Kazuhide; Yokomizo, Tomomasa; Watanabe, Naoki; Tanaka, Yosuke; Osato, Motomi; Takaku, Tomoiku; Komatsu, Norio

2016-01-01

During mouse ontogeny, hematopoietic cells arise from specialized endothelial cells, i.e., the hemogenic endothelium, and form clusters in the lumen of arterial vessels. Hemogenic endothelial cells have been observed in several embryonic tissues, such as the dorsal aorta, the placenta and the yolk sac. Recent work suggests that the mouse embryonic head also produces hematopoietic stem cells (HSCs)/progenitors. However, a histological basis for HSC generation in the head has not yet been determined because the hematopoietic clusters and hemogenic endothelium in the head region have not been well characterized. In this study, we used whole-mount immunostaining and 3D confocal reconstruction techniques to analyze both c-Kit+ hematopoietic clusters and Runx1+ hemogenic endothelium in the whole-head vasculature. The number of c-Kit+ hematopoietic cells was 20-fold less in the head arteries than in the dorsal aorta. In addition, apparent nascent hematopoietic cells, which are characterized by a "budding" structure and a Runx1+ hemogenic endothelium, were not observed in the head. These results suggest that head HSCs may not be or are rarely generated from the endothelium in the same manner as aortic HSCs.

Lack of Phenotypical and Morphological Evidences of Endothelial to Hematopoietic Transition in the Murine Embryonic Head during Hematopoietic Stem Cell Emergence

PubMed Central

Iizuka, Kazuhide; Yokomizo, Tomomasa; Watanabe, Naoki; Tanaka, Yosuke; Osato, Motomi; Takaku, Tomoiku; Komatsu, Norio

2016-01-01

During mouse ontogeny, hematopoietic cells arise from specialized endothelial cells, i.e., the hemogenic endothelium, and form clusters in the lumen of arterial vessels. Hemogenic endothelial cells have been observed in several embryonic tissues, such as the dorsal aorta, the placenta and the yolk sac. Recent work suggests that the mouse embryonic head also produces hematopoietic stem cells (HSCs)/progenitors. However, a histological basis for HSC generation in the head has not yet been determined because the hematopoietic clusters and hemogenic endothelium in the head region have not been well characterized. In this study, we used whole-mount immunostaining and 3D confocal reconstruction techniques to analyze both c-Kit+ hematopoietic clusters and Runx1+ hemogenic endothelium in the whole-head vasculature. The number of c-Kit+ hematopoietic cells was 20-fold less in the head arteries than in the dorsal aorta. In addition, apparent nascent hematopoietic cells, which are characterized by a “budding” structure and a Runx1+ hemogenic endothelium, were not observed in the head. These results suggest that head HSCs may not be or are rarely generated from the endothelium in the same manner as aortic HSCs. PMID:27227884

TGFβ restores hematopoietic homeostasis after myelosuppressive chemotherapy

PubMed Central

Brenet, Fabienne; Kermani, Pouneh; Spektor, Roman; Rafii, Shahin

2013-01-01

Myelosuppression is a life-threatening complication of antineoplastic therapy, but treatment is restricted to a few cytokines with unilineage hematopoietic activity. Although hematopoietic stem cells (HSCs) are predominantly quiescent during homeostasis, they are rapidly recruited into cell cycle by stresses, including myelosuppressive chemotherapy. Factors that induce HSCs to proliferate during stress have been characterized, but it is not known how HSC quiescence is then reestablished. In this study, we show that TGFβ signaling is transiently activated in hematopoietic stem and progenitor cells (HSPCs) during hematopoietic regeneration. Blockade of TGFβ signaling after chemotherapy accelerates hematopoietic reconstitution and delays the return of cycling HSCs to quiescence. In contrast, TGFβ blockade during homeostasis fails to induce cycling of HSPCs. We identified the cyclin-dependent kinase inhibitor Cdkn1c (p57) as a key downstream mediator of TGFβ during regeneration because the recovery of chimeric mice, incapable of expressing p57 in HSPCs, phenocopies blockade of TGFβ signaling after chemotherapy. This study demonstrates that context-dependent activation of TGFβ signaling is central to an unrecognized counterregulatory mechanism that promotes homeostasis once hematopoiesis has sufficiently recovered from myelosuppressive chemotherapy. These results open the door to new, potentially superior, approaches to promote multilineage hematopoietic recovery by blocking the TGFβ signaling that dampens regeneration. PMID:23440043

Circulating hematopoietic progenitor cells in patients affected by Chornobyl accident.

PubMed

Bilko, N M; Dyagil, I S; Russu, I Z; Bilko, D I

2016-12-01

High radiation sensitivity of stem cells and their ability to accumulate sublethal radiation damage provides the basis for investigation of hematopoietic progenitors using in vivo culture methodology. Unique samples of peripheral blood and bone marrow were derived from the patients affected by Chornobyl accident during liquidation campaign. To investigate functional activity of circulating hematopoietic progenitor cells from peripheral blood and bone marrow of cleanup workers in early and remote periods after the accident at Chornobyl nuclear power plant (CNPP). The assessment of the functional activity of circulating hematopoietic progenitor cells was performed in samples of peripheral blood and bone marrow of 46 cleanup workers, who were treated in the National Scientific Center for Radiation Medicine of the Academy of Medical Sciences of Ukraine alongside with 35 non radiated patients, who served as a control. Work was performed by culturing peripheral blood and bone marrow mononuclear cells in the original gel diffusion capsules, implanted into the peritoneal cavity of CBA mice. It was shown that hematopoietic progenitor cells could be identified in the peripheral blood of liquidators of CNPP accident. At the same time the number of functionally active progenitor cells of the bone marrow was significantly decreased and during the next 10 years after the accident, counts of circulating progenitor cells in the peripheral blood as well as functionally active hematopoietic cells in bone marrow returned to normal levels. It was shown that hematopoietic progenitor cells are detected not only in the bone marrow but also in the peripheral blood of liquidators as a consequence of radiation exposure associated with CNPP accident. This article is a part of a Special Issue entitled "The Chornobyl Nuclear Accident: Thirty Years After".

Small Molecule Protection of Bone Marrow Hematopoietic Stem Cells

DTIC Science & Technology

2015-10-01

several recently identified small molecules can protect hematopoietic stem cells (HSCs) from damage or killing by endogenous aldehydes . Proof-of-concept...anemia bone marrow failure CD34+ hematopoietic stem cells aldehydes formaldehyde DNA damage DNA base adduct DNA-protein crosslink mass...below. Revised Specific Aim 1: Small molecule protection of human cells from aldehyde - induced killing (in vitro studies - no mice or human subjects

Graft failure after allogeneic hematopoietic stem cell transplantation.

PubMed

Ozdemir, Zehra Narli; Civriz Bozdağ, Sinem

2018-04-18

Graft failure is a serious complication of allogeneic hematopoietic stem cell transplantation (allo-HSCT) defined as either lack of initial engraftment of donor cells (primary graft failure) or loss of donor cells after initial engraftment (secondary graft failure). Successful transplantation depends on the formation of engrafment, in which donor cells are integrated into the recipient's cell population. In this paper, we distinguish two different entities, graft failure (GF) and poor graft function (PGF), and review the current comprehensions of the interactions between the immune and hematopoietic compartments in these conditions. Factors associated with graft failure include histocompatibility locus antigen (HLA)-mismatched grafts, underlying disease, type of conditioning regimen and stem cell source employed, low stem cell dose, ex vivo T-cell depletion, major ABO incompatibility, female donor grafts for male recipients, disease status at transplantation. Although several approaches have been developed which aimed to prevent graft rejection, establish successful engraftment and treat graft failure, GF remains a major obstacle to the success of allo-HSCT. Allogeneic hematopoietic stem cell transplantation (allo-HSCT) still remains to be the curative treatment option for various non-malignant and malignant hematopoietic diseases. The outcome of allo-HSCT primarily depends on the engraftment of the graft. Graft failure (GF), is a life-threatening complication which needs the preferential therapeutic manipulation. In this paper, we focused on the definitions of graft failure / poor graft function and also we reviewed the current understanding of the pathophysiology, risk factors and treatment approaches for these entities. Copyright © 2018. Published by Elsevier Ltd.

The potential benefits of nicaraven to protect against radiation-induced injury in hematopoietic stem/progenitor cells with relative low dose exposures

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ali, Haytham; Department of Medical Physiology and Cell Biology, Qena Faculty of Medicine, South Valley University; Galal, Omima

Highlights: • Nicaraven mitigated the radiation-induced reduction of c-kit{sup +} stem cells. • Nicaraven enhanced the function of hematopoietic stem/progenitor cells. • Complex mechanisms involved in the protection of nicaraven to radiation injury. - Abstract: Nicaraven, a hydroxyl radical-specific scavenger has been demonstrated to attenuate radiation injury in hematopoietic stem cells with 5 Gy γ-ray exposures. We explored the effect and related mechanisms of nicaraven for protecting radiation injury induced by sequential exposures to a relatively lower dose γ-ray. C57BL/6 mice were given nicaraven or placebo within 30 min before exposure to 50 mGy γ-ray daily for 30 days inmore » sequences (cumulative dose of 1.5 Gy). Mice were victimized 24 h after the last radiation exposure, and the number, function and oxidative stress of hematopoietic stem cells were quantitatively estimated. We also compared the gene expression in these purified stem cells from mice received nicaraven and placebo treatment. Nicaraven increased the number of c-kit{sup +} stem/progenitor cells in bone marrow and peripheral blood, with a recovery rate around 60–90% of age-matched non-irradiated healthy mice. The potency of colony forming from hematopoietic stem/progenitor cells as indicator of function was completely protected with nicaraven treatment. Furthermore, nicaraven treatment changed the expression of many genes associated to DNA repair, inflammatory response, and immunomodulation in c-kit{sup +} stem/progenitor cells. Nicaraven effectively protected against damages of hematopoietic stem/progenitor cells induced by sequential exposures to a relatively low dose radiation, via complex mechanisms.« less

Downstream targets of HOXB4 in a cell line model of primitive hematopoietic progenitor cells.

PubMed

Lee, Han M; Zhang, Hui; Schulz, Vincent; Tuck, David P; Forget, Bernard G

2010-08-05

Enforced expression of the homeobox transcription factor HOXB4 has been shown to enhance hematopoietic stem cell self-renewal and expansion ex vivo and in vivo. To investigate the downstream targets of HOXB4 in hematopoietic progenitor cells, HOXB4 was constitutively overexpressed in the primitive hematopoietic progenitor cell line EML. Two genome-wide analytical techniques were used: RNA expression profiling using microarrays and chromatin immunoprecipitation (ChIP)-chip. RNA expression profiling revealed that 465 gene transcripts were differentially expressed in KLS (c-Kit(+), Lin(-), Sca-1(+))-EML cells that overexpressed HOXB4 (KLS-EML-HOXB4) compared with control KLS-EML cells that were transduced with vector alone. In particular, erythroid-specific gene transcripts were observed to be highly down-regulated in KLS-EML-HOXB4 cells. ChIP-chip analysis revealed that the promoter region for 1910 genes, such as CD34, Sox4, and B220, were occupied by HOXB4 in KLS-EML-HOXB4 cells. Side-by-side comparison of the ChIP-chip and RNA expression profiling datasets provided correlative information and identified Gp49a and Laptm4b as candidate "stemness-related" genes. Both genes were highly ranked in both dataset lists and have been previously shown to be preferentially expressed in hematopoietic stem cells and down-regulated in mature hematopoietic cells, thus making them attractive candidates for future functional studies in hematopoietic cells.

SBR-Blood: systems biology repository for hematopoietic cells.

PubMed

Lichtenberg, Jens; Heuston, Elisabeth F; Mishra, Tejaswini; Keller, Cheryl A; Hardison, Ross C; Bodine, David M

2016-01-04

Extensive research into hematopoiesis (the development of blood cells) over several decades has generated large sets of expression and epigenetic profiles in multiple human and mouse blood cell types. However, there is no single location to analyze how gene regulatory processes lead to different mature blood cells. We have developed a new database framework called hematopoietic Systems Biology Repository (SBR-Blood), available online at http://sbrblood.nhgri.nih.gov, which allows user-initiated analyses for cell type correlations or gene-specific behavior during differentiation using publicly available datasets for array- and sequencing-based platforms from mouse hematopoietic cells. SBR-Blood organizes information by both cell identity and by hematopoietic lineage. The validity and usability of SBR-Blood has been established through the reproduction of workflows relevant to expression data, DNA methylation, histone modifications and transcription factor occupancy profiles. Published by Oxford University Press on behalf of Nucleic Acids Research 2015. This work is written by (a) US Government employee(s) and is in the public domain in the US.

Dynamic equilibrium of heterogeneous and interconvertible multipotent hematopoietic cell subsets

PubMed Central

Weston, Wendy; Zayas, Jennifer; Perez, Ruben; George, John; Jurecic, Roland

2014-01-01

Populations of hematopoietic stem cells and progenitors are quite heterogeneous and consist of multiple cell subsets with distinct phenotypic and functional characteristics. Some of these subsets also appear to be interconvertible and oscillate between functionally distinct states. The multipotent hematopoietic cell line EML has emerged as a unique model to study the heterogeneity and interconvertibility of multipotent hematopoietic cells. Here we describe extensive phenotypic and functional heterogeneity of EML cells which stems from the coexistence of multiple cell subsets. Each of these subsets is phenotypically and functionally heterogeneous, and displays distinct multilineage differentiation potential, cell cycle profile, proliferation kinetics, and expression pattern of HSC markers and some of the key lineage-associated transcription factors. Analysis of their maintenance revealed that on a population level all EML cell subsets exhibit cell-autonomous interconvertible properties, with the capacity to generate all other subsets and re-establish complete parental EML cell population. Moreover, all EML cell subsets generated during multiple cell generations maintain their distinct phenotypic and functional signatures and interconvertible properties. The model of EML cell line suggests that interconvertible multipotent hematopoietic cell subsets coexist in a homeostatically maintained dynamic equilibrium which is regulated by currently unknown cell-intrinsic mechanisms. PMID:24903657

A diffusible signal derived from hematopoietic cells supports the survival and proliferation of regenerative cells during zebrafish fin fold regeneration.

PubMed

Hasegawa, Tomoya; Nakajima, Teruhiro; Ishida, Takashi; Kudo, Akira; Kawakami, Atsushi

2015-03-01

Multicellular organisms maintain body integrity by constantly regenerating tissues throughout their lives; however, the overall mechanism for regulating regeneration remains an open question. Studies of limb and fin regeneration in teleost fish and urodeles have shown the involvement of a number of locally activated signals at the wounded site during regeneration. Here, we demonstrate that a diffusible signal from a distance also play an essential role for regeneration. Among a number of zebrafish mutants, we found that the zebrafish cloche (clo) and tal1 mutants, which lack most hematopoietic tissues, displayed a unique regeneration defect accompanying apoptosis in primed regenerative tissue. Our analyses of the mutants showed that the cells in the primed regenerative tissue are susceptible to apoptosis, but their survival is normally supported by the presence of hematopoietic tissues, mainly the myeloid cells. We further showed that a diffusible factor in the wild-type body fluid mediates this signal. Thus, our study revealed a novel mechanism that the hematopoietic tissues regulate tissue regeneration through a diffusible signal. Copyright © 2014 Elsevier Inc. All rights reserved.

What do we know about the participation of hematopoietic stem cells in hematopoiesis?

PubMed

Drize, Nina; Petinati, Nataliya

2015-01-01

The demonstrated presence in adult tissues of cells with sustained tissue regenerative potential has given rise to the concept of tissue stem cells. Assays to detect and measure such cells indicate that they have enormous proliferative potential and usually an ability to produce all or many of the mature cell types that define the specialized functionality of the tissue. In the hematopoietic system, one or only a few cells can restore lifelong hematopoiesis of the whole organism. To what extent is the maintenance of hematopoietic stem cells required during normal hematopoiesis? How does the constant maintenance of hematopoiesis occur and what is the behavior of the hematopoietic stem cells in the normal organism? How many of the hematopoietic stem cells are created during the development of the organism? How many hematopoietic stem cells are generating more mature progeny at any given moment? What happens to the population of hematopoietic stem cells in aging? This review will attempt to describe the results of recent research which contradict some of the ideas established over the past 30 years about how hematopoiesis is regulated.

Plasticity and Maintenance of Hematopoietic Stem Cells During Development

PubMed Central

Kanji, Suman; Pompili, Vincent J.; Das, Hiranmoy

2012-01-01

Maintenance of hematopoietic stem cells (HSCs) pool depends on fine balance between self-renewal and differentiation of HSCs. HSCs normally reside within the bone marrow niche of an adult mammal. The embryonic development of HSCs is a complex process that involves the migration of developing HSCs in multiple anatomical sites. Throughout the process, developing HSCs receive internal (transcriptional program) and external (HSC niche) signals, which direct them to maintain balance between self-renewal and differentiation, also to generate a pool of HSCs. In physiological condition HSCs differentiate into all mature cell types present in the blood. However, in pathological condition they may differentiate into non-hematological cells according to the need of the body. It was shown that HSCs can transdifferentiate into cell types that do not belong to the hematopoietic system suggests a complete paradigm shift of the hierarchical hematopoietic tree. This review describes the developmental origins and regulation of HSCs focusing on developmental signals that induce the adult hematopoietic stem cell program, as these informations are very critical for manipulating conditions for expansion of HSCs in ex vivo condition. This review also states clinical application and related patents using HSC. PMID:21517745

Integration of adeno-associated virus vectors in CD34+ human hematopoietic progenitor cells after transduction.

PubMed

Fisher-Adams, G; Wong, K K; Podsakoff, G; Forman, S J; Chatterjee, S

1996-07-15

Gene transfer vectors based on adeno-associated virus (AAV) appear promising because of their high transduction frequencies regardless of cell cycle status and ability to integrate into chromosomal DNA. We tested AAV-mediated gene transfer into a panel of human bone marrow or umbilical cord-derived CD34+ hematopoietic progenitor cells, using vectors encoding several transgenes under the control of viral and cellular promoters. Gene transfer was evaluated by (1) chromosomal integration of vector sequences and (2) analysis of transgene expression. Southern hybridization and fluorescence in situ hybridization analysis of transduced CD34 genomic DNA showed the presence of integrated vector sequences in chromosomal DNA in a portion of transduced cells and showed that integrated vector sequences were replicated along with cellular DNA during mitosis. Transgene expression in transduced CD34 cells in suspension cultures and in myeloid colonies differentiating in vitro from transduced CD34 cells approximated that predicted by the multiplicity of transduction. This was true in CD34 cells from different donors, regardless of the transgene or selective pressure. Comparisons of CD34 cell transduction either before or after cytokine stimulation showed similar gene transfer frequencies. Our findings suggest that AAV transduction of CD34+ hematopoietic progenitor cells is efficient, can lead to stable integration in a population of transduced cells, and may therefore provide the basis for safe and efficient ex vivo gene therapy of the hematopoietic system.

Biologic activity of irradiated, autologous, GM-CSF-secreting leukemia cell vaccines early after allogeneic stem cell transplantation

PubMed Central

Ho, Vincent T.; Vanneman, Matthew; Kim, Haesook; Sasada, Tetsuro; Kang, Yoon Joong; Pasek, Mildred; Cutler, Corey; Koreth, John; Alyea, Edwin; Sarantopoulos, Stefanie; Antin, Joseph H.; Ritz, Jerome; Canning, Christine; Kutok, Jeffery; Mihm, Martin C.; Dranoff, Glenn; Soiffer, Robert

2009-01-01

Through an immune-mediated graft-versus-leukemia effect, allogeneic hematopoietic stem cell transplantation (HSCT) affords durable clinical benefits for many patients with hematologic malignancies. Nonetheless, subjects with high-risk acute myeloid leukemia or advanced myelodysplasia often relapse, underscoring the need to intensify tumor immunity within this cohort. In preclinical models, allogeneic HSCT followed by vaccination with irradiated tumor cells engineered to secrete GM-CSF generates a potent antitumor effect without exacerbating the toxicities of graft-versus-host disease (GVHD). To test whether this strategy might be similarly active in humans, we conducted a Phase I clinical trial in which high-risk acute myeloid leukemia or myelodysplasia patients were immunized with irradiated, autologous, GM-CSF-secreting tumor cells early after allogeneic, nonmyeloablative HSCT. Despite the administration of a calcineurin inhibitor as prophylaxis against GVHD, vaccination elicited local and systemic reactions that were qualitatively similar to those previously observed in nontransplanted, immunized solid-tumor patients. While the frequencies of acute and chronic GVHD were not increased, 9 of 10 subjects who completed vaccination achieved durable complete remissions, with a median follow-up of 26 months (range 12–43 months). Six long-term responders showed marked decreases in the levels of soluble NKG2D ligands, and 3 demonstrated normalization of cytotoxic lymphocyte NKG2D expression as a function of treatment. Together, these results establish the safety and immunogenicity of irradiated, autologous, GM-CSF-secreting leukemia cell vaccines early after allogeneic HSCT, and raise the possibility that this combinatorial immunotherapy might potentiate graft-versus-leukemia in patients. PMID:19717467

Uhrf1 controls the self-renewal versus differentiation of hematopoietic stem cells by epigenetically regulating the cell-division modes

PubMed Central

Zhao, Jingyao; Chen, Xufeng; Song, Guangrong; Zhang, Jiali; Liu, Haifeng; Liu, Xiaolong

2017-01-01

Hematopoietic stem cells (HSCs) are able to both self-renew and differentiate. However, how individual HSC makes the decision between self-renewal and differentiation remains largely unknown. Here we report that ablation of the key epigenetic regulator Uhrf1 in the hematopoietic system depletes the HSC pool, leading to hematopoietic failure and lethality. Uhrf1-deficient HSCs display normal survival and proliferation, yet undergo erythroid-biased differentiation at the expense of self-renewal capacity. Notably, Uhrf1 is required for the establishment of DNA methylation patterns of erythroid-specific genes during HSC division. The expression of these genes is enhanced in the absence of Uhrf1, which disrupts the HSC-division modes by promoting the symmetric differentiation and suppressing the symmetric self-renewal. Moreover, overexpression of one of the up-regulated genes, Gata1, in HSCs is sufficient to phenocopy Uhrf1-deficient HSCs, which show impaired HSC symmetric self-renewal and increased differentiation commitment. Taken together, our findings suggest that Uhrf1 controls the self-renewal versus differentiation of HSC through epigenetically regulating the cell-division modes, thus providing unique insights into the relationship among Uhrf1-mediated DNA methylation, cell-division mode, and HSC fate decision. PMID:27956603

Notch2 blockade enhances hematopoietic stem cell mobilization and homing.

PubMed

Wang, Weihuan; Yu, Shuiliang; Myers, Jay; Wang, Yiwei; Xin, William W; Albakri, Marwah; Xin, Alison W; Li, Ming; Huang, Alex Y; Xin, Wei; Siebel, Christian W; Lazarus, Hillard M; Zhou, Lan

2017-10-01

Despite use of newer approaches, some patients being considered for autologous hematopoietic cell transplantation (HCT) may only mobilize limited numbers of hematopoietic progenitor cells (HPCs) into blood, precluding use of the procedure, or being placed at increased risk of complications due to slow hematopoietic reconstitution. Developing more efficacious HPC mobilization regimens and strategies may enhance the mobilization process and improve patient outcome. Although Notch signaling is not essential for homeostasis of adult hematopoietic stem cells (HSCs), Notch-ligand adhesive interaction maintains HSC quiescence and niche retention. Using Notch receptor blocking antibodies, we report that Notch2 blockade, but not Notch1 blockade, sensitizes hematopoietic stem cells and progenitors (HSPCs) to mobilization stimuli and leads to enhanced egress from marrow to the periphery. Notch2 blockade leads to transient myeloid progenitor expansion without affecting HSC homeostasis and self-renewal. We show that transient Notch2 blockade or Notch2-loss in mice lacking Notch2 receptor lead to decreased CXCR4 expression by HSC but increased cell cycling with CXCR4 transcription being directly regulated by the Notch transcriptional protein RBPJ. In addition, we found that Notch2-blocked or Notch2-deficient marrow HSPCs show an increased homing to the marrow, while mobilized Notch2-blocked, but not Notch2-deficient stem cells and progenitors, displayed a competitive repopulating advantage and enhanced hematopoietic reconstitution. These findings suggest that blocking Notch2 combined with the current clinical regimen may further enhance HPC mobilization and improve engraftment during HCT. Copyright© 2017 Ferrata Storti Foundation.

The Gottingen minipig is a model of the hematopoietic acute radiation syndrome: G-CSF stimulates hematopoiesis and enhances survival from lethal total-body gamma-irradiation

PubMed Central

Moroni, Maria; Ngudiankama, Barbara F.; Christensen, Christine; Olsen, Cara H.; Owens, Rossitsa; Lombardini, Eric D.; Holt, Rebecca K.; Whitnall, Mark H.

2013-01-01

Purpose We are characterizing the Gottingen minipig as an additional large animal model for advanced drug testing for the Acute Radiation Syndrome (ARS), to enhance discovery and development of novel radiation countermeasures. Among the advantages provided by this model, the similarities to human hematological parameters and dynamics of cell loss/recovery following irradiation provide a convenient means to compare efficacy of drugs known to affect bone marrow cellularity and hematopoiesis. Methods and Materials Male Gottingen minipigs, 4–5 months old and weighing 9–11 kg were used for this study. We tested the standard off-label treatment for ARS, rhG-CSF (Neupogen®, 10 μg/kg/day for 17 days), at the estimated LD70/30 total-body gamma-irradiation (TBI) radiation dose for the hematopoietic syndrome, starting 24 hours after irradiation. Results Results indicate G-CSF enhanced survival, stimulated recovery from neutropenia, and induced mobilization of hematopoietic progenitor cells. In addition, administration of G-CSF resulted in maturation of monocytes/macrophages. Conclusion These results support continuing efforts toward validation of the minipig as a large animal model for advanced testing of radiation countermeasures and characterization of the pathophysiology of ARS, and suggest that the efficacy of G-CSF in improving survival after total body irradiation may involve mechanisms other than increasing numbers of circulating granulocytes. PMID:23845847

Hematopoietic stem cell origin of connective tissues.

PubMed

Ogawa, Makio; Larue, Amanda C; Watson, Patricia M; Watson, Dennis K

2010-07-01

Connective tissue consists of "connective tissue proper," which is further divided into loose and dense (fibrous) connective tissues and "specialized connective tissues." Specialized connective tissues consist of blood, adipose tissue, cartilage, and bone. In both loose and dense connective tissues, the principal cellular element is fibroblasts. It has been generally believed that all cellular elements of connective tissue, including fibroblasts, adipocytes, chondrocytes, and bone cells, are generated solely by mesenchymal stem cells. Recently, a number of studies, including those from our laboratory based on transplantation of single hematopoietic stem cells, strongly suggested a hematopoietic stem cell origin of these adult mesenchymal tissues. This review summarizes the experimental evidence for this new paradigm and discusses its translational implications. Copyright 2010 ISEH - Society for Hematology and Stem Cells. All rights reserved.

Hematopoietic stem cell engineering at a crossroads.

PubMed

Rivière, Isabelle; Dunbar, Cynthia E; Sadelain, Michel

2012-02-02

The genetic engineering of hematopoietic stem cells is the basis for potentially treating a large array of hereditary and acquired diseases, and stands as the paradigm for stem cell engineering in general. Recent clinical reports support the formidable promise of this approach but also highlight the limitations of the technologies used to date, which have on occasion resulted in clonal expansion, myelodysplasia, or leukemogenesis. New research directions, predicated on improved vector designs, targeted gene delivery or the therapeutic use of pluripotent stem cells, herald the advent of safer and more effective hematopoietic stem cell therapies that may transform medical practice. In this review, we place these recent advances in perspective, emphasizing the solutions emerging from a wave of new technologies and highlighting the challenges that lie ahead.

Single high-dose irradiation aggravates eosinophil-mediated fibrosis through IL-33 secreted from impaired vessels in the skin compared to fractionated irradiation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Eun-Jung, E-mail: forejs2@yuhs.ac; Kim, Jun Won, E-mail: JUNWON@yuhs.ac; Yoo, Hyun, E-mail: gochunghee@yuhs.ac

We have revealed in a porcine skin injury model that eosinophil recruitment was dose-dependently enhanced by a single high-dose irradiation. In this study, we investigated the underlying mechanism of eosinophil-associated skin fibrosis and the effect of high-dose-per-fraction radiation. The dorsal skin of a mini-pig was divided into two sections containing 4-cm{sup 2} fields that were irradiated with 30 Gy in a single fraction or 5 fractions and biopsied regularly over 14 weeks. Eosinophil-related Th2 cytokines such as interleukin (IL)-4, IL-5, and C–C motif chemokine-11 (CCL11/eotaxin) were evaluated by quantitative real-time PCR. RNA-sequencing using 30 Gy-irradiated mouse skin and functional assays in amore » co-culture system of THP-1 and irradiated-human umbilical vein endothelial cells (HUVECs) were performed to investigate the mechanism of eosinophil-mediated radiation fibrosis. Single high-dose-per-fraction irradiation caused pronounced eosinophil accumulation, increased profibrotic factors collagen and transforming growth factor-β, enhanced production of eosinophil-related cytokines including IL-4, IL-5, CCL11, IL-13, and IL-33, and reduced vessels compared with 5-fraction irradiation. IL-33 notably increased in pig and mouse skin vessels after single high-dose irradiation of 30 Gy, as well as in irradiated HUVECs following 12 Gy. Blocking IL-33 suppressed the migration ability of THP-1 cells and cytokine secretion in a co-culture system of THP-1 cells and irradiated HUVECs. Hence, high-dose-per-fraction irradiation appears to enhance eosinophil-mediated fibrotic responses, and IL-33 may be a key molecule operating in eosinophil-mediated fibrosis in high-dose-per fraction irradiated skin. - Highlights: • Single high-dose irradiation aggravates eosinophil-mediated fibrosis through IL-33. • Vascular endothelial cells damaged by high-dose radiation secrete IL-33. • Blocking IL-33 suppressed migration of inflammatory cells and cytokine secretion

Bag1 is essential for differentiation and survival of hematopoietic and neuronal cells.

PubMed

Götz, Rudolf; Wiese, Stefan; Takayama, Shinichi; Camarero, Guadalupe C; Rossoll, Wilfried; Schweizer, Ulrich; Troppmair, Jakob; Jablonka, Sibylle; Holtmann, Bettina; Reed, John C; Rapp, Ulf R; Sendtner, Michael

2005-09-01

Bag1 is a cochaperone for the heat-shock protein Hsp70 that interacts with C-Raf, B-Raf, Akt, Bcl-2, steroid hormone receptors and other proteins. Here we use targeted gene disruption in mice to show that Bag1 has an essential role in the survival of differentiating neurons and hematopoietic cells. Cells of the fetal liver and developing nervous system in Bag1-/- mice underwent massive apoptosis. Lack of Bag1 did not disturb the primary function of Akt or Raf, as phosphorylation of the forkhead transcription factor FKHR and activation of extracellular signal-regulated kinase (Erk)-1/2 were not affected. However, the defect was associated with the disturbance of a tripartite complex formed by Akt, B-Raf and Bag1, in addition to the absence of Bad phosphorylation at Ser136. We also observed reduced expression of members of the inhibitor of apoptosis (IAP) family. Our data show that Bag1 is a physiological mediator of extracellular survival signals linked to the cellular mechanisms that prevent apoptosis in hematopoietic and neuronal progenitor cells.

Hematopoietic Stem Cells in Regenerative Medicine: Astray or on the Path?

PubMed Central

Müller, Albrecht M.; Huppertz, Sascha; Henschler, Reinhard

2016-01-01

Hematopoietic stem cells (HSCs) are the best characterized adult stem cells and the only stem cell type in routine clinical use. The concept of stem cell transplantation laid the foundations for the development of novel cell therapies within, and even outside, the hematopoietic system. Here, we report on the history of hematopoietic cell transplantation (HCT) and of HSC isolation, we briefly summarize the capabilities of HSCs to reconstitute the entire hemato/lymphoid cell system, and we assess current indications for HCT. We aim to draw the lines between areas where HCT has been firmly established, areas where HCT can in the future be expected to be of clinical benefit using their regenerative functions, and areas where doubts persist. We further review clinical trials for diverse approaches that are based on HCT. Finally, we highlight the advent of genome editing in HSCs and critically view the use of HSCs in non-hematopoietic tissue regeneration. PMID:27721700

Efficient ablation of genes in human hematopoietic stem and effector cells using CRISPR/Cas9

PubMed Central

Mandal, Pankaj K.; Ferreira, Leonardo M. R.; Collins, Ryan; Meissner, Torsten B.; Boutwell, Christian L.; Friesen, Max; Vrbanac, Vladimir; Garrison, Brian S.; Stortchevoi, Alexei; Bryder, David; Musunuru, Kiran; Brand, Harrison; Tager, Andrew M.; Allen, Todd M.; Talkowski, Michael E.; Rossi, Derrick J.; Cowan, Chad A.

2014-01-01

SUMMARY Genome editing via CRISPR/Cas9 has rapidly become the tool of choice by virtue of its efficacy and ease of use. However, CRISPR/Cas9 mediated genome editing in clinically relevant human somatic cells remains untested. Here, we report CRISPR/Cas9 targeting of two clinically relevant genes, B2M and CCR5, in primary human CD4+ T cells and CD34+ hematopoietic stem and progenitor cells (HSPCs). Use of single RNA guides led to highly efficient mutagenesis in HSPCs but not in T cells. A dual guide approach improved gene deletion efficacy in both cell types. HSPCs that had undergone genome editing with CRISPR/Cas9 retained multi-lineage potential. We examined predicted on- and off-target mutations via target capture sequencing in HSPCs and observed low levels of off-target mutagenesis at only one site. These results demonstrate that CRISPR/Cas9 can efficiently ablate genes in HSPCs with minimal off-target mutagenesis, which could have broad applicability for hematopoietic cell-based therapy. PMID:25517468

Hematopoietic progenitor migration to the adult thymus

PubMed Central

Zlotoff, Daniel A.; Bhandoola, Avinash

2010-01-01

While most hematopoietic lineages develop in the bone marrow (BM), T cells uniquely complete their development in the specialized environment of the thymus. Hematopoietic stem cells with long-term self-renewal capacity are not present in the thymus. As a result, continuous T cell development requires that BM-derived progenitors be imported into the thymus throughout adult life. The process of thymic homing begins with the mobilization of progenitors out of the bone marrow, continues with their circulation in the bloodstream, and concludes with their settling in the thymus. This review will discuss each of these steps as they occur in the unirradiated and post-irradiation scenarios, focusing on the molecular mechanisms of regulation. Improved knowledge about these early steps in T cell generation may accelerate the development of new therapeutic options in patients with impaired T cell number or function. PMID:21251013

BAF180 regulates cellular senescence and hematopoietic stem cell homeostasis through p21

PubMed Central

Lee, Hyemin; Dai, Fangyan; Zhuang, Li; Xiao, Zhen-Dong; Kim, Jongchan; Zhang, Yilei; Ma, Li; You, M. James; Wang, Zhong; Gan, Boyi

2016-01-01

BAF180 (also called PBRM1), a subunit of the SWI/SNF complex, plays critical roles in the regulation of chromatin remodeling and gene transcription, and is frequently mutated in several human cancers. However, the role of mammalian BAF180 in tumor suppression and tissue maintenance in vivo remains largely unknown. Here, using a conditional somatic knockout approach, we explored the cellular and organismal functions of BAF180 in mouse. BAF180 deletion in primary mouse embryonic fibroblasts (MEFs) triggers profound cell cycle arrest, premature cellular senescence, without affecting DNA damage response or chromosomal integrity. While somatic deletion of BAF180 in adult mice does not provoke tumor development, BAF180 deficient mice exhibit defects in hematopoietic system characterized by progressive reduction of hematopoietic stem cells (HSCs), defective long-term repopulating potential, and hematopoietic lineage developmental aberrations. BAF180 deletion results in elevated p21 expression in both MEFs and HSCs. Mechanistically, we showed that BAF180 binds to p21 promoter, and BAF180 deletion enhances the binding of modified histones associated with transcriptional activation on p21 promoter. Deletion of p21 rescues cell cycle arrest and premature senescence in BAF180 deficient MEFs, and partially rescues hematopoietic defects in BAF180 deficient mice. Together, our study identifies BAF180 as a critical regulator of cellular senescence and HSC homeostasis, which is at least partially regulated through BAF180-mediated suppression of p21 expression. Our results also suggest that senescence triggered by BAF180 inactivation may serve as a failsafe mechanism to restrain BAF180 deficiency-associated tumor development, providing a conceptual framework to further understand BAF180 function in tumor biology. PMID:26992241

Neurotrophins promote revascularization by local recruitment of TrkB+ endothelial cells and systemic mobilization of hematopoietic progenitors

PubMed Central

Kermani, Pouneh; Rafii, Dahlia; Jin, David K.; Whitlock, Paul; Schaffer, Wendy; Chiang, Anne; Vincent, Loic; Friedrich, Matthias; Shido, Koji; Hackett, Neil R.; Crystal, Ronald G.; Rafii, Shahin; Hempstead, Barbara L.

2005-01-01

The neurotrophin brain-derived neurotrophic factor (BDNF) is required for the maintenance of cardiac vessel wall stability during embryonic development through direct angiogenic actions on endothelial cells expressing the tropomysin receptor kinase B (TrkB). However, the role of BDNF and a related neurotrophin ligand, neurotrophin-4 (NT-4), in the regulation of revascularization of the adult tissues is unknown. To study the potential angiogenic capacity of BDNF in mediating the neovascularization of ischemic and non-ischemic adult mouse tissues, we utilized a hindlimb ischemia and a subcutaneous Matrigel model. Recruitment of endothelial cells and promotion of channel formation within the Matrigel plug by BDNF and NT-4 was comparable to that induced by VEGF-A. The introduction of BDNF into non-ischemic ears or ischemic limbs induced neoangiogenesis, with a 2-fold increase in the capillary density. Remarkably, treatment with BDNF progressively increased blood flow in the ischemic limb over 21 days, similar to treatment with VEGF-A. The mechanism by which BDNF enhances capillary formation is mediated in part through local activation of the TrkB receptor and also by recruitment of Sca-1+CD11b+ pro-angiogenic hematopoietic cells. BDNF induces a potent direct chemokinetic action on subsets of marrow-derived Sca-1+ hematopoietic cells co-expressing TrkB. These studies suggest that local regional delivery of BDNF may provide a novel mechanism for inducing neoangiogenesis through both direct actions on local TrkB-expressing endothelial cells in skeletal muscle and recruitment of specific subsets of TrkB+ bone marrow–derived hematopoietic cells to provide peri-endothelial support for the newly formed vessels. PMID:15765148

Memory T and memory B cells share a transcriptional program of self-renewal with long-term hematopoietic stem cells

PubMed Central

Luckey, Chance John; Bhattacharya, Deepta; Goldrath, Ananda W.; Weissman, Irving L.; Benoist, Christophe; Mathis, Diane

2006-01-01

The only cells of the hematopoietic system that undergo self-renewal for the lifetime of the organism are long-term hematopoietic stem cells and memory T and B cells. To determine whether there is a shared transcriptional program among these self-renewing populations, we first compared the gene-expression profiles of naïve, effector and memory CD8+ T cells with those of long-term hematopoietic stem cells, short-term hematopoietic stem cells, and lineage-committed progenitors. Transcripts augmented in memory CD8+ T cells relative to naïve and effector T cells were selectively enriched in long-term hematopoietic stem cells and were progressively lost in their short-term and lineage-committed counterparts. Furthermore, transcripts selectively decreased in memory CD8+ T cells were selectively down-regulated in long-term hematopoietic stem cells and progressively increased with differentiation. To confirm that this pattern was a general property of immunologic memory, we turned to independently generated gene expression profiles of memory, naïve, germinal center, and plasma B cells. Once again, memory-enriched and -depleted transcripts were also appropriately augmented and diminished in long-term hematopoietic stem cells, and their expression correlated with progressive loss of self-renewal function. Thus, there appears to be a common signature of both up- and down-regulated transcripts shared between memory T cells, memory B cells, and long-term hematopoietic stem cells. This signature was not consistently enriched in neural or embryonic stem cell populations and, therefore, appears to be restricted to the hematopoeitic system. These observations provide evidence that the shared phenotype of self-renewal in the hematopoietic system is linked at the molecular level. PMID:16492737

Correction of mutant Fanconi anemia gene by homologous recombination in human hematopoietic cells using adeno-associated virus vector.

PubMed

Paiboonsukwong, Kittiphong; Ohbayashi, Fumi; Shiiba, Haruka; Aizawa, Emi; Yamashita, Takayuki; Mitani, Kohnosuke

2009-11-01

Adeno-associated virus (AAV) vectors have been shown to correct a variety of mutations in human cells by homologous recombination (HR) at high rates, which can overcome insertional mutagenesis and transgene silencing, two of the major hurdles in conventional gene addition therapy of inherited diseases. We examined an ability of AAV vectors to repair a mutation in human hematopoietic cells by HR. We infected a human B-lymphoblastoid cell line (BCL) derived from a normal subject with an AAV, which disrupts the hypoxanthine phosphoribosyl transferase1 (HPRT1) locus, to measure the frequency of AAV-mediated HR in BCL cells. We subsequently constructed an AAV vector encoding the normal sequences from the Fanconi anemia group A (FANCA) locus to correct a mutation in the gene in BCL derived from a FANCA patient. Under optimal conditions, approximately 50% of BCL cells were transduced with an AAV serotype 2 (AAV-2) vector. In FANCA BCL cells, up to 0.016% of infected cells were gene-corrected by HR. AAV-mediated restoration of normal genotypic and phenotypic characteristics in FANCA-mutant cells was confirmed at the DNA, protein and functional levels. The results obtained in the present study indicate that AAV vectors may be applicable for gene correction therapy of inherited hematopoietic disorders.

Direct Reprogramming of Murine Fibroblasts to Hematopoietic Progenitor Cells

PubMed Central

Batta, Kiran; Florkowska, Magdalena; Kouskoff, Valerie; Lacaud, Georges

2014-01-01

Summary Recent reports have shown that somatic cells, under appropriate culture conditions, could be directly reprogrammed to cardiac, hepatic, or neuronal phenotype by lineage-specific transcription factors. In this study, we demonstrate that both embryonic and adult somatic fibroblasts can be efficiently reprogrammed to clonal multilineage hematopoietic progenitors by the ectopic expression of the transcription factors ERG, GATA2, LMO2, RUNX1c, and SCL. These reprogrammed cells were stably expanded on stromal cells and possessed short-term reconstitution ability in vivo. Loss of p53 function facilitated reprogramming to blood, and p53−/− reprogrammed cells efficiently generated erythroid, megakaryocytic, myeloid, and lymphoid lineages. Genome-wide analyses revealed that generation of hematopoietic progenitors was preceded by the appearance of hemogenic endothelial cells expressing endothelial and hematopoietic genes. Altogether, our findings suggest that direct reprogramming could represent a valid alternative approach to the differentiation of embryonic stem cells (ESCs) or induced pluripotent stem cells (iPSCs) for disease modeling and autologous blood cell therapies. PMID:25466247

Are neural crest stem cells the missing link between hematopoietic and neurogenic niches?

PubMed

Coste, Cécile; Neirinckx, Virginie; Gothot, André; Wislet, Sabine; Rogister, Bernard

2015-01-01

Hematopoietic niches are defined as cellular and molecular microenvironments that regulate hematopoietic stem cell (HSC) function together with stem cell autonomous mechanisms. Many different cell types have been characterized as contributors to the formation of HSC niches, such as osteoblasts, endothelial cells, Schwann cells, and mesenchymal progenitors. These mesenchymal progenitors have themselves been classified as CXC chemokine ligand (CXCL) 12-abundant reticular (CAR) cells, stem cell factor expressing cells, or nestin-positive mesenchymal stem cells (MSCs), which have been recently identified as neural crest-derived cells (NCSCs). Together, these cells are spatially associated with HSCs and believed to provide appropriate microenvironments for HSC self-renewal, differentiation, mobilization and hibernation both by cell-cell contact and soluble factors. Interestingly, it appears that regulatory pathways governing the hematopoietic niche homeostasis are operating in the neurogenic niche as well. Therefore, this review paper aims to compare both the regulation of hematopoietic and neurogenic niches, in order to highlight the role of NCSCs and nervous system components in the development and the regulation of the hematopoietic system.

[Hematopoietic reconstitution after transplantation of uncontrolled-rate cryopreservation autologous peripheral blood hematopoietic stem cells using -80 °C mechanical freezer].

PubMed

Liu, Mo; Zhao, Yu; Sun, Jing-Fen; Zhao, Wei; Wang, Li-Li; Yu, Li

2015-02-01

This study was to identify the efficacy of -80°C cryopreservated peripheral blood hemato-poietic stem cell (PBHSC) transplantation for hematopoietic reanstitution in patients. The efficacy of 104 patients underwent autologous peripheral blood hematopoietic stem cell transplantation using uncontrolled-rate freezing and storage at -80°C was evaluated. This cryopreservation method could effectively cryopreserve peripheral blood stem cells. Out of 104 patients only 2 patients died, other patients got hematologic reconstition satisfactorily, the median engrafement times of neutrophils and platelet were 12 and 14 days respectively, the activity of cells after rehabilitation was 94%, the mean recovery rates of CD34(+) cells and mononuclear cells (MNC) were 86% and 80.3% respectively. There were no significant influences on engrafement time in sex, chemotherapy circles and radiotherapy. The engrafement of leukocytes associated with amount of CD34(+) cells. This simple uncontrolled-rate freezing PBHSC at -80°C is safe, effective and economic, and can meet clinical needs. As compared with the classical cryopreservation, there were no significant differences in hematopoietic reconstitution. Therefore, this method worth to popularize and apply in clinic.

Hematopoietic stem cell engineering at a crossroads

PubMed Central

Rivière, Isabelle; Dunbar, Cynthia E.

2012-01-01

The genetic engineering of hematopoietic stem cells is the basis for potentially treating a large array of hereditary and acquired diseases, and stands as the paradigm for stem cell engineering in general. Recent clinical reports support the formidable promise of this approach but also highlight the limitations of the technologies used to date, which have on occasion resulted in clonal expansion, myelodysplasia, or leukemogenesis. New research directions, predicated on improved vector designs, targeted gene delivery or the therapeutic use of pluripotent stem cells, herald the advent of safer and more effective hematopoietic stem cell therapies that may transform medical practice. In this review, we place these recent advances in perspective, emphasizing the solutions emerging from a wave of new technologies and highlighting the challenges that lie ahead. PMID:22096239

Identifying States along the Hematopoietic Stem Cell Differentiation Hierarchy with Single Cell Specificity via Raman Spectroscopy.

PubMed

Ilin, Yelena; Choi, Ji Sun; Harley, Brendan A C; Kraft, Mary L

2015-11-17

A major challenge for expanding specific types of hematopoietic cells ex vivo for the treatment of blood cell pathologies is identifying the combinations of cellular and matrix cues that direct hematopoietic stem cells (HSC) to self-renew or differentiate into cell populations ex vivo. Microscale screening platforms enable minimizing the number of rare HSCs required to screen the effects of numerous cues on HSC fate decisions. These platforms create a strong demand for label-free methods that accurately identify the fate decisions of individual hematopoietic cells at specific locations on the platform. We demonstrate the capacity to identify discrete cells along the HSC differentiation hierarchy via multivariate analysis of Raman spectra. Notably, cell state identification is accurate for individual cells and independent of the biophysical properties of the functionalized polyacrylamide gels upon which these cells are cultured. We report partial least-squares discriminant analysis (PLS-DA) models of single cell Raman spectra enable identifying four dissimilar hematopoietic cell populations across the HSC lineage specification. Successful discrimination was obtained for a population enriched for long-term repopulating HSCs (LT-HSCs) versus their more differentiated progeny, including closely related short-term repopulating HSCs (ST-HSCs) and fully differentiated lymphoid (B cells) and myeloid (granulocytes) cells. The lineage-specific differentiation states of cells from these four subpopulations were accurately identified independent of the stiffness of the underlying biomaterial substrate, indicating subtle spectral variations that discriminated these populations were not masked by features from the culture substrate. This approach enables identifying the lineage-specific differentiation stages of hematopoietic cells on biomaterial substrates of differing composition and may facilitate correlating hematopoietic cell fate decisions with the extrinsic cues that

In vivo engineering of bone tissues with hematopoietic functions and mixed chimerism

PubMed Central

Shih, Yu-Ru; Kang, Heemin; Rao, Vikram; Chiu, Yu-Jui; Kwon, Seong Keun; Varghese, Shyni

2017-01-01

Synthetic biomimetic matrices with osteoconductivity and osteoinductivity have been developed to regenerate bone tissues. However, whether such systems harbor donor marrow in vivo and support mixed chimerism remains unknown. We devised a strategy to engineer bone tissues with a functional bone marrow (BM) compartment in vivo by using a synthetic biomaterial with spatially differing cues. Specifically, we have developed a synthetic matrix recapitulating the dual-compartment structures by modular assembly of mineralized and nonmineralized macroporous structures. Our results show that these matrices incorporated with BM cells or BM flush transplanted into recipient mice matured into functional bone displaying the cardinal features of both skeletal and hematopoietic compartments similar to native bone tissue. The hematopoietic function of bone tissues was demonstrated by its support for a higher percentage of mixed chimerism compared with i.v. injection and donor hematopoietic cell mobilization in the circulation of nonirradiated recipients. Furthermore, hematopoietic cells sorted from the engineered bone tissues reconstituted the hematopoietic system when transplanted into lethally irradiated secondary recipients. Such engineered bone tissues could potentially be used as ectopic BM surrogates for treatment of nonmalignant BM diseases and as a tool to study hematopoiesis, donor–host cell dynamics, tumor tropism, and hematopoietic cell transplantation. PMID:28484009

In vivo engineering of bone tissues with hematopoietic functions and mixed chimerism.

PubMed

Shih, Yu-Ru; Kang, Heemin; Rao, Vikram; Chiu, Yu-Jui; Kwon, Seong Keun; Varghese, Shyni

2017-05-23

Synthetic biomimetic matrices with osteoconductivity and osteoinductivity have been developed to regenerate bone tissues. However, whether such systems harbor donor marrow in vivo and support mixed chimerism remains unknown. We devised a strategy to engineer bone tissues with a functional bone marrow (BM) compartment in vivo by using a synthetic biomaterial with spatially differing cues. Specifically, we have developed a synthetic matrix recapitulating the dual-compartment structures by modular assembly of mineralized and nonmineralized macroporous structures. Our results show that these matrices incorporated with BM cells or BM flush transplanted into recipient mice matured into functional bone displaying the cardinal features of both skeletal and hematopoietic compartments similar to native bone tissue. The hematopoietic function of bone tissues was demonstrated by its support for a higher percentage of mixed chimerism compared with i.v. injection and donor hematopoietic cell mobilization in the circulation of nonirradiated recipients. Furthermore, hematopoietic cells sorted from the engineered bone tissues reconstituted the hematopoietic system when transplanted into lethally irradiated secondary recipients. Such engineered bone tissues could potentially be used as ectopic BM surrogates for treatment of nonmalignant BM diseases and as a tool to study hematopoiesis, donor-host cell dynamics, tumor tropism, and hematopoietic cell transplantation.

Genotoxicity of retroviral hematopoietic stem cell gene therapy

PubMed Central

Trobridge, Grant D

2012-01-01

Introduction Retroviral vectors have been developed for hematopoietic stem cell (HSC) gene therapy and have successfully cured X-linked severe combined immunodeficiency (SCID-X1), adenosine deaminase deficiency (ADA-SCID), adrenoleukodystrophy, and Wiskott-Aldrich syndrome. However, in HSC gene therapy clinical trials, genotoxicity mediated by integrated vector proviruses has led to clonal expansion, and in some cases frank leukemia. Numerous studies have been performed to understand the molecular basis of vector-mediated genotoxicity with the aim of developing safer vectors and safer gene therapy protocols. These genotoxicity studies are critical to advancing HSC gene therapy. Areas covered This review provides an introduction to the mechanisms of retroviral vector genotoxicity. It also covers advances over the last 20 years in designing safer gene therapy vectors, and in integration site analysis in clinical trials and large animal models. Mechanisms of retroviral-mediated genotoxicity, and the risk factors that contribute to clonal expansion and leukemia in HSC gene therapy are introduced. Expert opinion Continued research on virus–host interactions and next-generation vectors should further improve the safety of future HSC gene therapy vectors and protocols. PMID:21375467

Adult hematopoietic stem cells lacking Hif-1α self-renew normally

PubMed Central

Vukovic, Milica; Sepulveda, Catarina; Subramani, Chithra; Guitart, Amélie V.; Mohr, Jasmine; Allen, Lewis; Panagopoulou, Theano I.; Paris, Jasmin; Lawson, Hannah; Villacreces, Arnaud; Armesilla-Diaz, Alejandro; Gezer, Deniz; Holyoake, Tessa L.; Ratcliffe, Peter J.

2016-01-01

The hematopoietic stem cell (HSC) pool is maintained under hypoxic conditions within the bone marrow microenvironment. Cellular responses to hypoxia are largely mediated by the hypoxia-inducible factors, Hif-1 and Hif-2. The oxygen-regulated α subunits of Hif-1 and Hif-2 (namely, Hif-1α and Hif-2α) form dimers with their stably expressed β subunits and control the transcription of downstream hypoxia-responsive genes to facilitate adaptation to low oxygen tension. An initial study concluded that Hif-1α is essential for HSC maintenance, whereby Hif-1α–deficient HSCs lost their ability to self-renew in serial transplantation assays. In another study, we demonstrated that Hif-2α is dispensable for cell-autonomous HSC maintenance, both under steady-state conditions and following transplantation. Given these unexpected findings, we set out to revisit the role of Hif-1α in cell-autonomous HSC functions. Here we demonstrate that inducible acute deletion of Hif-1α has no impact on HSC survival. Notably, unstressed HSCs lacking Hif-1α efficiently self-renew and sustain long-term multilineage hematopoiesis upon serial transplantation. Finally, Hif-1α–deficient HSCs recover normally after hematopoietic injury induced by serial administration of 5-fluorouracil. We therefore conclude that despite the hypoxic nature of the bone marrow microenvironment, Hif-1α is dispensable for cell-autonomous HSC maintenance. PMID:27060169

Early osteoinductive human bone marrow mesenchymal stromal/stem cells support an enhanced hematopoietic cell expansion with altered chemotaxis- and adhesion-related gene expression profiles

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sugino, Noriko; Department of Transfusion Medicine and Cell Therapy, Kyoto University Hospital, Kyoto 606-8507; Miura, Yasuo, E-mail: ym58f5@kuhp.kyoto-u.ac.jp

Bone marrow (BM) microenvironment has a crucial role in supporting hematopoiesis. Here, by using a microarray analysis, we demonstrate that human BM mesenchymal stromal/stem cells (MSCs) in an early osteoinductive stage (e-MSCs) are characterized by unique hematopoiesis-associated gene expression with an enhanced hematopoiesis-supportive ability. In comparison to BM-MSCs without osteoinductive treatment, gene expression in e-MSCs was significantly altered in terms of their cell adhesion- and chemotaxis-related profiles, as identified with Gene Ontology and Gene Set Enrichment Analysis. Noteworthy, expression of the hematopoiesis-associated molecules CXCL12 and vascular cell adhesion molecule 1 was remarkably decreased in e-MSCs. e-MSCs supported an enhanced expansionmore » of CD34{sup +} hematopoietic stem and progenitor cells, and generation of myeloid lineage cells in vitro. In addition, short-term osteoinductive treatment favored in vivo hematopoietic recovery in lethally irradiated mice that underwent BM transplantation. e-MSCs exhibited the absence of decreased stemness-associated gene expression, increased osteogenesis-associated gene expression, and apparent mineralization, thus maintaining the ability to differentiate into adipogenic cells. Our findings demonstrate the unique biological characteristics of e-MSCs as hematopoiesis-regulatory stromal cells at differentiation stage between MSCs and osteoprogenitor cells and have significant implications in developing new strategy for using pharmacological osteoinductive treatment to support hematopoiesis in hematopoietic stem and progenitor cell transplantation. - Highlights: • Human BM-MSCs in an early osteoinductive stage (e-MSCs) support hematopoiesis. • Adhesion- and chemotaxis-associated gene signatures are altered in e-MSCs. • Expression of CXCL12 and VCAM1 is remarkably decreased in e-MSCs. • e-MSCs are at differentiation stage between MSCs and osteoprogenitor cells. • Osteoinductive

Long-term culture and differentiation of porcine red bone marrow hematopoietic cells co-cultured with immortalized mesenchymal cells.

PubMed

Garba, Abubakar; Acar, Delphine D; Roukaerts, Inge D M; Desmarets, Lowiese M B; Devriendt, Bert; Nauwynck, Hans J

2017-09-01

Mesenchymal cells are multipotent stromal cells with self-renewal, differentiation and immunomodulatory capabilities. We aimed to develop a co-culture model for differentiating hematopoietic cells on top of immortalized mesenchymal cells for studying interactions between hematopoietic and mesenchymal cells, useful for adequately exploring the therapeutic potential of mesenchymal cells. In this study, we investigated the survival, proliferation and differentiation of porcine red bone marrow hematopoietic cells co-cultured with immortalized porcine bone marrow mesenchymal cells for a period of five weeks. Directly after collection, primary porcine bone marrow mesenchymal cells adhered firmly to the bottom of the culture plates and showed a fibroblast-like appearance, one week after isolation. Upon immortalization, porcine bone marrow mesenchymal cells were continuously proliferating. They were positive for simian virus 40 (SV40) large T antigen and the mesenchymal cell markers CD44 and CD55. Isolated red bone marrow cells were added to these immortalized mesenchymal cells. Five weeks post-seeding, 92±6% of the red bone marrow hematopoietic cells were still alive and their number increased 3-fold during five weekly subpassages on top of the immortalized mesenchymal cells. The red bone marrow hematopoietic cells were originally small and round; later, the cells increased in size. Some of them became elongated, while others remained round. Tiny dendrites appeared attaching hematopoietic cells to the underlying immortalized mesenchymal cells. Furthermore, weekly differential-quick staining of the cells indicated the presence of monoblasts, monocytes, macrophages and lymphocytes in the co-cultures. At three weeks of co-culture, flow cytometry analysis showed an increased surface expression of CD172a, CD14, CD163, CD169, CD4 and CD8 up to 37±0.8%, 40±8%, 41±4%, 23±3% and 19±5% of the hematopoietic cells, respectively. In conclusion, continuous mesenchymal cell

Culture materials affect ex vivo expansion of hematopoietic progenitor cells.

PubMed

LaIuppa, J A; McAdams, T A; Papoutsakis, E T; Miller, W M

1997-09-05

Ex vivo expansion of hematopoietic cells is important for applications such as cancer treatment, gene therapy, and transfusion medicine. While cell culture systems are widely used to evaluate the biocompatibility of materials for implantation, the ability of materials to support proliferation of primary human cells in cultures for reinfusion into patients has not been addressed. We screened a variety of commercially available polymer (15 types), metal (four types), and glass substrates for their ability to support expansion of hematopoietic cells when cultured under conditions that would be encountered in a clinical setting. Cultures of peripheral blood (PB) CD34+ cells and mononuclear cells (MNC) were evaluated for expansion of total cells and colony-forming unit-granulocyte monocyte (CFU-GM; progenitors committed to the granulocyte and/or monocyte lineage). Human hematopoietic cultures in serum-free medium were found to be extremely sensitive to the substrate material. The only materials tested that supported expansion at or near the levels of polystyrene were tissue culture polystyrene, Teflon perfluoroalkoxy, Teflon fluorinated ethylene propylene, cellulose acetate, titanium, new polycarbonate, and new polymethylpentene. MNC were less sensitive to the substrate materials than the primitive CD34+ progenitors, although similar trends were seen for expansion of the two cell populations on the substrates tested. CFU-GM expansion was more sensitive to substrate materials than was total cell expansion. The detrimental effects of a number of the materials on hematopoietic cultures appear to be caused by protein adsorption and/or leaching of toxins. Factors such as cleaning, sterilization, and reuse significantly affected the performance of some materials as culture substrates. We also used PB CD34+ cell cultures to examine the biocompatibility of gas-permeable cell culture and blood storage bags and several types of tubing commonly used with biomedical equipment

Airway structural cells regulate TLR5-mediated mucosal adjuvant activity.

PubMed

Van Maele, L; Fougeron, D; Janot, L; Didierlaurent, A; Cayet, D; Tabareau, J; Rumbo, M; Corvo-Chamaillard, S; Boulenouar, S; Jeffs, S; Vande Walle, L; Lamkanfi, M; Lemoine, Y; Erard, F; Hot, D; Hussell, T; Ryffel, B; Benecke, A G; Sirard, J-C

2014-05-01

Antigen-presenting cell (APC) activation is enhanced by vaccine adjuvants. Most vaccines are based on the assumption that adjuvant activity of Toll-like receptor (TLR) agonists depends on direct, functional activation of APCs. Here, we sought to establish whether TLR stimulation in non-hematopoietic cells contributes to flagellin's mucosal adjuvant activity. Nasal administration of flagellin enhanced T-cell-mediated immunity, and systemic and secretory antibody responses to coadministered antigens in a TLR5-dependent manner. Mucosal adjuvant activity was not affected by either abrogation of TLR5 signaling in hematopoietic cells or the presence of flagellin-specific, circulating neutralizing antibodies. We found that flagellin is rapidly degraded in conducting airways, does not translocate into lung parenchyma and stimulates an early immune response, suggesting that TLR5 signaling is regionalized. The flagellin-specific early response of lung was regulated by radioresistant cells expressing TLR5 (particularly the airway epithelial cells). Flagellin stimulated the epithelial production of a small set of mediators that included the chemokine CCL20, which is known to promote APC recruitment in mucosal tissues. Our data suggest that (i) the adjuvant activity of TLR agonists in mucosal vaccination may require TLR stimulation of structural cells and (ii) harnessing the effect of adjuvants on epithelial cells can improve mucosal vaccines.

Graft-versus-host disease is independent of innate signaling pathways triggered by pathogens in host hematopoietic cells.

PubMed

Li, Hongmei; Matte-Martone, Catherine; Tan, Hung Sheng; Venkatesan, Srividhya; McNiff, Jennifer; Demetris, Anthony J; Jain, Dhanpat; Lakkis, Fadi; Rothstein, David; Shlomchik, Warren D

2011-01-01

Graft-versus-host disease (GVHD) is initiated by APCs that prime alloreactive donor T cells. In antipathogen responses, Ag-bearing APCs receive signals through pattern-recognition receptors, including TLRs, which induce the expression of costimulatory molecules and production of inflammatory cytokines, which in turn mold the adaptive T cell response. However, in allogeneic hematopoietic stem cell transplantation (alloSCT), there is no specific pathogen, alloantigen is ubiquitous, and signals that induce APC maturation are undefined. To investigate APC activation in GVHD, we used recipient mice with hematopoietic cells genetically deficient in pathways critical for APC maturation in models in which host APCs are absolutely required. Strikingly, CD8-mediated and CD4-mediated GVHD were similar whether host APCs were wild-type or deficient in MyD88, TRIF, or MyD88 and TRIF, which excludes essential roles for TLRs and IL-1β, the key product of inflammasome activation. Th1 differentiation was if anything augmented when APCs were MyD88/TRIF(-/-), and T cell production of IFN-γ did not require host IL-12. GVHD was also intact when APCs lacked the type I IFNR, which amplifies APC activation pathways that induce type I IFNs. Thus in GVHD, alloreactive T cells can be activated when pathways critical for antipathogen T cell responses are impaired.

Generating Human Hematopoietic Stem Cells In Vitro: Exploring Endothelial To Hematopoietic Transition As A Portal For Stemness Acquisition

PubMed Central

Slukvin, Igor I.

2016-01-01

Advances in cellular reprogramming technologies have created alternative platforms for the production of blood cells, either through inducing pluripotency in somatic cells or by way of direct conversion of non-hematopoietic cells into blood cells. However, de novo generation of hematopoietic stem cells (HSCs) with robust and sustained multilineage engraftment potential remains a significant challenge. Hemogenic endothelium (HE) has been recognized as a unique transitional stage of blood development from mesoderm at which HSCs arise in certain embryonic locations. The major aim of this review is to summarize historical perspectives and recent advances in the investigation of endothelial-hematopoietic transition (EHT) and HSC formation in the context of aiding in vitro approaches to instruct HSC fate from human pluripotent stem cells. In addition, direct conversion of somatic cells to blood and HSCs and progression of this conversion through HE stage are discussed. A thorough understanding of the intrinsic and microenvironmental regulators of EHT that lead to the acquisition of self-renewal potential by emerging blood cells, is essential to advance the technologies for HSC production and expansion. PMID:27391301

Donor Dependent Variations in Hematopoietic Differentiation among Embryonic and Induced Pluripotent Stem Cell Lines

PubMed Central

Féraud, Olivier; Valogne, Yannick; Melkus, Michael W.; Zhang, Yanyan; Oudrhiri, Noufissa; Haddad, Rima; Daury, Aurélie; Rocher, Corinne; Larbi, Aniya; Duquesnoy, Philippe; Divers, Dominique; Gobbo, Emilie; Brunet de la Grange, Philippe; Louache, Fawzia; Bennaceur-Griscelli, Annelise; Mitjavila-Garcia, Maria Teresa

2016-01-01

Hematopoiesis generated from human embryonic stem cells (ES) and induced pluripotent stem cells (iPS) are unprecedented resources for cell therapy. We compared hematopoietic differentiation potentials from ES and iPS cell lines originated from various donors and derived them using integrative and non-integrative vectors. Significant differences in differentiation toward hematopoietic lineage were observed among ES and iPS. The ability of engraftment of iPS or ES-derived cells in NOG mice varied among the lines with low levels of chimerism. iPS generated from ES cell-derived mesenchymal stem cells (MSC) reproduce a similar hematopoietic outcome compared to their parental ES cell line. We were not able to identify any specific hematopoietic transcription factors that allow to distinguish between good versus poor hematopoiesis in undifferentiated ES or iPS cell lines. There is a relatively unpredictable variation in hematopoietic differentiation between ES and iPS cell lines that could not be predicted based on phenotype or gene expression of the undifferentiated cells. These results demonstrate the influence of genetic background in variation of hematopoietic potential rather than the reprogramming process. PMID:26938212

A model for hematopoietic death in man from irradiation of bone marrow during radioimmunotherapy.

PubMed

Scott, B R; Dillehay, L E

1990-11-01

There are numerous institutions worldwide performing clinical trials of radioimmunotherapy (RIT) for cancer. For RIT, an exponentially decaying radionuclide is attached by using a chelating agent to a specific monoclonal or polyclonal tumour antibody (e.g. antiferritin IgG). The major limitation to RIT is toxicity to normal tissue in organs other than the one containing the tumour (e.g. bone marrow). The focus of this manuscript is on modelling the risk (or probability) of hematopoietic death in man for exponentially decaying patterns of high-energy beta irradiation (e.g. 90Y) of bone marrow by radioimmunoglobulin injected into the blood. The analytical solutions presented are only applicable to protocols for which significant uptake of radioactivity by the bone marrow does not occur, and only for high energy beta emitters. However, the generic equation used to obtain the analytical solutions is applicable to any continuous pattern of high energy beta irradiation. A model called the "normalized dose model" was used to generate calculated values for the LD50 as a function of the effective half-time for the radioimmunoglobulin in the blood. A less complicated empirical model was used to describe the calculated values. This model is presumed to be valid for effective half-times in blood of up to about 20 days. For longer effective half-times, the LD50 can be estimated using the normalized-dose model presented. In this manuscript, we also provide a modified Weibull model that allows estimation of the risk of hematopoietic death for single or multiple injections (in one cycle) of radioimmunoglobulin, for patients with normal susceptibility to irradiation and for patients with heightened susceptibility. With the modified Weibull model, the risk of hematopoietic death depends on the level of medical treatment provided to mitigate radiation injuries.

Embryonic hematopoiesis in vertebrate somites gives rise to definitive hematopoietic stem cells

PubMed Central

Qiu, Juhui; Fan, Xiaoying; Wang, Yixia; Jin, Hongbin; Song, Yixiao; Han, Yang; Huang, Shenghong; Meng, Yaping; Tang, Fuchou; Meng, Anming

2016-01-01

Hematopoietic stem cells (HSCs) replenish all types of blood cells. It is debating whether HSCs in adults solely originate from the aorta-gonad-mesonephros (AGM) region, more specifically, the dorsal aorta, during embryogenesis. Here, we report that somite hematopoiesis, a previously unwitnessed hematopoiesis, can generate definitive HSCs (dHSCs) in zebrafish. By transgenic lineage tracing, we found that a subset of cells within the forming somites emigrate ventromedially and mix with lateral plate mesoderm-derived primitive hematopoietic cells before the blood circulation starts. These somite-derived hematopoietic precursors and stem cells (sHPSCs) subsequently enter the circulation and colonize the kidney of larvae and adults. RNA-seq analysis reveals that sHPSCs express hematopoietic genes with sustained expression of many muscle/skeletal genes. Embryonic sHPSCs transplanted into wild-type embryos expand during growth and survive for life time with differentiation into various hematopoietic lineages, indicating self-renewal and multipotency features. Therefore, the embryonic origin of dHSCs in adults is not restricted to the AGM. PMID:27252540

Hematopoietic stem cells: can old cells learn new tricks?

PubMed

Ho, Anthony D; Punzel, Michael

2003-05-01

Since the establishment of cell lines derived from human embryonic stem (ES) cells, it has been speculated that out of such "raw material," we could some day produce all sorts of replacement parts for the human body. Human pluripotent stem cells can be isolated from embryonic, fetal, or adult tissues. Enormous self-renewal capacity and developmental potential are the characteristics of ES cells. Somatic stem cells, especially those derived from hematopoietic tissues, have also been reported to exhibit developmental potential heretofore not considered possible. The initial evidences for the plasticity potential of somatic stem cells were so encouraging that the opponents of ES cell research used them as arguments for restricting ES cell research. In the past months, however, critical issues have been raised challenging the validity and the interpretation of the initial data. Whereas hematopoietic stem-cell therapy has been a clinical reality for almost 40 years, there is still a long way to go in basic research before novel therapy strategies with stem cells as replacement for other organ systems can be established. Given the present status, we should keep all options open for research in ES cells and adult stem cells to appreciate the complexity of their differentiation pathways and the relative merits of various types of stem cells for regenerative medicine.

Melanoma Stem Cells and Metastasis: Mimicking Hematopoietic Cell Trafficking?

PubMed Central

Lee, Nayoung; Barthel, Steven R.; Schatton, Tobias

2014-01-01

Malignant melanoma is a highly metastatic cancer that bears responsibility for the majority of skin cancer-related deaths. Amidst the research efforts to better understand melanoma progression, there has been increasing evidence that hints at a role for a subpopulation of virulent cancer cells, termed malignant melanoma stem or initiating cells (MMICs), in metastasis formation. MMICs are characterized by their preferential ability to initiate and propagate tumor growth and their selective capacity for self-renewal and differentiation into less tumorigenic melanoma cells. The frequency of MMICs has been shown to correlate with poor clinical prognosis in melanoma. Additionally, MMICs are enriched among circulating tumor cells (CTCs) in the peripheral blood of cancer patients, suggesting that MMICs may be a critical player in the metastatic cascade. Although these links exist between MMICs and metastatic disease, the mechanisms by which MMICs may advance metastatic progression are only beginning to be elucidated. Recent studies have shown that MMICs express molecules critical for hematopoietic cell maintenance and trafficking, providing a possible explanation for how circulating MMICs could drive melanoma dissemination. We therefore propose that MMICs might fuel melanoma metastasis by exploiting homing mechanisms commonly utilized by hematopoietic cells. Here we review the biological properties of MMICs and the existing literature on their metastatic potential. We will discuss possible mechanisms by which MMICs might initiate metastases in the context of established knowledge of cancer stem cells (CSCs) in other cancers and of hematopoietic homing molecules, with a particular focus on selectins, integrins, chemokines, and chemokine receptors known to be expressed by melanoma cells. Biological understanding of how these molecules might be utilized by MMICs to propel the metastatic cascade could critically impact the development of more effective therapies for advanced

HIERARCHICAL ORGANIZATION OF OSTEOBLASTS REVEALS THE SIGNIFICANT ROLE OF CD166 IN HEMATOPOIETIC STEM CELL MAINTANANCE AND FUNCTION

PubMed Central

Chitteti, Brahmananda R.; Cheng, Ying-Hua; Kacena, Melissa A.; Srour, Edward F.

2013-01-01

The role of osteoblasts (OB) in maintaining hematopoietic stem cells (HSC) in their niche is well elucidated, but the exact definition, both phenotypically and hierarchically of OB responsible for these functions is not clearly known. We previously demonstrated that OB maturational status influences HSC function whereby immature OB with high Runx2 expression promote hematopoietic expansion. Here, we show that Activated Leukocyte Cell Adhesion Molecule (ALCAM) or CD166 expression on OB is directly correlated with Runx2 expression and high hematopoiesis enhancing activity (HEA). Fractionation of OB with lineage markers: Sca1, osteopontin (OPN), CD166, CD44, and CD90 revealed that Lin-Sca1-OPN+CD166+ cells (CD166+) and their subpopulations fractionated with CD44 and CD90 expressed high levels of Runx2 and low levels of osteocalcin (OC) demonstrating the relatively immature status of these cells. Conversely, the majority of the Lin-Sca1-OPN+CD166− cells (CD166−) expressed high OC levels suggesting that CD166− OB are more mature. In vitro hematopoietic potential of LSK cells co-cultured for 7 days with fresh OB or OB pre-cultured for 1, 2, or 3 weeks declined precipitously with increasing culture duration concomitant with loss of CD166 expression. Importantly, LSK cells co-cultured with CD166+CD44+CD90+ OB maintained their in vivo repopulating potential through primary and secondary transplantation, suggesting that robust HEA activity is best mediated by immature CD166+ OB with high Runx2 and low OC expression. These studies begin to define the hierarchical organization of osteoblastic cells and provide a more refined definition of OB that can mediate HEA. PMID:23369988

Subregional localization and characterization of Ly6aGFP-expressing hematopoietic cells in the mouse embryonic head.

PubMed

Li, Zhuan; Vink, Chris S; Mariani, Samanta A; Dzierzak, Elaine

2016-08-01

Hematopoietic cell generation in the midgestation mouse embryo occurs through the natural transdifferentiation of temporally and spatially restricted set of hemogenic endothelial cells. These cells take on hematopoietic fate in the aorta, vitelline and umbilical arteries and appear as hematopoietic cell clusters that emerge from the vascular wall. Genetic and live imaging data have supported this. Recently, the embryonic head has been shown to contain fully functional hematopoietic stem cells (HSC). By lineage tracing, cerebrovascular specific endothelial cells were shown to contribute to the postnatal mouse hematopoietic system. Since Ly6aGFP is a marker of all HSCs, some hematopoietic cluster cells and hemogenic endothelial cells in the midgestation mouse aorta, we examine here whether embryonic head HSCs and vascular endothelial cells are positive for this marker. Whereas some head vasculature, single hematopoietic cells and all HSCs are Ly6aGFP expressing, we do not find clusters of hematopoietic cells emerging from the cerebrovasculature that are characteristic of endothelial-to-hematopoietic transition. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Engraftment and reconstitution of hematopoiesis is dependent on VEGFR2 mediated regeneration of sinusoidal endothelial cells

PubMed Central

Hooper, Andrea T.; Butler, Jason M.; Nolan, Daniel J; Kranz, Andrea; Iida, Kaoruko; Kobayashi, Mariko; Kopp, Hans-Georg; Shido, Koji; Petit, Isabelle; Yanger, Kilangsungla; James, Daylon; Witte, Larry; Zhu, Zhenping; Wu, Yan; Pytowski, Bronislaw; Rosenwaks, Zev; Mittal, Vivek; Sato, Thomas N.; Rafii, Shahin

2011-01-01

SUMMARY The phenotypic attributes and molecular determinants for the regeneration of bone marrow (BM) sinusoidal endothelial cells (SECs) and their contribution to hematopoiesis are unknown. We show that after myelosuppression VEGFR2 activation promotes reassembly of regressed SECs, reconstituting hematopoietic stem and progenitor cells (HSPCs). VEGFR2 and VEGFR3 expression are restricted to BM vasculature, demarcating a continuous network of VEGFR2+VEGFR3+Sca1− SECs and VEGFR2+VEGFR3−Sca1+ arterioles. While chemotherapy (5FU) and sublethal irradiation (650 rad) induce minor SEC regression, lethal irradiation (950 rad) induces severe regression of SECs requiring BM transplantation (BMT) for regeneration. Conditional deletion of VEGFR2 in adult mice blocks regeneration of SECs in sublethally irradiated animals, preventing hematopoietic reconstitution. Inhibition of VEGFR2 signaling in lethally irradiated wild type mice rescued with BMT severely impairs SEC reconstruction, preventing engraftment and reconstitution of HSPCs. Therefore, activation of VEGFR2 is critical for regeneration of VEGFR3+Sca1− SECs that are essential for engraftment and restoration of HSPCs and hematopoiesis. PMID:19265665

The Gottingen minipig is a model of the hematopoietic acute radiation syndrome: G-colony stimulating factor stimulates hematopoiesis and enhances survival from lethal total-body γ-irradiation.

PubMed

Moroni, Maria; Ngudiankama, Barbara F; Christensen, Christine; Olsen, Cara H; Owens, Rossitsa; Lombardini, Eric D; Holt, Rebecca K; Whitnall, Mark H

2013-08-01

We are characterizing the Gottingen minipig as an additional large animal model for advanced drug testing for the acute radiation syndrome (ARS) to enhance the discovery and development of novel radiation countermeasures. Among the advantages provided by this model, the similarities to human hematologic parameters and dynamics of cell loss/recovery after irradiation provide a convenient means to compare the efficacy of drugs known to affect bone marrow cellularity and hematopoiesis. Male Gottingen minipigs, 4 to 5 months old and weighing 9 to 11 kg, were used for this study. We tested the standard off-label treatment for ARS, rhG-CSF (Neupogen, 10 μg/kg/day for 17 days), at the estimated LD70/30 total-body γ-irradiation (TBI) radiation dose for the hematopoietic syndrome, starting 24 hours after irradiation. The results indicated that granulocyte colony stimulating factor (G-CSF) enhanced survival, stimulated recovery from neutropenia, and induced mobilization of hematopoietic progenitor cells. In addition, the administration of G-CSF resulted in maturation of monocytes/macrophages. These results support continuing efforts toward validation of the minipig as a large animal model for advanced testing of radiation countermeasures and characterization of the pathophysiology of ARS, and they suggest that the efficacy of G-CSF in improving survival after total body irradiation may involve mechanisms other than increasing the numbers of circulating granulocytes. Published by Elsevier Inc.

The Hematopoietic Stem Cell Therapy for Exploration of Space

NASA Astrophysics Data System (ADS)

Ohi, S.

Departments of Biochemistry &Molecular Biology, Genetics &Human Genetics, Pediatrics &Child Long-duration space missions require countermeasures against severe/invasive disorders in astronauts that are caused by space environments, such as hematological/cardiac abnormalities, bone/muscle losses, immunodeficiency, neurological disorders, and cancer. Some, if not all, of these disorders may be amenable to hematopoietic stem cell therapy and gene therapy. Growing evidence indicates that hematopoietic stem cells (HSCs) possess extraordinary plasticity to differentiate not only to all types of blood cells but also to various tissues, including bone, muscle, skin, liver and neuronal cells. Therefore, our working hypothesis is that the hematopoietic stem cell-based therapy, herein called as the hematopoietic stem cell therapy (HSCT), might provide countermeasure/prevention for hematological abnormalities, bone and muscle losses in space, thereby maintaining astronauts' homeostasis. Our expertise lies in recombinant adeno-associated virus (rAAV)-mediated gene therapy for the hemoglobinopathies, -thalassemia and sickle cell disease (Ohi S, Kim BC, J Pharm Sci 85: 274-281, 1996; Ohi S, et al. Grav Space Biol Bull 14: 43, 2000). As the requisite steps in this protocol, we established procedures for purification of HSCs from both mouse and human bone marrow in 1 G. Furthermore, we developed an easily harvestable, long-term liquid suspension culture system, which lasts more than one year, for growing/expanding HSCs without stromal cells. Human globin cDNAs/gene were efficiently expressed from the rAAVs in the mouse HSCs in culture. Additionally, the NASA Rotating Wall Vessel (RWV) culture system is being optimized for the HSC growth/expansion. Thus, using these technologies, the above hypothesis is being investigated by the ground-based experiments as follows: 1) -thalassemic mice (C57BL/6-Hbbth/Hbbth, Hbd-minor) are transplanted with normal isologous HSCs to correct the

Protein Tyrosine Phosphatase PRL2 Mediates Notch and Kit Signals in Early T Cell Progenitors.

PubMed

Kobayashi, Michihiro; Nabinger, Sarah C; Bai, Yunpeng; Yoshimoto, Momoko; Gao, Rui; Chen, Sisi; Yao, Chonghua; Dong, Yuanshu; Zhang, Lujuan; Rodriguez, Sonia; Yashiro-Ohtani, Yumi; Pear, Warren S; Carlesso, Nadia; Yoder, Mervin C; Kapur, Reuben; Kaplan, Mark H; Daniel Lacorazza, Hugo; Zhang, Zhong-Yin; Liu, Yan

2017-04-01

The molecular pathways regulating lymphoid priming, fate, and development of multipotent bone marrow hematopoietic stem and progenitor cells (HSPCs) that continuously feed thymic progenitors remain largely unknown. While Notch signal is indispensable for T cell specification and differentiation, the downstream effectors are not well understood. PRL2, a protein tyrosine phosphatase that regulates hematopoietic stem cell proliferation and self-renewal, is highly expressed in murine thymocyte progenitors. Here we demonstrate that protein tyrosine phosphatase PRL2 and receptor tyrosine kinase c-Kit are critical downstream targets and effectors of the canonical Notch/RBPJ pathway in early T cell progenitors. While PRL2 deficiency resulted in moderate defects of thymopoiesis in the steady state, de novo generation of T cells from Prl2 null hematopoietic stem cells was significantly reduced following transplantation. Prl2 null HSPCs also showed impaired T cell differentiation in vitro. We found that Notch/RBPJ signaling upregulated PRL2 as well as c-Kit expression in T cell progenitors. Further, PRL2 sustains Notch-mediated c-Kit expression and enhances stem cell factor/c-Kit signaling in T cell progenitors, promoting effective DN1-DN2 transition. Thus, we have identified a critical role for PRL2 phosphatase in mediating Notch and c-Kit signals in early T cell progenitors. Stem Cells 2017;35:1053-1064. © 2016 AlphaMed Press.

Hematopoietic Stem Cell Transplantation in Primary Immunodeficiency Patients in the Black Sea Region of Turkey.

PubMed

Yıldıran, Alişan; Çeliksoy, Mehmet Halil; Borte, Stephan; Güner, Şükrü Nail; Elli, Murat; Fışgın, Tunç; Özyürek, Emel; Sancak, Recep; Oğur, Gönül

2017-12-01

Hematopoietic stem cell transplantation is a promising curative therapy for many combined primary immunodeficiencies and phagocytic disorders. We retrospectively reviewed pediatric cases of patients diagnosed with primary immunodeficiencies and scheduled for hematopoietic stem cell transplantation. We identified 22 patients (median age, 6 months; age range, 1 month to 10 years) with various diagnoses who received hematopoietic stem cell transplantation. The patient diagnoses included severe combined immunodeficiency (n=11), Chediak-Higashi syndrome (n=2), leukocyte adhesion deficiency (n=2), MHC class 2 deficiency (n=2), chronic granulomatous syndrome (n=2), hemophagocytic lymphohistiocytosis (n=1), Wiskott-Aldrich syndrome (n=1), and Omenn syndrome (n=1). Of the 22 patients, 7 received human leukocyte antigen-matched related hematopoietic stem cell transplantation, 12 received haploidentical hematopoietic stem cell transplantation, and 2 received matched unrelated hematopoietic stem cell transplantation. The results showed that 5 patients had graft failure. Fourteen patients survived, yielding an overall survival rate of 67%. Screening newborn infants for primary immunodeficiency diseases may result in timely administration of hematopoietic stem cell transplantation.

[Clinical significance of autologous transplantation with hematopoietic stem cells in leukemia and solid tumors].

PubMed

Hinterberger, W; Adler, V; Bauer, K; Haberhauer, G; Habertheuer, K H; Höniger, S; Huber, K; Kier, P; Kittel, E; Ruckser, R

1995-01-01

Autologous Transplantation of hematopoietic tissue with frozen hematopoietic stem cells is increasingly used for leukemias and lymphomas, but also for some solid tumors. In the past, autotransplants have been performed with bone marrow as the source of hematopoietic stem cells. Circulating, blood derived hematopoietic stem cells, however, allow safe engraftment of all cell lines after supralethal chemo-radiotherapy. This survey describes the role of autologous stem cell transplantation in disorders that are currently in the center of clinical and scientific interest. This estimation is based on the proportion of protocols dealing with, and centering on, autologous stem cell transplantation in the context of treatment for leukemias and solid tumors ("Oncodisc", "PDQ").

Intravenous infusion of apoptotic cells simultaneously with allogeneic hematopoietic grafts alters anti-donor humoral immune responses.

PubMed

Perruche, Sylvain; Kleinclauss, François; Bittencourt, Marcelo de Carvalho; Paris, Dominique; Tiberghien, Pierre; Saas, Philippe

2004-08-01

Intravenous infusion of apoptotic donor or third-party leukocytes simultaneously with an allogeneic donor bone marrow (BM) graft favors engraftment across major histocompatibility barriers. While verifying that such apoptotic cell infusion might not also be associated with antibody (Ab)-mediated allo-immune responses, we found, rather strikingly, that apoptotic cell infusion could in fact successfully prevent a humoral allo-immunization against a BM graft in mice. Indeed, among recipients having rejected their BM graft, prior apoptotic cell infusion was associated with a near absence of Ab-mediated allo-responses, while such an immunization was frequently observed in the absence of apoptotic cell infusion. This was also observed when infusing host apoptotic cells, thus showing that the prevention of immunization was linked to the apoptotic state of the cells rather than mediated by residual anti-recipient activity. In vivo anti-transforming growth factor-beta (TGF-beta) treatment resulted in the loss of this apoptotic cell infusion-associated protective effect on humoral allo-responses. Further studies will determine whether apoptotic cell infusion, in addition to hematopoietic graft facilitation might also contribute to preventing deleterious Ab-mediated allo-responses in various transplantation settings.

Bone marrow adipocytes as negative regulators of the hematopoietic microenvironment

PubMed Central

Naveiras, Olaia; Nardi, Valentina; Wenzel, Pamela L.; Fahey, Frederic; Daley, George Q.

2009-01-01

Osteoblasts and endothelium constitute functional niches that support hematopoietic stem cells (HSC) in mammalian bone marrow (BM) 1,2,3 . Adult BM also contains adipocytes, whose numbers correlate inversely with the hematopoietic activity of the marrow. Fatty infiltration of hematopoietic red marrow follows irradiation or chemotherapy and is a diagnostic feature in biopsies from patients with marrow aplasia 4. To explore whether adipocytes influence hematopoiesis or simply fill marrow space, we compared the hematopoietic activity of distinct regions of the mouse skeleton that differ in adiposity. By flow cytometry, colony forming activity, and competitive repopulation assay, HSCs and short-term progenitors are reduced in frequency in the adipocyte-rich vertebrae of the mouse tail relative to the adipocyte-free vertebrae of the thorax. In lipoatrophic A-ZIP/F1 “fatless” mice, which are genetically incapable of forming adipocytes8, and in mice treated with the PPARγ inhibitor Bisphenol-A-DiGlycidyl-Ether (BADGE), which inhibits adipogenesis9, post-irradiation marrow engraftment is accelerated relative to wild type or untreated mice. These data implicate adipocytes as predominantly negative regulators of the bone marrow microenvironment, and suggest that antagonizingmarrow adipogenesis may enhance hematopoietic recovery in clinical bone marrow transplantation. PMID:19516257

Hematopoietic stem cell transplantation for HIV cure.

PubMed

Kuritzkes, Daniel R

2016-02-01

The apparent cure of an HIV-infected person following hematopoietic stem cell transplantation (HSCT) from an allogeneic donor homozygous for the ccr5Δ32 mutation has stimulated the search for strategies to eradicate HIV or to induce long-term remission without requiring ongoing antiretroviral therapy. A variety of approaches, including allogeneic HSCT from CCR5-deficient donors and autologous transplantation of genetically modified hematopoietic stem cells, are currently under investigation. This Review covers the experience with HSCT in HIV infection to date and provides a survey of ongoing work in the field. The challenges of developing HSCT for HIV cure in the context of safe, effective, and convenient once-daily antiretroviral therapy are also discussed.

Chromosome Instability Underlies Hematopoietic Stem Cell Dysfunction and Lymphoid Neoplasia Associated with Impaired Fbw7-Mediated Cyclin E Regulation

PubMed Central

Siu, Ka Tat; Xu, Yanfei; Swartz, Kelsey L.; Bhattacharyya, Mitra; Gurbuxani, Sandeep; Hua, Youjia

2014-01-01

The Fbw7 ubiquitin ligase critically regulates hematopoietic stem cell (HSC) function, though the precise contribution of individual substrate ubiquitination pathways to HSC homeostasis is unknown. In the work reported here, we used a mouse model in which we introduced two knock-in mutations (T74A and T393A [changes of T to A at positions 74 and 393]) to disrupt Fbw7-dependent regulation of cyclin E, its prototypic substrate, and to examine the consequences of cyclin E dysregulation for HSC function. Serial transplantation revealed that cyclin ET74A T393A HSCs self-renewed normally; however, we identified defects in their multilineage reconstituting capacity. By inducing hematologic stress, we exposed an impaired self-renewal phenotype in cyclin E knock-in HSCs that was associated with defective cell cycle exit and the emergence of chromosome instability (CIN). Importantly, p53 deletion induced both defects in self-renewal and multilineage reconstitution in cyclin E knock-in HSCs with serial transplantation and CIN in hematopoietic stem and progenitor cells. Moreover, CIN was a feature of fatal T-cell malignancies that ultimately developed in recipients of cyclin ET74A T393A; p53-null HSCs. Together, our findings demonstrate the importance of Fbw7-dependent cyclin E control to the hematopoietic system and highlight CIN as a characteristic feature of HSC dysfunction and malignancy induced by deregulated cyclin E. PMID:24958101

Vitamin D deficiency in children and adolescents submitted to hematopoietic stem cell transplantation.

PubMed

Campos, Denise Johnsson; Biagini, Gleyne Lopes Kujew; Funke, Vaneuza Araujo Moreira; Bonfim, Carmem Maria Sales; Boguszewski, César Luiz; Borba, Victória Zeghbi Cochenski

2014-03-01

Sub-optimal levels of vitamin D have been found to be highly prevalent in all age groups, with epidemiologic studies demonstrating a link between vitamin D deficiency and disease susceptibility, such as infection and cancer, and mortality rates. In adult transplant patients, it has been suggested that the immunomodulatory properties of vitamin D may have an important role in the prevention and treatment of graft-versus-host disease. The objective of this study was to assess serum 25-hydroxyvitamin D levels of children and adolescents submitted to allogeneic hematopoietic stem cell transplantation. Serum 25-hydroxyvitamin D levels of 66 patients, aged 4-20 years, were assessed at three stages: before hospitalization for hematopoietic stem cell transplantation and at 30 and 180 days after hematopoietic stem cell transplantation. The control group consisted of 25 healthy children. At the pre-hematopoietic stem cell transplantation stage, patients had lower levels of 25-hydroxyvitamin D compared to controls (25.7 ± 12.3 ng/mL vs. 31.9 ± 9.9 ng/mL; p-value = 0.01), and a higher prevalence of 25-hydroxyvitamin D deficiency (32% vs. 8%; p-value = 0.01). Prevalence increased significantly after hematopoietic stem cell transplantation (p-value = 0.01) with half of the patients having vitamin D deficiency at 180 days after transplantation. At this stage, mean serum 25-hydroxyvitamin D levels were 20.9 ± 10.9 ng/mL, a significant decline in relation to baseline (p-value = 0.01). No correlation was found between 25-hydroxyvitamin D levels and vitamin D intake, graft-versus-host disease, corticoid use or survival rates. Low levels of 25-hydroxyvitamin D were detected even before hematopoietic stem cell transplantation and were significantly lower at 180 days after hematopoietic stem cell transplantation, thus recommending vitamin D supplementation for children and adolescents submitted to hematopoietic stem cell transplantation.

A reciprocal regulatory interaction between megakaryocytes, bone cells, and hematopoietic stem cells.

PubMed

Kacena, Melissa A; Gundberg, Caren M; Horowitz, Mark C

2006-11-01

A growing body of evidence suggests that megakaryocytes (MK) or their growth factors play a role in skeletal homeostasis. MK have been shown to express and/or secrete several bone-related proteins including osteocalcin, osteonectin, bone sialoprotein, osteopontin, bone morphogenetic proteins, and osteoprotegerin. In addition, at least 3 mouse models have been described in which MK number was significantly elevated with an accompanying marked increase in bone mineral density. Mice overexpressing thrombopoietin, the major MK growth factor, have an osteosclerotic bone phenotype. Mice deficient in transcription factors GATA-1 and NF-E2, which are required for the differentiation of MK, exhibited a strikingly increased bone mass. Importantly, recent studies have demonstrated that MK can stimulate osteoblast (OB) proliferation and differentiation in vitro and that they can also inhibit osteoclast (OC) formation in vitro. These findings suggest that MK play a dual role in skeletal homeostasis by stimulating formation while simultaneously inhibiting resorption. Conversely, cells of the osteoblast lineage support hematopoiesis, including megakaryopoiesis. Postnatal hematopoiesis occurs almost solely in the bone marrow (BM), close to or on endosteal surfaces. This finding, in conjunction with the observed contact of OB with hematopoietic cells, has lead investigators to explore the molecular and cellular interactions between hematopoietic cells and cells of the OB lineage. Importantly, it has been shown that many of the cytokines that are critical for normal hematopoiesis and megakaryopoiesis are produced by OB. Indeed, culturing osteoblasts with CD34+ BM cells significantly enhances hematopoietic cell number by both enhancing the proliferation of long-term culture initiating cells and the proliferation and differentiation of MK. These data are consistent with cells in the OB lineage playing a critical role in the hematopoietic niche. Overall, these observations demonstrate

Expansion of Human and Murine Hematopoietic Stem and Progenitor Cells Ex Vivo without Genetic Modification Using MYC and Bcl-2 Fusion Proteins

PubMed Central

Bird, Gregory A.; Polsky, Avital; Estes, Patricia; Hanlon, Teri; Hamilton, Haley; Morton, John J.; Gutman, Jonathan; Jimeno, Antonio

2014-01-01

The long-term repopulating hematopoietic stem cell (HSC) population can self-renew in vivo, support hematopoiesis for the lifetime of the individual, and is of critical importance in the context of bone marrow stem cell transplantation. The mechanisms that regulate the expansion of HSCs in vivo and in vitro remain unclear to date. Since the current set of surface markers only allow for the identification of a population of cells that is highly enriched for HSC activity, we will refer to the population of cells we expand as Hematopoietic Stem and Progenitor cells (HSPCs). We describe here a novel approach to expand a cytokine-dependent Hematopoietic Stem and Progenitor Cell (HSPC) population ex vivo by culturing primary adult human or murine HSPCs with fusion proteins including the protein transduction domain of the HIV-1 transactivation protein (Tat) and either MYC or Bcl-2. HSPCs obtained from either mouse bone marrow, human cord blood, human G-CSF mobilized peripheral blood, or human bone marrow were expanded an average of 87 fold, 16.6 fold, 13.6 fold, or 10 fold, respectively. The expanded cell populations were able to give rise to different types of colonies in methylcellulose assays in vitro, as well as mature hematopoietic populations in vivo upon transplantation into irradiated mice. Importantly, for both the human and murine case, the ex vivo expanded cells also gave rise to a self-renewing cell population in vivo, following initial transplantation, that was able to support hematopoiesis upon serial transplantation. Our results show that a self-renewing cell population, capable of reconstituting the hematopoietic compartment, expanded ex vivo in the presence of Tat-MYC and Tat-Bcl-2 suggesting that this may be an attractive approach to expand human HSPCs ex vivo for clinical use. PMID:25170611

Regulation of Hematopoietic Stem Cell Behavior by the Nanostructured Presentation of Extracellular Matrix Components

PubMed Central

Muth, Christine Anna; Steinl, Carolin; Klein, Gerd; Lee-Thedieck, Cornelia

2013-01-01

Hematopoietic stem cells (HSCs) are maintained in stem cell niches, which regulate stem cell fate. Extracellular matrix (ECM) molecules, which are an essential part of these niches, can actively modulate cell functions. However, only little is known on the impact of ECM ligands on HSCs in a biomimetic environment defined on the nanometer-scale level. Here, we show that human hematopoietic stem and progenitor cell (HSPC) adhesion depends on the type of ligand, i.e., the type of ECM molecule, and the lateral, nanometer-scaled distance between the ligands (while the ligand type influenced the dependency on the latter). For small fibronectin (FN)–derived peptide ligands such as RGD and LDV the critical adhesive interligand distance for HSPCs was below 45 nm. FN-derived (FN type III 7–10) and osteopontin-derived protein domains also supported cell adhesion at greater distances. We found that the expression of the ECM protein thrombospondin-2 (THBS2) in HSPCs depends on the presence of the ligand type and its nanostructured presentation. Functionally, THBS2 proved to mediate adhesion of HSPCs. In conclusion, the present study shows that HSPCs are sensitive to the nanostructure of their microenvironment and that they are able to actively modulate their environment by secreting ECM factors. PMID:23405094

Extramedullary Relapse Following Total Marrow and Lymphoid Irradiation in Patients Undergoing Allogeneic Hematopoietic Cell Transplantation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Ji Hyun; Stein, Anthony; Tsai, Nicole

Purpose: Approximately 5% to 20% of patients who undergo total body irradiation (TBI) in preparation for hematopoietic cell transplantation (HCT) can develop extramedullary (EM) relapse. Whereas total marrow and lymphoid irradiation (TMLI) provides a more conformally targeted radiation therapy for patients, organ sparing has the potential to place the patient at a higher risk for EM relapse than TBI. This study evaluated EM relapse in patients treated with TMLI at our institution. Methods and Materials: Patients eligible for analysis had been enrolled in 1 of 3 prospective TMLI trials between 2006 and 2012. The TMLI targeted bones, major lymph nodemore » chains, liver, spleen, testes, and brain, using image-guided tomotherapy with total dose ranging from 12 to 15 Gy. Results: A total of 101 patients with a median age of 47 years were studied. The median follow-up was 12.8 months. Incidence of EM relapse and bone marrow (BM) relapse were 12.9% and 25.7%, respectively. Of the 13 patients who had EM relapse, 4 also had BM relapse, and 7 had EM disease prior to HCT. There were a total of 19 EM relapse sites as the site of initial recurrence: 11 soft tissue, 6 lymph node, 2 skin. Nine of these sites were within the target region and received ≥12 Gy. Ten initial EM relapse sites were outside of the target region: 5 sites received 10.1 to 11.4 Gy while 5 sites received <10 Gy. Pretransplantation EM was the only significant predictor of subsequent EM relapse. The cumulative incidence of EM relapse was 4% at 1 year and 11.4% at 2 years. Conclusions: EM relapse incidence was as frequent in regions receiving ≥10 Gy as those receiving <10 Gy. EM relapse rates following TMLI that included HCT regimens were comparable to published results with regimens including TBI and suggest that TMLI is not associated with an increased EM relapse risk.« less

Rutin-Enriched Extract from Coriandrum sativum L. Ameliorates Ionizing Radiation-Induced Hematopoietic Injury

PubMed Central

Han, Xiaodan; Xue, Xiaolei; Zhao, Yu; Li, Yuan; Liu, Weili; Zhang, Junling; Fan, Saijun

2017-01-01

Hematopoietic injury is a major cause of mortality in radiation accidents and a primary side effect in patients undergoing radiotherapy. Ionizing radiation (IR)-induced myelosuppression is largely attributed to the injury of hematopoietic stem and progenitor cells (HSPCs). Coriander is a culinary herb with multiple pharmacological effects and has been widely used in traditional medicine. In this study, flavonoids were identified as the main component of coriander extract with rutin being the leading compound (rutin-enriched coriander extract; RE-CE). We evaluated the radioprotective effect of RE-CE against IR-induced HSPCs injury. Results showed that RE-CE treatment markedly improved survival, ameliorated organ injuries and myelosuppression, elevated HSPCs frequency, and promoted differentiation and proliferation of HSPCs in irradiated mice. The protective role of RE-CE in hematopoietic injury is probably attributed to its anti-apoptotic and anti-DNA damage effect in irradiated HSPCs. Moreover, these changes were associated with reduced reactive oxygen species (ROS) and enhanced antioxidant enzymatic activities in irradiated HSPCs. Collectively, these findings demonstrate that RE-CE is able to ameliorate IR-induced hematopoietic injury partly by reducing IR-induced oxidative stress. PMID:28468251

Short Stat5-Interacting Peptide Derived from Phospholipase C-β3 Inhibits Hematopoietic Cell Proliferation and Myeloid Differentiation

PubMed Central

Yasudo, Hiroki; Ando, Tomoaki; Xiao, Wenbin; Kawakami, Yuko; Kawakami, Toshiaki

2011-01-01

Constitutive activation of the transcription factor Stat5 in hematopoietic stem/progenitor cells leads to various hematopoietic malignancies including myeloproliferative neoplasm (MPN). Our recent study found that phospholipase C (PLC)-β3 is a novel tumor suppressor involved in MPN, lymphoma and other tumors. Stat5 activity is negatively regulated by the SH2 domain-containing protein phosphatase SHP-1 in a PLC-β3-dependent manner. PLC-β3 can form the multimolecular SPS complex together with SHP-1 and Stat5. The close physical proximity of SHP-1 and Stat5 brought about by interacting with the C-terminal segment of PLC-β3 (PLC-β3-CT) accelerates SHP-1-mediated dephosphorylation of Stat5. Here we identify the minimal sequences within PLC-β3-CT required for its tumor suppressor function. Two of the three Stat5-binding noncontiguous regions, one of which also binds SHP-1, substantially inhibited in vitro proliferation of Ba/F3 cells. Surprisingly, an 11-residue Stat5-binding peptide (residues 988-998) suppressed Stat5 activity in Ba/F3 cells and in vivo proliferation and myeloid differentiation of hematopoietic stem/progenitor cells. Therefore, this study further defines PLC-β3-CT as the Stat5- and SHP-1-binding domain by identifying minimal functional sequences of PLC-β3 for its tumor suppressor function and implies their potential utility in the control of hematopoietic malignancies. PMID:21949826

Adult hematopoietic stem cells lacking Hif-1α self-renew normally.

PubMed

Vukovic, Milica; Sepulveda, Catarina; Subramani, Chithra; Guitart, Amélie V; Mohr, Jasmine; Allen, Lewis; Panagopoulou, Theano I; Paris, Jasmin; Lawson, Hannah; Villacreces, Arnaud; Armesilla-Diaz, Alejandro; Gezer, Deniz; Holyoake, Tessa L; Ratcliffe, Peter J; Kranc, Kamil R

2016-06-09

The hematopoietic stem cell (HSC) pool is maintained under hypoxic conditions within the bone marrow microenvironment. Cellular responses to hypoxia are largely mediated by the hypoxia-inducible factors, Hif-1 and Hif-2. The oxygen-regulated α subunits of Hif-1 and Hif-2 (namely, Hif-1α and Hif-2α) form dimers with their stably expressed β subunits and control the transcription of downstream hypoxia-responsive genes to facilitate adaptation to low oxygen tension. An initial study concluded that Hif-1α is essential for HSC maintenance, whereby Hif-1α-deficient HSCs lost their ability to self-renew in serial transplantation assays. In another study, we demonstrated that Hif-2α is dispensable for cell-autonomous HSC maintenance, both under steady-state conditions and following transplantation. Given these unexpected findings, we set out to revisit the role of Hif-1α in cell-autonomous HSC functions. Here we demonstrate that inducible acute deletion of Hif-1α has no impact on HSC survival. Notably, unstressed HSCs lacking Hif-1α efficiently self-renew and sustain long-term multilineage hematopoiesis upon serial transplantation. Finally, Hif-1α-deficient HSCs recover normally after hematopoietic injury induced by serial administration of 5-fluorouracil. We therefore conclude that despite the hypoxic nature of the bone marrow microenvironment, Hif-1α is dispensable for cell-autonomous HSC maintenance. © 2016 by The American Society of Hematology.

[Protective effects of WR2721 on early bone marrow hematopoietic function in mice exposed to 6.5 Gy of (60)Co γ-rays].

PubMed

Deng, Zi-Liang; Zhang, Liu-Zhen; Cong, Yue; Liu, Xiao-Lan; Yu, Zu-Ying; Shan, Ya-Jun; Cui, Yu; Wang, Li-Mei; Xing, Shuang; Cong, Yu-Wen; Luo, Qing-Liang

2014-06-01

The aim of this study was to investigate the effect of WR2721(amifostine) against bone marrow hematopoietic damage of mice exposed to 6.5 Gy of (60)Co-γ ray. A total of 60 C57/BL6J mice was divided into 3 groups:normal group (mice were injected with physiological salt solution), irradiation group (mice were injected with physiologic salt solution before irradiation) and WR2721 group (mice were injected with WR2721 before irradiation). The WBC, neutrophil (Neut), Plt and RBC levels in peripheral blood of 3 group mice were counted within 60 days after irradiation; the bone marrow nuclear cells (BMNC) were counted at 2 and 24 hours after irradiation; the hematopoietic stem/progenitor cell (LK/LSK) level and colony formation capability were detected by flow cytometry at 2 and 24 hours after irradiation. The results indicated that the counts of WBC and neut at 4 and 18 days, Plt at 7-18 days and RBC at 10-30 day after irradiation in WR2721 group were higher than those in irradiation group (P < 0.05); the BMNC, LSK and LK levels obviously increased at 24 hours after irradiation (P < 0.05), the CFU-GEMM, CFU-GM, CFU-MK BFU-E and CFU-E all significantly increased at 2 and 24 hours after irradiation (P < 0.01), as compared with irradiation group. It is concluded that WR2721 can effectively alleviate early hematopoietic damage and promote the fast recovery of peripheral blood cells in mice exposed to γ-ray, suggesting that the WR2721 has significant radioprotective effect on hematopoietic system.

Allometric Scaling of the Active Hematopoietic Stem Cell Pool across Mammals

PubMed Central

Dingli, David; Pacheco, Jorge M.

2006-01-01

Background Many biological processes are characterized by allometric relations of the type Y = Y 0 Mb between an observable Y and body mass M, which pervade at multiple levels of organization. In what regards the hematopoietic stem cell pool, there is experimental evidence that the size of the hematopoietic stem cell pool is conserved in mammals. However, demands for blood cell formation vary across mammals and thus the size of the active stem cell compartment could vary across species. Methodology/Principle Findings Here we investigate the allometric scaling of the hematopoietic system in a large group of mammalian species using reticulocyte counts as a marker of the active stem cell pool. Our model predicts that the total number of active stem cells, in an adult mammal, scales with body mass with the exponent ¾. Conclusion/Significance The scaling predicted here provides an intuitive justification of the Hayflick hypothesis and supports the current view of a small active stem cell pool supported by a large, quiescent reserve. The present scaling shows excellent agreement with the available (indirect) data for smaller mammals. The small size of the active stem cell pool enhances the role of stochastic effects in the overall dynamics of the hematopoietic system. PMID:17183646

Ability of circulating human hematopoietic lineage negative cells to support hematopoiesis.

PubMed

Peris, Pilar; Roforth, Matthew M; Nicks, Kristy M; Fraser, Daniel; Fujita, Koji; Jilka, Robert L; Khosla, Sundeep; McGregor, Ulrike

2015-01-01

Hematopoietic stem cell (HSC) self-renewal is regulated by osteoblast and/or endothelial cells within the hematopoietic niche. However, the true identity of the supporting cells and the nature of the secreted factors remain uncertain. We developed a novel mouse model and analyzed whether circulating human peripheral hematopoietic lineage negative/AP+ (lin-/AP+) cells support hematopoiesis in vivo. Thus, immunocompromised (Rag) mice expressing thymidine kinase (Tk) under the control of the 3.6Col1α1 promoter (Tk-Rag) were treated with ganciclovir, resulting in osteoblast progenitor cell ablation and subsequent loss of hematopoiesis (evaluated by measuring mouse Ter119+ erythroid cells). Following hematopoietic cell depletion, human bone marrow-derived marrow stromal cells (MSCs) or lin-/AP+ cells were infused into Tk-Rag mice and compared with saline infusions. Ganciclovir significantly reduced (7.4-fold) Ter119+ cells in the bone marrow of Tk-Rag mice compared to saline injections. Infusion of either MSCs or lin-/AP+ cells into ganciclovir-treated mice resulted in a 3.3-fold and 2.7-fold increase (P < 0.01), respectively, in Ter119+ cells compared to mice receiving saline. Relative to lin-/AP- cells, lin-/AP+ cells expressed high levels of mesenchymal, endothelial, and hematopoiesis supporting genes. Thus, human peripheral blood lin-/AP+ cells represent a novel cell type capable of supporting hematopoiesis in a manner comparable to MSCs. © 2014 Wiley Periodicals, Inc.

The effect of lithium on hematopoietic, mesenchymal and neural stem cells.

PubMed

Ferensztajn-Rochowiak, Ewa; Rybakowski, Janusz K

2016-04-01

Lithium has been used in modern psychiatry for more than 65 years, constituting a cornerstone for the long-term treatment of bipolar disorder. A number of biological properties of lithium have been discovered, including its hematological, antiviral and neuroprotective effects. In this article, a systematic review of the effect of lithium on hematopoietic, mesenchymal and neural stem cells is presented. The beneficial effects of lithium on the level of hematopoietic stem cells (HSC) and growth factors have been reported since 1970s. Lithium improves homing of stem cells, the ability to form colonies and HSC self-renewal. Lithium also exerts a favorable influence on the proliferation and maintenance of mesenchymal stem cells (MSC). Studies on the effect of lithium on neurogenesis have indicated an increased proliferation of progenitor cells in the dentate gyrus of the hippocampus and enhanced mitotic activity of Schwann cells. This may be connected with the neuroprotective and neurotrophic effects of lithium, reflected in an improvement in synaptic plasticity promoting cell survival and inhibiting apoptosis. In clinical studies, lithium treatment increases cerebral gray matter, mainly in the frontal lobes, hippocampus and amygdala. Recent findings also suggest that lithium may reduce the risk of dementia and exert a beneficial effect in neurodegenerative diseases. The most important mediators and signaling pathways of lithium action are the glycogen synthase kinase-3 and Wnt/β-catenin pathways. Recently, to study of bipolar disorder pathogenesis and the mechanism of lithium action, the induced pluripotent stem cells (iPSC) obtained from bipolar patients have been used. Copyright © 2015 Institute of Pharmacology, Polish Academy of Sciences. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.

Epigenomics in hematopoietic transplantation: novel treatment strategies.

PubMed

Engel, Nicole; Rank, Andreas

2011-10-01

Allogeneic hematopoietic stem cell transplantation is a high risk but curative treatment option for leukemia, myelodysplasia and other hematological malignancies. After high dose radio- or chemo-therapy, recipient's hematopoiesis is replaced by a new immunosystem and residual malignant cells are eliminated by the graft-versus-leukemia reaction. The benefit of this immunological effect is limited by the most frequent complication of hematopoietic stem cell transplantation: graft-versus-host disease. In addition to their well-known anti-tumor activity, epigenetic drugs mediate immunotolerance without reducing alloreactivity or even enhance graft-versus-leukemia effect without inducing graft-versus-host disease by regulating cytokine release, increasing the circulating number of regulatory T cells and interacting with natural killer cells. We focus on the use of epigenetic drugs in the allogeneic transplantation setting in relation to their anti-tumor and immunomodulatory potential.

Clonal precursor of bone, cartilage, and hematopoietic niche stromal cells

PubMed Central

Chan, Charles K. F.; Lindau, Paul; Jiang, Wen; Chen, James Y.; Zhang, Lillian F.; Chen, Ching-Cheng; Seita, Jun; Sahoo, Debashis; Kim, Jae-Beom; Lee, Andrew; Park, Sujin; Nag, Divya; Gong, Yongquan; Kulkarni, Subhash; Luppen, Cynthia A.; Theologis, Alexander A.; Wan, Derrick C.; DeBoer, Anthony; Seo, Eun Young; Vincent-Tompkins, Justin D.; Loh, Kyle; Walmsley, Graham G.; Kraft, Daniel L.; Wu, Joseph C.; Longaker, Michael T.; Weissman, Irving L.

2013-01-01

Organs are composites of tissue types with diverse developmental origins, and they rely on distinct stem and progenitor cells to meet physiological demands for cellular production and homeostasis. How diverse stem cell activity is coordinated within organs is not well understood. Here we describe a lineage-restricted, self-renewing common skeletal progenitor (bone, cartilage, stromal progenitor; BCSP) isolated from limb bones and bone marrow tissue of fetal, neonatal, and adult mice. The BCSP clonally produces chondrocytes (cartilage-forming) and osteogenic (bone-forming) cells and at least three subsets of stromal cells that exhibit differential expression of cell surface markers, including CD105 (or endoglin), Thy1 [or CD90 (cluster of differentiation 90)], and 6C3 [ENPEP glutamyl aminopeptidase (aminopeptidase A)]. These three stromal subsets exhibit differential capacities to support hematopoietic (blood-forming) stem and progenitor cells. Although the 6C3-expressing subset demonstrates functional stem cell niche activity by maintaining primitive hematopoietic stem cell (HSC) renewal in vitro, the other stromal populations promote HSC differentiation to more committed lines of hematopoiesis, such as the B-cell lineage. Gene expression analysis and microscopic studies further reveal a microenvironment in which CD105-, Thy1-, and 6C3-expressing marrow stroma collaborate to provide cytokine signaling to HSCs and more committed hematopoietic progenitors. As a result, within the context of bone as a blood-forming organ, the BCSP plays a critical role in supporting hematopoiesis through its generation of diverse osteogenic and hematopoietic-promoting stroma, including HSC supportive 6C3(+) niche cells. PMID:23858471

Proliferation of multipotent hematopoietic cells controlled by a truncated erythropoietin receptor transgene.

PubMed Central

Kirby, S L; Cook, D N; Walton, W; Smithies, O

1996-01-01

The long-term efficacy of gene therapy using bone marrow transplantation requires the engraftment of genetically altered totipotent hematopoietic stem cells (THSCs). Ex vivo expansion of corrected THSCs is one way to increase the efficiency of the procedure. Similarly, selective in vivo expansion of the therapeutic THSCs rather than the endogenous THSCs could favor the transplant. To test whether a conferred proliferative advantage gene can facilitate the in vitro and in vivo expansion of hematopoietic stem cells, we have generated transgenic mice expressing a truncated receptor for the growth factor erythropoietin. These mice are phenotypically normal, but when treated in vivo with exogenous erythropoietin they exhibit a marked increase in multipotent, clonogenic hematopoietic cells [colony-forming units in the spleen (CFU-S) and CFUs that give rise to granulocytes, erythroid cells, macrophages, and megakaryocytes within the same colony (CFU-GEMM)] in comparison with the wild-type mice. In addition, long-term in vitro culture of tEpoR transgenic bone marrow in the presence of erythropoietin induces exponential expansion of trilineage hematopoietic stem cells not seen with wild-type bone marrow. Thus, the truncated erythropoietin receptor gene shows promise as a means for obtaining cytokine-inducible hematopoietic stem cell proliferation to facilitate the direct targeting of THSCs and to provide a competitive repopulation advantage for transplanted therapeutic stem cells. Images Fig. 3 PMID:8790342

Comparison of Cyclophosphamide Combined with Total Body Irradiation, Oral Busulfan, or Intravenous Busulfan for Allogeneic Hematopoietic Cell Transplantation in Adults with Acute Lymphoblastic Leukemia.

PubMed

Mitsuhashi, Kenjiro; Kako, Shinichi; Shigematsu, Akio; Atsuta, Yoshiko; Doki, Noriko; Fukuda, Takahiro; Kanamori, Heiwa; Onizuka, Makoto; Takahashi, Satoshi; Ozawa, Yukiyasu; Kurokawa, Mineo; Inoue, Yoshiko; Nagamura-Inoue, Tokiko; Morishima, Yasuo; Mizuta, Shuichi; Tanaka, Junji

2016-12-01

We conducted a retrospective analysis to compare outcomes in adult patients with acute lymphoblastic leukemia (ALL) who underwent allogeneic hematopoietic cell transplantation (allo-HCT) with conditioning regimens containing cyclophosphamide (CY) in combination with total body irradiation (TBI), oral busulfan (p.o. BU), or intravenous busulfan (i.v. BU). We used data for January 2000 to December 2012 from the Transplant Registry Unified Management Program of the Japan Society of Hematopoietic Cell Transplantation. We identified 2130 patients treated with TBI/CY (n = 2028), p.o. BU/CY (n = 60), or i.v. BU/CY (n = 42). Two-year overall survival (OS) and 2-year relapse-free survival rates were 69.0% and 62.1%, respectively, in the TBI/CY group, 55.9% and 54.2% in the p.o. BU/CY group, and 71.0% and 46.8% in the i.v. BU/CY group. In multivariate analysis, compared with TBI/CY, p.o. BU/CY, but not i.v. BU/CY, was associated with lower OS (hazard ratio [HR], 1.46; P = .047) and a higher incidence of sinusoidal obstruction syndrome (HR, 3.36; P = .030). No between-group differences were seen in the incidence of nonrelapse mortality, relapse, acute graft-versus-host disease (GVHD), or chronic GVHD. We suggest that i.v. BU/CY might be a possible alternative allo-HCT conditioning regimen for adults with ALL who are not suitable for TBI. Copyright © 2016 The American Society for Blood and Marrow Transplantation. Published by Elsevier Inc. All rights reserved.

Gadd45a deletion aggravates hematopoietic stem cell dysfunction in ATM-deficient mice.

PubMed

Chen, Yulin; Yang, Runan; Guo, Peng; Ju, Zhenyu

2014-01-01

Ataxia telangiectasia mutated (ATM) kinase plays an essential role in the maintenance of genomic stability. ATM-deficient (ATM(-/-)) mice exhibit hematopoietic stem cell (HSC) dysfunction and a high incidence of lymphoma. Gadd45a controls cell cycle arrest, apoptosis and DNA repair, and is involved in the ATM-p53 mediated DNA damage response. However, the role of Gadd45a in regulating the functionality of ATM(-/-) HSCs is unknown. Here we report that Gadd45a deletion did not rescue the defects of T-cells and B-cells development in ATM(-/-) mice. Instead, ATM and Gadd45a double knockout (ATM(-/-) Gadd45a(-/-)) HSCs exhibited an aggravated defect in long-term self-renewal capacity compared to ATM(-/-) HSCs in HSC transplantation experiments. Further experiments revealed that the aggravated defect of ATM(-/-) Gadd45a(-/-) HSCs was due to a reduction of cell proliferation, associated with an accumulation of DNA damage and subsequent activation of DNA damage response including an up-regulation of p53-p21 signaling pathway. Additionally, ATM(-/-) Gadd45a(-/-) mice showed an increased incidence of hematopoietic malignancies, as well as an increased rate of metastasis than ATM(-/-) mice. In conclusion, Gadd45a deletion aggravated the DNA damage accumulation, which subsequently resulted in a further impaired self-renewal capacity and an increased malignant transformation in ATM(-/-) HSCs.

Enforced hematopoietic cell E- and L-selectin ligand (HCELL) expression primes transendothelial migration of human mesenchymal stem cells.

PubMed

Thankamony, Sai P; Sackstein, Robert

2011-02-08

According to the multistep model of cell migration, chemokine receptor engagement (step 2) triggers conversion of rolling interactions (step 1) into firm adhesion (step 3), yielding transendothelial migration. We recently reported that glycosyltransferase-programmed stereosubstitution (GPS) of CD44 on human mesenchymal stem cells (hMSCs) creates the E-selectin ligand HCELL (hematopoietic cell E-selectin/L-selectin ligand) and, despite absence of CXCR4, systemically administered HCELL(+)hMSCs display robust osteotropism visualized by intravital microscopy. Here we performed studies to define the molecular effectors of this process. We observed that engagement of hMSC HCELL with E-selectin triggers VLA-4 adhesiveness, resulting in shear-resistant adhesion to ligand VCAM-1. This VLA-4 activation is mediated via a Rac1/Rap1 GTPase signaling pathway, resulting in transendothelial migration on stimulated human umbilical vein endothelial cells without chemokine input. These findings indicate that hMSCs coordinately integrate CD44 ligation and integrin activation, circumventing chemokine-mediated signaling, yielding a step 2-bypass pathway of the canonical multistep paradigm of cell migration.

An Analysis of microRNA Expression in the Myelodysplastic Syndromes Using Hematopoietic Stem Cells

DTIC Science & Technology

2015-10-01

AWARD NUMBER: W81XWH-13-1-0082 TITLE: An Analysis of microRNA Expression in the Myelodysplastic Syndromes Using Hematopoietic Stem Cells ...syndromes (MDS) to identify microRNAs (miRNAs) dysregulated in MDS hematopoietic stem cells (MDS HSCs) as compared with normal HSCs. MiRNAs differentially...the age-related predisposition for the development of MDS. 15. SUBJECT TERMS MicroRNAs, the myelodysplastic syndromes, hematopoietic stem cells

An Analysis of MicroRNA Expression in the Myelodysplastic Syndromes Using Hematopoietic Stem Cells

DTIC Science & Technology

2015-10-01

AWARD NUMBER: W81XWH-13-1-0082 TITLE: An Analysis of microRNA Expression in the Myelodysplastic Syndromes Using Hematopoietic Stem Cells ...syndromes (MDS) to identify microRNAs (miRNAs) dysregulated in MDS hematopoietic stem cells (MDS HSCs) as compared with normal HSCs. MiRNAs differentially...the age-related predisposition for the development of MDS. 15. SUBJECT TERMS MicroRNAs, the myelodysplastic syndromes, hematopoietic stem cells

The Gottingen Minipig Is a Model of the Hematopoietic Acute Radiation Syndrome: G-Colony Stimulating Factor Stimulates Hematopoiesis and Enhances Survival From Lethal Total-Body γ-Irradiation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Moroni, Maria, E-mail: maria.moroni@usuhs.edu; Ngudiankama, Barbara F.; Christensen, Christine

Purpose: We are characterizing the Gottingen minipig as an additional large animal model for advanced drug testing for the acute radiation syndrome (ARS) to enhance the discovery and development of novel radiation countermeasures. Among the advantages provided by this model, the similarities to human hematologic parameters and dynamics of cell loss/recovery after irradiation provide a convenient means to compare the efficacy of drugs known to affect bone marrow cellularity and hematopoiesis. Methods and Materials: Male Gottingen minipigs, 4 to 5 months old and weighing 9 to 11 kg, were used for this study. We tested the standard off-label treatment formore » ARS, rhG-CSF (Neupogen, 10 μg/kg/day for 17 days), at the estimated LD70/30 total-body γ-irradiation (TBI) radiation dose for the hematopoietic syndrome, starting 24 hours after irradiation. Results: The results indicated that granulocyte colony stimulating factor (G-CSF) enhanced survival, stimulated recovery from neutropenia, and induced mobilization of hematopoietic progenitor cells. In addition, the administration of G-CSF resulted in maturation of monocytes/macrophages. Conclusions: These results support continuing efforts toward validation of the minipig as a large animal model for advanced testing of radiation countermeasures and characterization of the pathophysiology of ARS, and they suggest that the efficacy of G-CSF in improving survival after total body irradiation may involve mechanisms other than increasing the numbers of circulating granulocytes.« less

Stromal Progenitor Cells in Mitigation of Non-Hematopoietic Radiation Injuries

PubMed Central

Kulkarni, Shilpa; Wang, Timothy C.; Guha, Chandan

2016-01-01

Purpose of review Therapeutic exposure to high doses of radiation can severely impair organ function due to ablation of stem cells. Normal tissue injury is a dose-limiting toxicity for radiation therapy (RT). Although advances in the delivery of high precision conformal RT has increased normal tissue sparing, mitigating and therapeutic strategies that could alleviate early and chronic radiation effects are urgently needed in order to deliver curative doses of RT, especially in abdominal, pelvic and thoracic malignancies. Radiation-induced gastrointestinal injury is also a major cause of lethality from accidental or intentional exposure to whole body irradiation in the case of nuclear accidents or terrorism. This review examines the therapeutic options for mitigation of non-hematopoietic radiation injuries. Recent findings We have developed stem cell based therapies for the mitigation of acute radiation syndrome (ARS) and radiation-induced gastrointestinal syndrome (RIGS). This is a promising option because of the robustness of standardized isolation and transplantation of stromal cells protocols, and their ability to support and replace radiation-damaged stem cells and stem cell niche. Stromal progenitor cells (SPC) represent a unique multipotent and heterogeneous cell population with regenerative, immunosuppressive, anti-inflammatory, and wound healing properties. SPC are also known to secrete various key cytokines and growth factors such as platelet derived growth factors (PDGF), keratinocyte growth factor (KGF), R-spondins (Rspo), and may consequently exert their regenerative effects via paracrine function. Additionally, secretory vesicles such as exosomes or microparticles can potentially be a cell-free alternative replacing the cell transplant in some cases. Summary This review highlights the beneficial effects of SPC on tissue regeneration with their ability to (a) target the irradiated tissues, (b) recruit host stromal cells, (c) regenerate endothelium and

Hematopoietic Stem Cell Transplant in Adolescent and Young Adults With Fanconi Anemia Is Feasible With Acceptable Toxicity, With Those Surviving 100 Days Posttransplant Having Excellent Outcomes.

PubMed

Alhuraiji, Ahmad; Alzahrani, Hazza; Al Mohareb, Fahad; Chaudhri, Naeem; Alsharif, Fahad; Mohamed, Said; Rasheed, Walid; Aldawsari, Ghuzayel; Ahmed, Syed Osman; Aljurf, Mahmoud

2016-12-01

Fanconi anemia is a congenital bone marrow failure syndrome that is associated with congenital anomalies and increased risk of cancer. Hematopoietic stem cell transplant is a potentially curative modality for bone marrow failure in Fanconi anemia patients. Here, we report our center's experience regarding adolescent and young adult patients with Fanconi anemia and hematopoietic stem cell transplant. We conducted a retrospective patient record analyses of patients who presented at our center from 1988 to 2014. We included patients greater than 14 years old with confirmed Fanconi anemia based on positive chromosome breakage study and who underwent hematopoietic stem cell transplant at our institution. Our study group comprised 12 patients with Fanconi anemia who underwent hematopoietic stem cell transplant at our institution. The median age was 20 years (range, 14-31 y) with a female predominance of 83%. Low-dose cyclophosphamide (20-80 mg/kg)-based conditioning regimens were used with different combinations that included fludarabine, antithymocyte globulin, or total body irradiation. All patients had HLA-matched sibling grafts. In all patients, stem cell source was the bone marrow. All patients showed engraftment. Four patients (33%) developed acute graft-versus-host disease. Three patients (25%) died early before day 100 after hematopoietic stem cell transplant due to infectious complications, with 1 patient having steroid refractory acute graft-versus-host disease. Overall survival was 75% at a median follow-up of 43 months. All patients who survived are well and remained transfusion independent without evidence of secondary malignancy. Our findings support the feasibility of reduced intensity conditioning allogeneic hematopoietic stem cell transplant in older and more heavily pretreated patients with Fanconi anemia, especially for those who are engrafted.

miR-146a deficiency in hematopoietic cells is not involved in the development of atherosclerosis.

PubMed

Del Monte, Alberto; Arroyo, Ana B; Andrés-Manzano, María J; García-Barberá, Nuria; Caleprico, María S; Vicente, Vicente; Roldán, Vanessa; González-Conejero, Rocío; Martínez, Constantino; Andrés, Vicente

2018-01-01

Atherosclerosis involves activation of the IRAK1/TRAF6/NF-κB inflammatory cascade, which is negatively regulated by miR146a. Previous studies showed that the TT genotype of rs2431697, located near the miR-146a gene, drives lower miR-146a transcription and predicts adverse cardiovascular events in anticoagulated atrial fibrillation patients. Moreover, systemic miR-146a administration protects mice from atherosclerosis. Here we evaluated the ability of miR-146a expression in the hematopoietic component to regulate atherosclerosis in low-density lipoprotein receptor-null mice (Ldlr-/-). Lethally-irradiated Ldlr-/- mice transplanted with bone marrow from wild-type or miR-146a-null mice were fed an atherogenic diet for 8 and 20 weeks. Irak1, Traf6 and MIR146A expression were quantified in thoracic aorta by qRT-PCR and Western blot. Aortic plaque size and composition were characterized by Oil-Red staining and immunohistochemistry and leukocyte recruitment by intravital microscopy. Blood cell counts were similar in fat-fed Ldlr-/-mice with or without hematopoietic miR-146a expression. However, plasma cholesterol decreased in fat-fed Ldlr-/-mice transplanted with bone marrow deficient for miR-146a. Finally, aortic atherosclerosis burden and recruitment of leukocytes into the vessel wall were undistinguishable between the two groups, despite higher levels of Irak1 and Traf6 mRNA and protein in the aorta of fat-fed mice lacking hematopoietic miR-146a expression. miR-146a deficiency exclusively in hematopoietic cells modulates cholesterol levels in plasma and the expression of its targets in the artery wall of fat-fed Ldlr-/- mice, but does not accelerate atherosclerosis. Atheroprotection upon systemic miR-146a administration may therefore be caused by specific effects on vascular cells.

Hematopoietic stem cell fate through metabolic control.

PubMed

Ito, Kyoko; Ito, Keisuke

2018-05-25

Hematopoietic stem cells (HSCs) maintain a quiescent state in the bone marrow to preserve their self-renewal capacity, but also undergo cell divisions as required. Organelles such as the mitochondria sustain cumulative damage during these cell divisions, and this damage may eventually compromise the cells' self-renewal capacity. HSC divisions result in either self-renewal or differentiation, with the balance between the two directly impacting hematopoietic homeostasis; but the heterogeneity of available HSC-enriched fractions, together with the technical challenges of observing HSC behavior, has long hindered the analysis of individual HSCs, and prevented the elucidation of this process. However, recent advances in genetic models, metabolomics analyses and single-cell approaches have revealed the contributions made to HSC self-renewal by metabolic cues, mitochondrial biogenesis, and autophagy/mitophagy, which have highlighted mitochondrial quality as a key control factor in the equilibrium of HSCs. A deeper understanding of precisely how specific modes of metabolism control HSC fate at the single cell level is therefore not only of great biological interest, but will have clear clinical implications for the development of therapies for hematological disease. Copyright © 2018. Published by Elsevier Inc.

Aging, Clonality and Rejuvenation of Hematopoietic Stem Cells

PubMed Central

Akunuru, Shailaja; Geiger, Hartmut

2016-01-01

Aging is associated with reduced organ function and increased disease incidence. Hematopoietic stem cell (HSC) aging driven by both cell intrinsic and extrinsic factors is linked to impaired HSC self-renewal and regeneration, aging-associated immune remodeling, and increased leukemia incidence. Compromised DNA damage responses and increased production of reactive oxygen species have been previously causatively attributed to HSC aging. However, recent paradigm-shifting concepts such as global epigenetic and cytoskeletal polarity shifts, cellular senescence, as well as clonal selection of HSCs upon aging provide new insights into HSC aging mechanisms. Rejuvenating agents that can reprogram the epigenetic status of aged HSCs or senolytic drugs that selectively deplete senescent cells provide promising translational avenues for attenuating hematopoietic aging and potentially, alleviating aging-associated immune remodeling and myeloid malignancies. PMID:27380967

Wnt5a Regulates Hematopoietic Stem Cell Proliferation and Repopulation Through the Ryk Receptor

PubMed Central

Povinelli, Benjamin J.; Nemeth, Michael J.

2017-01-01

Proper regulation of the balance between hematopoietic stem cell (HSC) proliferation, self-renewal, and differentiation is necessary to maintain hematopoiesis throughout life. The Wnt family of ligands has been implicated as critical regulators of these processes through a network of signaling pathways. Previously, we have demonstrated that the Wnt5a ligand can induce HSC quiescence through a noncanonical Wnt pathway, resulting in an increased ability to reconstitute hematopoiesis. In this study, we tested the hypothesis that the Ryk protein, a Wnt ligand receptor that can bind the Wnt5a ligand, regulated the response of HSCs to Wnt5a. We observed that inhibiting Ryk blocked the ability of Wnt5a to induce HSC quiescence and enhance short-term and long-term hematopoietic repopulation. We found that Wnt5a suppressed production of reactive oxygen species, a known inducer of HSC proliferation. The ability of Wnt5a to inhibit ROS production was also regulated by Ryk. From these data, we propose that Wnt5a regulates HSC quiescence and hematopoietic repopulation through the Ryk receptor and that this process is mediated by suppression of reactive oxygen species. PMID:23939973

Wnt5a regulates hematopoietic stem cell proliferation and repopulation through the Ryk receptor.

PubMed

Povinelli, Benjamin J; Nemeth, Michael J

2014-01-01

Proper regulation of the balance between hematopoietic stem cell (HSC) proliferation, self-renewal, and differentiation is necessary to maintain hematopoiesis throughout life. The Wnt family of ligands has been implicated as critical regulators of these processes through a network of signaling pathways. Previously, we have demonstrated that the Wnt5a ligand can induce HSC quiescence through a noncanonical Wnt pathway, resulting in an increased ability to reconstitute hematopoiesis. In this study, we tested the hypothesis that the Ryk protein, a Wnt ligand receptor that can bind the Wnt5a ligand, regulated the response of HSCs to Wnt5a. We observed that inhibiting Ryk blocked the ability of Wnt5a to induce HSC quiescence and enhance short-term and long-term hematopoietic repopulation. We found that Wnt5a suppressed production of reactive oxygen species, a known inducer of HSC proliferation. The ability of Wnt5a to inhibit ROS production was also regulated by Ryk. From these data, we propose that Wnt5a regulates HSC quiescence and hematopoietic repopulation through the Ryk receptor and that this process is mediated by suppression of reactive oxygen species. © 2013 AlphaMed Press.

An Overview of Kidney Disease Following Hematopoietic Cell Transplantation.

PubMed

Ando, Minoru

2018-06-01

Hematopoietic stem cell transplantation (SCT) recipients are exposed to a large amount of anti-cancer drugs, immunosuppressors, and irradiation during the peri-SCT period. Thus, they have to overcome serious adverse events related to unavoidable but toxic procedures, including organ disorders. In particular, acute kidney injury (AKI) is one of the most critical complications, because it influences the mortality of patients. A few patients who survive AKI may develop nephrotic syndrome, and precedent AKI is also closely associated with chronic and progressive loss of the renal function in post-SCT patients. These kidney diseases place a heavy burden on SCT patients, both medically and economically. Therefore, hematologists who evaluate SCT should be fully aware of the development of these kidney diseases after SCT. We herein review the common course of kidney disease development following allogeneic SCT to provide healthcare professionals with practical information on renal disease in SCT patients.

Hematopoietic Kit Deficiency, rather than Lack of Mast Cells, Protects Mice from Obesity and Insulin Resistance.

PubMed

Gutierrez, Dario A; Muralidhar, Sathya; Feyerabend, Thorsten B; Herzig, Stephan; Rodewald, Hans-Reimer

2015-05-05

Obesity, insulin resistance, and related pathologies are associated with immune-mediated chronic inflammation. Kit mutant mice are protected from diet-induced obesity and associated co-morbidities, and this phenotype has previously been attributed to their lack of mast cells. We performed a comprehensive metabolic analysis of Kit-dependent Kit(W/Wv) and Kit-independent Cpa3(Cre/+) mast-cell-deficient mouse strains, employing diet-induced or genetic (Lep(Ob/Ob) background) models of obesity. Our results show that mast cell deficiency, in the absence of Kit mutations, plays no role in the regulation of weight gain or insulin resistance. Moreover, we provide evidence that the metabolic phenotype observed in Kit mutant mice, while independent of mast cells, is immune regulated. Our data underscore the value of definitive mast cell deficiency models to conclusively test the involvement of this enigmatic cell in immune-mediated pathologies and identify Kit as a key hematopoietic factor in the pathogenesis of metabolic syndrome. Copyright © 2015 Elsevier Inc. All rights reserved.

Large-scale multiplex polymerase chain reaction assay for diagnosis of viral reactivations after allogeneic hematopoietic stem cell transplantation.

PubMed

Inazawa, Natsuko; Hori, Tsukasa; Hatakeyama, Naoki; Yamamoto, Masaki; Yoto, Yuko; Nojima, Masanori; Suzuki, Nobuhiro; Shimizu, Norio; Tsutsumi, Hiroyuki

2015-08-01

Viral reactivations following hematopoietic stem cell transplantation are thought to result from the breakdown of both cell-mediated and humoral immunity. As a result, many viruses could be reactivated individually or simultaneously. Using a multiplex polymerase chain reaction (PCR), we prospectively examined many kinds of viral DNAs at a time in 105 patients who underwent allogeneic hematopoietic stem cell transplantation. In total, 591 whole blood samples were collected weekly from pre- to 42 days post-transplantation and the following 13 viruses were tested; herpes simplex virus 1 (HSV-1), HSV-2, varicella-zoster virus (VZV), Epstein-Barr virus (EBV), cytomegalovirus (CMV), human herpes virus 6 (HHV-6), HHV-7, HHV-8, adenovirus, BK virus (BKV), JC virus (JCV), parvovirus B19, and hepatitis B virus (HBV). Several viral DNAs were detected in 12 patients before hematopoietic stem cell transplantation. The detection rate gradually increased after transplantation and peaked at 21 days. The most frequently detected virus was HHV-6 (n = 63; 60.0%), followed by EBV (n = 11; 10.5%), CMV (n = 11; 10.5%), and HHV-7 (n = 9; 8.6%). Adenovirus and HBV were each detected in one patient (1.0%). Detection of HHV-6 DNA was significantly more common among patients undergoing cord blood transplantation or with steroid treatment. EBV DNA tended to be more common in patients treated with anti-thymocyte globulin. Multiplex PCR was useful for detecting many viral reactivations after hematopoietic stem cell transplantation, simultaneously. Cord blood transplantation, steroid treatment, or anti-thymocyte globulin use was confirmed to be risk factors after transplantation. © 2015 Wiley Periodicals, Inc.

Consequences of irradiation on bone and marrow phenotypes, and its relation to disruption of hematopoietic precursors

PubMed Central

Green, Danielle E.; Rubin, Clinton T.

2014-01-01

The rising levels of radiation exposure, specifically for medical treatments and accidental exposures, have added great concern for the long term risks of bone fractures. Both the bone marrow and bone architecture are devastated following radiation exposure. Even sub-lethal doses cause a deficit to the bone marrow microenvironment, including a decline in hematopoietic cells, and this deficit occurs in a dose dependent fashion. Certain cell phenotypes though are more susceptible to radiation damage, with mesenchymal stem cells being more resilient than the hematopoietic stem cells. The decline in total bone marrow hematopoietic cells is accompanied with elevated adipocytes into the marrow cavity, thereby inhibiting hematopoiesis and recovery of the bone marrow microenvironment. Poor bone marrow is also associated with a decline in bone architectural quality. Therefore, the ability to maintain the bone marrow microenvironment would hinder much of the trabecular bone loss caused by radiation exposure, ultimately decreasing some comorbidities in patients exposed to radiation. PMID:24607941

Coordination of receptor signaling in multiple hematopoietic cell lineages by the adaptor protein SLP-76.

PubMed

Jordan, Martha S; Koretzky, Gary A

2010-04-01

The adaptor protein SLP-76 is expressed in multiple hematopoietic lineages including T cells, platelets, and neutrophils. SLP-76 mediated signaling is dependent on its multiple protein interaction domains, as it creates a scaffold on which key signaling complexes are built. SLP-76 is critical for supporting signaling downstream of both immunoreceptors and integrins. The signaling molecules used both upstream and downstream of SLP-76 are similar among these receptors and across cell types; however, important differences exist. Appreciating how SLP-76 coordinates signal transduction across different cell and receptor types provides insights into the complex interplay of pathways critical for activation of cells of the immune system that are essential for host defense.

B Cell allogeneic responses after hematopoietic cell transplantation: is it time to address this issue?

PubMed

Perruche, Sylvain; Kleinclauss, François; Tiberghien, Pierre; Saas, Philippe

2005-02-15

To date, B cell responses have retained less attention than T, natural killer or dendritic cell responses in the alloreactive conflict after allogeneic hematopoietic cell transplantation (HCT). Here, we discuss recent clinical and experimental data supporting a role of allogeneic B cell responses in graft-host interactions after HCT. We report results in a murine model of reduced intensity conditioning transplantation (RICT) showing that host B cells can be involved in chronic graft-versus-host disease occurrence. We also describe the control of antidonor alloresponses by intravenous simultaneous infusion of apoptotic cells with allogeneic hematopoietic grafts.

Transduction of Human Primitive Repopulating Hematopoietic Cells With Lentiviral Vectors Pseudotyped With Various Envelope Proteins

PubMed Central

Kim, Yoon-Sang; Wielgosz, Matthew M; Hargrove, Phillip; Kepes, Steven; Gray, John; Persons, Derek A; Nienhuis, Arthur W

2010-01-01

Lentiviral vectors are useful for transducing primitive hematopoietic cells. We examined four envelope proteins for their ability to mediate lentiviral transduction of mobilized human CD34+ peripheral blood cells. Lentiviral particles encoding green fluorescent protein (GFP) were pseudotyped with the vesicular stomatitis virus envelope glycoprotein (VSV-G), the amphotropic (AMPHO) murine leukemia virus envelope protein, the endogenous feline leukemia viral envelope protein or the feline leukemia virus type C envelope protein. Because the relative amount of genome RNA per ml was similar for each pseudotype, we transduced CD34+ cells with a fixed volume of each vector preparation. Following an overnight transduction, CD34+ cells were transplanted into immunodeficient mice which were sacrificed 12 weeks later. The average percentages of engrafted human CD45+ cells in total bone marrow were comparable to that of the control, mock-transduced group (37–45%). Lenti-particles pseudotyped with the VSV-G envelope protein transduced engrafting cells two- to tenfold better than particles pseudotyped with any of the γ-retroviral envelope proteins. There was no correlation between receptor mRNA levels for the γ-retroviral vectors and transduction efficiency of primitive hematopoietic cells. These results support the use of the VSV-G envelope protein for the development of lentiviral producer cell lines for manufacture of clinical-grade vector. PMID:20372106

An Analysis of microRNA Expression in the Myelodysplastic Syndromes Using Hematopoietic Stem Cells

DTIC Science & Technology

2014-08-01

AWARD NUMBER: W81XWH-13-1-0082 TITLE: An Analysis of microRNA Expression in the Myelodysplastic Syndromes Using Hematopoietic Stem Cells ... Hematopoietic Stem Cells 5b. GRANT NUMBER W81XWH-13-1-0082 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S) 5d. PROJECT NUMBER Dr. Stephen Chung 5e. TASK...in MDS hematopoietic stem cells (MDS HSCs) as compared with normal HSCs. MiRNAs differentially expressed between MDS HSCs and normal HSCs overlapped

Hematopoietic cell transplantation and HIV cure: where we are and what next?

PubMed

Zou, Shimian; Glynn, Simone; Kuritzkes, Daniel; Shah, Monica; Cook, Nakela; Berliner, Nancy

2013-10-31

The report of the so-called Berlin patient cured of HIV with hematopoietic stem cell transplantation and a few other studies raised tremendous hope, excitement, and curiosity in the field. The National Heart, Lung and Blood Institute of the National Institutes of Health convened a Working Group to address emerging heart, lung, and blood research priorities related to HIV infection. Hematopoietic cells could contribute to HIV cure through allogeneic or autologous transplantation of naturally occurring or engineered cells with anti-HIV moieties. Protection of central memory T cells from HIV infection could be a critical determinant of achieving a functional cure. HIV cure can only be achieved if the virus is eradicated from reservoirs in resting T cells and possibly other hematopoietic cells. The Working Group recommended multidisciplinary efforts leveraging HIV and cell therapy expertise to answer the critical need to support research toward an HIV cure.

Regulation of HDL on hematopoietic stem/progenitor cells in atherosclerosis requires SR-BI expression

PubMed Central

Gao, Mingming; Zhao, Dong; Schouteden, Sarah; Sorci-Thomas, Mary G.; Van Veldhoven, Paul P.; Eggermont, Kristel; Liu, George; Verfaillie, Catherine M.; Feng, Yingmei

2014-01-01

Objective Recently we demonstrated that scavenger receptor type BI (SR-BI), a HDL receptor, was expressed on murine hematopoietic stem/progenitor cells (HSPC) and infusion of reconstituted HDL and purified human apoA-I suppressed HSPC proliferation. We hypothesized that SR-B1 expression is required for the observed anti-proliferative effects of HDL on HSPC. Approach and Results SR-BI deficient (SR-BI−/−) mice and wild type (WT) controls were fed on chow or HFD (HFD) for 8–10 weeks. Under chow diet, a significant increase in Lin-Sca1+cKit+ cells (LSK cells, so called HSPC) was found in the BM of SR-BI−/− mice compared with WT mice. HFD induced a further expansion of CD150+CD48− LSK cells (HSCs), HSPCs, and granulocyte monocyte progenitors (GMPs) in SR-BI−/− mice. Injection of reactive oxygen species (ROS) inhibitor N-acetylcysteine attenuated HFD-induced HSPC expansion, leukocytosis and atherosclerosis in SR-BI−/− mice. ApoA-I infusion inhibited HSPC cell proliferation, Akt phosphorylation and ROS production in HSPC and plaque progression in low density lipoprotein receptor knockout (LDLr−/−) apoA-I−/− mice on HFD but had no effect on SR-BI−/− mice on HFD. Transplantation of SR-BI−/− BM cells into irradiated LDLr−/− recipients resulted in enhanced white blood cells (WBC) reconstitution, inflammatory cell production and plaque development. In patients with coronary heart disease, HDL levels were negatively correlated with WBC count and HSPC frequency in the peripheral blood. By flow cytometry, SR-BI expression was detected on human HSPC. Conclusions SR-BI plays a critical role in the HDL-mediated regulation HSPC proliferation and differentiation which is associated with atherosclerosis progression. PMID:24969774

Eosinophils from hematopoietic stem cell recipients suppress allogeneic T cell proliferation.

PubMed

Andersson, Jennie; Cromvik, Julia; Ingelsten, Madeleine; Lingblom, Christine; Andersson, Kerstin; Johansson, Jan-Erik; Wennerås, Christine

2014-12-01

Eosinophilia has been associated with less severe graft-versus-host disease (GVHD), but the underlying mechanism is unknown. We hypothesized that eosinophils diminish allogeneic T cell activation in patients with chronic GVHD. The capacity of eosinophils derived from healthy subjects and hematopoietic stem cell (HSC) transplant recipients, with or without chronic GVHD, to reduce allogeneic T cell proliferation was evaluated using a mixed leukocyte reaction. Eosinophil-mediated inhibition of proliferation was observed for the eosinophils of both healthy subjects and patients who underwent HSC transplantation. Eosinophils from patients with and without chronic GVHD were equally suppressive. Healthy eosinophils required cell-to-cell contact for their suppressive capacity, which was directed against CD4(+) T cells and CD8(+) T cells. Neither eosinophilic cationic protein, eosinophil-derived neurotoxin, indoleamine 2,3-dioxygenase, or increased numbers of regulatory T cells could account for the suppressive effect of healthy eosinophils. Real-time quantitative PCR analysis revealed significantly increased mRNA levels of the immunoregulatory protein galectin-10 in the eosinophils of both chronic GVHD patients and patients without GVHD, as compared with those from healthy subjects. The upregulation of galectin-10 expression in eosinophils from patients suggests a stimulatory effect of HSC transplantation in itself on eosinophilic galectin-10 expression, regardless of chronic GVHD status. To conclude, eosinophils from HSC transplant recipients and healthy subjects have a T cell suppressive capacity. Copyright © 2014 American Society for Blood and Marrow Transplantation. Published by Elsevier Inc. All rights reserved.

Image-guided total-marrow irradiation using helical tomotherapy in patients with multiple myeloma and acute leukemia undergoing hematopoietic cell transplantation.

PubMed

Wong, Jeffrey Y C; Rosenthal, Joseph; Liu, An; Schultheiss, Timothy; Forman, Stephen; Somlo, George

2009-01-01

Total-body irradiation (TBI) has an important role in patients undergoing hematopoietic cell transplantation (HCT), but is associated with significant toxicities. Targeted TBI using helical tomotherapy results in reduced doses to normal organs, which predicts for reduced toxicities compared with standard TBI. Thirteen patients with multiple myeloma were treated in an autologous tandem transplantation Phase I trial with high-dose melphalan, followed 6 weeks later by total-marrow irradiation (TMI) to skeletal bone. Dose levels were 10, 12, 14, and 16 Gy at 2 Gy daily/twice daily. In a separate allogeneic HCT trial, 8 patients (5 with acute myelogenous leukemia, 1 with acute lymphoblastic leukemia, 1 with non-Hodgkin's lymphoma, and 1 with multiple myeloma) were treated with TMI plus total lymphoid irradiation plus splenic radiotherapy to 12 Gy (1.5 Gy twice daily) combined with fludarabine/melphalan. For the 13 patients in the tandem autologous HCT trial, median age was 54 years (range, 42-66 years). Median organ doses were 15-65% that of the gross target volume dose. Primarily Grades 1-2 acute toxicities were observed. Six patients reported no vomiting; 9 patients, no mucositis; 6 patients, no fatigue; and 8 patients, no diarrhea. For the 8 patients in the allogeneic HCT trial, median age was 52 years (range, 24-61 years). Grades 2-3 nausea, vomiting, mucositis, and diarrhea were observed. In both trials, no Grade 4 nonhematologic toxicity was observed, and all patients underwent successful engraftment. This study shows that TMI using helical tomotherapy is clinically feasible. The reduced acute toxicities observed compare favorably with those seen with standard TBI. Initial results are encouraging and warrant further evaluation as a method to dose escalate with acceptable toxicity or to offer TBI-containing regimens to patients unable to tolerate standard approaches.

Investigating the feasibility of stem cell enrichment mediated by immobilized selectins.

PubMed

Charles, Nichola; Liesveld, Jane L; King, Michael R

2007-01-01

Hematopoietic stem cell therapy is used to treat both malignant and non-malignant diseases, and enrichment of the hematopoietic stem and progenitor cells (HSPCs) has the potential to reduce the likelihood of graft vs host disease or relapse, potentially fatal complications associated with the therapy. Current commercial HSPC isolation technologies rely solely on the CD34 surface marker, and while they have proven to be invaluable, they can be time-consuming with variable recoveries reported. We propose that selectin-mediated enrichment could prove to be a quick and effective method for recovering HSPCs from adult bone marrow (ABM) on the basis of differences in rolling velocities and independently of CD34 expression. Purified CD34+ ABM cells and the unselected CD34- ABM cells were perfused over immobilized P-, E-, and L-selectin-IgG at physiologic wall shear stresses, and rolling velocities and cell retention data were collected. CD34+ ABM cells generally exhibited lower rolling velocities and higher retention than the unselected CD34- ABM cells on all three selectins. For initial CD34+ ABM cell concentrations ranging from 1% to 5%, we predict an increase in purity ranging from 5.2% to 36.1%, depending on the selectin used. Additionally, selectin-mediated cell enrichment is not limited to subsets of cells with inherent differences in rolling velocities. CD34+ KG1a cells and CD34- HL60 cells exhibited nearly identical rolling velocities on immobilized P-selectin-IgG over the entire range of shear stresses studied. However, when anti-CD34 antibody was co-immobilized with the P-selectin-IgG, the rolling velocity of the CD34+ KG1a cells was significantly reduced, making selectin-mediated cell enrichment a feasible option. Optimal cell enrichment in immobilized selectin surfaces can be achieved within 10 min, much faster than most current commercially available systems.

The continuum of stem cell transdifferentiation: possibility of hematopoietic stem cell plasticity with concurrent CD45 expression.

PubMed

Udani, V M

2006-02-01

Recent years have seen a surge of scientific research examining adult stem cell plasticity. For example, the hematopoietic stem cell has been shown to give rise to skin, respiratory epithelium, intestinal epithelium, renal epithelium, liver parenchyma, pancreas, skeletal muscle, vascular endothelium, myocardium, and central nervous system (CNS) neurons. The potential for such stem cell plasticity seems to be enhanced by stressors such as injury and neoplasia. Interestingly, recent studies have demonstrated that hematopoietic stem cells may be able to adopt certain nonhematopoietic phenotypes, such as endothelial, neural, or skeletal muscle phenotypes, without entirely losing their initial hematopoietic identity. We propose that transdifferentiation can, in certain conditions, be a partial rather than a complete event, and we encourage further investigation into the phenomenon of a stem cell simultaneously expressing phenotypic features of two distinct cell fates.

The effect of the immunophilin ligands rapamycin and FK506 on proliferation of mast cells and other hematopoietic cell lines.

PubMed Central

Hultsch, T; Martin, R; Hohman, R J

1992-01-01

The immunosuppressive drugs FK506 and cyclosporin A have an identical spectrum of activities with respect to IgE receptor (Fc epsilon RI)-mediated exocytosis from mast cells and T cell receptor-mediated transcription of IL-2. These findings suggest a common step in receptor-mediated signal transduction leading to exocytosis and transcription and imply that immunosuppressive drugs target specific signal transduction pathways, rather than specific cell types. This hypothesis is supported by studies on the effect of rapamycin on IL-3 dependent proliferation of the rodent mast cell line PT18. Rapamycin inhibits proliferation of PT18 cells, achieving a plateau of 80% inhibition at 1 nM. This inhibition is prevented in a competitive manner by FK506, a structural analogue of rapamycin. Proliferation of rat basophilic leukemia cells and WEHI-3 cells was also inhibited, at doses comparable to those shown previously to inhibit IL-2-dependent proliferation of cytotoxic T lymphocyte line (CTLL) cells. In contrast, proliferation of A-431 cells, a epidermoid cell line, was not affected by rapamycin. DNA histograms indicate that complexes formed between the rapamycin-FK506-binding protein (FKBP) and rapamycin arrest-proliferating PT18 cells in the G0/G1-phase. It is concluded that FKBP-rapamycin complexes may inhibit proliferative signals emanating from IL-3 receptors, resulting in growth arrest of cytokine-dependent, hematopoietic cells. PMID:1384815

Differences in lymphocyte developmental potential between human embryonic stem cell and umbilical cord blood-derived hematopoietic progenitor cells.

PubMed

Martin, Colin H; Woll, Petter S; Ni, Zhenya; Zúñiga-Pflücker, Juan Carlos; Kaufman, Dan S

2008-10-01

Hematopoietic progenitor cells derived from human embryonic stem cells (hESCs) develop into diverse mature hematopoietic lineages, including lymphocytes. Whereas functional natural killer (NK) cells can be efficiently generated in vitro from hESC-derived CD34(+) cells, studies of T- and B-cell development from hESCs have been much more limited. Here, we demonstrate that despite expressing functional Notch-1, CD34(+) cells from hESCs did not derive T cells when cocultured with OP9 cells expressing Delta-like 1, or in fetal thymus organ culture. hESC-derived CD34(+) cells also did not produce B cells in vitro. In contrast, CD34(+) cells isolated from UCB routinely generated T and B cells when cultured in the same conditions. Notably, both undifferentiated hESCs, and sorted hESC-derived populations with hematopoietic developmental potential exhibited constitutive expression of ID family genes and of transcriptional targets of stem cell factor-induced signaling. These pathways both inhibit T-cell development and promote NK-cell development. Together, these results demonstrate fundamental differences between hESC-derived hematopoietic progenitors and analogous primary human cells. Therefore, hESCs can be more readily supported to differentiate into certain cell types than others, findings that have important implications for derivation of defined lineage-committed populations from hESCs.

Depleting dietary valine permits nonmyeloablative mouse hematopoietic stem cell transplantation.

PubMed

Taya, Yuki; Ota, Yasunori; Wilkinson, Adam C; Kanazawa, Ayano; Watarai, Hiroshi; Kasai, Masataka; Nakauchi, Hiromitsu; Yamazaki, Satoshi

2016-12-02

A specialized bone marrow microenvironment (niche) regulates hematopoietic stem cell (HSC) self-renewal and commitment. For successful donor-HSC engraftment, the niche must be emptied via myeloablative irradiation or chemotherapy. However, myeloablation can cause severe complications and even mortality. Here we report that the essential amino acid valine is indispensable for the proliferation and maintenance of HSCs. Both mouse and human HSCs failed to proliferate when cultured in valine-depleted conditions. In mice fed a valine-restricted diet, HSC frequency fell dramatically within 1 week. Furthermore, dietary valine restriction emptied the mouse bone marrow niche and afforded donor-HSC engraftment without chemoirradiative myeloablation. These findings indicate a critical role for valine in HSC maintenance and suggest that dietary valine restriction may reduce iatrogenic complications in HSC transplantation. Copyright © 2016, American Association for the Advancement of Science.

Sowing the Seeds of a Fruitful Harvest: Hematopoietic Stem Cell Mobilization

PubMed Central

Hoggatt, Jonathan; Speth, Jennifer M.; Pelus, Louis M.

2014-01-01

Hematopoietic stem cell transplantation is the only curative option for a number of malignant and non-malignant diseases. As the use of hematopoietic transplant has expanded, so too has the source of stem and progenitor cells. The predominate source of stem and progenitors today, particularly in settings of autologous transplantation, is mobilized peripheral blood. This review will highlight the historical advances which lead to the widespread use of peripheral blood stem cells for transplantation, with a look towards future enhancements to mobilization strategies. PMID:24123398

Fanconi anemia genes are highly expressed in primitive CD34+ hematopoietic cells

PubMed Central

Aubé, Michel; Lafrance, Matthieu; Brodeur, Isabelle; Delisle, Marie-Chantal; Carreau, Madeleine

2003-01-01

Background Fanconi anemia (FA) is a complex recessive genetic disease characterized by progressive bone marrow failure (BM) and a predisposition to cancer. We have previously shown using the Fancc mouse model that the progressive BM failure results from a hematopoietic stem cell defect suggesting that function of the FA genes may reside in primitive hematopoietic stem cells. Methods Since genes involved in stem cell differentiation and/or maintenance are usually regulated at the transcription level, we used a semiquantitative RT-PCR method to evaluate FA gene transcript levels in purified hematopoietic stem cells. Results We show that most FA genes are highly expressed in primitive CD34-positive and negative cells compared to lower levels in more differentiated cells. However, in CD34- stem cells the Fancc gene was found to be expressed at low levels while Fancg was undetectable in this population. Furthermore, Fancg expression is significantly decreased in Fancc -/- stem cells as compared to wild-type cells while the cancer susceptibility genes Brca1 and Fancd1/Brac2 are upregulated in Fancc-/- hematopoietic cells. Conclusions These results suggest that FA genes are regulated at the mRNA level, that increased Fancc expression in LTS-CD34+ cells correlates with a role at the CD34+ differentiation stage and that lack of Fancc affects the expression of other FA gene, more specifically Fancg and Fancd1/Brca2, through an unknown mechanism. PMID:12809565

Mapping gene expression patterns during myeloid differentiation using the EML hematopoietic progenitor cell line.

PubMed

Du, Yang; Campbell, Janee L; Nalbant, Demet; Youn, Hyewon; Bass, Ann C Hughes; Cobos, Everardo; Tsai, Schickwann; Keller, Jonathan R; Williams, Simon C

2002-07-01

The detailed examination of the molecular events that control the early stages of myeloid differentiation has been hampered by the relative scarcity of hematopoietic stem cells and the lack of suitable cell line models. In this study, we examined the expression of several myeloid and nonmyeloid genes in the murine EML hematopoietic stem cell line. Expression patterns for 19 different genes were examined by Northern blotting and RT-PCR in RNA samples from EML, a variety of other immortalized cell lines, and purified murine hematopoietic stem cells. Representational difference analysis (RDA) was performed to identify differentially expressed genes in EML. Expression patterns of genes encoding transcription factors (four members of the C/EBP family, GATA-1, GATA-2, PU.1, CBFbeta, SCL, and c-myb) in EML were examined and were consistent with the proposed functions of these proteins in hematopoietic differentiation. Expression levels of three markers of terminal myeloid differentiation (neutrophil elastase, proteinase 3, and Mac-1) were highest in EML cells at the later stages of differentiation. In a search for genes that were differentially expressed in EML cells during myeloid differentiation, six cDNAs were isolated. These included three known genes (lysozyme, histidine decarboxylase, and tryptophan hydroxylase) and three novel genes. Expression patterns of known genes in differentiating EML cells accurately reflected their expected expression patterns based on previous studies. The identification of three novel genes, two of which encode proteins that may act as regulators of hematopoietic differentiation, suggests that EML is a useful model system for the molecular analysis of hematopoietic differentiation.

Longitudinal trajectory of sexual functioning after hematopoietic cell transplantation: impact of chronic graft-versus-host disease and total body irradiation.

PubMed

Wong, F Lennie; Francisco, Liton; Togawa, Kayo; Kim, Heeyoung; Bosworth, Alysia; Atencio, Liezl; Hanby, Cara; Grant, Marcia; Kandeel, Fouad; Forman, Stephen J; Bhatia, Smita

2013-12-05

This prospective study described the trajectory of sexual well-being from before hematopoietic cell transplantation (HCT) to 3 years after in 131 allogeneic and 146 autologous HCT recipients using Derogatis Interview for Sexual Function and Derogatis Global Sexual Satisfaction Index. Sixty-one percent of men and 37% of women were sexually active pre-HCT; the prevalence declined to 51% (P = .01) in men and increased to 48% (P = .02) in women at 3 years post-HCT. After HCT, sexual satisfaction declined in both sexes (P < .001). All sexual function domains were worse in women compared with men (P ≤ .001). Orgasm (P = .002) and drive/relationship (P < .001) declined in men, but sexual cognition/fantasy (P = .01) and sexual behavior/experience (P = .01) improved in women. Older age negatively impacted sexual function post-HCT in both sexes (P < .01). Chronic graft-versus-host disease was associated with lower sexual cognition/fantasy (P = .003) and orgasm (P = .006) in men and sexual arousal (P = .05) and sexual satisfaction (P = .005) in women. All male sexual function domains declined after total body irradiation (P < .05). This study identifies vulnerable subpopulations that could benefit from interventional strategies to improve sexual well-being.

Aging of hematopoietic stem cells: DNA damage and mutations?

PubMed

Moehrle, Bettina M; Geiger, Hartmut

2016-10-01

Aging in the hematopoietic system and the stem cell niche contributes to aging-associated phenotypes of hematopoietic stem cells (HSCs), including leukemia and aging-associated immune remodeling. Among others, the DNA damage theory of aging of HSCs is well established, based on the detection of a significantly larger amount of γH2AX foci and a higher tail moment in the comet assay, both initially thought to be associated with DNA damage in aged HSCs compared with young cells, and bone marrow failure in animals devoid of DNA repair factors. Novel data on the increase in and nature of DNA mutations in the hematopoietic system with age, the quality of the DNA damage response in aged HSCs, and the nature of γH2AX foci question a direct link between DNA damage and the DNA damage response and aging of HSCs, and rather favor changes in epigenetics, splicing-factors or three-dimensional architecture of the cell as major cell intrinsic factors of HSCs aging. Aging of HSCs is also driven by a strong contribution of aging of the niche. This review discusses the DNA damage theory of HSC aging in the light of these novel mechanisms of aging of HSCs. Copyright © 2016 ISEH - International Society for Experimental Hematology. Published by Elsevier Inc. All rights reserved.

[Effect of different oxygen concentrations on biological properties of bone marrow hematopoietic stem cells of mice].

PubMed

Ma, Yi-Ran; Ren, Si-Hua; He, Yu-Xin; Wang, Lin-Lin; Jin, Li; Hao, Yi-Wen

2012-10-01

This study purposed to investigate the effects of different oxygen concentrations and reactive oxygen species (ROS) on the biological characteristics of hematopoietic stem cells (HSC) and their possible mechanisms through simulating oxygen environment to which the peripheral blood HSC are subjected in peripheral blood HSCT. The proliferation ability, cell cycle, directed differentiation ability, ROS level and hematopoietic reconstitution ability of Lin(-)c-kit(+)Sca-1(+) BMHSC were detected by using in vitro amplification test, directional differentiation test, cell cycle analysis, ROS assay and transplantation of Lin(-)c-kit(+)Sca-1(+) HSC from sublethally irradiated mice respectively. The results showed that oxygen concentrations lower than normal oxygen concentration, especially in hypoxic oxygen environment, could reduce ROS generation and amplify more primitive CD34(+)AC133(+) HSC and active CD34(+) HSC, and maintain more stem cells in the G(0)/G(1) phase, which is more helpful to the growth of CFU-S and viability of mice. At the same time, BMHSC exposed to normal oxygen level or inconstant and greatly changed oxygen concentrations could produce a high level of ROS, and the above-mentioned features and functional indicators are relatively low. It is concluded that ROS levels of HSC in BMHSCT are closely related with the oxygen concentration surrounding the cells and its stability. Low oxygen concentration and antioxidant intervention are helpful to transplantation of BMHSC.

Single-Cell RNA-Sequencing Reveals a Continuous Spectrum of Differentiation in Hematopoietic Cells.

PubMed

Macaulay, Iain C; Svensson, Valentine; Labalette, Charlotte; Ferreira, Lauren; Hamey, Fiona; Voet, Thierry; Teichmann, Sarah A; Cvejic, Ana

2016-02-02

The transcriptional programs that govern hematopoiesis have been investigated primarily by population-level analysis of hematopoietic stem and progenitor cells, which cannot reveal the continuous nature of the differentiation process. Here we applied single-cell RNA-sequencing to a population of hematopoietic cells in zebrafish as they undergo thrombocyte lineage commitment. By reconstructing their developmental chronology computationally, we were able to place each cell along a continuum from stem cell to mature cell, refining the traditional lineage tree. The progression of cells along this continuum is characterized by a highly coordinated transcriptional program, displaying simultaneous suppression of genes involved in cell proliferation and ribosomal biogenesis as the expression of lineage specific genes increases. Within this program, there is substantial heterogeneity in the expression of the key lineage regulators. Overall, the total number of genes expressed, as well as the total mRNA content of the cell, decreases as the cells undergo lineage commitment. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Hematopoietic stem cell transplantation in thalassemia major and sickle cell disease: indications and management recommendations from an international expert panel

PubMed Central

Angelucci, Emanuele; Matthes-Martin, Susanne; Baronciani, Donatella; Bernaudin, Françoise; Bonanomi, Sonia; Cappellini, Maria Domenica; Dalle, Jean-Hugues; Di Bartolomeo, Paolo; de Heredia, Cristina Díaz; Dickerhoff, Roswitha; Giardini, Claudio; Gluckman, Eliane; Hussein, Ayad Achmed; Kamani, Naynesh; Minkov, Milen; Locatelli, Franco; Rocha, Vanderson; Sedlacek, Petr; Smiers, Frans; Thuret, Isabelle; Yaniv, Isaac; Cavazzana, Marina; Peters, Christina

2014-01-01

Thalassemia major and sickle cell disease are the two most widely disseminated hereditary hemoglobinopathies in the world. The outlook for affected individuals has improved in recent years due to advances in medical management in the prevention and treatment of complications. However, hematopoietic stem cell transplantation is still the only available curative option. The use of hematopoietic stem cell transplantation has been increasing, and outcomes today have substantially improved compared with the past three decades. Current experience world-wide is that more than 90% of patients now survive hematopoietic stem cell transplantation and disease-free survival is around 80%. However, only a few controlled trials have been reported, and decisions on patient selection for hematopoietic stem cell transplantation and transplant management remain principally dependent on data from retrospective analyses and on the clinical experience of the transplant centers. This consensus document from the European Blood and Marrow Transplantation Inborn Error Working Party and the Paediatric Diseases Working Party aims to report new data and provide consensus-based recommendations on indications for hematopoietic stem cell transplantation and transplant management. PMID:24790059

Allogeneic disparities in immunoglobulin-like transcript 5 induce potent antibody responses in hematopoietic stem cell transplant recipients.

PubMed

Pfistershammer, Katharina; Lawitschka, Anita; Klauser, Christoph; Leitner, Judith; Weigl, Roman; Heemskerk, Mirjam H M; Pickl, Winfried F; Majdic, Otto; Böhmig, Georg A; Fischer, Gottfried F; Greinix, Hildegard T; Steinberger, Peter

2009-09-10

In hematopoietic stem cell transplant (HSCT) recipients, the recognition of polymorphic antigens by the donor-derived immune system is an important mechanism underlying both graft-versus-host disease and graft-versus-leukemia (GVL) effect. Here we show that a subset of HSCT recipients (13.9%, n = 108) have antibodies directed to surface molecules of dendritic cells. We have used one such serum in conjunction with retroviral expression cloning to identify the highly polymorphic surface molecule immunoglobulin-like transcript 5 (ILT5) as one of the targets of dendritic cell-reactive antibodies. ILT5 reactive antibodies were found in 5.4% of HSCT patients but not in solid organ transplantation recipients, patients with collagen diseases, multiparous women, or polytransfused or healthy persons. We show that ILT5-specific antibodies can mediate killing of ILT5-bearing cells and furthermore demonstrate ILT5 expression in some leukemic cells, indicating that it might be a target for GVL effects. Thus, our results represent the first description of potent allogeneic antibody responses to a non-major histocompatibility complex cell surface molecule in hematopoietic stem cell transplanted patients and warrant further studies to elucidate the role of antibodies to polymorphic cell surface molecules in GVL and graft-versus-host responses.

Functional dissection of hematopoietic stem cell populations with a stemness-monitoring system based on NS-GFP transgene expression.

PubMed

Ali, Mohamed A E; Fuse, Kyoko; Tadokoro, Yuko; Hoshii, Takayuki; Ueno, Masaya; Kobayashi, Masahiko; Nomura, Naho; Vu, Ha Thi; Peng, Hui; Hegazy, Ahmed M; Masuko, Masayoshi; Sone, Hirohito; Arai, Fumio; Tajima, Atsushi; Hirao, Atsushi

2017-09-12

Hematopoietic stem cells (HSCs) in a steady state can be efficiently purified by selecting for a combination of several cell surface markers; however, such markers do not consistently reflect HSC activity. In this study, we successfully enriched HSCs with a unique stemness-monitoring system using a transgenic mouse in which green florescence protein (GFP) is driven by the promoter/enhancer region of the nucleostemin (NS) gene. We found that the phenotypically defined long-term (LT)-HSC population exhibited the highest level of NS-GFP intensity, whereas NS-GFP intensity was strongly downregulated during differentiation in vitro and in vivo. Within the LT-HSC population, NS-GFP high cells exhibited significantly higher repopulating capacity than NS-GFP low cells. Gene expression analysis revealed that nine genes, including Vwf and Cdkn1c (p57), are highly expressed in NS-GFP high cells and may represent a signature of HSCs, i.e., a stemness signature. When LT-HSCs suffered from remarkable stress, such as transplantation or irradiation, NS-GFP intensity was downregulated. Finally, we found that high levels of NS-GFP identified HSC-like cells even among CD34 + cells, which have been considered progenitor cells without long-term reconstitution ability. Thus, high NS-GFP expression represents stem cell characteristics in hematopoietic cells, making this system useful for identifying previously uncharacterized HSCs.

Intrathymic injection of hematopoietic progenitor cells establishes functional T cell development in a mouse model of severe combined immunodeficiency.

PubMed

Tuckett, Andrea Z; Thornton, Raymond H; O'Reilly, Richard J; van den Brink, Marcel R M; Zakrzewski, Johannes L

2017-05-16

Even though hematopoietic stem cell transplantation can be curative in patients with severe combined immunodeficiency, there is a need for additional strategies boosting T cell immunity in individuals suffering from genetic disorders of lymphoid development. Here we show that image-guided intrathymic injection of hematopoietic stem and progenitor cells in NOD-scid IL2rγ null mice is feasible and facilitates the generation of functional T cells conferring protective immunity. Hematopoietic stem and progenitor cells were isolated from the bone marrow of healthy C57BL/6 mice (wild-type, Luciferase + , CD45.1 + ) and injected intravenously or intrathymically into both male and female, young or aged NOD-scid IL2rγ null recipients. The in vivo fate of injected cells was analyzed by bioluminescence imaging and flow cytometry of thymus- and spleen-derived T cell populations. In addition to T cell reconstitution, we evaluated mice for evidence of immune dysregulation based on diabetes development and graft-versus-host disease. T cell immunity following intrathymic injection of hematopoietic stem and progenitor cells in NOD-scid IL2rγ null mice was assessed in a B cell lymphoma model. Despite the small size of the thymic remnant in NOD-scid IL2rγ null mice, we were able to accomplish precise intrathymic delivery of hematopoietic stem and progenitor cells by ultrasound-guided injection. Thymic reconstitution following intrathymic injection of healthy allogeneic hematopoietic cells was most effective in young male recipients, indicating that even in the setting of severe immunodeficiency, sex and age are important variables for thymic function. Allogeneic T cells generated in intrathymically injected NOD-scid IL2rγ null mice displayed anti-lymphoma activity in vivo, but we found no evidence for severe auto/alloreactivity in T cell-producing NOD-scid IL2rγ null mice, suggesting that immune dysregulation is not a major concern. Our findings suggest that intrathymic

Oxidative stress in normal hematopoietic stem cells and leukemia.

PubMed

Samimi, Azin; Kalantari, Heybatullah; Lorestani, Marzieh Zeinvand; Shirzad, Reza; Saki, Najmaldin

2018-04-01

Leukemia is developed following the abnormal proliferation of immature hematopoietic cells in the blood when hematopoietic stem cells lose the ability to turn into mature cells at different stages of maturation and differentiation. Leukemia initiating cells are specifically dependent upon the suppression of oxidative stress in the hypoglycemic bone marrow (BM) environment to be able to start their activities. Relevant literature was identified by a PubMed search (2000-2017) of English-language literature using the terms 'oxidative stress,' 'reactive oxygen species,' 'hematopoietic stem cell,' and 'leukemia.' The generation and degradation of free radicals is a main component of the metabolism in aerobic organisms. A certain level of ROS is required for proper cellular function, but values outside this range will result in oxidative stress (OS). Long-term overactivity of reactive oxygen species (ROS) has harmful effects on the function of cells and their vital macromolecules, including the transformation of proteins into autoantigens and increased degradation of protein/DNA, which eventually leads to the change in pathways involved in the development of cancer and several other disorders. According to the metabolic disorders of cancer, the relationship between OS changes, the viability of cancer cells, and their response to chemotherapeutic agents affecting this pathway are undeniable. Recently, studies have been conducted to determine the effect of herbal agents and cancer chemotherapy drugs on oxidative stress pathways. By emphasizing the role of oxidative stress on stem cells in the incidence of leukemia, this paper attempts to state and summarize this subject. © 2018 APMIS. Published by John Wiley & Sons Ltd.

Hematopoietic cell transplantation in Fanconi anemia: current evidence, challenges and recommendations.

PubMed

Ebens, Christen L; MacMillan, Margaret L; Wagner, John E

2017-01-01

Hematopoietic cell transplantation for Fanconi Anemia (FA) has improved dramatically over the past 40 years. With an enhanced understanding of the intrinsic DNA-repair defect and pathophysiology of hematopoietic failure and leukemogenesis, sequential changes to conditioning and graft engineering have significantly improved the expectation of survival after allogeneic hematopoietic cell transplantation (alloHCT) with incidence of graft failure decreased from 35% to <10% and acute graft-versus-host disease (GVHD) from >40% to <10%. Today, five-year overall survival exceeds 90% in younger FA patients with bone marrow failure but remains about 50% in those with hematologic malignancy. Areas covered: We review the evolution of alloHCT contributing to decreased rates of transplant related complications; highlight current challenges including poorer outcomes in cases of clonal hematologic disorders, alloHCT impact on endocrine function and intrinsic FA risk of epithelial malignancies; and describe investigational therapies for prevention and treatment of the hematologic manifestations of FA. Expert commentary: Current methods allow for excellent survival following alloHCT for FA associated BMF irrespective of donor hematopoietic cell source. Alternative curative approaches, such as gene therapy, are being explored to eliminate the risks of GVHD and minimize therapy-related adverse effects.

VE-cadherin expression allows identification of a new class of hematopoietic stem cells within human embryonic liver.

PubMed

Oberlin, Estelle; Fleury, Maud; Clay, Denis; Petit-Cocault, Laurence; Candelier, Jean-Jacques; Mennesson, Benoît; Jaffredo, Thierry; Souyri, Michèle

2010-11-25

Edification of the human hematopoietic system during development is characterized by the production of waves of hematopoietic cells separated in time, formed in distinct embryonic sites (ie, yolk sac, truncal arteries including the aorta, and placenta). The embryonic liver is a major hematopoietic organ wherein hematopoietic stem cells (HSCs) expand, and the future, adult-type, hematopoietic cell hierarchy becomes established. We report herein the identification of a new, transient, and rare cell population in the human embryonic liver, which coexpresses VE-cadherin, an endothelial marker, CD45, a pan-hematopoietic marker, and CD34, a common endothelial and hematopoietic marker. This population displays an outstanding self-renewal, proliferation, and differentiation potential, as detected by in vitro and in vivo hematopoietic assays compared with its VE-cadherin negative counterpart. Based on VE-cadherin expression, our data demonstrate the existence of 2 phenotypically and functionally separable populations of multipotent HSCs in the human embryo, the VE-cadherin(+) one being more primitive than the VE-cadherin(-) one, and shed a new light on the hierarchical organization of the embryonic liver HSC compartment.

Fancb deficiency impairs hematopoietic stem cell function

PubMed Central

Du, Wei; Amarachintha, Surya; Erden, Ozlem; Wilson, Andrew; Meetei, Amom Ruhikanta; Andreassen, Paul R.; Namekawa, Satoshi H.; Pang, Qishen

2015-01-01

Fanconi anemia (FA) is a genetic disorder characterized by bone marrow failure, variable congenital malformations and a predisposition to malignancies. FANCB (also known as FAAP95), is the only X-linked FA gene discovered thus far. In the present study, we investigated hematopoiesis in adult Fancb deficient (Fancb−/y) mice and found that Fancb−/y mice have decreased hematopoietic stem cell (HSC) quiescence accompanied by reduced progenitor activity in vitro and reduced repopulating capacity in vivo. Like other FA mouse models previously reported, the hematopoietic system of Fancb−/y mice is hypersensitive to DNA cross-linking agent mitomycin C (MMC), which induces bone marrow failure in Fancb−/y mice. Furthermore, Fancb−/y BM exhibits slower recovery kinetics and less tolerance to myelotoxic stress induced by 5-fluorouracil than wild-type littermates. RNA-seq analysis reveals altered expression of genes involved in HSC function and cell cycle regulation in Fancb−/y HSC and progenitor cells. Thus, this Fancb−/y mouse model provides a novel approach for studying the critical role of the FA pathway not only in germ cell development but also in the maintenance of HSC function. PMID:26658157

Childhood Hematopoietic Cell Transplantation (PDQ®)—Health Professional Version

Cancer.gov

Childhood hematopoietic cell transplantation involves the infusion of blood stem cells into a patient to reconstitute the blood system. Get detailed information about autologous and allogeneic transplant, HLA matching, preparative regimens, and complications in this summary for clinicians.

Differences in lymphocyte developmental potential between human embryonic stem cell and umbilical cord blood–derived hematopoietic progenitor cells

PubMed Central

Martin, Colin H.; Woll, Petter S.; Ni, Zhenya; Zúñiga-Pflücker, Juan Carlos

2008-01-01

Hematopoietic progenitor cells derived from human embryonic stem cells (hESCs) develop into diverse mature hematopoietic lineages, including lymphocytes. Whereas functional natural killer (NK) cells can be efficiently generated in vitro from hESC-derived CD34+ cells, studies of T- and B-cell development from hESCs have been much more limited. Here, we demonstrate that despite expressing functional Notch-1, CD34+ cells from hESCs did not derive T cells when cocultured with OP9 cells expressing Delta-like 1, or in fetal thymus organ culture. hESC-derived CD34+ cells also did not produce B cells in vitro. In contrast, CD34+ cells isolated from UCB routinely generated T and B cells when cultured in the same conditions. Notably, both undifferentiated hESCs, and sorted hESC-derived populations with hematopoietic developmental potential exhibited constitutive expression of ID family genes and of transcriptional targets of stem cell factor–induced signaling. These pathways both inhibit T-cell development and promote NK-cell development. Together, these results demonstrate fundamental differences between hESC-derived hematopoietic progenitors and analogous primary human cells. Therefore, hESCs can be more readily supported to differentiate into certain cell types than others, findings that have important implications for derivation of defined lineage-committed populations from hESCs. PMID:18621931

Multilineage hematopoietic recovery by a single injection of pegylated recombinant human megakaryocyte growth and development factor in myelosuppressed mice.

PubMed

Shibuya, K; Akahori, H; Takahashi, K; Tahara, E; Kato, T; Miyazaki, H

1998-01-01

Previous studies have shown that daily multiple administration of pegylated recombinant human megakaryocyte growth and development factor (PEG-rHuMGDF) markedly stimulates thrombopoiesis and effectively ameliorates thrombocytopenia, and in most cases anemia and neutropenia, in myelosuppressed animals. In this study, we evaluated the effects of a single intravenous injection of PEG-rHuMGDF on hematopoietic recovery after sublethal total-body irradiation in mice. A single injection of PEG-rHuMGDF (1 to 640 microg/kg) 1 hour after irradiation accelerated platelet, red blood cell (RBC), and white blood cell (WBC) recovery in a dose-dependent fashion. In the bone marrow of vehicle-treated mice, megakaryocytic, erythroid, and myeloid progenitors, as well as day 12 colony-forming unit-spleen (CFU-S), were dramatically decreased much earlier than the nadirs of peripheral blood cells, whereas megakaryocytes were modestly decreased. Treatment with PEG-rHuMGDF (80 microg/kg, an optimal dose) 1 hour after irradiation resulted in more rapid recovery of these four hematopoietic progenitors and also significantly facilitated megakaryocyte recovery. In addition, the same PEG-rHuMGDF administration schedule expanded bone marrow cells capable of rescuing lethally irradiated recipient mice. As the interval between irradiation and PEG-rHuMGDF treatment was longer, its effects on hematopoietic recovery were attenuated. In contrast to the effects of PEG-rHuMGDF, a single injection of recombinant human granulocyte colony-stimulating factor (rhG-CSF) 1 hour after irradiation exclusively accelerated WBC recovery, but only to a similar extent as PEG-rHuMGDF (80 microg/kg) treatment even when rhG-CSF doses were escalated to 1,000 microg/kg. This appeared related to different pharmacokinetics of these two factors after a single injection in irradiated mice. The concentrations of PEG-rHuMGDF after injection persisted in the plasma for a longer time compared with rhG-CSF. These results indicate

Long-term in vivo provision of antigen-specific T cell immunity by programming hematopoietic stem cells

NASA Astrophysics Data System (ADS)

Yang, Lili; Baltimore, David

2005-03-01

A method to genetically program mouse hematopoietic stem cells to develop into functional CD8 or CD4 T cells of defined specificity in vivo is described. For this purpose, a bicistronic retroviral vector was engineered that efficiently delivers genes for both and chains of T cell receptor (TCR) to hematopoietic stem cells. When modified cell populations were used to reconstruct the hematopoietic lineages of recipient mice, significant percentages of antigen-specific CD8 or CD4 T cells were observed. These cells expressed normal surface markers and responded to peptide antigen stimulation by proliferation and cytokine production. Moreover, they could mature into memory cells after peptide stimulation. Using TCRs specific for a model tumor antigen, we found that the recipient mice were able to partially resist a challenge with tumor cells carrying the antigen. By combining cells modified with CD8- and CD4-specific TCRs, and boosting with dendritic cells pulsed with cognate peptides, complete suppression of tumor could be achieved and even tumors that had become established would regress and be eliminated after dendritic cell/peptide immunization. This methodology of "instructive immunotherapy" could be developed for controlling the growth of human tumors and attacking established pathogens.

T-cell and natural killer cell therapies for hematologic malignancies after hematopoietic stem cell transplantation: enhancing the graft-versus-leukemia effect

PubMed Central

Cruz, C. Russell; Bollard, Catherine M.

2015-01-01

Hematopoietic stem cell transplantation has revolutionized the treatment of hematologic malignancies, but infection, graft-versus-host disease and relapse are still important problems. Calcineurin inhibitors, T-cell depletion strategies, and immunomodulators have helped to prevent graft-versus-host disease, but have a negative impact on the graft-versus-leukemia effect. T cells and natural killer cells are both thought to be important in the graft-versus-leukemia effect, and both cell types are amenable to ex vivo manipulation and clinical manufacture, making them versatile immunotherapeutics. We provide an overview of these immunotherapeutic strategies following hematopoietic stem cell transplantation, with discussions centered on natural killer and T-cell biology. We discuss the contributions of each cell type to graft-versus-leukemia effects, as well as the current research directions in the field as related to adoptive cell therapy after hematopoietic stem cell transplantation. PMID:26034113

Breaking the rules? X-ray examination of hematopoietic stem cell grafts at international airports.

PubMed

Petzer, Andreas L; Speth, Hans-Georg; Hoflehner, Elisabeth; Clausen, Johannes; Nachbaur, David; Gastl, Günther; Gunsilius, Eberhard

2002-06-15

Hematopoietic stem cell grafts from unrelated donors are commonly transported by aircraft. They must not be subjected to x-rays during security checks, which may cause inconvenient discussions between the courier and the airport security staff. We exposed hematopoietic stem cells from mobilized peripheral blood to a widely used x-ray hand-luggage control system. Cell viability as well as growth in vitro of mature progenitor cells (colony-forming cells), primitive progenitor cells (long-term culture-initiating cells), and lymphocytes were not altered even after 10 passages through the hand-luggage control system. Thus, repeated exposure to the low radiation dose of hand-luggage control systems (1.5 +/- 0.6 microSv per exposure) seems to be harmless for hematopoietic stem cells, which should simplify the international transport of stem cell grafts.

Bone marrow adipocytes promote the regeneration of stem cells and hematopoiesis by secreting SCF

PubMed Central

Zhou, Bo O.; Yu, Hua; Yue, Rui; Zhao, Zhiyu; Rios, Jonathan J.; Naveiras, Olaia; Morrison, Sean J.

2017-01-01

Endothelial cells and Leptin Receptor+ (LepR+) stromal cells are critical sources of haematopoietic stem cell (HSC) niche factors, including Stem Cell Factor (SCF), in bone marrow. After irradiation or chemotherapy, these cells are depleted while adipocytes become abundant. We discovered that bone marrow adipocytes synthesize SCF. They arise from Adipoq-Cre/ER+ progenitors, which represent ~5% of LepR+ cells, and proliferate after irradiation. Scf deletion using Adipoq-Cre/ER inhibited hematopoietic regeneration after irradiation or 5-fluorouracil treatment, depleting HSCs and reducing mouse survival. Scf from LepR+ cells, but not endothelial, hematopoietic, or osteoblastic cells, also promoted regeneration. In non-irradiated mice, Scf deletion using Adipoq-Cre/ER did not affect HSC frequency in long bones, which have few adipocytes, but depleted HSCs in tail vertebrae, which have abundant adipocytes. A-ZIP/F1 ‘fatless” mice exhibited delayed hematopoietic regeneration in long bones but not in tail vertebrae, where adipocytes inhibited vascularization. Adipocytes are a niche component that promotes hematopoietic regeneration. PMID:28714970

Recombinant adeno-associated virus mediates a high level of gene transfer but less efficient integration in the K562 human hematopoietic cell line.

PubMed Central

Malik, P; McQuiston, S A; Yu, X J; Pepper, K A; Krall, W J; Podsakoff, G M; Kurtzman, G J; Kohn, D B

1997-01-01

We tested the ability of a recombinant adeno-associated virus (rAAV) vector to express and integrate exogenous DNA into human hematopoietic cells in the absence of selection. We developed an rAAV vector, AAV-tNGFR, carrying a truncated rat nerve growth factor receptor (tNGFR) cDNA as a cell surface reporter under the control of the Moloney murine leukemia virus (MoMuLV) long terminal repeat. An analogous MoMuLV-based retroviral vector (L-tNGFR) was used in parallel, and gene transfer and expression in human hematopoietic cells were assessed by flow cytometry and DNA analyses. Following gene transfer into K562 cells with AAV-tNGFR at a multiplicity of infection (MOI) of 13 infectious units (IU), 26 to 38% of cells expressed tNGFR on the surface early after transduction, but the proportion of tNGFR expressing cells steadily declined to 3.0 to 3.5% over 1 month of culture. At an MOI of 130 IU, nearly all cells expressed tNGFR immediately posttransduction, but the proportion of cells expressing tNGFR declined to 62% over 2 months of culture. The decline in the proportion of AAV-tNGFR-expressing cells was associated with ongoing losses of vector genomes. In contrast, K562 cells transduced with the retroviral vector L-tNGFR expressed tNGFR in a constant fraction. Integration analyses on clones showed that integration occurred at different sites. Integration frequencies were estimated at about 49% at an MOI of 130 and 2% at an MOI of 1.3. Transduction of primary human CD34+ progenitor cells by AAV-tNGFR was less efficient than with K562 cells and showed a declining percentage of cells expressing tNGFR over 2 weeks of culture. Thus, purified rAAV caused very high gene transfer and expression in human hematopoietic cells early after transduction, which steadily declined during cell passage in the absence of selection. Although the efficiency of integration was low, overall integration was markedly improved at a high MOI. While prolonged episomal persistence may be adequate

Recombinant adeno-associated virus mediates a high level of gene transfer but less efficient integration in the K562 human hematopoietic cell line.

PubMed

Malik, P; McQuiston, S A; Yu, X J; Pepper, K A; Krall, W J; Podsakoff, G M; Kurtzman, G J; Kohn, D B

1997-03-01

We tested the ability of a recombinant adeno-associated virus (rAAV) vector to express and integrate exogenous DNA into human hematopoietic cells in the absence of selection. We developed an rAAV vector, AAV-tNGFR, carrying a truncated rat nerve growth factor receptor (tNGFR) cDNA as a cell surface reporter under the control of the Moloney murine leukemia virus (MoMuLV) long terminal repeat. An analogous MoMuLV-based retroviral vector (L-tNGFR) was used in parallel, and gene transfer and expression in human hematopoietic cells were assessed by flow cytometry and DNA analyses. Following gene transfer into K562 cells with AAV-tNGFR at a multiplicity of infection (MOI) of 13 infectious units (IU), 26 to 38% of cells expressed tNGFR on the surface early after transduction, but the proportion of tNGFR expressing cells steadily declined to 3.0 to 3.5% over 1 month of culture. At an MOI of 130 IU, nearly all cells expressed tNGFR immediately posttransduction, but the proportion of cells expressing tNGFR declined to 62% over 2 months of culture. The decline in the proportion of AAV-tNGFR-expressing cells was associated with ongoing losses of vector genomes. In contrast, K562 cells transduced with the retroviral vector L-tNGFR expressed tNGFR in a constant fraction. Integration analyses on clones showed that integration occurred at different sites. Integration frequencies were estimated at about 49% at an MOI of 130 and 2% at an MOI of 1.3. Transduction of primary human CD34+ progenitor cells by AAV-tNGFR was less efficient than with K562 cells and showed a declining percentage of cells expressing tNGFR over 2 weeks of culture. Thus, purified rAAV caused very high gene transfer and expression in human hematopoietic cells early after transduction, which steadily declined during cell passage in the absence of selection. Although the efficiency of integration was low, overall integration was markedly improved at a high MOI. While prolonged episomal persistence may be adequate

HEMATOPOIETIC STEM CELL INFUSION/TRANSPLANTATION FOR INDUCTION OF ALLOGRAFT TOLERANCE

PubMed Central

Granados, Jose M. Marino; Benichou, Gilles; Kawai, Tatsuo

2015-01-01

Purpose of review This review updates the current status of basic, preclinical, and clinical research on donor hematopoietic stem cell infusion for allograft tolerance induction. Recent findings Recent basic studies in mice provide evidence of significant involvement of both central deletional and peripheral regulatory mechanisms in induction and maintenance of allograft tolerance effected through a mixed chimerism approach with donor hematopoietic stem cell infusion. The presence of heterologous memory T cells in primates hampers the induction of persistent chimerism. Durable mixed chimerism, however, now has been recently induced in inbred major histocompatibility complex-mismatched swine, resulting in tolerance of vascularized composite tissue allografts. In clinical transplantation, allograft tolerance has been achieved in human leukocyte antigen-mismatched kidney transplantation after the induction of transient mixed chimerism or persistent full donor chimerism. Summary Tolerance induction in clinical kidney transplantation has been achieved by donor hematopoietic stem cell infusion. Improving the consistency and safety of tolerance induction and extending successful protocols to other organs, as well as to organs from deceased donors, are critical next steps to bringing tolerance to a wider range of clinical applications. PMID:25563992

Evidence that β7 Integrin Regulates Hematopoietic Stem Cell Homing and Engraftment Through Interaction with MAdCAM-1.

PubMed

Murakami, Jodi L; Xu, Baohui; Franco, Christopher B; Hu, Xingbin; Galli, Stephen J; Weissman, Irving L; Chen, Ching-Cheng

2016-01-01

α4β7 integrin is a cell adhesion receptor that is crucial for the migration of hematopoietic progenitors and mature effector cells in the periphery, but its role in adult hematopoiesis is controversial. We identified a subset of hematopoietic stem cells (HSCs) in the bone marrow (BM) that expressed β7 integrin. These β7(+) HSCs were capable of multilineage, long-term reconstitution and had an inherent competitive advantage over β7(-) HSCs. On the other hand, HSCs that lacked β7 integrin (β7KO) had reduced engraftment potential. Interestingly, quantitative RT-PCR and flow cytometry revealed that β7KO HSCs expressed lower levels of the chemokine receptor CXCR4. Accordingly, β7KO HSCs exhibited impaired migration abilities in vitro and BM homing capabilities in vivo. Lethal irradiation induced expression of the α4β7 integrin ligand-mucosal addressin cell adhesion molecule-1 (MAdCAM-1) on BM endothelial cells. Moreover, blocking MAdCAM-1 reduced the homing of HSCs and impaired the survival of recipient mice. Altogether, these data indicate that β7 integrin, when expressed by HSCs, interacted with its endothelial ligand MAdCAM-1 in the BM microenvironment, thereby promoting HSC homing and engraftment.

[Allogeneic hematopoietic stem cell transplantation using myeloablative conditioning including total body irradiation for pediatric acute lymphoblastic leukemia: a single-center retrospective analysis].

PubMed

Honda, Mamoru; Arakawa, Yuki; Kawakami, Ryota; Itabashi, Toshikazu; Yanagi, Masato; Sasaki, Koji; Watanabe, Kentaro; Isobe, Kiyotaka; Mori, Makiko; Hanada, Ryoji; Koh, Katsuyoshi

2018-01-01

This study aimed to investigate the clinical outcomes of hematopoietic stem cell transplantation (HSCT) with total body irradiation-based myeloablative conditioning (TBI-MAC) in pediatric patients with acute lymphoblastic leukemia (ALL). We retrospectively examined patients with ALL who underwent HSCT with TBI-MAC from January 2000 to August 2016 at our institute. We enrolled 67 patients with a median follow-up period of 8 years. The 5-year event-free survival (EFS) and overall survival (OS) were 51.2% and 59.6%, respectively. At the first complete remission, HSCT exhibited significantly superior EFS and OS in our patients than that in patients with other diseases. We encountered 57.9% of patients with at least one late complication. Major late complications were short stature (26.3%) and hypogonadism (18.4%). While late complications were observed in several recipients of HSCT, late complication-related deaths occurred in three patients. The TBI-MAC regimen led to favorable clinical outcomes in pediatric patients with ALL who underwent HSCT. Thus, proper evaluation and management of late complications are mandatory.

MERP1: a mammalian ependymin-related protein gene differentially expressed in hematopoietic cells.

PubMed

Gregorio-King, Claudia C; McLeod, Janet L; Collier, Fiona McL; Collier, Gregory R; Bolton, Karyn A; Van Der Meer, Gavin J; Apostolopoulos, Jim; Kirkland, Mark A

2002-03-20

We have utilized differential display polymerase chain reaction to investigate the gene expression of hematopoietic progenitor cells from adult bone marrow and umbilical cord blood. A differentially expressed gene was identified in CD34+ hematopoietic progenitor cells, with low expression in CD34- cells. We have obtained the full coding sequence of this gene which we designated human mammalian ependymin-related protein 1 (MERP1). Expression of MERP1 was found in a variety of normal human tissues, and is 4- and 10-fold higher in adult bone marrow and umbilical cord blood CD34+ cells, respectively, compared to CD34- cells. Additionally, MERP1 expression in a hematopoietic stem cell enriched population was down-regulated with proliferation and differentiation. Conceptual translation of the MERP1 open reading frame reveals significant homology to two families of glycoprotein calcium-dependant cell adhesion molecules: ependymins and protocadherins.

Amino acid–insensitive mTORC1 regulation enables nutritional stress resilience in hematopoietic stem cells

PubMed Central

Kalaitzidis, Demetrios; Efeyan, Alejo; Kfoury, Youmna; Nayyar, Naema; Sykes, David B.; Mercier, Francois E.; Papazian, Ani; Baryawno, Ninib; Victora, Gabriel D.; Sabatini, David M.; Scadden, David T.

2017-01-01

The mTOR pathway is a critical determinant of cell persistence and growth wherein mTOR complex 1 (mTORC1) mediates a balance between growth factor stimuli and nutrient availability. Amino acids or glucose facilitates mTORC1 activation by inducing RagA GTPase recruitment of mTORC1 to the lysosomal outer surface, enabling activation of mTOR by the Ras homolog Rheb. Thereby, RagA alters mTORC1-driven growth in times of nutrient abundance or scarcity. Here, we have evaluated differential nutrient-sensing dependence through RagA and mTORC1 in hematopoietic progenitors, which dynamically drive mature cell production, and hematopoietic stem cells (HSC), which provide a quiescent cellular reserve. In nutrient-abundant conditions, RagA-deficient HSC were functionally unimpaired and upregulated mTORC1 via nutrient-insensitive mechanisms. RagA was also dispensable for HSC function under nutritional stress conditions. Similarly, hyperactivation of RagA did not affect HSC function. In contrast, RagA deficiency markedly altered progenitor population function and mature cell output. Therefore, RagA is a molecular mechanism that distinguishes the functional attributes of reactive progenitors from a reserve stem cell pool. The indifference of HSC to nutrient sensing through RagA contributes to their molecular resilience to nutritional stress, a characteristic that is relevant to organismal viability in evolution and in modern HSC transplantation approaches. PMID:28319048

Development and Function of Myeloid-Derived Suppressor Cells Generated From Mouse Embryonic and Hematopoietic Stem Cells

PubMed Central

Zhou, Zuping; French, Deborah L.; Ma, Ge; Eisenstein, Samuel; Chen, Ying; Divino, Celia M.; Keller, Gordon; Chen, Shu-Hsia; Pan, Ping-Ying

2015-01-01

Emerging evidence suggests that myeloid-derived suppressor cells (MDSCs) have great potential as a novel immune intervention modality in the fields of transplantation and autoimmune diseases. Thus far, efforts to develop MDSC-based therapeutic strategies have been hampered by the lack of a reliable source of MDSCs. Here we show that functional MDSCs can be efficiently generated from mouse embryonic stem (ES) cells and bone marrow hematopoietic stem (HS) cells. In vitro-derived MDSCs encompass two homogenous subpopulations: CD115+Ly-6C+ and CD115+Ly-6C− cells. The CD115+Ly-6C+ subset is equivalent to the monocytic Gr-1+CD115+F4/80+ MDSCs found in tumor-bearing mice. In contrast, the CD115+Ly-6C− cells, a previously unreported population of MDSCs, resemble the granulocyte/macrophage progenitors developmentally. In vitro, ES- and HS-MDSCs exhibit robust suppression against T-cell proliferation induced by polyclonal stimuli or alloantigens via multiple mechanisms involving nitric oxide synthase-mediated NO production and interleukin (IL)-10. Impressively, they display even stronger suppressive activity and significantly enhance ability to induce CD4+CD25+Foxp3+ regulatory T-cell development compared with tumor-derived MDSCs. Furthermore, adoptive transfer of ES-MDSCs can effectively prevent alloreactive T-cell-mediated lethal graft-versus-host disease, leading to nearly 82% long-term survival among treated mice. The successful in vitro generation of MDSCs may represent a critical step toward potential clinical application of MDSCs. PMID:20073041

Studies on the organization and regeneration of bone marrow: origin, growth, and differentiation of endocloned hematopoietic colonies

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lambertsen, R.H.; Weiss, L.

1983-04-01

Hematopoietic colonies were studied by light microscopy in the marrow of alternate fraction x-irradiated mice (C576J/B1) to investigate the microenvironmental organization of marrow and identify early hematopoietic cell-stromal cell interactions. Undifferentiated colonies (UC) were detected at 3 days postirradiation, showed a marked predilection for bone surfaces, and disappeared as differentiated colonies developed. Some UC occurred along marrow arteries. Neutrophilic granulocyte colonies (GC) occurred in all areas at 3 days but grew rapidly only subosteally. Few eosinophilic colonies (GCe) occurred. Erythrocytic colonies (EC) appeared at 4 days as dispersed populations of motile cells within a localized area of marrow; these tendedmore » to proliferate initially in intermediate and central marrow zones. Macrophage colonies (M phi C) of two ''subtypes'' were detected, peaking in relative frequency at 4 days. These appeared active in stromal repair and monocytopoiesis. Megakaryocyte colonies (MC) originated along bone and differentiated away from bone. These results were interpreted as evidence that in x-irradiated marrow: (1) hematopoietic microenvironments (HMs) for stem-cell proliferation and commitment to differentiation, with the possible exception of HMs determining erythroid differentiation, occur in endosteal and periarterial regions; (2) a proliferative and/or chemotactic stimulus to erythroid progenitors exists in intermediate and central marrow regions; and (3) some subosteal regions may exclude erythropoiesis, or preferentially support nonerythroid differentiation. Elaborate associations occurred between macrophages and early UC, GC, and EC, but not MC hematopoietic cells. UC and GC often associated with osteoclasts. Reticular and other fibroblastic cells associated with the cells of all colony types.« less

HRV signaling in airway epithelial cells is regulated by ITAM-mediated recruitment and activation of Syk.

PubMed

Lau, Christine; Castellanos, Patricia; Ranev, Dimitre; Wang, Xiaomin; Chow, Chung-Wai

2011-05-01

Human rhinovirus (HRV), cause of the common cold, is a leading cause of exacerbations of asthma and chronic obstruction pulmonary disease (COPD). Binding of HRV to ICAM (intercellular adhesion molecule)-1, its major receptor, induces a profound inflammatory response from airway epithelial cells. My laboratory has identified Syk tyrosine kinase to be an early regulator of HRV-ICAM-1 signalling: Syk mediates replication-independent p38 mitogen-activated protein (MAP) kinase and phosphatidyl-inositol 3 (PI3)-kinase activation, interleukin (IL)-8 expression, as well as HRV internalization via clathrin-mediated endocytosis. Syk activation is accompanied by formation of a protein complex consisting of ICAM-1, ezrin and Syk at the plasma membrane. However, the molecular mechanisms that regulate this process are not understood. In this report, we investigated the role of the Syk-SH2 domains and the ezrin ITAM (immuno-tyrosine activation motif)-like motif in HRV-induced cell activation using the human BEAS-2B airway epithelial cells. Our observations suggest that the ezrin-ITAM plays a role in Syk recruitment and activation by binding to the Syk tandem SH2 domains, as originally described in the canonical ITAM-mediating signal transduction pathway in hematopoietic cells. This report is the first to demonstrate ITAM-mediated signaling in non-hematopoietic cells, suggesting that this signaling paradigm may be more ubiquitous than previously recognized.

Pre-malignant lymphoid cells arise from hematopoietic stem/progenitor cells in chronic lymphocytic leukemia.

PubMed

Kikushige, Yoshikane; Miyamoto, Toshihiro

2015-11-01

Human malignancies progress through a multistep process that includes the development of critical somatic mutations over the clinical course. Recent novel findings have indicated that hematopoietic stem cells (HSCs), which have the potential to self-renew and differentiate into multilineage hematopoietic cells, are an important cellular target for the accumulation of critical somatic mutations in hematological malignancies and play a central role in myeloid malignancy development. In contrast to myeloid malignancies, mature lymphoid malignancies, such as chronic lymphocytic leukemia (CLL), are thought to originate directly from differentiated mature lymphocytes; however, recent compelling data have shown that primitive HSCs and hematopoietic progenitor cells contribute to the pathogenesis of mature lymphoid malignancies. Several representative mutations of hematological malignancies have been identified within the HSCs of CLL and lymphoma patients, indicating that the self-renewing long-lived fraction of HSCs can serve as a reservoir for the development of oncogenic events. Novel mice models have been established as human mature lymphoma models, in which specific oncogenic events target the HSCs and immature progenitor cells. These data collectively suggest that HSCs can be the cellular target involved in the accumulation of oncogenic events in the pathogenesis of mature lymphoid and myeloid malignancies.

Congenital amegakaryocytic thrombocytopenia iPS cells exhibit defective MPL-mediated signaling

PubMed Central

Hirata, Shinji; Takayama, Naoya; Jono-Ohnishi, Ryoko; Endo, Hiroshi; Nakamura, Sou; Dohda, Takeaki; Nishi, Masanori; Hamazaki, Yuhei; Ishii, Ei-ichi; Kaneko, Shin; Otsu, Makoto; Nakauchi, Hiromitsu; Kunishima, Shinji; Eto, Koji

2013-01-01

Congenital amegakaryocytic thrombocytopenia (CAMT) is caused by the loss of thrombopoietin receptor–mediated (MPL-mediated) signaling, which causes severe pancytopenia leading to bone marrow failure with onset of thrombocytopenia and anemia prior to leukopenia. Because Mpl–/– mice do not exhibit the human disease phenotype, we used an in vitro disease tracing system with induced pluripotent stem cells (iPSCs) derived from a CAMT patient (CAMT iPSCs) and normal iPSCs to investigate the role of MPL signaling in hematopoiesis. We found that MPL signaling is essential for maintenance of the CD34+ multipotent hematopoietic progenitor (MPP) population and development of the CD41+GPA+ megakaryocyte-erythrocyte progenitor (MEP) population, and its role in the fate decision leading differentiation toward megakaryopoiesis or erythropoiesis differs considerably between normal and CAMT cells. Surprisingly, complimentary transduction of MPL into normal or CAMT iPSCs using a retroviral vector showed that MPL overexpression promoted erythropoiesis in normal CD34+ hematopoietic progenitor cells (HPCs), but impaired erythropoiesis and increased aberrant megakaryocyte production in CAMT iPSC–derived CD34+ HPCs, reflecting a difference in the expression of the transcription factor FLI1. These results demonstrate that impaired transcriptional regulation of the MPL signaling that normally governs megakaryopoiesis and erythropoiesis underlies CAMT. PMID:23908116

Congenital amegakaryocytic thrombocytopenia iPS cells exhibit defective MPL-mediated signaling.

PubMed

Hirata, Shinji; Takayama, Naoya; Jono-Ohnishi, Ryoko; Endo, Hiroshi; Nakamura, Sou; Dohda, Takeaki; Nishi, Masanori; Hamazaki, Yuhei; Ishii, Ei-ichi; Kaneko, Shin; Otsu, Makoto; Nakauchi, Hiromitsu; Kunishima, Shinji; Eto, Koji

2013-09-01

Congenital amegakaryocytic thrombocytopenia (CAMT) is caused by the loss of thrombopoietin receptor-mediated (MPL-mediated) signaling, which causes severe pancytopenia leading to bone marrow failure with onset of thrombocytopenia and anemia prior to leukopenia. Because Mpl(-/-) mice do not exhibit the human disease phenotype, we used an in vitro disease tracing system with induced pluripotent stem cells (iPSCs) derived from a CAMT patient (CAMT iPSCs) and normal iPSCs to investigate the role of MPL signaling in hematopoiesis. We found that MPL signaling is essential for maintenance of the CD34+ multipotent hematopoietic progenitor (MPP) population and development of the CD41+GPA+ megakaryocyte-erythrocyte progenitor (MEP) population, and its role in the fate decision leading differentiation toward megakaryopoiesis or erythropoiesis differs considerably between normal and CAMT cells. Surprisingly, complimentary transduction of MPL into normal or CAMT iPSCs using a retroviral vector showed that MPL overexpression promoted erythropoiesis in normal CD34+ hematopoietic progenitor cells (HPCs), but impaired erythropoiesis and increased aberrant megakaryocyte production in CAMT iPSC-derived CD34+ HPCs, reflecting a difference in the expression of the transcription factor FLI1. These results demonstrate that impaired transcriptional regulation of the MPL signaling that normally governs megakaryopoiesis and erythropoiesis underlies CAMT.

Erythro-Myeloid Progenitors: “definitive” hematopoiesis in the conceptus prior to the emergence of hematopoietic stem cells

PubMed Central

Frame, Jenna M.; McGrath, Kathleen E.; Palis, James

2013-01-01

Erythro-myeloid progenitors (EMP) serve as a major source of hematopoiesis in the developing conceptus prior to the formation of a permanent blood system. In this review, we summarize the current knowledge regarding the emergence, fate, and potential of this hematopoietic stem cell (HSC)-independent wave of hematopoietic progenitors, focusing on the murine embryo as a model system. A better understanding of the temporal and spatial control of hematopoietic emergence in the embryo will ultimately improve our ability to derive hematopoietic stem and progenitor cells from embryonic stem cells and induced pluripotent stem cells to serve therapeutic purposes. PMID:24095199

Rescue of CD8+ T cell vaccine memory following sublethal γ irradiation.

PubMed

McFarland, Hugh I; Berkson, Julia D; Lee, Jay P; Elkahloun, Abdel G; Mason, Karen P; Rosenberg, Amy S

2015-07-31

Sublethal γ irradiation eliminates CD8+ T cell mediated memory responses. In this work, we explored how these memory responses could be rescued in the aftermath of such exposure. We utilized two models of CD8+ T cell mediated immunity: a mouse model of Listeria monocytogenes (LM) infection in which CD8+ T cells specific for LM expressed antigens (Listeriolysin O, LLO) can be tracked, and a murine skin graft model in which CD8+ T cells mediate rejection across a MHC class I (D(d)) disparity. In the LM immunized mice, LL0 specific CD8+ T memory cells were lost on irradiation, preserved with rapid revaccination with an attenuated strain 1-3 days post-irradiation (PI), and these mice survived a subsequent wild type LM challenge. A genetic "signature of rescue" identified a group of immune-associated mRNA maintained or upregulated following irradiation and rescue. A number of these factors, including IL-36γ, dectin-2 (Clec4n), and mir101c are upregulated rapidly after exposure of mice to sublethal γ radiation alone and are sustained by early, but not later rescue. Such factors will be evaluated as potential therapeutics to replace individual vaccines for global rescue of CD8+ T memory cell responses following sublethal γ irradiation. The skin allograft model mirrored that of the LM model in that the accelerated D(d) skin allograft rejection response was lost in mice exposed to sublethal γ radiation, but infusion of allogeneic D(d) expressing bone marrow cells 1-4 days PI preserved the CD8+ T memory mediated accelerated rejection response, further suggesting that innate immune responses may not always be essential to rescue of CD8+ memory T cells following γ irradiation. Published by Elsevier Ltd.

Hematopoietic stem cells are acutely sensitive to Acd shelterin gene inactivation

PubMed Central

Jones, Morgan; Osawa, Gail; Regal, Joshua A.; Weinberg, Daniel N.; Taggart, James; Kocak, Hande; Friedman, Ann; Ferguson, David O.; Keegan, Catherine E.; Maillard, Ivan

2013-01-01

The shelterin complex plays dual functions in telomere homeostasis by recruiting telomerase and preventing the activation of a DNA damage response at telomeric ends. Somatic stem cells require telomerase activity, as evidenced by progressive stem cell loss leading to bone marrow failure in hereditary dyskeratosis congenita. Recent work demonstrates that dyskeratosis congenita can also arise from mutations in specific shelterin genes, although little is known about shelterin functions in somatic stem cells. We found that mouse hematopoietic stem cells (HSCs) are acutely sensitive to inactivation of the shelterin gene Acd, encoding TPP1. Homozygosity for a hypomorphic acd allele preserved the emergence and expansion of fetal HSCs but led to profoundly defective function in transplantation assays. Upon complete Acd inactivation, HSCs expressed p53 target genes, underwent cell cycle arrest, and were severely depleted within days, leading to hematopoietic failure. TPP1 loss induced increased telomeric fusion events in bone marrow progenitors. However, unlike in epidermal stem cells, p53 deficiency did not rescue TPP1-deficient HSCs, indicating that shelterin dysfunction has unique effects in different stem cell populations. Because the consequences of telomere shortening are progressive and unsynchronized, acute loss of shelterin function represents an attractive alternative for studying telomere crisis in hematopoietic progenitors. PMID:24316971

Hematopoietic stem cells can differentiate into restricted myeloid progenitors before cell division in mice.

PubMed

Grinenko, Tatyana; Eugster, Anne; Thielecke, Lars; Ramasz, Beáta; Krüger, Anja; Dietz, Sevina; Glauche, Ingmar; Gerbaulet, Alexander; von Bonin, Malte; Basak, Onur; Clevers, Hans; Chavakis, Triantafyllos; Wielockx, Ben

2018-05-15

Hematopoietic stem cells (HSCs) continuously replenish all blood cell types through a series of differentiation steps and repeated cell divisions that involve the generation of lineage-committed progenitors. However, whether cell division in HSCs precedes differentiation is unclear. To this end, we used an HSC cell-tracing approach and Ki67 RFP knock-in mice, in a non-conditioned transplantation model, to assess divisional history, cell cycle progression, and differentiation of adult HSCs. Our results reveal that HSCs are able to differentiate into restricted progenitors, especially common myeloid, megakaryocyte-erythroid and pre-megakaryocyte progenitors, without undergoing cell division and even before entering the S phase of the cell cycle. Additionally, the phenotype of the undivided but differentiated progenitors correlated with the expression of lineage-specific genes and loss of multipotency. Thus HSC fate decisions can be uncoupled from physical cell division. These results facilitate a better understanding of the mechanisms that control fate decisions in hematopoietic cells.

Quantitative stability of hematopoietic stem and progenitor cell clonal output in rhesus macaques receiving transplants

PubMed Central

Koelle, Samson J.

2017-01-01

Autologous transplantation of hematopoietic stem and progenitor cells lentivirally labeled with unique oligonucleotide barcodes flanked by sequencing primer targets enables quantitative assessment of the self-renewal and differentiation patterns of these cells in a myeloablative rhesus macaque model. Compared with other approaches to clonal tracking, this approach is highly quantitative and reproducible. We documented stable multipotent long-term hematopoietic clonal output of monocytes, granulocytes, B cells, and T cells from a polyclonal pool of hematopoietic stem and progenitor cells in 4 macaques observed for up to 49 months posttransplantation. A broad range of clonal behaviors characterized by contribution level and biases toward certain cell types were extremely stable over time. Correlations between granulocyte and monocyte clonalities were greatest, followed by correlations between these cell types and B cells. We also detected quantitative expansion of T cell–biased clones consistent with an adaptive immune response. In contrast to recent data from a nonquantitative murine model, there was little evidence for clonal succession after initial hematopoietic reconstitution. These findings have important implications for human hematopoiesis, given the similarities between macaque and human physiologies. PMID:28087539

Challenges and Opportunities to Harnessing the (Hematopoietic) Stem Cell Niche

PubMed Central

Choi, Ji Sun; Harley, Brendan A. C.

2016-01-01

In our body, stem cells reside in a microenvironment termed the niche. While the exact composition and therefore the level of complexity of a stem cell niche can vary significantly tissue-to-tissue, the stem cell niche microenvironment is dynamic, typically containing spatial and temporal variations in both cellular, extracellular matrix, and biomolecular components. This complex flow of secreted or bound biomolecules, cytokines, extracellular matrix components, and cellular constituents all contribute to the regulation of stem cell fate specification events, making engineering approaches at the nano- and micro-scale of particular interest for creating an artificial niche environment in vitro. Recent advances in fabrication approaches have enabled biomedical researchers to capture and recreate the complexity of stem cell niche microenvironments in vitro. Such engineered platforms show promise as a means to enhance our understanding of the mechanisms underlying niche-mediated stem cell regulation as well as offer opportunities to precisely control stem cell expansion and differentiation events for clinical applications. While these principles generally apply to all adult stem cells and niches, in this review, we focus on recent developments in engineering synthetic niche microenvironments for one of the best-characterized stem cell populations, hematopoietic stem cells (HSC). Specifically, we highlight recent advances in platforms designed to facilitate the extrinsic control of HSC fate decisions. PMID:27134819

Supportive Care of Hematopoietic Cell Transplant Patients

PubMed Central

Jim, Heather S. L.; Syrjala, Karen L.; Rizzo, Doug

2012-01-01

Hematopoietic cell transplant survivors face a number of challenges including low energy and stamina, “chemo-brain” and emotional distress, and late effects that can compromise functioning or lead to early mortality. This session will review the most recent interventions and recommendations to avoid or mitigate these complications. PMID:22226095

Childhood Hematopoietic Cell Transplantation (PDQ®)—Health Professional Version

Cancer.gov

Hematopoietic cell transplantation involves the infusion of blood stem cells (peripheral/umbilical cord blood, bone marrow) into a patient to reconstitute the blood system. Get detailed information about autologous and allogeneic transplant, including cell selection, HLA matching, and preparative regimens, and the acute complications and late effects of treatment in this summary for clinicians.

CXCR1 remodels the vascular niche to promote hematopoietic stem and progenitor cell engraftment

PubMed Central

Blaser, Bradley W.; Moore, Jessica L.; Hagedorn, Elliott J.; Li, Brian; Riquelme, Raquel; Yang, Song; Zhou, Yi; Tamplin, Owen J.; Binder, Vera

2017-01-01

The microenvironment is an important regulator of hematopoietic stem and progenitor cell (HSPC) biology. Recent advances marking fluorescent HSPCs have allowed exquisite visualization of HSPCs in the caudal hematopoietic tissue (CHT) of the developing zebrafish. Here, we show that the chemokine cxcl8 and its receptor, cxcr1, are expressed by zebrafish endothelial cells, and we identify cxcl8/cxcr1 signaling as a positive regulator of HSPC colonization. Single-cell tracking experiments demonstrated that this is a result of increases in HSPC–endothelial cell “cuddling,” HSPC residency time within the CHT, and HSPC mitotic rate. Enhanced cxcl8/cxcr1 signaling was associated with an increase in the volume of the CHT and induction of cxcl12a expression. Finally, using parabiotic zebrafish, we show that cxcr1 acts HSPC nonautonomously to improve the efficiency of donor HSPC engraftment. This work identifies a mechanism by which the hematopoietic niche remodels to promote HSPC engraftment and suggests that cxcl8/cxcr1 signaling is a potential therapeutic target in patients undergoing hematopoietic stem cell transplantation. PMID:28351983

C/EBPδ deficiency sensitizes mice to ionizing radiation-induced hematopoietic and intestinal injury.

PubMed

Pawar, Snehalata A; Shao, Lijian; Chang, Jianhui; Wang, Wenze; Pathak, Rupak; Zhu, Xiaoyan; Wang, Junru; Hendrickson, Howard; Boerma, Marjan; Sterneck, Esta; Zhou, Daohong; Hauer-Jensen, Martin

2014-01-01

Knowledge of the mechanisms involved in the radiation response is critical for developing interventions to mitigate radiation-induced injury to normal tissues. Exposure to radiation leads to increased oxidative stress, DNA-damage, genomic instability and inflammation. The transcription factor CCAAT/enhancer binding protein delta (Cebpd; C/EBPδ is implicated in regulation of these same processes, but its role in radiation response is not known. We investigated the role of C/EBPδ in radiation-induced hematopoietic and intestinal injury using a Cebpd knockout mouse model. Cebpd-/- mice showed increased lethality at 7.4 and 8.5 Gy total-body irradiation (TBI), compared to Cebpd+/+ mice. Two weeks after a 6 Gy dose of TBI, Cebpd-/- mice showed decreased recovery of white blood cells, neutrophils, platelets, myeloid cells and bone marrow mononuclear cells, decreased colony-forming ability of bone marrow progenitor cells, and increased apoptosis of hematopoietic progenitor and stem cells compared to Cebpd+/+ controls. Cebpd-/- mice exhibited a significant dose-dependent decrease in intestinal crypt survival and in plasma citrulline levels compared to Cebpd+/+ mice after exposure to radiation. This was accompanied by significantly decreased expression of γ-H2AX in Cebpd-/- intestinal crypts and villi at 1 h post-TBI, increased mitotic index at 24 h post-TBI, and increase in apoptosis in intestinal crypts and stromal cells of Cebpd-/- compared to Cebpd+/+ mice at 4 h post-irradiation. This study uncovers a novel biological function for C/EBPδ in promoting the response to radiation-induced DNA-damage and in protecting hematopoietic and intestinal tissues from radiation-induced injury.

Radiation-induced hematopoietic myelosuppression and genotoxicity get significantly countered by active principles of Podophyllum hexandrum: A study in strain 'A' mice.

PubMed

Verma, Savita; Gupta, Manju Lata

2015-01-01

To investigate the protective role of a novel formulation, prepared by a combination of three active principles isolated from Podophyllum hexandrum (G-002M), against radiation- mediated hematopoietic suppression and cytogenetic aberrations in lethally irradiated mice. G-002M, a combination of podophyllotoxin, podophyllotoxin-β-D glucoside and rutin, was administered intramuscularly in mice (- 1 h) to radiation (9 Gy) exposure. The animals were autopsied at different time intervals for further studies. Loss of bone marrow progenitor cells, altered myeloid/erythroid ratio, serum erythropoietin and pancytopenia in irradiated mice was found significantly (p < 0.001) ameliorated in G-002M pre-administered mice within 30 d. Bcl-2 (B-cell lymphoma 2) and BAX (Bcl-2-associated X) protein expression was also positively (p < 0.001) countered in these mice. Chromosomal aberrations in 30 d were found remarkably (p < 0.001) reduced in marrow of G-002M pretreated mice. Accelerated antioxidants, reduced DNA damage, stimulated lymphocyte proliferation and minimal cellular atrophy in spleen were some of the other key features observed in G-002M administered mice. Reduction in hematopoietic aplasia and chromosomal aberrations, besides, early recovery in bone marrow and spleen of G-002M pretreated mice, could be attributed to its free radical scavenging, DNA protecting and apoptotic proteins modulating ability against radiation.

Current state of hematopoietic cell transplantation in CLL as smart therapies emerge.

PubMed

Kharfan-Dabaja, Mohamed A; El-Asmar, Jessica; Awan, Farrukh T; Hamadani, Mehdi; Ayala, Ernesto

2016-03-01

Novel therapies targeting various kinases downstream of the B-cell receptor have emerged along with monoclonal antibodies and BCL-2 antagonists, and are changing the therapeutic landscape of chronic lymphocytic leukemia. However, cure remains unattainable unless eligible patients are offered an allogeneic hematopoietic cell transplant. Access to allogeneic hematopoietic cell transplantation has expanded considerably with availability of reduced intensity conditioning regimens which is capable offering durable remissions even in poor-risk disease. Encouraging data from ibrutinib and venetoclax in Del17p is challenging the notion of disease eradication as the ultimate therapeutic goal to a new concept of merely disease control. By favoring the non-transplant approach, patients should be aware that there are no established salvage therapies, yet, to rescue disease progression after ibrutinib. When disease eradication is the desirable approach, a reduced intensity conditioning allogeneic hematopoietic cell transplant is the preferred choice at this time. Copyright © 2016 Elsevier Ltd. All rights reserved.

Time lapse video recordings of highly purified human hematopoietic progenitor cells in culture.

PubMed

Denkers, I A; Dragowska, W; Jaggi, B; Palcic, B; Lansdorp, P M

1993-05-01

Major hurdles in studies of stem cell biology include the low frequency and heterogeneity of human hematopoietic precursor cells in bone marrow and the difficulty of directly studying the effect of various culture conditions and growth factors on such cells. We have adapted the cell analyzer imaging system for monitoring and recording the morphology of limited numbers of cells under various culture conditions. Hematopoietic progenitor cells with a CD34+ CD45RAlo CD71lo phenotype were purified from previously frozen organ donor bone marrow by fluorescence activated cell sorting. Cultures of such cells were analyzed with the imaging system composed of an inverted microscope contained in an incubator, a video camera, an optical memory disk recorder and a computer-controlled motorized microscope XYZ precision stage. Fully computer-controlled video images at defined XYZ positions were captured at selected time intervals and recorded at a predetermined sequence on an optical memory disk. In this study, the cell analyzer system was used to obtain descriptions and measurements of hematopoietic cell behavior, like cell motility, cell interactions, cell shape, cell division, cell cycle time and cell size changes under different culture conditions.

Suppression of HLA Expression by Lentivirus-mediated Gene Transfer of siRNA Cassettes and In Vivo Chemoselection to Enhance Hematopoietic Stem Cell Transplantation

PubMed Central

Hacke, Katrin; Falahati, Rustom; Flebbe-Rehwaldt, Linda; Kasahara, Noriyuki; Gaensler, Karin M. L.

2010-01-01

Current approaches for hematopoietic stem cell (HSC) and organ transplantation are limited by donor and host-mediated immune responses to allo-antigens. Application of these therapies is limited by the toxicity of preparative and post-transplant immunosuppressive regimens and a shortage of appropriate HLA-matched donors. We have been exploring two complementary approaches for genetically modifying donor cells that achieve long-term suppression of cellular proteins that elicit host immune responses to mismatched donor antigens, and provide a selective advantage to genetically engineered donor cells after transplantation. The first approach is based on recent advances that make feasible targeted down-regulation of HLA expression. Suppression of HLA expression could help to overcome limitations imposed by extensive HLA polymorphisms that restrict the availability of suitable donors. Accordingly, we have recently investigated whether knockdown of HLA by RNA interference (RNAi) enables allogeneic cells to evade immune recognition. For efficient and stable delivery of short hairpin-type RNAi constructs (shRNA), we employed lentivirus-based gene transfer vectors that integrate into genomic DNA, thereby permanently modifying transduced donor cells. Lentivirus-mediated delivery of shRNA targeting pan-Class I and allele-specific HLA achieved efficient and dose-dependent reduction in surface expression of HLA in human cells, and enhanced resistance to allo-reactive T lymphocyte-mediated cytotoxicity, while avoiding non-MHC restricted killing. Complementary strategies for genetic engineering of HSC that would provide a selective advantage for transplanted donor cells and enable successful engraftment with less toxic preparative and immunosuppressive regimens would increase the numbers of individuals to whom HLA suppression therapy could be offered. Our second strategy is to provide a mechanism for in vivo selection of genetically modified HSC and other donor cells. We have

Acquired initiating mutations in early hematopoietic cells of CLL patients.

PubMed

Damm, Frederik; Mylonas, Elena; Cosson, Adrien; Yoshida, Kenichi; Della Valle, Véronique; Mouly, Enguerran; Diop, M'boyba; Scourzic, Laurianne; Shiraishi, Yuichi; Chiba, Kenichi; Tanaka, Hiroko; Miyano, Satoru; Kikushige, Yoshikane; Davi, Frederick; Lambert, Jérôme; Gautheret, Daniel; Merle-Béral, Hélène; Sutton, Laurent; Dessen, Philippe; Solary, Eric; Akashi, Koichi; Vainchenker, William; Mercher, Thomas; Droin, Nathalie; Ogawa, Seishi; Nguyen-Khac, Florence; Bernard, Olivier A

2014-09-01

Appropriate cancer care requires a thorough understanding of the natural history of the disease, including the cell of origin, the pattern of clonal evolution, and the functional consequences of the mutations. Using deep sequencing of flow-sorted cell populations from patients with chronic lymphocytic leukemia (CLL), we established the presence of acquired mutations in multipotent hematopoietic progenitors. Mutations affected known lymphoid oncogenes, including BRAF, NOTCH1, and SF3B1. NFKBIE and EGR2 mutations were observed at unexpectedly high frequencies, 10.7% and 8.3% of 168 advanced-stage patients, respectively. EGR2 mutations were associated with a shorter time to treatment and poor overall survival. Analyses of BRAF and EGR2 mutations suggest that they result in deregulation of B-cell receptor (BCR) intracellular signaling. Our data propose disruption of hematopoietic and early B-cell differentiation through the deregulation of pre-BCR signaling as a phenotypic outcome of CLL mutations and show that CLL develops from a pre-leukemic phase. The origin and pathogenic mechanisms of CLL are not fully understood. The current work indicates that CLL develops from pre-leukemic multipotent hematopoietic progenitors carrying somatic mutations. It advocates for abnormalities in early B-cell differentiation as a phenotypic convergence of the diverse acquired mutations observed in CLL. ©2014 American Association for Cancer Research.

Open the gates: vascular neurocrine signaling mobilizes hematopoietic stem and progenitor cells.

PubMed

Itkin, Tomer; Gómez-Salinero, Jesús María; Rafii, Shahin

2017-12-01

Mobilization of hematopoietic stem and progenitor cells (HSPCs) from the bone marrow (BM) into the peripheral blood is a complex process that is enhanced dramatically under stress-induced conditions. A better understanding of how the mobilization process is regulated will likely facilitate the development of improved clinical protocols for stem cell harvesting and transplantation. In this issue of the JCI, Singh et al. (1) showed that the truncated cleaved form of neurotransmitter neuropeptide Y (NPY) actively promotes a breach of BM vascular sinusoidal portals, thereby augmenting HSPC trafficking to the circulation. The authors report a previously unrecognized axis, in which expression of the enzyme dipeptidylpeptidase-4 (DPP4)/CD26 by endothelial cells activates NPY-mediated signaling by increasing the bioavailability of the truncated form of NPY. These findings underscore the importance of and urgency to develop pharmacological therapies that target the vasculature and regulate diverse aspects of hematopoiesis, such as HSPC trafficking, in steady-state and stress-induced conditions.

Long survival and immunologic reconstitution following transplantation with syngeneic or allogeneic fetal liver and neonatal spleen cells. [X radiation, mice

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yunis, E.J.; Fernandes, G.; Smith, J.

1976-12-01

Spleen cells from newborn syngeneic and allogeneic mice that lack fully differentiated T lymphocytes can be used as a hematopoietic source to reconstitute both hematopoietic and lymphoid systems of lethally irradiated mice without producing a GVHR. Fetal liver cells from syngeneic and allogeneic mice that lack postthymic T lymphocytes can also be used for hematopoietic and immunologic reconstitution of lethally irradiated mice without producing GVHR. Immunologic deficiency is observed in some experiments in mice given supralethal irradiation (1000 R) and fetal liver as reconstituting hematopoietic tissue. The findings suggest that T cells, at an early stage of differentiation, are moremore » susceptible to tolerance induction than are T lymphocytes at later stages of differentiation and do not, in general, produce GVHR. It is postulated that hematopoietic cells, free of postthymic lymphoid cells, can be used for hematopoietic or immunologic reconstitution and celular engineering without producing GVHD.« less

Longitudinal trajectory of sexual functioning after hematopoietic cell transplantation: impact of chronic graft-versus-host disease and total body irradiation

PubMed Central

Wong, F. Lennie; Francisco, Liton; Togawa, Kayo; Kim, Heeyoung; Bosworth, Alysia; Atencio, Liezl; Hanby, Cara; Grant, Marcia; Kandeel, Fouad; Forman, Stephen J.

2013-01-01

This prospective study described the trajectory of sexual well-being from before hematopoietic cell transplantation (HCT) to 3 years after in 131 allogeneic and 146 autologous HCT recipients using Derogatis Interview for Sexual Function and Derogatis Global Sexual Satisfaction Index. Sixty-one percent of men and 37% of women were sexually active pre-HCT; the prevalence declined to 51% (P = .01) in men and increased to 48% (P = .02) in women at 3 years post-HCT. After HCT, sexual satisfaction declined in both sexes (P < .001). All sexual function domains were worse in women compared with men (P ≤ .001). Orgasm (P = .002) and drive/relationship (P < .001) declined in men, but sexual cognition/fantasy (P = .01) and sexual behavior/experience (P = .01) improved in women. Older age negatively impacted sexual function post-HCT in both sexes (P < .01). Chronic graft-versus-host disease was associated with lower sexual cognition/fantasy (P = .003) and orgasm (P = .006) in men and sexual arousal (P = .05) and sexual satisfaction (P = .005) in women. All male sexual function domains declined after total body irradiation (P < .05). This study identifies vulnerable subpopulations that could benefit from interventional strategies to improve sexual well-being. PMID:24159171

Direct Toll-like receptor-mediated stimulation of hematopoietic stem and progenitor cells occurs in vivo and promotes differentiation toward macrophages.

PubMed

Megías, Javier; Yáñez, Alberto; Moriano, Silvia; O'Connor, José-Enrique; Gozalbo, Daniel; Gil, María-Luisa

2012-07-01

As Toll-like receptors (TLRs) are expressed by hematopoietic stem and progenitor cells (HSPCs), they may play a role in hematopoiesis in response to pathogens during infection. We show here that TLR2, TLR4, and TLR9 agonists (tripalmitoyl-S-glyceryl-L-Cys-Ser-(Lys)4 [Pam3CSK4], lipopolysaccharide [LPS], and CpG oligodeoxynucleotide [ODN]) induce the in vitro differentiation of purified murine lineage negative cells (Lin(-) ) as well as HSPCs (identified as Lin(-) c-Kit(+) Sca-1(+) IL-7Rα(-) [LKS] cells) toward macrophages (Mph), through a myeloid differentiation factor 88 (MyD88)-dependent pathway. In order to investigate the possible direct interaction of soluble microorganism-associated molecular patterns and TLRs on HSPCs in vivo, we designed a new experimental approach: purified Lin(-) and LKS cells from bone marrow of B6Ly5.1 mice (CD45.1 alloantigen) were transplanted into TLR2(-/-) , TLR4(-/-) , or MyD88(-/-) mice (CD45.2 alloantigen), which were then injected with soluble TLR ligands (Pam3CSK4, LPS, or ODN, respectively). As recipient mouse cells do not recognize the TLR ligands injected, interference by soluble mediators secreted by recipient cells is negligible. Transplanted cells were detected in the spleen and bone marrow of recipient mice, and in response to soluble TLR ligands, cells differentiated preferentially to Mph. These results show, for the first time, that HSPCs may be directly stimulated by TLR agonists in vivo, and that the engagement of these receptors induces differentiation toward Mph. Therefore, HSPCs may sense pathogen or pathogen-derived products directly during infection, inducing a rapid generation of cells of the innate immune system. Copyright © 2012 AlphaMed Press.

Nutraceutical augmentation of circulating endothelial progenitor cells and hematopoietic stem cells in human subjects.

PubMed

Mikirova, Nina A; Jackson, James A; Hunninghake, Ron; Kenyon, Julian; Chan, Kyle W H; Swindlehurst, Cathy A; Minev, Boris; Patel, Amit N; Murphy, Michael P; Smith, Leonard; Ramos, Famela; Ichim, Thomas E; Riordan, Neil H

2010-04-08

The medical significance of circulating endothelial or hematopoietic progenitors is becoming increasing recognized. While therapeutic augmentation of circulating progenitor cells using G-CSF has resulted in promising preclinical and early clinical data for several degenerative conditions, this approach is limited by cost and inability to perform chronic administration. Stem-Kine is a food supplement that was previously reported to augment circulating EPC in a pilot study. Here we report a trial in 18 healthy volunteers administered Stem-Kine twice daily for a 2 week period. Significant increases in circulating CD133 and CD34 cells were observed at days 1, 2, 7, and 14 subsequent to initiation of administration, which correlated with increased hematopoietic progenitors as detected by the HALO assay. Augmentation of EPC numbers in circulation was detected by KDR-1/CD34 staining and colony forming assays. These data suggest Stem-Kine supplementation may be useful as a stimulator of reparative processes associated with mobilization of hematopoietic and endothelial progenitors.

Spheroid Coculture of Hematopoietic Stem/Progenitor Cells and Monolayer Expanded Mesenchymal Stem/Stromal Cells in Polydimethylsiloxane Microwells Modestly Improves In Vitro Hematopoietic Stem/Progenitor Cell Expansion.

PubMed

Futrega, Kathryn; Atkinson, Kerry; Lott, William B; Doran, Michael R

2017-04-01

While two-dimensional (2D) monolayers of mesenchymal stem/stromal cells (MSCs) have been shown to enhance hematopoietic stem/progenitor cell (HSPC) expansion in vitro, expanded cells do not engraft long term in human recipients. This outcome is attributed to the failure of 2D culture to recapitulate the bone marrow (BM) niche signal milieu. Herein, we evaluated the capacity of a novel three-dimensional (3D) coculture system to support HSPC expansion in vitro. A high-throughput polydimethylsiloxane (PDMS) microwell platform was used to manufacture thousands of uniform 3D multicellular coculture spheroids. Relative gene expression in 3D spheroid versus 2D adherent BM-derived MSC cultures was characterized and compared with literature reports. We evaluated coculture spheroids, each containing 25-400 MSCs and 10 umbilical cord blood (CB)-derived CD34 + progenitor cells. At low exogenous cytokine concentrations, 2D and 3D MSC coculture modestly improved overall hematopoietic cell and CD34 + cell expansion outcomes. By contrast, a substantial increase in CD34 + CD38 - cell yield was observed in PDMS microwell cultures, regardless of the presence or absence of MSCs. This outcome indicated that CD34 + CD38 - cell culture yield could be increased using the microwell platform alone, even without MSC coculture support. We found that the increase in CD34 + CD38 - cell yield observed in PDMS microwell cultures did not translate to enhanced engraftment in NOD/SCID gamma (NSG) mice or a modification in the relative human hematopoietic lineages established in engrafted mice. In summary, there was no statistical difference in CD34 + cell yield from 2D or 3D cocultures, and MSC coculture support provided only modest benefit in either geometry. While the high-throughput 3D microwell platform may provide a useful model system for studying cells in coculture, further optimization will be required to generate HSPC yields suitable for use in clinical applications.

Spheroid Coculture of Hematopoietic Stem/Progenitor Cells and Monolayer Expanded Mesenchymal Stem/Stromal Cells in Polydimethylsiloxane Microwells Modestly Improves In Vitro Hematopoietic Stem/Progenitor Cell Expansion

PubMed Central

Futrega, Kathryn; Atkinson, Kerry; Lott, William B.

2017-01-01

While two-dimensional (2D) monolayers of mesenchymal stem/stromal cells (MSCs) have been shown to enhance hematopoietic stem/progenitor cell (HSPC) expansion in vitro, expanded cells do not engraft long term in human recipients. This outcome is attributed to the failure of 2D culture to recapitulate the bone marrow (BM) niche signal milieu. Herein, we evaluated the capacity of a novel three-dimensional (3D) coculture system to support HSPC expansion in vitro. A high-throughput polydimethylsiloxane (PDMS) microwell platform was used to manufacture thousands of uniform 3D multicellular coculture spheroids. Relative gene expression in 3D spheroid versus 2D adherent BM-derived MSC cultures was characterized and compared with literature reports. We evaluated coculture spheroids, each containing 25–400 MSCs and 10 umbilical cord blood (CB)-derived CD34+ progenitor cells. At low exogenous cytokine concentrations, 2D and 3D MSC coculture modestly improved overall hematopoietic cell and CD34+ cell expansion outcomes. By contrast, a substantial increase in CD34+CD38− cell yield was observed in PDMS microwell cultures, regardless of the presence or absence of MSCs. This outcome indicated that CD34+CD38− cell culture yield could be increased using the microwell platform alone, even without MSC coculture support. We found that the increase in CD34+CD38− cell yield observed in PDMS microwell cultures did not translate to enhanced engraftment in NOD/SCID gamma (NSG) mice or a modification in the relative human hematopoietic lineages established in engrafted mice. In summary, there was no statistical difference in CD34+ cell yield from 2D or 3D cocultures, and MSC coculture support provided only modest benefit in either geometry. While the high-throughput 3D microwell platform may provide a useful model system for studying cells in coculture, further optimization will be required to generate HSPC yields suitable for use in clinical applications. PMID:28406754

Investigating Cell Surface Markers on Normal Hematopoietic Stem Cells in Three Different Niche Conditions

PubMed Central

Garg, Swati; Madkaikar, Manisha

2013-01-01

Hematopoietic stem cells are of therapeutic interest to the clinicians and researchers due to their promising assistance in management of malignant and inherited hematological conditions. Evaluation of cell surface markers using multiparametric flow cytometry is a well adapted qualitative measure of cells in question for many years. An artillery of these markers has been studied in hematological malignancies and related disorders. However, their role and differential expression on normal hematopoietic stem cells from clinically available sources is not always described carefully. In the present study, we attempted to evaluate expression of CD44, CD90, CD96 and CD123 in three clinically available sources of normal HSCs (Hematopoietic stem cells). Sources of HSCs in the present study involved umbilical cord blood (UCB), normal bone marrow (NBM) and bone marrow from idiopathic thrombocytopenic purpura (ITP) patients (IBM). CD44 is an important homing receptor while CD90 is involved in maintaining stem cell quiescent. CD96 is known to be leukemia specific marker and CD123 is involved in stem cell differentiation and survival. We observed a significant difference in expression CD44, CD90 and CD123 on normal HSCs derived from umbilical cord and ITP marrow. CD96 was highly expressed on HSCs obtained from ITP marrow. Investigating expression of these markers on normal HSCs in different niches will be helpful in correlating their function with niche condition and delineating their ‘abnormal’ expression from the normal. PMID:24386557

Investigating cell surface markers on normal hematopoietic stem cells in three different niche conditions.

PubMed

Garg, Swati; Madkaikar, Manisha; Ghosh, Kanjaksha

2013-11-01

Hematopoietic stem cells are of therapeutic interest to the clinicians and researchers due to their promising assistance in management of malignant and inherited hematological conditions. Evaluation of cell surface markers using multiparametric flow cytometry is a well adapted qualitative measure of cells in question for many years. An artillery of these markers has been studied in hematological malignancies and related disorders. However, their role and differential expression on normal hematopoietic stem cells from clinically available sources is not always described carefully. In the present study, we attempted to evaluate expression of CD44, CD90, CD96 and CD123 in three clinically available sources of normal HSCs (Hematopoietic stem cells). Sources of HSCs in the present study involved umbilical cord blood (UCB), normal bone marrow (NBM) and bone marrow from idiopathic thrombocytopenic purpura (ITP) patients (IBM). CD44 is an important homing receptor while CD90 is involved in maintaining stem cell quiescent. CD96 is known to be leukemia specific marker and CD123 is involved in stem cell differentiation and survival. We observed a significant difference in expression CD44, CD90 and CD123 on normal HSCs derived from umbilical cord and ITP marrow. CD96 was highly expressed on HSCs obtained from ITP marrow. Investigating expression of these markers on normal HSCs in different niches will be helpful in correlating their function with niche condition and delineating their 'abnormal' expression from the normal.

Effect of Calcium-Infiltrated Hydroxyapatite Scaffolds on the Hematopoietic Fate of Human Umbilical Vein Endothelial Cells.

PubMed

Zhang, Qinghao; Gerlach, Jörg C; Schmelzer, Eva; Nettleship, Ian

2017-01-01

Foamed hydroxyapatite offers a three-dimensional scaffold for the development of bone constructs, mimicking perfectly the in vivo bone structure. In vivo, calcium release at the surface is assumed to provide a locally increased gradient supporting the maintenance of the hematopoietic stem cells niche. We fabricated hydroxyapatite scaffolds with high surface calcium concentration by infiltration, and used human umbilical vein endothelial cells (HUVECs) as a model to study the effects on hematopoietic lineage direction. HUVECs are umbilical vein-derived and thus possess progenitor characteristics, with a prospective potential to give rise to hematopoietic lineages. HUVECs were cultured for long term on three-dimensional porous hydroxyapatite scaffolds, which were either infiltrated biphasic foams or untreated. Controls were cultured in two-dimensional dishes. The release of calcium into culture medium was determined, and cells were analyzed for typical hematopoietic and endothelial gene expressions, surface markers by flow cytometry, and hematopoietic potential using colony-forming unit assays. Our results indicate that the biphasic foams promoted a hematopoietic lineage direction of HUVECs, suggesting an improved in vivo-like scaffold for hematopoietic bone tissue engineering. © 2017 S. Karger AG, Basel.

Correction of the sickle cell disease mutation in human hematopoietic stem/progenitor cells.

PubMed

Hoban, Megan D; Cost, Gregory J; Mendel, Matthew C; Romero, Zulema; Kaufman, Michael L; Joglekar, Alok V; Ho, Michelle; Lumaquin, Dianne; Gray, David; Lill, Georgia R; Cooper, Aaron R; Urbinati, Fabrizia; Senadheera, Shantha; Zhu, Allen; Liu, Pei-Qi; Paschon, David E; Zhang, Lei; Rebar, Edward J; Wilber, Andrew; Wang, Xiaoyan; Gregory, Philip D; Holmes, Michael C; Reik, Andreas; Hollis, Roger P; Kohn, Donald B

2015-04-23

Sickle cell disease (SCD) is characterized by a single point mutation in the seventh codon of the β-globin gene. Site-specific correction of the sickle mutation in hematopoietic stem cells would allow for permanent production of normal red blood cells. Using zinc-finger nucleases (ZFNs) designed to flank the sickle mutation, we demonstrate efficient targeted cleavage at the β-globin locus with minimal off-target modification. By co-delivering a homologous donor template (either an integrase-defective lentiviral vector or a DNA oligonucleotide), high levels of gene modification were achieved in CD34(+) hematopoietic stem and progenitor cells. Modified cells maintained their ability to engraft NOD/SCID/IL2rγ(null) mice and to produce cells from multiple lineages, although with a reduction in the modification levels relative to the in vitro samples. Importantly, ZFN-driven gene correction in CD34(+) cells from the bone marrow of patients with SCD resulted in the production of wild-type hemoglobin tetramers. © 2015 by The American Society of Hematology.

ADAM17 limits the expression of CSF1R on murine hematopoietic progenitors

PubMed Central

Becker, Amy M.; Walcheck, Bruce; Bhattacharya, Deepta

2014-01-01

All-lymphoid progenitors (ALPs) yield few myeloid cells in vivo, but readily generate such cells in vitro. The basis for this difference remains unknown. We hypothesized that ALPs limit responsiveness to in vivo concentrations of myeloid-promoting cytokines by reducing expression of the corresponding receptors, potentially through post-transcriptional mechanisms. Consistent with such a mechanism, ALPs express higher levels of Csf1r transcripts than their upstream precursors, yet show limited cell surface protein expression of CSF1R. ALPs and other hematopoietic progenitors deficient in ADAM17, a metalloprotease that can cleave CSF1R, display elevated cell surface CSF1R expression. Adam17−/− ALPs, however, fail to yield myeloid cells upon transplantation into irradiated recipients. Moreover, Adam17−/− ALPs yield fewer macrophages in vitro than control ALPs at high concentrations of M-CSF. Mice with hematopoietic-specific deletion of Adam17 have grossly normal numbers of myeloid and lymphoid progenitors and mature cells in vivo. These data demonstrate that ADAM17 limits CSF1R protein expression on hematopoietic progenitors, but that compensatory mechanisms prevent elevated CSF1R levels from altering lymphoid progenitor potential. PMID:25308957

Reduced incidence of interstitial pneumonitis after allogeneic hematopoietic stem cell transplantation using a modified technique of total body irradiation.

PubMed

Chiang, Yun; Tsai, Cheng-Hong; Kuo, Sung-Hsin; Liu, Chieh-Yu; Yao, Ming; Li, Chi-Cheng; Huang, Shang-Yi; Ko, Bor-Sheng; Lin, Chien-Ting; Hou, Hsin-An; Chou, Wen-Chien; Liu, Jia-Hau; Lin, Chien-Chin; Wu, Shang-Ju; Hsu, Szu-Chun; Chen, Yao-Chang; Lin, Kai-Hsin; Lin, Dong-Tsamn; Chou, Hsien-Tang; Lu, Meng-Yu; Yang, Yung-Li; Chang, Hsiu-Hao; Liu, Ming-Chih; Liao, Xiu-Wen; Wu, Jian-Kuen; Chou, Sheng-Chieh; Cheng, Chieh-Lung; Chen, Chien-Yuan; Tsay, Woei; Tien, Hwei-Fang; Tang, Jih-Luh; Chen, Yu-Hsuan

2016-11-10

Allogeneic hematopoietic stem cell transplantation is a curative-intent treatment for patients with high-risk hematologic diseases. However, interstitial pneumonitis (IP) and other toxicities remain major concerns after total body irradiation (TBI). We have proposed using linear accelerators with rice-bag compensators for intensity modulation (IM-TBI), as an alternative to the traditional cobalt-60 teletherapy with lung-shielding technique (Co-TBI). Patients who received a TBI-based myeloablative conditioning regimen between 1995 and 2014 were recruited consecutively. Before March 2007, TBI was delivered using Co-TBI (n = 181); afterward, TBI was administered using IM-TBI (n = 126). Forty-four patients developed IP; of these cases, 19 were idiopathic. The IP-related mortality rate was 50% in the total IP cohort and 63% in the idiopathic subgroup. The 1-year cumulative incidences of IP and idiopathic IP were 16.5% and 7.4%, respectively; both rates were significantly higher in the Co-TBI group than in the IM-TBI group. Multivariate analysis revealed that Co-TBI was an independent prognostic factor for both total and idiopathic IP. In the acute myeloid leukemia subgroup, patients with different TBI techniques had similar outcomes for both overall and relapse-free survival. In conclusion, IM-TBI is an easy and effective TBI technique that could substantially reduce the complication rate of IP without compromising treatment efficacy.

Impact of Cranial Irradiation Added to Intrathecal Conditioning in Hematopoietic Cell Transplantation in Adult Acute Myeloid Leukemia With Central Nervous System Involvement

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mayadev, Jyoti S.; Department of Radiation Oncology University of California-Davis Medical Center, Davis, CA; Douglas, James G., E-mail: drjay@u.washington.ed

Purpose: Neither the prognostic importance nor the appropriate management of central nervous system (CNS) involvement is known for patients with acute myeloid leukemia (AML) undergoing hematopoietic cell transplantation (HCT). We examined the impact of a CNS irradiation boost to standard intrathecal chemotherapy (ITC). Methods and Materials: From 1995 to 2005, a total of 648 adult AML patients received a myeloablative HCT: 577 patients were CNS negative (CNS-), and 71 were CNS positive (CNS+). Of the 71 CNS+ patients, 52 received intrathecal chemotherapy alone (CNS+ITC), and 19 received ITC plus an irradiation boost (CNS+RT). Results: The CNS-, CNS+ITC, and CNS+RT patientsmore » had 1- and 5-year relapse-free survivals (RFS) of 43% and 35%, 15% and 6%, and 37% and 32%, respectively. CNS+ITC patients had a statistically significant worse RFS compared with CNS- patients (hazard ratio [HR], 2.65; 95% confidence interval [CI], 2.0-3.6; p < 0.0001). CNS+RT patients had improved relapse free survival over that of CNS+ITC patients (HR, 0.45; 95% CI, 0.2-0.8; p = 0.01). The 1- and 5-year overall survivals (OS) of patients with CNS-, CNS+ITC, and CNS+RT, were 50% and 38%, 21% and 6%, and 53% and 42%, respectively. The survival of CNS+RT were significantly better than CNS+ITC patients (p = 0.004). After adjusting for known risk factors, CNS+RT patients had a trend toward lower relapse rates and reduced nonrelapse mortality. Conclusions: CNS+ AML is associated with a poor prognosis. The role of a cranial irradiation boost to intrathecal chemotherapy appears to mitigate the risk of CNS disease, and needs to be further investigated to define optimal treatment strategies.« less

Identification of the Niche and Phenotype of the First Human Hematopoietic Stem Cells

PubMed Central

Ivanovs, Andrejs; Rybtsov, Stanislav; Anderson, Richard A.; Turner, Marc L.; Medvinsky, Alexander

2014-01-01

Summary In various vertebrate species, the dorsal aorta (Ao) is the site of specification of adult hematopoietic stem cells (HSCs). It has been observed that the upregulation of essential hematopoietic transcription factors and the formation of specific intra-aortic hematopoietic cell clusters occur predominantly in the ventral domain of the Ao (AoV). In the mouse, the first HSCs emerge in the AoV. Here, we demonstrate that in the human embryo the first definitive HSCs also emerge asymmetrically and are localized to the AoV, which thus identifies a functional niche for developing human HSCs. Using magnetic cell separation and xenotransplantations, we show that the first human HSCs are CD34+VE-cadherin+CD45+C-KIT+THY-1+Endoglin+RUNX1+CD38−/loCD45RA−. This population harbors practically all committed hematopoietic progenitors and is underrepresented in the dorsal domain of the Ao (AoD) and urogenital ridges (UGRs). The present study provides a foundation for analysis of molecular mechanisms underpinning embryonic specification of human HSCs. PMID:24749070

Do autologous peripheral blood cell transplants provide more than hematopoietic recovery?

PubMed

Kessinger, A

1995-07-01

Bone marrow damage caused by myeloablative radiation therapy and/or chemotherapy can be repaired by intravenously infusing viable stem/progenitor cells collected from either blood or bone marrow. The hematopoietic graft product contains both stem/progenitor cells and populations of hematopoietic and nonhematopoietic (accessory) cells. The frequency of accessory cell types varies with the source of the graft product; marrow or blood. Reinfusion of these accessory cells causes effects other than the hematopoietic restoration provided by the stem/progenitor cells such as graft versus host disease and graft versus leukemia effect after allogeneic transplants. Effects of infused accessory cells in the autologous setting are less well studied and could provide ancillary advantages and/or disadvantages to the patient. Do these additional effects actually occur, and, if they do, are they more likely to appear following peripheral blood cell transplants (PBCT) or after autologous bone marrow transplants (AMBT)? Preliminary data are beginning to accumulate which suggest that reinfusion of occult tumor cells is less likely with PBCT, that immune reconstitution is different depending on the source of the autograft and that, for certain diseases, patient event-free survival following PBCT rather than ABMT may be better. However, infusion of occult tumor cells may result in re-establishment of the malignancy. If the accessory cells (including potential occult tumor cells) are eliminated from the product before transplant, will the patient have a better clinical outcome, or would benefits provided by infused accessory cells outweigh the risks of infused occult tumor cells? These controversial issues are in the very early stages of investigation.

c-Kit-Mediated Functional Positioning of Stem Cells to Their Niches Is Essential for Maintenance and Regeneration of Adult Hematopoiesis

PubMed Central

Kimura, Yuki; Ding, Bisen; Imai, Norikazu; Nolan, Daniel J.; Butler, Jason M.; Rafii, Shahin

2011-01-01

The mechanism by which hematopoietic stem and progenitor cells (HSPCs) through interaction with their niches maintain and reconstitute adult hematopoietic cells is unknown. To functionally and genetically track localization of HSPCs with their niches, we employed novel mutant loxPs, lox66 and lox71 and Cre-recombinase technology to conditionally delete c-Kit in adult mice, while simultaneously enabling GFP expression in the c-Kit-deficient cells. Conditional deletion of c-Kit resulted in hematopoietic failure and splenic atrophy both at steady state and after marrow ablation leading to the demise of the treated adult mice. Within the marrow, the c-Kit-expressing GFP+ cells were positioned to Kit ligand (KL)-expressing niche cells. This c-Kit-mediated cellular adhesion was essential for long-term maintenance and expansion of HSPCs. These results lay the foundation for delivering KL within specific niches to maintain and restore hematopoiesis. PMID:22046410

PPARγ antagonist attenuates mouse immune-mediated bone marrow failure by inhibition of T cell function

PubMed Central

Sato, Kazuya; Feng, Xingmin; Chen, Jichun; Li, Jungang; Muranski, Pawel; Desierto, Marie J.; Keyvanfar, Keyvan; Malide, Daniela; Kajigaya, Sachiko; Young, Neal S.

2016-01-01

Acquired aplastic anemia is an immune-mediated disease, in which T cells target hematopoietic cells; at presentation, the bone marrow is replaced by fat. It was reported that bone marrow adipocytes were negative regulators of hematopoietic microenvironment. To examine the role of adipocytes in bone marrow failure, we investigated peroxisomal proliferator-activated receptor gamma, a key transcription factor in adipogenesis, utilizing an antagonist of this factor called bisphenol-A-diglycidyl-ether. While bisphenol-A-diglycidyl-ether inhibited adipogenesis as expected, it also suppressed T cell infiltration of bone marrow, reduced plasma inflammatory cytokines, decreased expression of multiple inflammasome genes, and ameliorated marrow failure. In vitro, bisphenol-A-diglycidyl-ether suppressed activation and proliferation, and reduced phospholipase C gamma 1 and nuclear factor of activated T-cells 1 expression, as well as inhibiting calcium flux in T cells. The in vivo effect of bisphenol-A-diglycidyl-ether on T cells was confirmed in a second immune-mediated bone marrow failure model, using different strains and non-major histocompatibility antigen mismatched: bisphenol-A-diglycidyl-ether ameliorated marrow failure by inhibition of T cell infiltration of bone marrow. Our data indicate that peroxisomal proliferator-activated receptor gamma antagonists may attenuate murine immune-mediated bone marrow failure, at least in part, by suppression of T cell activation, which might hold implications in the application of peroxisomal proliferator-activated receptor gamma antagonists in immune-mediated pathophysiologies, both in the laboratory and in the clinic. Genetically “fatless” mice developed bone marrow failure with accumulation of marrow adipocytes in our model, even in the absence of body fat, suggesting different mechanisms of systematic and marrow adipogenesis and physiologic versus pathophysiologic fat accumulation. PMID:26589913

Oncogenic Kras initiates leukemia in hematopoietic stem cells.

PubMed

Sabnis, Amit J; Cheung, Laurene S; Dail, Monique; Kang, Hio Chung; Santaguida, Marianne; Hermiston, Michelle L; Passegué, Emmanuelle; Shannon, Kevin; Braun, Benjamin S

2009-03-17

How oncogenes modulate the self-renewal properties of cancer-initiating cells is incompletely understood. Activating KRAS and NRAS mutations are among the most common oncogenic lesions detected in human cancer, and occur in myeloproliferative disorders (MPDs) and leukemias. We investigated the effects of expressing oncogenic Kras(G12D) from its endogenous locus on the proliferation and tumor-initiating properties of murine hematopoietic stem and progenitor cells. MPD could be initiated by Kras(G12D) expression in a highly restricted population enriched for hematopoietic stem cells (HSCs), but not in common myeloid progenitors. Kras(G12D) HSCs demonstrated a marked in vivo competitive advantage over wild-type cells. Kras(G12D) expression also increased the fraction of proliferating HSCs and reduced the overall size of this compartment. Transplanted Kras(G12D) HSCs efficiently initiated acute T-lineage leukemia/lymphoma, which was associated with secondary Notch1 mutations in thymocytes. We conclude that MPD-initiating activity is restricted to the HSC compartment in Kras(G12D) mice, and that distinct self-renewing populations with cooperating mutations emerge during cancer progression.

Generation of induced pluripotent stem cells as a potential source of hematopoietic stem cells for transplant in PNH patients.

PubMed

Phondeechareon, Tanapol; Wattanapanitch, Methichit; U-Pratya, Yaowalak; Damkham, Chanapa; Klincumhom, Nuttha; Lorthongpanich, Chanchao; Kheolamai, Pakpoom; Laowtammathron, Chuti; Issaragrisil, Surapol

2016-10-01

Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired hemolytic anemia caused by lack of CD55 and CD59 on blood cell membrane leading to increased sensitivity of blood cells to complement. Hematopoietic stem cell transplantation (HSCT) is the only curative therapy for PNH, however, lack of HLA-matched donors and post-transplant complications are major concerns. Induced pluripotent stem cells (iPSCs) derived from patients are an attractive source for generating autologous HSCs to avoid adverse effects resulting from allogeneic HSCT. The disease involves only HSCs and their progeny; therefore, other tissues are not affected by the mutation and may be used to produce disease-free autologous HSCs. This study aimed to derive PNH patient-specific iPSCs from human dermal fibroblasts (HDFs), characterize and differentiate to hematopoietic cells using a feeder-free protocol. Analysis of CD55 and CD59 expression was performed before and after reprogramming, and hematopoietic differentiation. Patients' dermal fibroblasts expressed CD55 and CD59 at normal levels and the normal expression remained after reprogramming. The iPSCs derived from PNH patients had typical pluripotent properties and differentiation capacities with normal karyotype. After hematopoietic differentiation, the differentiated cells expressed early hematopoietic markers (CD34 and CD43) with normal CD59 expression. The iPSCs derived from HDFs of PNH patients have normal levels of CD55 and CD59 expression and hold promise as a potential source of HSCs for autologous transplantation to cure PNH patients.

Mice with targeted inactivation of ppap2b in endothelial and hematopoietic cells display enhanced vascular inflammation and permeability.

PubMed

Panchatcharam, Manikandan; Salous, Abdel K; Brandon, Jason; Miriyala, Sumitra; Wheeler, Jessica; Patil, Pooja; Sunkara, Manjula; Morris, Andrew J; Escalante-Alcalde, Diana; Smyth, Susan S

2014-04-01

Lipid phosphate phosphatase 3 (LPP3), encoded by the PPAP2B gene, is an integral membrane enzyme that dephosphorylates, and thereby terminates, the G-protein-coupled receptor-mediated signaling actions of lysophosphatidic acid (LPA) and sphingosine-1-phosphate. LPP3 is essential for normal vascular development in mice, and a common PPAP2B polymorphism is associated with increased risk of coronary artery disease in humans. Herein, we investigate the function of endothelial LPP3 to understand its role in the development and human disease. We developed mouse models with selective LPP3 deficiency in endothelial and hematopoietic cells. Tyrosine kinase Tek promoter-mediated inactivation of Ppap2b resulted in embryonic lethality because of vascular defects. LPP3 deficiency in adult mice, achieved using a tamoxifen-inducible Cre transgene under the control of the Tyrosine kinase Tek promoter, enhanced local and systemic inflammatory responses. Endothelial, but not hematopoietic, cell LPP3 deficiency led to significant increases in vascular permeability at baseline and enhanced sensitivity to inflammation-induced vascular leak. Endothelial barrier function was restored by pharmacological or genetic inhibition of either LPA production by the circulating lysophospholipase D autotaxin or of G-protein-coupled receptor-dependent LPA signaling. Our results identify a role for the autotaxin/LPA-signaling nexus as a mediator of endothelial permeability in inflammation and demonstrate that LPP3 limits these effects. These findings have implications for therapeutic targets to maintain vascular barrier function in inflammatory states.

Oral Complications in Hematopoietic Stem Cell Recipients: The Role of Inflammation

PubMed Central

Haverman, T. M.; Raber-Durlacher, J. E.; Rademacher, W. M. H.; Vokurka, S.; Epstein, J. B.; Huisman, C.; Hazenberg, M. D.; de Soet, J. J.; de Lange, J.; Rozema, F. R.

2014-01-01

Hematopoietic stem cell transplantation (HSCT) is widely used as a potentially curative treatment for patients with various hematological malignancies, bone marrow failure syndromes, and congenital immune deficiencies. The prevalence of oral complications in both autologous and allogeneic HSCT recipients remains high, despite advances in transplant medicine and in supportive care. Frequently encountered oral complications include mucositis, infections, oral dryness, taste changes, and graft versus host disease in allogeneic HSCT. Oral complications are associated with substantial morbidity and in some cases with increased mortality and may significantly affect quality of life, even many years after HSCT. Inflammatory processes are key in the pathobiology of most oral complications in HSCT recipients. This review article will discuss frequently encountered oral complications associated with HSCT focusing on the inflammatory pathways and inflammatory mediators involved in their pathogenesis. PMID:24817792

Genetic modification of hematopoietic stem cells with nonviral systems: past progress and future prospects.

PubMed

Papapetrou, E P; Zoumbos, N C; Athanassiadou, A

2005-10-01

Serious unwanted complications provoked by retroviral gene transfer into hematopoietic stem cells (HSCs) have recently raised the need for the development and assessment of alternative gene transfer vectors. Within this context, nonviral gene transfer systems are attracting increasing interest. Their main advantages include low cost, ease of handling and large-scale production, large packaging capacity and, most importantly, biosafety. While nonviral gene transfer into HSCs has been restricted in the past by poor transfection efficiency and transient maintenance, in recent years, biotechnological developments are converting nonviral transfer into a realistic approach for genetic modification of cells of hematopoietic origin. Herein we provide an overview of past accomplishments in the field of nonviral gene transfer into hematopoietic progenitor/stem cells and we point at future challenges. We argue that episomally maintained self-replicating vectors combined with physical methods of delivery show the greatest promise among nonviral gene transfer strategies for the treatment of disorders of the hematopoietic system.

Accelerating immune reconstitution after hematopoietic stem cell transplantation

PubMed Central

Tzannou, Ifigeneia; Leen, Ann M

2014-01-01

Viral infections remain a significant cause of morbidity and mortality after hematopoietic stem cell transplantation. Pharmacologic agents are effective against some pathogens, but they are costly and can be associated with significant toxicities. Thus, many groups have investigated adoptive T-cell transfer as a means of hastening immune reconstitution and preventing and treating viral infections. This review discusses the immunotherapeutic strategies that have been explored. PMID:25505959

Epigenetic regulation of hematopoietic stem cell aging

DOE Office of Scientific and Technical Information (OSTI.GOV)

Beerman, Isabel, E-mail: isabel.beerman@childrens.harvard.edu; Department of Pediatrics, Harvard Medical School, Boston, MA 02115; Program in Cellular and Molecular Medicine, Division of Hematology/Oncology, Boston Children's Hospital, MA 02116

2014-12-10

Aging is invariably associated with alterations of the hematopoietic stem cell (HSC) compartment, including loss of functional capacity, altered clonal composition, and changes in lineage contribution. Although accumulation of DNA damage occurs during HSC aging, it is unlikely such consistent aging phenotypes could be solely attributed to changes in DNA integrity. Another mechanism by which heritable traits could contribute to the changes in the functional potential of aged HSCs is through alterations in the epigenetic landscape of adult stem cells. Indeed, recent studies on hematopoietic stem cells have suggested that altered epigenetic profiles are associated with HSC aging and playmore » a key role in modulating the functional potential of HSCs at different stages during ontogeny. Even small changes of the epigenetic landscape can lead to robustly altered expression patterns, either directly by loss of regulatory control or through indirect, additive effects, ultimately leading to transcriptional changes of the stem cells. Potential drivers of such changes in the epigenetic landscape of aged HSCs include proliferative history, DNA damage, and deregulation of key epigenetic enzymes and complexes. This review will focus largely on the two most characterized epigenetic marks – DNA methylation and histone modifications – but will also discuss the potential role of non-coding RNAs in regulating HSC function during aging.« less

Autologous hematopoietic stem cells for refractory Crohn's disease.

PubMed

DiNicola, C A; Zand, A; Hommes, D W

2017-05-01

Autologous hematopoietic stem cells are gaining ground as an effective and safe treatment for treating severe refractory Crohn's disease (CD). Autologous hematopoietic stem cell therapy (AHSCT) induces resetting of the immune system by de novo regeneration of T-cell repertoire and repopulation of epithelial cells by bone-marrow derived cells to help patients achieve clinical and endoscopic remission. Areas covered: Herein, the authors discuss the use of AHSCT in treating patients with CD. Improvements in disease activity have been seen in patients with severe autoimmune disease and patients with severe CD who underwent AHSCT for a concomitant malignant hematological disease. Clinical and endoscopic remission has been achieved in patients treated with AHSCT for CD. The only randomized trial published to date, the ASTIC Trial, did not support further use of AHSCT to treat CD. Yet, critics of this trial have deemed AHSCT as a promising treatment for severe refractory CD. Expert opinion: Even with the promising evidence presented for HSCT for refractory CD, protocols need to be refined through the collaboration of GI and hemato-oncology professionals. The goal is to incorporate safe AHSCT and restore tolerance by delivering an effective immune 'cease fire' as a treatment option for severe refractory CD.

AML1/ETO accelerates cell migration and impairs cell-to-cell adhesion and homing of hematopoietic stem/progenitor cells

PubMed Central

Saia, Marco; Termanini, Alberto; Rizzi, Nicoletta; Mazza, Massimiliano; Barbieri, Elisa; Valli, Debora; Ciana, Paolo; Gruszka, Alicja M.; Alcalay, Myriam

2016-01-01

The AML1/ETO fusion protein found in acute myeloid leukemias functions as a transcriptional regulator by recruiting co-repressor complexes to its DNA binding site. In order to extend the understanding of its role in preleukemia, we expressed AML1/ETO in a murine immortalized pluripotent hematopoietic stem/progenitor cell line, EML C1, and found that genes involved in functions such as cell-to-cell adhesion and cell motility were among the most significantly regulated as determined by RNA sequencing. In functional assays, AML1/ETO-expressing cells showed a decrease in adhesion to stromal cells, an increase of cell migration rate in vitro, and displayed an impairment in homing and engraftment in vivo upon transplantation into recipient mice. Our results suggest that AML1/ETO expression determines a more mobile and less adherent phenotype in preleukemic cells, therefore altering the interaction with the hematopoietic niche, potentially leading to the migration across the bone marrow barrier and to disease progression. PMID:27713544

The actin polymerization regulator WAVE2 is required for early bone marrow repopulation by hematopoietic stem cells.

PubMed

Ogaeri, Takunori; Eto, Koji; Otsu, Makoto; Ema, Hideo; Nakauchi, Hiromitsu

2009-05-01

The Rho GTPase family members play essential roles in hematopoiesis. Of these, Rac1 is thought to be required for the appropriate spatial localization of hematopoietic stem and/or progenitor cells (HSPCs) within the bone marrow (BM), whereas Rac2 likely plays a role in BM retention of HSPCs. To elucidate the molecular mechanisms underlying Rac-mediated functions in hematopoietic stem cells (HSCs), we studied Wiskott-Aldrich syndrome protein family verprolin-homologous proteins (WAVEs), the specific effectors downstream of the Rac GTPases in actin polymerization. We here showed that CD34(-/low)c-Kit(+)Sca-1(+)lineage(-) HSCs (CD34(-)KSL HSCs) express WAVE2 but neither WAVE1 nor WAVE3. Because WAVE2 knockout mice are embryonic-lethal, we utilized HSCs in which the expression of WAVE2 was reduced by small interfering RNA. We found that knockdown (KD) of WAVE2 in HSCs affected neither in vitro colony formation nor cell proliferation but did impair in vivo long-term reconstitution. Interestingly, WAVE2 KD HSCs exhibited unaltered homing but showed poor BM repopulation detected as early as day 5 after transplantation. The mechanistic studies on WAVE2 KD HSCs revealed modest but significant impairment in both cobblestone-like area-forming on stromal layers and actin polymerization upon integrin ligation by fibronectin. These results suggested that WAVE2-mediated actin polymerization, potentially downstream of Rac1, plays an important role in intramarrow mobilization and proliferation of HSCs, which are believed to be crucial steps for long-term marrow reconstitution after transplantation.

Vascular niche promotes hematopoietic multipotent progenitor formation from pluripotent stem cells

PubMed Central

Gori, Jennifer L.; Butler, Jason M.; Chan, Yan-Yi; Chandrasekaran, Devikha; Poulos, Michael G.; Ginsberg, Michael; Nolan, Daniel J.; Elemento, Olivier; Wood, Brent L.; Adair, Jennifer E.; Rafii, Shahin; Kiem, Hans-Peter

2015-01-01

Pluripotent stem cells (PSCs) represent an alternative hematopoietic stem cell (HSC) source for treating hematopoietic disease. The limited engraftment of human PSC–derived (hPSC-derived) multipotent progenitor cells (MPP) has hampered the clinical application of these cells and suggests that MPP require additional cues for definitive hematopoiesis. We hypothesized that the presence of a vascular niche that produces Notch ligands jagged-1 (JAG1) and delta-like ligand-4 (DLL4) drives definitive hematopoiesis. We differentiated hes2 human embryonic stem cells (hESC) and Macaca nemestrina–induced PSC (iPSC) line-7 with cytokines in the presence or absence of endothelial cells (ECs) that express JAG1 and DLL4. Cells cocultured with ECs generated substantially more CD34+CD45+ hematopoietic progenitors compared with cells cocultured without ECs or with ECs lacking JAG1 or DLL4. EC-induced cells exhibited Notch activation and expressed HSC-specific Notch targets RUNX1 and GATA2. EC-induced PSC-MPP engrafted at a markedly higher level in NOD/SCID/IL-2 receptor γ chain–null (NSG) mice compared with cytokine-induced cells, and low-dose chemotherapy-based selection further increased engraftment. Long-term engraftment and the myeloid-to-lymphoid ratio achieved with vascular niche induction were similar to levels achieved for cord blood–derived MPP and up to 20-fold higher than those achieved with hPSC-derived MPP engraftment. Our findings indicate that endothelial Notch ligands promote PSC-definitive hematopoiesis and production of long-term engrafting CD34+ cells, suggesting these ligands are critical for HSC emergence. PMID:25664855

The Hematopoietic Stem Cell Therapy for Exploration of Space

NASA Technical Reports Server (NTRS)

Roach, Allana Nicole; Brezo, Jelena

2002-01-01

Astronauts experience severe/invasive disorders caused by space environments. These include hematological/cardiac abnormalities, bone and muscle losses, immunodeficiency, neurological disorders and cancer. While the cause of these symptoms are not yet fully delineated, one possible explanation could be the inhibition of hematopoietic stem cell (HSC) growth and hematopoiesis in space. HSCs differentiate into all types of blood cells, and growing evidence indicates that the HSCs also have the ability to transdifferentiate to various tissues, including muscle, skin, liver, neuronal cells and possibly bone. Therefore, a hypothesis was advanced in this laboratory that the hematopoietic stem cell-based therapy, herein called the hematopoietic stem cell therapy (HSCT), could mitigate some of the disorders described above. Due to the magnitude of this project our laboratory has subdivided it into 3 sections: a) HSCT for space anemia; b) HSCT for muscle and bone losses; and c) HSCT for immunodeficiency. Toward developing the HSCT protocol for space anemia, the HSC transplantation procedure was established using a mouse model of beta thalassemia. In addition, the NASA Rotating Wall Vessel (RWV) culture system was used to grow HSCs in space condition. To investigate the HSCT for muscle loss and bone loss, donor HSCs were genetically marked either by transfecting the beta-galactosidase-containing plasmid, pCMV.SPORT-beta-gal or by preparing from b-galactosidase transgenic mice. The transdifferentiation of HSCs to muscle is traced by the reporter gene expression in the hindlimb suspended mice with some positive outcome, as studied by the X-gal staining procedure. The possible structural contribution of HSCs against muscle loss is being investigated histochemically.

Intravenous apoptotic cell infusion as a cell-based therapy toward improving hematopoietic cell transplantation outcome.

PubMed

Saas, Philippe; Gaugler, Béatrice; Perruche, Sylvain

2010-10-01

Allogeneic hematopoietic cell transplantation (AHCT) is an efficient therapy for different malignant and nonmalignant hematological diseases. However, the use of this therapeutic approach is still limited by some severe toxic side effects, mainly graft-versus-host disease (GvHD). Today, the risk of fatal GvHD restrains the wider application of AHCT to many patients in need of an effective therapy for their high-risk hematologic malignancies. Thus, new strategies, including cell-based therapy approaches, are required. We propose to use intravenous donor apoptotic leukocyte infusion to improve AHCT outcome. In experimental AHCT models, we demonstrated that intravenous apoptotic leukocyte infusion, simultaneously with allogeneic bone marrow grafts, favors hematopoietic engraftment, prevents allo-immunization, and delays acute GvHD onset. Here, we review the different mechanisms and the potential beneficial effects associated with the immunomodulatory properties of apoptotic cells in the AHCT setting. © 2010 New York Academy of Sciences.

Thrombopoietin contributes to the formation and the maintenance of hematopoietic progenitor-containing cell clusters in the aorta-gonad-mesonephros region.

PubMed

Harada, Kaho; Nobuhisa, Ikuo; Anani, Maha; Saito, Kiyoka; Taga, Tetsuya

2017-07-01

In the midgestation mouse embryo, hematopoietic cell clusters containing hematopoietic stem/progenitor cells arise in the aorta-gonad-mesonephros (AGM) region. We have previously reported that forced expression of the Sox17 transcription factor in CD45 low c-Kit high AGM cells, which are the hematopoietic cellular component of the cell clusters, and subsequent coculture with OP9 stromal cells in the presence of three cytokines, stem cell factor (SCF), interleukin-3 (IL-3), and thrombopoietin (TPO), led to the formation and the maintenance of cell clusters with cells at an undifferentiated state in vitro. In this study, we investigated the role of each cytokine in the formation of hematopoietic cell clusters. We cultured Sox17-transduced AGM cells with each of the 7 possible combinations of the three cytokines. The size and the number of Sox17-transduced cell clusters in the presence of TPO, either alone or in combination, were comparable to that observed with the complete set of the three cytokines. Expression of TPO receptor, c-Mpl was almost ubiquitously expressed and maintained in Sox17-transduced hematopoietic cell clusters. In addition, the expression level of c-Mpl was highest in the CD45 low c-Kit high cells among the Sox17-transduced cell clusters. Moreover, c-Mpl protein was highly expressed in the intra-aortic hematopoietic cell clusters in comparison with endothelial cells of dorsal aorta. Finally, stimulation of the endothelial cells prepared from the AGM region by TPO induced the production of hematopoietic cells. These results suggest that TPO contributes to the formation and the maintenance of hematopoietic cell clusters in the AGM region. Copyright © 2017 Elsevier Ltd. All rights reserved.