Defective endochondral ossification-derived matrix and bone cells alter the lymphopoietic niche in collagen X mouse models.

PubMed

Sweeney, Elizabeth; Roberts, Douglas; Lin, Angela; Guldberg, Robert; Jacenko, Olena

2013-10-01

Despite the appreciated interdependence of skeletal and hematopoietic development, the cell and matrix components of the hematopoietic niche remain to be fully defined. Utilizing mice with disrupted function of collagen X (ColX), a major hypertrophic cartilage matrix protein associated with endochondral ossification, our data identified a cytokine defect in trabecular bone cells at the chondro-osseous hematopoietic niche as a cause for aberrant B lymphopoiesis in these mice. Specifically, analysis of ColX transgenic and null mouse chondro-osseous regions via micro-computed tomography revealed an altered trabecular bone environment. Additionally, cocultures with hematopoietic and chondro-osseous cell types highlighted impaired hematopoietic support by ColX transgenic and null mouse derived trabecular bone cells. Further, cytokine arrays with conditioned media from the trabecular osteoblast cocultures suggested an aberrant hematopoietic cytokine milieu within the chondro-osseous niche of the ColX deficient mice. Accordingly, B lymphopoiesis was rescued in the ColX mouse derived trabecular osteoblast cocultures with interlukin-7, stem cell factor, and stromal derived factor-1 supplementation. Moreover, B cell development was restored in vivo after injections of interlukin-7. These data support our hypothesis that endrochondrally-derived trabecular bone cells and matrix constituents provide cytokine-rich niches for hematopoiesis. Furthermore, this study contributes to the emerging concept that niche defects may underlie certain immuno-osseous and hematopoietic disorders.

Regulatory coding of lymphoid lineage choice by hematopoietic transcription factors

NASA Technical Reports Server (NTRS)

Warren, Luigi A.; Rothenberg, Ellen V.

2003-01-01

During lymphopoiesis, precursor cells negotiate a complex regulatory space, defined by the levels of several competing and cross-regulating transcription factors, before arriving at stable states of commitment to the B-, T- and NK-specific developmental programs. Recent perturbation experiments provide evidence that this space has three major axes, corresponding to the PU.1 versus GATA-1 balance, the intensity of Notch signaling through the CSL pathway, and the ratio of E-box transcription factors to their Id protein antagonists.

Impact of immune-metabolic interactions on age-related thymic demise and T cell senescence.

PubMed

Dixit, Vishwa Deep

2012-10-01

Emerging evidence indicates that the immune and metabolic interactions control several aspects of the aging process and associated chronic diseases. Among several sites of immune-metabolic interactions, thymic demise represents a particularly puzzling phenomenon because even in metabolically healthy middle-aged individuals the majority of thymic space is replaced with ectopic lipids. The new T cell specificities can only be generated in a functional thymus and, peripheral proliferation of pre-existing T cell clones provides limited immune-vigilance in the elderly. Therefore, it is hypothesized that the strategies that enhance thymic-lymphopoiesis may extend healthspan. Recent data suggest that byproducts of thymic fatty acids and lipids result in accumulation of 'lipotoxic DAMPs' (damage associated molecular patterns), which triggers the innate immune-sensing mechanism like inflammasome activation which links aging to thymic demise. The immune-metabolic interaction within the aging thymus produces a local pro-inflammatory state that directly compromises the thymic stromal microenvironment, thymic-lymphopoiesis and serves a precursor of systemic immune-dysregulation in the elderly. New evidence also suggests that ectopic thymic adipocytes may develop from specific intrathymic stromal cell precursors instead of a passive process that is simply a consequence of thymic lymphopenia. Thus the complex bidirectional interactions between metabolic and immune systems may link aging to health, T cell senescence, and associated diseases. This review discusses the immune-metabolic mechanisms during aging - with implications for developing future therapeutic strategies for living well beyond the expected. Copyright © 2012 Elsevier Ltd. All rights reserved.

B lymphocyte selection and age-related changes in VH gene usage in mutant Alicia rabbits.

PubMed

Zhu, X; Boonthum, A; Zhai, S K; Knight, K L

1999-09-15

Young Alicia rabbits use VHa-negative genes, VHx and VHy, in most VDJ genes, and their serum Ig is VHa negative. However, as Alicia rabbits age, VHa2 allotype Ig is produced at high levels. We investigated which VH gene segments are used in the VDJ genes of a2 Ig-secreting hybridomas and of a2 Ig+ B cells from adult Alicia rabbits. We found that 21 of the 25 VDJ genes used the a2-encoding genes, VH4 or VH7; the other four VDJ genes used four unknown VH gene segments. Because VH4 and VH7 are rarely found in VDJ genes of normal or young Alicia rabbits, we investigated the timing of rearrangement of these genes in Alicia rabbits. During fetal development, VH4 was used in 60-80% of nonproductively rearranged VDJ genes, and VHx and VHy together were used in 10-26%. These data indicate that during B lymphopoiesis VH4 is preferentially rearranged. However, the percentage of productive VHx- and VHy-utilizing VDJ genes increased from 38% at day 21 of gestation to 89% at birth (gestation day 31), whereas the percentage of VH4-utilizing VDJ genes remained at 15%. These data suggest that during fetal development, either VH4-utilizing B-lineage cells are selectively eliminated, or B cells with VHx- and VHy-utilizing VDJ genes are selectively expanded, or both. The accumulation of peripheral VH4-utilizing a2 B cells with age indicates that these B cells might be selectively expanded in the periphery. We discuss the possible selection mechanisms that regulate VH gene segment usage in rabbit B cells during lymphopoiesis and in the periphery.

Labeling single cell for in-vivo study of cell fate mapping and lineage tracing

NASA Astrophysics Data System (ADS)

He, Sicong; Xu, Jin; Wu, Yi; Tian, Ye; Sun, Qiqi; Wen, Zilong; Qu, Jianan Y.

2018-02-01

Cell fate mapping and lineage tracing are significant ways to understanding the developmental origins of biological tissues. It requires labeling individual cells and tracing the development of their progeny. We develop an infrared laser-evoked gene operator heat-shock microscope system to achieve single-cell labeling in zebrafish. With a fluorescent thermometry technique, we measure the temperature increase in zebrafish tissues induced by infrared laser and identify the optimal heat shock conditions for single-cell gene induction in different types of zebrafish cells. We use this technique to study the fate mapping of T lymphocytes and discover the distinct waves of lymphopoiesis during the zebrafish development.

Neuro-immune modulation of the thymus microenvironment (review).

PubMed

Mignini, Fiorenzo; Sabbatini, Maurizio; Mattioli, Laura; Cosenza, Monica; Artico, Marco; Cavallotti, Carlo

2014-06-01

The thymus is the primary site for T-cell lympho-poiesis. Its function includes the maturation and selection of antigen specific T cells and selective release of these cells to the periphery. These highly complex processes require precise parenchymal organization and compartmentation where a plethora of signalling pathways occur, performing strict control on the maturation and selection processes of T lymphocytes. In this review, the main morphological characteristics of the thymus microenvironment, with particular emphasis on nerve fibers and neuropeptides were assessed, as both are responsible for neuro-immune‑modulation functions. Among several neurotransmitters that affect thymus function, we highlight the dopaminergic system as only recently has its importance on thymus function and lymphocyte physiology come to light.

The effect of continuous low dose-rate gamma irradiation on cell population kinetics of lymphoid tissue

NASA Technical Reports Server (NTRS)

Foster, B. R.

1974-01-01

Cellular response and cell population kinetics were studied during lymphopoiesis in the thymus of the mouse under continuous gamma irradiation using autoradiographic techniques and specific labeling with tritiated thymidine. On the basis of tissue weights, it is concluded that the response of both the thymus and spleen to continuous low dose-rate irradiation is multiphasic. That is, alternating periods of steady state growth, followed by collapse, which in turn is followed by another period of homeostasis. Since there are two populations of lymphocytes - short lived and long-lived, it may be that different phases of steady state growth are mediated by different lymphocytes. The spleen is affected to a greater extent with shorter periods of steady-state growth than exhibited by the thymus.

Molecular and Functional Characterization of Lymphoid Progenitor Subsets Reveals a Bipartite Architecture of Human Lymphopoiesis.

PubMed

Alhaj Hussen, Kutaiba; Vu Manh, Thien-Phong; Guimiot, Fabien; Nelson, Elisabeth; Chabaane, Emna; Delord, Marc; Barbier, Maxime; Berthault, Claire; Dulphy, Nicolas; Alberdi, Antonio José; Burlen-Defranoux, Odile; Socié, Gerard; Bories, Jean Christophe; Larghero, Jerôme; Vanneaux, Valérie; Verhoeyen, Els; Wirth, Thierry; Dalod, Marc; Gluckman, Jean Claude; Cumano, Ana; Canque, Bruno

2017-10-17

The classical model of hematopoiesis established in the mouse postulates that lymphoid cells originate from a founder population of common lymphoid progenitors. Here, using a modeling approach in humanized mice, we showed that human lymphoid development stemmed from distinct populations of CD127 - and CD127 + early lymphoid progenitors (ELPs). Combining molecular analyses with in vitro and in vivo functional assays, we demonstrated that CD127 - and CD127 + ELPs emerged independently from lympho-mono-dendritic progenitors, responded differently to Notch1 signals, underwent divergent modes of lineage restriction, and displayed both common and specific differentiation potentials. Whereas CD127 - ELPs comprised precursors of T cells, marginal zone B cells, and natural killer (NK) and innate lymphoid cells (ILCs), CD127 + ELPs supported production of all NK cell, ILC, and B cell populations but lacked T potential. On the basis of these results, we propose a "two-family" model of human lymphoid development that differs from the prevailing model of hematopoiesis. Copyright © 2017 Elsevier Inc. All rights reserved.

Dataset on granulopoiesis- and lymphopoiesis-stimulating cytokine levels in insulin secretagogue users with incident breast cancer.

PubMed

Wintrob, Zachary A P; Hammel, Jeffrey P; Nimako, George K; Gaile, Dan P; Forrest, Alan; Ceacareanu, Alice C

2017-04-01

GM-CSF and G-CSF are widely used for their benefit in reducing chemotherapy-associated neutropenia. However, whether GM- or G-CSF administration could have tumorigenic or pro-metastatic effects or whether insulin resistance could negatively impact such effects is not known. Their ability to stimulate monocyte production at the same time with the highly sought after neutrophils' production, enables an enhanced potential for activation of tumor-associated macrophages. At the same time, IL-7 remains the main driver of B and T cell differentiation and maturation, a process linked to the development of insulin resistance and response to diabetes pharmacotherapy. Insulin secretagogues have the potential to interfere with the hematopoiesis process, respectively with the formation of lineages that may lead to a tumorigenic or pro-metastatic phenotype, but this relationship has not been yet investigated. The data presented here shows the relationship between pre-existing use of insulin secretagogues in women diagnosed with breast cancer and type 2 diabetes mellitus, the GM-CSF, G-CSF and IL-7 cytokine profiles at the time of breast cancer diagnosis, and subsequent cancer outcomes. A Pearson correlation analysis evaluating the relationship between investigated cytokines stratified by secretagogue use and controls, and interferon is also provided.

Isolation and characterization of lymphocyte-like cells from a lamprey

PubMed Central

Mayer, Werner E.; Uinuk-ool, Tatiana; Tichy, Herbert; Gartland, Lanier A.; Klein, Jan; Cooper, Max D.

2002-01-01

Lymphocyte-like cells in the intestine of the sea lamprey, Petromyzon marinus, were isolated by flow cytometry under light-scatter conditions used for the purification of mouse intestinal lymphocytes. The purified lamprey cells were morphologically indistinguishable from mammalian lymphocytes. A cDNA library was prepared from the lamprey lymphocyte-like cells, and more than 8,000 randomly selected clones were sequenced. Homology searches comparing these ESTs with sequences deposited in the databases led to the identification of numerous genes homologous to those predominantly or characteristically expressed in mammalian lymphocytes, which included genes controlling lymphopoiesis, intracellular signaling, proliferation, migration, and involvement of lymphocytes in innate immune responses. Genes closely related to those that in gnathostomes control antigen processing and transport of antigenic peptides could be ascertained, although no sequences with significant similarity to MHC, T cell receptor, or Ig genes were found. The data suggest that the evolution of lymphocytes in the lamprey has reached a stage poised for the emergence of adaptive immunity. PMID:12388781

Inherited variation in immune response genes in follicular lymphoma and diffuse large B-cell lymphoma.

PubMed

Nielsen, Kaspar Rene; Steffensen, Rudi; Haunstrup, Thure Mors; Bødker, Julie Støve; Dybkær, Karen; Baech, John; Bøgsted, Martin; Johnsen, Hans Erik

2015-01-01

Diffuse large B-cell lymphoma (DLBCL) and follicular lymphoma (FL) both depend on immune-mediated survival and proliferation signals from the tumor microenvironment. Inherited genetic variation influences this complex interaction. A total of 89 studies investigating immune-response genes in DLBCL and FL were critically reviewed. Relatively consistent association exists for variation in the tumor necrosis factor alpha (TNFA) and interleukin-10 loci and DLBCL risk; for DLBCL outcome association with the TNFA locus exists. Variations at chromosome 6p31-32 were associated with FL risk. Importantly, individual risk alleles have been shown to interact with each other. We suggest that the pathogenetic impact of polymorphic genes should include gene-gene interaction analysis and should be validated in preclinical model systems of normal B lymphopoiesis and B-cell malignancies. In the future, large cohort studies of interactions and genome-wide association studies are needed to extend the present findings and explore new risk alleles to be studied in preclinical models.

Low doses of imatinib induce myelopoiesis and enhance host anti-microbial immunity.

PubMed

Napier, Ruth J; Norris, Brian A; Swimm, Alyson; Giver, Cynthia R; Harris, Wayne A C; Laval, Julie; Napier, Brooke A; Patel, Gopi; Crump, Ryan; Peng, Zhenghong; Bornmann, William; Pulendran, Bali; Buller, R Mark; Weiss, David S; Tirouvanziam, Rabindra; Waller, Edmund K; Kalman, Daniel

2015-03-01

Imatinib mesylate (Gleevec) inhibits Abl1, c-Kit, and related protein tyrosine kinases (PTKs) and serves as a therapeutic for chronic myelogenous leukemia and gastrointestinal stromal tumors. Imatinib also has efficacy against various pathogens, including pathogenic mycobacteria, where it decreases bacterial load in mice, albeit at doses below those used for treating cancer. We report that imatinib at such low doses unexpectedly induces differentiation of hematopoietic stem cells and progenitors in the bone marrow, augments myelopoiesis but not lymphopoiesis, and increases numbers of myeloid cells in blood and spleen. Whereas progenitor differentiation relies on partial inhibition of c-Kit by imatinib, lineage commitment depends upon inhibition of other PTKs. Thus, imatinib mimics "emergency hematopoiesis," a physiological innate immune response to infection. Increasing neutrophil numbers by adoptive transfer sufficed to reduce mycobacterial load, and imatinib reduced bacterial load of Franciscella spp., which do not utilize imatinib-sensitive PTKs for pathogenesis. Thus, potentiation of the immune response by imatinib at low doses may facilitate clearance of diverse microbial pathogens.

Early loss of Crebbp confers malignant stem cell properties on lymphoid progenitors.

PubMed

Horton, Sarah J; Giotopoulos, George; Yun, Haiyang; Vohra, Shabana; Sheppard, Olivia; Bashford-Rogers, Rachael; Rashid, Mamunur; Clipson, Alexandra; Chan, Wai-In; Sasca, Daniel; Yiangou, Loukia; Osaki, Hikari; Basheer, Faisal; Gallipoli, Paolo; Burrows, Natalie; Erdem, Ayşegül; Sybirna, Anastasiya; Foerster, Sarah; Zhao, Wanfeng; Sustic, Tonci; Petrunkina Harrison, Anna; Laurenti, Elisa; Okosun, Jessica; Hodson, Daniel; Wright, Penny; Smith, Ken G; Maxwell, Patrick; Fitzgibbon, Jude; Du, Ming Q; Adams, David J; Huntly, Brian J P

2017-09-01

Loss-of-function mutations of cyclic-AMP response element binding protein, binding protein (CREBBP) are prevalent in lymphoid malignancies. However, the tumour suppressor functions of CREBBP remain unclear. We demonstrate that loss of Crebbp in murine haematopoietic stem and progenitor cells (HSPCs) leads to increased development of B-cell lymphomas. This is preceded by accumulation of hyperproliferative lymphoid progenitors with a defective DNA damage response (DDR) due to a failure to acetylate p53. We identify a premalignant lymphoma stem cell population with decreased H3K27ac, which undergoes transcriptional and genetic evolution due to the altered DDR, resulting in lymphomagenesis. Importantly, when Crebbp is lost later in lymphopoiesis, cellular abnormalities are lost and tumour generation is attenuated. We also document that CREBBP mutations may occur in HSPCs from patients with CREBBP-mutated lymphoma. These data suggest that earlier loss of Crebbp is advantageous for lymphoid transformation and inform the cellular origins and subsequent evolution of lymphoid malignancies.

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

Differential effects of Notch ligands Delta-1 and Jagged-1 in human lymphoid differentiation.

PubMed

Jaleco, A C; Neves, H; Hooijberg, E; Gameiro, P; Clode, N; Haury, M; Henrique, D; Parreira, L

2001-10-01

Notch signaling is known to differentially affect the development of lymphoid B and T cell lineages, but it remains unclear whether such effects are specifically dependent on distinct Notch ligands. Using a cell coculture assay we observed that the Notch ligand Delta-1 completely inhibits the differentiation of human hematopoietic progenitors into the B cell lineage while promoting the emergence of cells with a phenotype of T cell/natural killer (NK) precursors. In contrast, Jagged-1 did not disturb either B or T cell/NK development. Furthermore, cells cultured in the presence of either Delta-1 or Jagged-1 can acquire a phenotype of NK cells, and Delta-1, but not Jagged-1, permits the emergence of a de novo cell population coexpressing CD4 and CD8. Our results thus indicate that distinct Notch ligands can mediate differential effects of Notch signaling and provide a useful system to further address cell-fate decision processes in lymphopoiesis.

Differential Effects of Notch Ligands Delta-1 and Jagged-1 in Human Lymphoid Differentiation

PubMed Central

Jaleco, Ana C.; Neves, Hélia; Hooijberg, Erik; Gameiro, Paula; Clode, Nuno; Haury, Matthias; Henrique, Domingos; Parreira, Leonor

2001-01-01

Notch signaling is known to differentially affect the development of lymphoid B and T cell lineages, but it remains unclear whether such effects are specifically dependent on distinct Notch ligands. Using a cell coculture assay we observed that the Notch ligand Delta-1 completely inhibits the differentiation of human hematopoietic progenitors into the B cell lineage while promoting the emergence of cells with a phenotype of T cell/natural killer (NK) precursors. In contrast, Jagged-1 did not disturb either B or T cell/NK development. Furthermore, cells cultured in the presence of either Delta-1 or Jagged-1 can acquire a phenotype of NK cells, and Delta-1, but not Jagged-1, permits the emergence of a de novo cell population coexpressing CD4 and CD8. Our results thus indicate that distinct Notch ligands can mediate differential effects of Notch signaling and provide a useful system to further address cell-fate decision processes in lymphopoiesis. PMID:11581320

Thymocyte Maturation Is Regulated by the Activity of the Helix-Loop-Helix Protein, E47

PubMed Central

Bain, Gretchen; Quong, Melanie W.; Soloff, Rachel S.; Hedrick, Stephen M.; Murre, Cornelis

1999-01-01

The E2A proteins, E12 and E47, are required for progression through multiple developmental pathways, including early B and T lymphopoiesis. Here, we provide in vitro and in vivo evidence demonstrating that E47 activity regulates double-positive thymocyte maturation. In the absence of E47 activity, positive selection of both major histocompatibility complex (MHC) class I– and class II–restricted T cell receptors (TCRs) is perturbed. Additionally, development of CD8 lineage T cells in an MHC class I–restricted TCR transgenic background is sensitive to the dosage of E47. Mice deficient for E47 display an increase in production of mature CD4 and CD8 lineage T cells. Furthermore, ectopic expression of an E2A inhibitor helix-loop-helix protein, Id3, promotes the in vitro differentiation of an immature T cell line. These results demonstrate that E2A functions as a regulator of thymocyte positive selection. PMID:10587351

Graft-versus-host disease causes failure of donor hematopoiesis and lymphopoiesis in interferon-gamma receptor-deficient hosts.

PubMed

Delisle, Jean-Sébastien; Gaboury, Louis; Bélanger, Marie-Pier; Tassé, Eliane; Yagita, Hideo; Perreault, Claude

2008-09-01

The immunopathologic condition known as graft-versus-host disease (GVHD) results from a type I T-cell process. However, a prototypical type I cytokine, interferon-gamma (IFN-gamma), can protect against several manifestations of GVHD in recipients of major histocompatibility complex (MHC)-mismatched hematopoietic cells. We transplanted hematopoietic cells from C3H.SW donors in wild-type (wt) and IFN-gamma-receptor-deficient (IFN-gammaRKO) MHC-matched C57BL/6 recipients. In IFN-gammaRKO recipients, host cells were unresponsive to IFN-gamma, whereas wt donor cells were exposed to exceptionally high levels of IFN-gamma. From an IFN-gamma perspective, we could therefore evaluate the impact of a loss-of-function on host cells and gain-of-function on donor cells. We found that lack of IFN-gammaR prevented up-regulation of MHC proteins on host cells but did not mitigate damage to most target organs. Two salient phenotypes in IFN-gammaRKO recipients involved donor cells: lymphoid hypoplasia and hematopoietic failure. Lymphopenia was due to FasL-induced apoptosis and decreased cell proliferation. Bone marrow aplasia resulted from a decreased proliferation of hematopoietic stem/progenitor cells that was associated with down-regulation of 2 genes negatively regulated by IFN-gamma: Ccnd1 and Myc. We conclude that IFN-gamma produced by alloreactive T cells may entail a severe graft-versus-graft reaction and could be responsible for cytopenias that are frequently observed in subjects with GVHD.

Combined effects of interleukin-7 and stem cell factor administration on lymphopoiesis after murine bone marrow transplantation.

PubMed

Chung, Brile; Min, Dullei; Joo, Lukas W; Krampf, Mark R; Huang, Jing; Yang, Yujun; Shashidhar, Sumana; Brown, Janice; Dudl, Eric P; Weinberg, Kenneth I

2011-01-01

The decreased ability of the thymus to generate T cells after bone marrow transplantation (BMT) is a clinically significant problem. Interleukin (IL)-7 and stem cell factor (SCF) induce proliferation, differentiation, and survival of thymocytes. Although previous studies have shown that administration of recombinant human IL-7 (rhIL-7) after murine and human BMT improves thymopoiesis and immune function, whether administration of SCF exerts similar effects is unclear. To evaluate independent or combinatorial effects of IL-7 and SCF in post-BMT thymopoiesis, bone marrow (BM)-derived mesenchymal stem cells transduced ex vivo with the rhIL-7 or murine SCF (mSCF) genes were cotransplanted with T cell-depleted BM cells into lethally irradiated mice. Although rhIL-7 and mSCF each improved immune reconstitution, the combination treatment had a significantly greater effect than either cytokine alone. Moreover, the combination treatment significantly increased donor-derived common lymphoid progenitors (CLPs) in BM, suggesting that transplanted CLPs expand more rapidly in response to IL-7 and SCF and may promote immune reconstitution. Our findings demonstrate that IL-7 and SCF might be therapeutically useful for enhancing de novo T cell development. Furthermore, combination therapy may allow the administration of lower doses of IL-7, thereby decreasing the likelihood of IL-7-mediated expansion of mature T cells. 2011. Published by Elsevier Inc.

African Trypanosomes Undermine Humoral Responses and Vaccine Development: Link with Inflammatory Responses?

PubMed Central

Stijlemans, Benoit; Radwanska, Magdalena; De Trez, Carl; Magez, Stefan

2017-01-01

African trypanosomosis is a debilitating disease of great medical and socioeconomical importance. It is caused by strictly extracellular protozoan parasites capable of infecting all vertebrate classes including human, livestock, and game animals. To survive within their mammalian host, trypanosomes have evolved efficient immune escape mechanisms and manipulate the entire host immune response, including the humoral response. This report provides an overview of how trypanosomes initially trigger and subsequently undermine the development of an effective host antibody response. Indeed, results available to date obtained in both natural and experimental infection models show that trypanosomes impair homeostatic B-cell lymphopoiesis, B-cell maturation and survival and B-cell memory development. Data on B-cell dysfunctioning in correlation with parasite virulence and trypanosome-mediated inflammation will be discussed, as well as the impact of trypanosomosis on heterologous vaccine efficacy and diagnosis. Therefore, new strategies aiming at enhancing vaccination efficacy could benefit from a combination of (i) early parasite diagnosis, (ii) anti-trypanosome (drugs) treatment, and (iii) anti-inflammatory treatment that collectively might allow B-cell recovery and improve vaccination. PMID:28596768

ZNF423 and ZNF521: EBF1 Antagonists of Potential Relevance in B-Lymphoid Malignancies

PubMed Central

Mesuraca, Maria; Chiarella, Emanuela; Scicchitano, Stefania; Codispoti, Bruna; Giordano, Marco; Nappo, Giovanna; Bond, Heather M.; Morrone, Giovanni

2015-01-01

The development of the B-lymphoid cell lineage is tightly controlled by the concerted action of a network of transcriptional and epigenetic regulators. EBF1, a central component of this network, is essential for B-lymphoid specification and commitment as well as for the maintenance of the B-cell identity. Genetic alterations causing loss of function of these B-lymphopoiesis regulators have been implicated in the pathogenesis of B-lymphoid malignancies, with particular regard to B-cell acute lymphoblastic leukaemias (B-ALLs), where their presence is frequently detected. The activity of the B-cell regulatory network may also be disrupted by the aberrant expression of inhibitory molecules. In particular, two multi-zinc finger transcription cofactors named ZNF423 and ZNF521 have been characterised as potent inhibitors of EBF1 and are emerging as potentially relevant contributors to the development of B-cell leukaemias. Here we will briefly review the current knowledge of these factors and discuss the importance of their functional cross talk with EBF1 in the development of B-cell malignancies. PMID:26788497

Molecular role of the PAX5-ETV6 oncoprotein in promoting B-cell acute lymphoblastic leukemia.

PubMed

Smeenk, Leonie; Fischer, Maria; Jurado, Sabine; Jaritz, Markus; Azaryan, Anna; Werner, Barbara; Roth, Mareike; Zuber, Johannes; Stanulla, Martin; den Boer, Monique L; Mullighan, Charles G; Strehl, Sabine; Busslinger, Meinrad

2017-03-15

PAX5 is a tumor suppressor in B-ALL, while the role of PAX5 fusion proteins in B-ALL development is largely unknown. Here, we studied the function of PAX5-ETV6 and PAX5-FOXP1 in mice expressing these proteins from the Pax5 locus. Both proteins arrested B-lymphopoiesis at the pro-B to pre-B-cell transition and, contrary to their proposed dominant-negative role, did not interfere with the expression of most regulated Pax5 target genes. Pax5-Etv6, but not Pax5-Foxp1, cooperated with loss of the Cdkna2a/b tumor suppressors in promoting B-ALL development. Regulated Pax5-Etv6 target genes identified in these B-ALLs encode proteins implicated in pre-B-cell receptor (BCR) signaling and migration/adhesion, which could contribute to the proliferation, survival, and tissue infiltration of leukemic B cells. Together with similar observations made in human PAX5-ETV6 + B-ALLs, these data identified PAX5-ETV6 as a potent oncoprotein that drives B-cell leukemia development. © 2017 The Authors.

Epigenetic repression of the Igk locus by STAT5-mediated Ezh2 recruitment

PubMed Central

Mandal, Malay; Powers, Sarah E.; Maienschein-Cline, Mark; Bartom, Elizabeth T.; Hamel, Keith M.; Kee, Barbara L.; Dinner, Aaron R.; Clark, Marcus R.

2011-01-01

During B lymphopoiesis, Igk recombination requires pre-B cell receptor (pre-BCR) expression and escape from interleukin 7 receptor (IL-7R) signaling. By activating the transcription factor STAT5, IL-7R signaling maintains proliferation and represses Igk germline transcription by unknown mechanisms. We demonstrate that STAT5 tetramer bound the Igk intronic enhancer (Eκi), leading to recruitment of the histone methyltransferase Ezh2. Ezh2 marked H3K27me3 throughout Jκ to Cκ. In the absence of Ezh2, IL-7 failed to repress Igk germline transcription. H3K27me3 modifications were lost after termination of IL-7R–STAT5 signaling and E2A bound Eκi, resulting in acquisition of H3K4me1 and H4Ac. Genome-wide analyses revealed a STAT5 tetrameric binding motif associated with transcriptional repression. These data demonstrate how IL-7R signaling represses Igk germline transcription and provide a general model for STAT5-mediated epigenetic transcriptional repression. PMID:22037603

The NLRP3 Inflammasome Promotes Age-related Thymic Demise and Immunosenescence

PubMed Central

Youm, Yun-Hee; Kanneganti, Thirumala-Devi; Vandanmagsar, Bolormaa; Zhu, Xuewei; Ravussin, Anthony; Adijiang, Ayinuer; Owen, John S.; Thomas, Michael J.; Francis, Joseph; Parks, John S.; Dixit, Vishwa Deep

2013-01-01

The collapse of thymic stromal cell microenvironment with age and resultant inability of the thymus to produce naïve T cells contributes to lower immune-surveillance in the elderly. Here we show that age-related increase in ‘lipotoxic danger signals’ such as free cholesterol (FC) and ceramides, leads to thymic caspase-1 activation via the Nlrp3 inflammasome. Elimination of Nlrp3 and Asc, a critical adaptor required for inflammasome assembly, reduces age-related thymic atrophy and results in an increase in cortical thymic epithelial cells, T cell progenitors and maintenance of T cell repertoire diversity. Using a mouse model of irradiation and hematopoietic stem cell transplantation (HSCT), we show that deletion of the Nlrp3 inflammasome accelerates T cell reconstitution and immune recovery in middle-aged animals. Collectively, these data demonstrate that lowering inflammasome-dependent caspase-1 activation increases thymic lymphopoiesis and suggest that Nlrp3 inflammasome inhibitors may aid the reestablishment of a diverse T cell repertoire in middle-aged or elderly patients undergoing HSCT. PMID:22832107

Low Doses of Imatinib Induce Myelopoiesis and Enhance Host Anti-microbial Immunity

PubMed Central

Swimm, Alyson; Giver, Cynthia R.; Harris, Wayne A. C.; Laval, Julie; Napier, Brooke A.; Patel, Gopi; Crump, Ryan; Peng, Zhenghong; Bornmann, William; Pulendran, Bali; Buller, R. Mark; Weiss, David S.; Tirouvanziam, Rabindra; Waller, Edmund K.; Kalman, Daniel

2015-01-01

Imatinib mesylate (Gleevec) inhibits Abl1, c-Kit, and related protein tyrosine kinases (PTKs) and serves as a therapeutic for chronic myelogenous leukemia and gastrointestinal stromal tumors. Imatinib also has efficacy against various pathogens, including pathogenic mycobacteria, where it decreases bacterial load in mice, albeit at doses below those used for treating cancer. We report that imatinib at such low doses unexpectedly induces differentiation of hematopoietic stem cells and progenitors in the bone marrow, augments myelopoiesis but not lymphopoiesis, and increases numbers of myeloid cells in blood and spleen. Whereas progenitor differentiation relies on partial inhibition of c-Kit by imatinib, lineage commitment depends upon inhibition of other PTKs. Thus, imatinib mimics “emergency hematopoiesis,” a physiological innate immune response to infection. Increasing neutrophil numbers by adoptive transfer sufficed to reduce mycobacterial load, and imatinib reduced bacterial load of Franciscella spp., which do not utilize imatinib-sensitive PTKs for pathogenesis. Thus, potentiation of the immune response by imatinib at low doses may facilitate clearance of diverse microbial pathogens. PMID:25822986

Rat Stem-Cell Factor Induces Splenocytes Capable Of Regenerating The Thymus

PubMed Central

Migita, Russell T.; Trebasky, Lisa D.; Housman, Jerry M.; Elliott, Gary S.; Hendren, R. Wayne; Deprince, Randolph B.; Greiner, Dale L.

1992-01-01

Cytokine regulation of prethymic T-lymphoid progenitor-cell proliferation and/or differentiation has not been well-defined, although much is known of cytokine regulation of hemopoietic stem- and progenitor-cell development. Here we use a recently identified hemopoietic growth factor, stem-cell factor (SCF) (a form of the c-kit ligand), and a transplant model of thymocyte regeneration to assess the effect of SCF on the in vivo generation of prethymic, thymocyte progenitor-cell activity. We show that recombinant rat SCF (rrSCF164 administered to weanling rats selectively induces an increase in thymocyte progenitor activity in the spleens of treated rats as compared to rats treated with vehicle, polyethylene glycol (PEG)-conjugated rat albumin, or recombinant human granulocyte colony-stimulating factor (rhG-CSF). These data demonstrate that administration of SCF in vivo affects extrathymic-origin thymocyte regenerating cells and may influence, directly or indirectly, early prethymic stages of T-cell lymphopoiesis in addition to its known effect on early stages of myelopoiesis and erythropoiesis. PMID:1285280

Effects of low-doses of Bacillus spp. from permafrost on differentiation of bone marrow cells.

PubMed

Kalyonova, L F; Novikova, M A; Kostolomova, E G

2015-01-01

The effects of a new microorganism species (Bacillus spp., strain M3) isolated from permafrost specimens from Central Yakutia (Mamontova Mountain) on the bone marrow hemopoiesis were studied on laboratory mice. Analysis of the count and immunophenotype of bone marrow cells indicated that even in low doses (1000-5000 microbial cells) these microorganisms modulated hemopoiesis and lymphopoiesis activity. The percentage of early hemopoietic precursors (CD117(+)CD34(-)) increased, intensity of lymphocyte precursor proliferation and differentiation (CD25(+)CD44(-)) decreased, and the percentage of lymphocytes released from the bone marrow (CD25(+)CD44(+)) increased on day 21 after injection of the bacteria. These changes in activity of hemopoiesis were associated with changes in the level of regulatory T lymphocytes (reduced expression of TCRαβ) and were most likely compensatory. The possibility of modulating hemopoiesis activity in the bone marrow by low doses of one microorganism strain isolated from the permafrost could be useful for evaluating the effects of other low dose bacteria on the bone marrow hemopoiesis.

The Nlrp3 inflammasome promotes age-related thymic demise and immunosenescence.

PubMed

Youm, Yun-Hee; Kanneganti, Thirumala-Devi; Vandanmagsar, Bolormaa; Zhu, Xuewei; Ravussin, Anthony; Adijiang, Ayinuer; Owen, John S; Thomas, Michael J; Francis, Joseph; Parks, John S; Dixit, Vishwa Deep

2012-01-26

The collapse of thymic stromal cell microenvironment with age and resultant inability of the thymus to produce naive T cells contributes to lower immune-surveillance in the elderly. Here we show that age-related increase in 'lipotoxic danger signals' such as free cholesterol (FC) and ceramides, leads to thymic caspase-1 activation via the Nlrp3 inflammasome. Elimination of Nlrp3 and Asc, a critical adaptor required for inflammasome assembly, reduces age-related thymic atrophy and results in an increase in cortical thymic epithelial cells, T cell progenitors and maintenance of T cell repertoire diversity. Using a mouse model of irradiation and hematopoietic stem cell transplantation (HSCT), we show that deletion of the Nlrp3 inflammasome accelerates T cell reconstitution and immune recovery in middle-aged animals. Collectively, these data demonstrate that lowering inflammasome-dependent caspase-1 activation increases thymic lymphopoiesis and suggest that Nlrp3 inflammasome inhibitors may aid the re-establishment of a diverse T cell repertoire in middle-aged or elderly patients undergoing HSCT. Copyright © 2012 The Authors. Published by Elsevier Inc. All rights reserved.

Immune Efficacy of a Genetically Engineered Vaccine against Lymphocystis Disease Virus: Analysis of Different Immunization Strategies

PubMed Central

Zheng, Fengrong; Sun, Xiuqin; Wu, Xing'an; Liu, Hongzhan; Li, Jiye; Wu, Suqi; Zhang, Jinxing

2011-01-01

Here, we report the construction of a vaccine against lymphocystis disease virus (LCDV) using nucleic acid vaccination technology. A fragment of the major capsid protein encoding gene from an LCDV isolated from China (LCDV-cn) was cloned into an eukaryotic expression vector pEGFP-N2, yielding a recombinant plasmid pEGFP-N2-LCDV-cn0.6 kb. This plasmid was immediately expressed after liposomal transfer into the Japanese flounder embryo cell line. The recombinant plasmid was inoculated into Japanese flounder via two routes (intramuscular injection and hypodermic injection) at three doses (0.1, 5, and 15 μg), and then T-lymphopoiesis in different tissues and antibodies raised against LCDV were evaluated. The results indicated that this recombinant plasmid induced unique humoral or cell-mediated immune responses depending on the inoculation route and conferred immune protection. Furthermore, the humoral immune responses and protective effects were significantly increased at higher vaccine doses via the two injection routes. Plasmid pEGFP-N2-LCDV0.6 kb is therefore a promising vaccine candidate against LCDV in Japanese flounder. PMID:21789044

Alpha-Particle-Induced Complex Chromosome Exchanges Transmitted through Extra-Thymic Lymphopoiesis In Vitro Show Evidence of Emerging Genomic Instability

PubMed Central

Sumption, Natalia; Goodhead, Dudley T.; Anderson, Rhona M.

2015-01-01

Human exposure to high-linear energy transfer α-particles includes environmental (e.g. radon gas and its decay progeny), medical (e.g. radiopharmaceuticals) and occupational (nuclear industry) sources. The associated health risks of α-particle exposure for lung cancer are well documented however the risk estimates for leukaemia remain uncertain. To further our understanding of α-particle effects in target cells for leukaemogenesis and also to seek general markers of individual exposure to α-particles, this study assessed the transmission of chromosomal damage initially-induced in human haemopoietic stem and progenitor cells after exposure to high-LET α-particles. Cells surviving exposure were differentiated into mature T-cells by extra-thymic T-cell differentiation in vitro. Multiplex fluorescence in situ hybridisation (M-FISH) analysis of naïve T-cell populations showed the occurrence of stable (clonal) complex chromosome aberrations consistent with those that are characteristically induced in spherical cells by the traversal of a single α-particle track. Additionally, complex chromosome exchanges were observed in the progeny of irradiated mature T-cell populations. In addition to this, newly arising de novo chromosome aberrations were detected in cells which possessed clonal markers of α-particle exposure and also in cells which did not show any evidence of previous exposure, suggesting ongoing genomic instability in these populations. Our findings support the usefulness and reliability of employing complex chromosome exchanges as indicators of past or ongoing exposure to high-LET radiation and demonstrate the potential applicability to evaluate health risks associated with α-particle exposure. PMID:26252014

Human SLP-65 isoforms contribute differently to activation and apoptosis of B lymphocytes.

PubMed

Grabbe, Annika; Wienands, Jürgen

2006-12-01

The SH2 domain-containing leukocyte adaptor protein of 65 kDa (SLP-65) is the key effector for signaling downstream of the B-cell antigen receptor (BCR). SLP-65 controls not only B lymphopoiesis and humoral immunity but also possesses a yet poorly defined tumor suppressor activity that is lost in many cases of acute lymphoblastic leukemia. We found that the 2 isoforms of human SLP-65 are differentially involved in positive and negative B-cell signaling. Reconstitution experiments revealed that an atypical SH3 domain-binding motif, which is present in the long but not in the short SLP-65 isoform, mediates association to Grb2 and suppresses activation of mitogen-activated protein kinases p38 and JNK as well as up-regulation of c-Fos expression. In turn, the short isoform activates not only AP1-driven but also NF-kappaB-driven gene transcription more potently than the long isoform. Conversely, the long rather than the short SLP-65 isoform promotes BCR-induced B-cell apoptosis. Our data further delineate the structural requirements of positive and negative SLP-65 signal transduction in normal and neoplastic cells.

Instruction of haematopoietic lineage choices, evolution of transcriptional landscapes and cancer stem cell hierarchies derived from an AML1-ETO mouse model

PubMed Central

Cabezas-Wallscheid, Nina; Eichwald, Victoria; de Graaf, Jos; Löwer, Martin; Lehr, Hans-Anton; Kreft, Andreas; Eshkind, Leonid; Hildebrandt, Andreas; Abassi, Yasmin; Heck, Rosario; Dehof, Anna Katharina; Ohngemach, Svetlana; Sprengel, Rolf; Wörtge, Simone; Schmitt, Steffen; Lotz, Johannes; Meyer, Claudius; Kindler, Thomas; Zhang, Dong-Er; Kaina, Bernd; Castle, John C; Trumpp, Andreas; Sahin, Ugur; Bockamp, Ernesto

2013-01-01

The t(8;21) chromosomal translocation activates aberrant expression of the AML1-ETO (AE) fusion protein and is commonly associated with core binding factor acute myeloid leukaemia (CBF AML). Combining a conditional mouse model that closely resembles the slow evolution and the mosaic AE expression pattern of human t(8;21) CBF AML with global transcriptome sequencing, we find that disease progression was characterized by two principal pathogenic mechanisms. Initially, AE expression modified the lineage potential of haematopoietic stem cells (HSCs), resulting in the selective expansion of the myeloid compartment at the expense of normal erythro- and lymphopoiesis. This lineage skewing was followed by a second substantial rewiring of transcriptional networks occurring in the trajectory to manifest leukaemia. We also find that both HSC and lineage-restricted granulocyte macrophage progenitors (GMPs) acquired leukaemic stem cell (LSC) potential being capable of initiating and maintaining the disease. Finally, our data demonstrate that long-term expression of AE induces an indolent myeloproliferative disease (MPD)-like myeloid leukaemia phenotype with complete penetrance and that acute inactivation of AE function is a potential novel therapeutic option. PMID:24124051

The impact of trisomy 21 on foetal haematopoiesis.

PubMed

Roberts, Irene; O'Connor, David; Roy, Anindita; Cowan, Gillian; Vyas, Paresh

2013-12-01

The high frequency of a unique neonatal preleukaemic syndrome, transient abnormal myelopoiesis (TAM), and subsequent acute myeloid leukaemia in early childhood in patients with trisomy 21 (Down syndrome) points to a specific role for trisomy 21 in transforming foetal haematopoietic cells. N-terminal truncating mutations in the key haematopoietic transcription factor GATA1 are acquired during foetal life in virtually every case. These mutations are not leukaemogenic in the absence of trisomy 21. In mouse models, deregulated expression of chromosome 21-encoded genes is implicated in leukaemic transformation, but does not recapitulate the effects of trisomy 21 in a human context. Recent work using primary human foetal liver and bone marrow cells, human embryonic stem cells and iPS cells shows that prior to acquisition of GATA1 mutations, trisomy 21 itself alters human foetal haematopoietic stem cell and progenitor cell biology causing multiple abnormalities in myelopoiesis and B-lymphopoiesis. The molecular basis by which trisomy 21 exerts these effects is likely to be extremely complex, to be tissue-specific and lineage-specific and to be dependent on ontogeny-related characteristics of the foetal microenvironment. © 2013.

The impact of trisomy 21 on foetal haematopoiesis

PubMed Central

Roberts, Irene; O'Connor, David; Roy, Anindita; Cowan, Gillian; Vyas, Paresh

2015-01-01

The high frequency of a unique neonatal preleukaemic syndrome, Transient Abnormal Myelopoiesis (TAM), and subsequent acute myeloid leukaemia in early childhood in patients with trisomy 21 (Down syndrome) points to a specific role for trisomy 21 in transforming foetal haematopoietic cells. N-terminal truncating mutations in the key haematopoietic transcription factor GATA1 are acquired during foetal life in virtually every case. These mutations are not leukaemogenic in the absence of trisomy 21. In mouse models, deregulated expression of chromosome 21-encoded genes is implicated in leukaemic transformation, but does not recapitulate the effects of trisomy 21 in a human context. Recent work using primary human foetal liver and bone marrow cells, human embryonic stem cells and iPS cells cells shows that prior to acquistion of GATA1 mutations, trisomy 21 itself alters human foetal haematopoietic stem cell and progenitor cell biology causing multiple abnormalities in myelopoiesis and B-lymphopoiesis. The molecular basis by which trisomy 21 exerts these effects is likely to be extremely complex, to be tissue- and lineage-specific and to be dependent on ontogeny-related characteristics of the foetal microenvironment. PMID:23932236

Developmentally induced Mll1 loss reveals defects in postnatal haematopoiesis.

PubMed

Gan, T; Jude, C D; Zaffuto, K; Ernst, P

2010-10-01

The mixed lineage leukemia (MLL) gene is disrupted by chromosomal translocations in acute leukemia, producing a fusion oncogene with altered properties relative to the wild-type gene. Murine loss-of-function studies have shown an essential role for Mll in developing the haematopoietic system, yet studies using different conditional knockout models have yielded conflicting results regarding the requirement for Mll during adult steady-state haematopoiesis. In this study, we used a loxP-flanked Mll allele (Mll(F)) and a developmentally regulated, haematopoietic-specific VavCre transgene to reassess the consequences of Mll loss in the haematopoietic lineage, without the need for inducers of Cre recombinase. We show that VavCre;Mll mutants exhibit phenotypically normal fetal haematopoiesis, but rarely survive past 3 weeks of age. Surviving animals are anemic, thrombocytopenic and exhibit a significant reduction in bone marrow haematopoietic stem/progenitor populations, consistent with our previous findings using the inducible Mx1Cre transgene. Furthermore, the analysis of VavCre mutants revealed additional defects in B-lymphopoiesis that could not be assessed using Mx1Cre-mediated Mll deletion. Collectively, these data support the conclusion that Mll has an essential role in sustaining postnatal haematopoiesis.

Use of Mutated Self-Cleaving 2A Peptides as a Molecular Rheostat to Direct Simultaneous Formation of Membrane and Secreted Anti-HIV Immunoglobulins

PubMed Central

Yu, Kenneth K.; Aguilar, Kiefer; Tsai, Jonathan; Galimidi, Rachel; Gnanapragasam, Priyanthi; Yang, Lili; Baltimore, David

2012-01-01

In nature, B cells produce surface immunoglobulin and secreted antibody from the same immunoglobulin gene via alternative splicing of the pre-messenger RNA. Here we present a novel system for genetically programming B cells to direct the simultaneous formation of membrane-bound and secreted immunoglobulins that we term a “Molecular Rheostat”, based on the use of mutated “self-cleaving” 2A peptides. The Molecular Rheostat is designed so that the ratio of secreted to membrane-bound immunoglobulins can be controlled by selecting appropriate mutations in the 2A peptide. Lentiviral transgenesis of Molecular Rheostat constructs into B cell lines enables the simultaneous expression of functional b12-based IgM-like BCRs that signal to the cells and mediate the secretion of b12 IgG broadly neutralizing antibodies that can bind and neutralize HIV-1 pseudovirus. We show that these b12-based Molecular Rheostat constructs promote the maturation of EU12 B cells in an in vitro model of B lymphopoiesis. The Molecular Rheostat offers a novel tool for genetically manipulating B cell specificity for B-cell based gene therapy. PMID:23209743

Leukemogenesis Induced by an Activating β-catenin mutation in Osteoblasts

PubMed Central

Kode, Aruna; Manavalan, John S.; Mosialou, Ioanna; Bhagat, Govind; Rathinam, Chozha V.; Luo, Na; Khiabanian, Hossein; Lee, Albert; Vundavalli, Murty; Friedman, Richard; Brum, Andrea; Park, David; Galili, Naomi; Mukherjee, Siddhartha; Teruya-Feldstein, Julie; Raza, Azra; Rabadan, Raul; Berman, Ellin; Kousteni, Stavroula

2014-01-01

Summary Cells of the osteoblast lineage affect homing, 1, 2 number of long term repopulating hematopoietic stem cells (HSCs) 3, 4, HSC mobilization and lineage determination and B lymphopoiesis 5-8. More recently osteoblasts were implicated in pre-leukemic conditions in mice 9, 10. Yet, it has not been shown that a single genetic event taking place in osteoblasts can induce leukemogenesis. We show here that in mice, an activating mutation of β-catenin in osteoblasts alters the differentiation potential of myeloid and lymphoid progenitors leading to development of acute myeloid leukemia (AML) with common chromosomal aberrations and cell autonomous progression. Activated β-catenin stimulates expression of the Notch ligand Jagged-1 in osteoblasts. Subsequent activation of Notch signaling in HSC progenitors induces the malignant changes. Demonstrating the pathogenetic role of the Notch pathway, genetic or pharmacological inhibition of Notch signaling ameliorates AML. Nuclear accumulation and increased β-catenin signaling in osteoblasts was also identified in 38% of patients with MDS/AML. These patients showed increased Notch signaling in hematopoietic cells. These findings demonstrate that genetic alterations in osteoblasts can induce AML, identify molecular signals leading to this transformation and suggest a potential novel pharmacotherapeutic approach to AML. PMID:24429522

Independent regulation of the two Pax5 alleles during B-cell development.

PubMed

Nutt, S L; Vambrie, S; Steinlein, P; Kozmik, Z; Rolink, A; Weith, A; Busslinger, M

1999-04-01

The developmental control genes of the Pax family are frequently associated with mouse mutants and human disease syndromes. The function of these transcription factors is sensitive to gene dosage, as mutation of one allele or a modest increase in gene number results in phenotypic abnormalities. Pax5 has an important role in B-cell and midbrain development. By following the expression of individual Pax5 alleles at the single-cell level, we demonstrate here that Pax5 is subject to allele-specific regulation during B-lymphopoiesis. Pax5 is predominantly transcribed from only one allele in early progenitors and mature B cells, whereas it switches to a biallelic transcription mode in immature B cells. The allele-specific regulation of Pax5 is stochastic, reversible, independent of parental origin and correlates with synchronous replication, in contrast with imprinted and other monoallelically expressed genes. As a consequence, B-lymphoid tissues are mosaics with respect to the transcribed Pax5 allele, and thus mutation of one allele in heterozygous mice results in deletion of the cell population expressing the mutant allele due to loss of Pax5 function at the single-cell level. Similar allele-specific regulation may be a common mechanism causing the haploinsufficiency and frequent association of other Pax genes with human disease.

Dietary restriction improves repopulation but impairs lymphoid differentiation capacity of hematopoietic stem cells in early aging

PubMed Central

Tang, Duozhuang; Tao, Si; Chen, Zhiyang; Koliesnik, Ievgen Oleksandrovich; Calmes, Philip Gerald; Hoerr, Verena; Han, Bing; Gebert, Nadja; Zörnig, Martin; Löffler, Bettina

2016-01-01

Dietary restriction (DR) improves health, delays tissue aging, and elongates survival in flies and worms. However, studies on laboratory mice and nonhuman primates revealed ambiguous effects of DR on lifespan despite improvements in health parameters. In this study, we analyzed consequences of adult-onset DR (24 h to 1 yr) on hematopoietic stem cell (HSC) function. DR ameliorated HSC aging phenotypes, such as the increase in number of HSCs and the skewing toward myeloid-biased HSCs during aging. Furthermore, DR increased HSC quiescence and improved the maintenance of the repopulation capacity of HSCs during aging. In contrast to these beneficial effects, DR strongly impaired HSC differentiation into lymphoid lineages and particularly inhibited the proliferation of lymphoid progenitors, resulting in decreased production of peripheral B lymphocytes and impaired immune function. The study shows that DR-dependent suppression of growth factors and interleukins mediates these divergent effects caused by DR. Supplementation of insulin-like growth factor 1 partially reverted the DR-induced quiescence of HSCs, whereas IL-6/IL-7 substitutions rescued the impairment of B lymphopoiesis exposed to DR. Together, these findings delineate positive and negative effects of long-term DR on HSC functionality involving distinct stress and growth signaling pathways. PMID:26951333

IL-7 receptor blockade following T cell depletion promotes long-term allograft survival

PubMed Central

Mai, Hoa-Le; Boeffard, Françoise; Longis, Julie; Danger, Richard; Martinet, Bernard; Haspot, Fabienne; Vanhove, Bernard; Brouard, Sophie; Soulillou, Jean-Paul

2014-01-01

T cell depletion is commonly used in organ transplantation for immunosuppression; however, a restoration of T cell homeostasis following depletion leads to increased memory T cells, which may promote transplant rejection. The cytokine IL-7 is important for controlling lymphopoiesis under both normal and lymphopenic conditions. Here, we investigated whether blocking IL-7 signaling with a mAb that targets IL-7 receptor α (IL-7Rα) alone or following T cell depletion confers an advantage for allograft survival in murine transplant models. We found that IL-7R blockade alone induced indefinite pancreatic islet allograft survival if anti–IL-7R treatment was started 3 weeks before graft. IL-7R blockade following anti-CD4– and anti-CD8–mediated T cell depletion markedly prolonged skin allograft survival. Furthermore, IL-7 inhibition in combination with T cell depletion synergized with either CTLA-4Ig administration or suboptimal doses of tacrolimus to induce long-term skin graft acceptance in this stringent transplant model. Together, these therapies inhibited T cell reconstitution, decreased memory T cell numbers, increased the relative frequency of Tregs, and abrogated both cellular and humoral alloimmune responses. Our data suggest that IL-7R blockade following T cell depletion has potential as a robust, immunosuppressive therapy in transplantation. PMID:24569454

Lentiviral Vector Induced Insertional Haploinsufficiency of Ebf1 Causes Murine Leukemia

PubMed Central

Heckl, Dirk; Schwarzer, Adrian; Haemmerle, Reinhard; Steinemann, Doris; Rudolph, Cornelia; Skawran, Britta; Knoess, Sabine; Krause, Johanna; Li, Zhixiong; Schlegelberger, Brigitte; Baum, Christopher; Modlich, Ute

2012-01-01

Integrating vectors developed on the basis of various retroviruses have demonstrated therapeutic potential following genetic modification of long-lived hematopoietic stem and progenitor cells. Lentiviral vectors (LV) are assumed to circumvent genotoxic events previously observed with γ-retroviral vectors, due to their integration bias to transcription units in comparison to the γ-retroviral preference for promoter regions and CpG islands. However, recently several studies have revealed the potential for gene activation by LV insertions. Here, we report a murine acute B-lymphoblastic leukemia (B-ALL) triggered by insertional gene inactivation. LV integration occurred into the 8th intron of Ebf1, a major regulator of B-lymphopoiesis. Various aberrant splice variants could be detected that involved splice donor and acceptor sites of the lentiviral construct, inducing downregulation of Ebf1 full-length message. The transcriptome signature was compatible with loss of this major determinant of B-cell differentiation, with partial acquisition of myeloid markers, including Csf1r (macrophage colony-stimulating factor (M-CSF) receptor). This was accompanied by receptor phosphorylation and STAT5 activation, both most likely contributing to leukemic progression. Our results highlight the risk of intragenic vector integration to initiate leukemia by inducing haploinsufficiency of a tumor suppressor gene. We propose to address this risk in future vector design. PMID:22472950

B Cell Development in the Bone Marrow Is Regulated by Homeostatic Feedback Exerted by Mature B Cells

PubMed Central

Shahaf, Gitit; Zisman-Rozen, Simona; Benhamou, David; Melamed, Doron; Mehr, Ramit

2016-01-01

Cellular homeostasis in the B cell compartment is strictly imposed to balance cell production and cell loss. However, it is not clear whether B cell development in the bone marrow is an autonomous process or subjected to regulation by the peripheral B cell compartment. To specifically address this question, we used mice transgenic for human CD20, where effective depletion of B lineage cells is obtained upon administration of mouse anti-human CD20 antibodies, in the absence of any effect on other cell lineages and/or tissues. We followed the kinetics of B cell return to equilibrium by BrdU labeling and flow cytometry and analyzed the resulting data by mathematical modeling. Labeling was much faster in depleted mice. Compared to control mice, B cell-depleted mice exhibited a higher proliferation rate in the pro-/pre-B compartment, and higher cell death and lower differentiation in the immature B cell compartment. We validated the first result by analysis of the expression of Ki67, the nuclear protein expressed in proliferating cells, and the second using Annexin V staining. Collectively, our results suggest that B lymphopoiesis is subjected to homeostatic feedback mechanisms imposed by mature B cells in the peripheral compartment. PMID:27047488

Aging-associated inflammation promotes selection for adaptive oncogenic events in B cell progenitors.

PubMed

Henry, Curtis J; Casás-Selves, Matias; Kim, Jihye; Zaberezhnyy, Vadym; Aghili, Leila; Daniel, Ashley E; Jimenez, Linda; Azam, Tania; McNamee, Eoin N; Clambey, Eric T; Klawitter, Jelena; Serkova, Natalie J; Tan, Aik Choon; Dinarello, Charles A; DeGregori, James

2015-12-01

The incidence of cancer is higher in the elderly; however, many of the underlying mechanisms for this association remain unexplored. Here, we have shown that B cell progenitors in old mice exhibit marked signaling, gene expression, and metabolic defects. Moreover, B cell progenitors that developed from hematopoietic stem cells (HSCs) transferred from young mice into aged animals exhibited similar fitness defects. We further demonstrated that ectopic expression of the oncogenes BCR-ABL, NRAS(V12), or Myc restored B cell progenitor fitness, leading to selection for oncogenically initiated cells and leukemogenesis specifically in the context of an aged hematopoietic system. Aging was associated with increased inflammation in the BM microenvironment, and induction of inflammation in young mice phenocopied aging-associated B lymphopoiesis. Conversely, a reduction of inflammation in aged mice via transgenic expression of α-1-antitrypsin or IL-37 preserved the function of B cell progenitors and prevented NRAS(V12)-mediated oncogenesis. We conclude that chronic inflammatory microenvironments in old age lead to reductions in the fitness of B cell progenitor populations. This reduced progenitor pool fitness engenders selection for cells harboring oncogenic mutations, in part due to their ability to correct aging-associated functional defects. Thus, modulation of inflammation--a common feature of aging--has the potential to limit aging-associated oncogenesis.

The transcriptional coactivator Maml1 is required for Notch2-mediated marginal zone B-cell development

PubMed Central

Maillard, Ivan; Nakamura, Makoto; Pear, Warren S.; Griffin, James D.

2007-01-01

Signaling mediated by various Notch receptors and their ligands regulates diverse biological processes, including lymphoid cell fate decisions. Notch1 is required during T-cell development, while Notch2 and the Notch ligand Delta-like1 control marginal zone B (MZB) cell development. We previously determined that Mastermind-like (MAML) transcriptional coactivators are required for Notchinduced transcription by forming ternary nuclear complexes with Notch and the transcription factor CSL. The 3 MAML family members (MAML1-MAML3) are collectively essential for Notch activity in vivo, but whether individual MAMLs contribute to the specificity of Notch functions is unknown. Here, we addressed this question by studying lymphopoiesis in the absence of the Maml1 gene. Since Maml1−/− mice suffered perinatal lethality, hematopoietic chimeras were generated with Maml1−/−, Maml1+/−, or wild-type fetal liver progenitors. Maml1 deficiency minimally affected T-cell development, but was required for the development of MZB cells, similar to the phenotype of Notch2 deficiency. Moreover, the number of MZB cells correlated with Maml1 gene dosage. Since all 3 Maml genes were expressed in MZB cells and their precursors, these results suggest that Maml1 is specifically required for Notch2 signaling in MZB cells. PMID:17699740

Impaired B cell development in the absence of Krüppel-like factor 3.

PubMed

Vu, Thi Thanh; Gatto, Dominique; Turner, Vivian; Funnell, Alister P W; Mak, Ka Sin; Norton, Laura J; Kaplan, Warren; Cowley, Mark J; Agenès, Fabien; Kirberg, Jörg; Brink, Robert; Pearson, Richard C M; Crossley, Merlin

2011-11-15

Krüppel-like factor 3 (Klf3) is a member of the Klf family of transcription factors. Klfs are widely expressed and have diverse roles in development and differentiation. In this study, we examine the function of Klf3 in B cell development by studying B lymphopoiesis in a Klf3 knockout mouse model. We show that B cell differentiation is significantly impaired in the bone marrow, spleen, and peritoneal cavity of Klf3 null mice and confirm that the defects are cell autonomous. In the bone marrow, there is a reduction in immature B cells, whereas recirculating mature cells are noticeably increased. Immunohistology of the spleen reveals a poorly structured marginal zone (MZ) that may in part be caused by deregulation of adhesion molecules on MZ B cells. In the peritoneal cavity, there are significant defects in B1 B cell development. We also report that the loss of Klf3 in MZ B cells is associated with reduced BCR signaling strength and an impaired ability to respond to LPS stimulation. Finally, we show increased expression of a number of Klf genes in Klf3 null B cells, suggesting that a Klf regulatory network may exist in B cells.

Depletion of stromal cells expressing fibroblast activation protein-α from skeletal muscle and bone marrow results in cachexia and anemia

PubMed Central

Roberts, Edward W.; Deonarine, Andrew; Jones, James O.; Denton, Alice E.; Feig, Christine; Lyons, Scott K.; Espeli, Marion; Kraman, Matthew; McKenna, Brendan; Wells, Richard J.B.; Zhao, Qi; Caballero, Otavia L.; Larder, Rachel; Coll, Anthony P.; O’Rahilly, Stephen; Brindle, Kevin M.; Teichmann, Sarah A.; Tuveson, David A.

2013-01-01

Fibroblast activation protein-α (FAP) identifies stromal cells of mesenchymal origin in human cancers and chronic inflammatory lesions. In mouse models of cancer, they have been shown to be immune suppressive, but studies of their occurrence and function in normal tissues have been limited. With a transgenic mouse line permitting the bioluminescent imaging of FAP+ cells, we find that they reside in most tissues of the adult mouse. FAP+ cells from three sites, skeletal muscle, adipose tissue, and pancreas, have highly similar transcriptomes, suggesting a shared lineage. FAP+ cells of skeletal muscle are the major local source of follistatin, and in bone marrow they express Cxcl12 and KitL. Experimental ablation of these cells causes loss of muscle mass and a reduction of B-lymphopoiesis and erythropoiesis, revealing their essential functions in maintaining normal muscle mass and hematopoiesis, respectively. Remarkably, these cells are altered at these sites in transplantable and spontaneous mouse models of cancer-induced cachexia and anemia. Thus, the FAP+ stromal cell may have roles in two adverse consequences of cancer: their acquisition by tumors may cause failure of immunosurveillance, and their alteration in normal tissues contributes to the paraneoplastic syndromes of cachexia and anemia. PMID:23712428

Site-Specific Expression of Polycomb-Group Genes Encoding the HPC-HPH/PRC1 Complex in Clinically Defined Primary Nodal and Cutaneous Large B-Cell Lymphomas

PubMed Central

Raaphorst, Frank M.; Vermeer, Maarten; Fieret, Elly; Blokzijl, Tjasso; Dukers, Danny; Sewalt, Richard G.A.B.; Otte, Arie P.; Willemze, Rein; Meijer, Chris J.L.M.

2004-01-01

Polycomb-group (PcG) genes preserve cell identity by gene silencing, and contribute to regulation of lymphopoiesis and malignant transformation. We show that primary nodal large B-cell lymphomas (LBCLs), and secondary cutaneous deposits from such lymphomas, abnormally express the BMI-1, RING1, and HPH1 PcG genes in cycling neoplastic cells. By contrast, tumor cells in primary cutaneous LBCLs lacked BMI-1 expression, whereas RING1 was variably detected. Lack of BMI-1 expression was characteristic for primary cutaneous LBCLs, because other primary extranodal LBCLs originating from brain, testes, and stomach were BMI-1-positive. Expression of HPH1 was rarely detected in primary cutaneous LBCLs of the head or trunk and abundant in primary cutaneous LBCLs of the legs, which fits well with its earlier recognition as a distinct clinical pathological entity with different clinical behavior. We conclude that clinically defined subclasses of primary LBCLs display site-specific abnormal expression patterns of PcG genes of the HPC-HPH/PRC1 PcG complex. Some of these patterns (such as the expression profile of BMI-1) may be diagnostically relevant. We propose that distinct expression profiles of PcG genes results in abnormal formation of HPC-HPH/PRC1 PcG complexes, and that this contributes to lymphomagenesis and different clinical behavior of clinically defined LBCLs. PMID:14742259

Congenital hypoplastic bone marrow failure associated with a de novo partial deletion of the MECOM gene at 3q26.2.

PubMed

Kjeldsen, Eigil; Veigaard, Christopher; Aggerholm, Anni; Hasle, Henrik

2018-05-20

Congenital hypoplastic bone marrow failure is a rare condition in neonates. The genetics and mechanisms behind are largely obscure. Here we characterize a neonate presenting with congenital thrombocytopenia and anemia. During the first 2-4 weeks after birth the neonate developed severe neutropenia while the lymphoid lineages were unaffected. The neonate was without dysmorphic signs. A de novo mono-allelic constitutional microdeletion of 175.1 kb at 3q26.2 affecting exon 2 of MECOM, involving MDS1 but not EVI1, was identified as the only copy number alteration by oligo-based array-CGH analysis. Expression analysis showed profoundly reduced expression of multiple MECOM transcripts in the bone marrow cells. Whole exome sequencing detected no pathogenic mutations in genes known to be associated with inherited bone marrow failure syndromes. The patient was successfully treated with hematopoietic stem cell transplantation at 5 months of age. Interstitial deletions encompassing the 3q26.2 region are very rare. A literature search revealed two previous cases with microdeletions involving this region, and the cases were associated with congenital thrombocytopenia and anemia, but unaffected lymphopoiesis. Together these data indicate that MECOM may be important for normal myeloid hematopoiesis in humans but dispensable for lymphoid differentiation. We suggest that partial deletion in MECOM may be a primary event associated with congenital pancytopenia. Copyright © 2018 Elsevier B.V. All rights reserved.

Superior Therapeutic Index in Lymphoma Therapy: CD30+ CD34+ Hematopoietic Stem Cells Resist a Chimeric Antigen Receptor T-cell Attack

PubMed Central

Hombach, Andreas A; Görgens, André; Chmielewski, Markus; Murke, Florian; Kimpel, Janine; Giebel, Bernd; Abken, Hinrich

2016-01-01

Recent clinical trials with chimeric antigen receptor (CAR) redirected T cells targeting CD19 revealed particular efficacy in the treatment of leukemia/lymphoma, however, were accompanied by a lasting depletion of healthy B cells. We here explored CD30 as an alternative target, which is validated in lymphoma therapy and expressed by a broad variety of Hodgkin's and non-Hodgkin's lymphomas. As a safty concern, however, CD30 is also expressed by lymphocytes and hematopoietic stem and progenitor cells (HSPCs) during activation. We revealed that HRS3scFv-derived CAR T cells are superior since they were not blocked by soluble CD30 and did not attack CD30+ HSPCs while eliminating CD30+ lymphoma cells. Consequently, normal hemato- and lymphopoiesis was not affected in the long-term in the humanized mouse; the number of blood B and T cells remained unchanged. We provide evidence that the CD30+ HSPCs are protected against a CAR T-cell attack by substantially lower CD30 levels than lymphoma cells and higher levels of the granzyme B inactivating SP6/PI9 serine protease, which furthermore increased upon activation. Taken together, adoptive cell therapy with anti-CD30 CAR T cells displays a superior therapeutic index in the treatment of CD30+ malignancies leaving healthy activated lymphocytes and HSPCs unaffected. PMID:27112062

Ibrutinib inhibits pre-BCR+ B-cell acute lymphoblastic leukemia progression by targeting BTK and BLK.

PubMed

Kim, Ekaterina; Hurtz, Christian; Koehrer, Stefan; Wang, Zhiqiang; Balasubramanian, Sriram; Chang, Betty Y; Müschen, Markus; Davis, R Eric; Burger, Jan A

2017-03-02

Targeting B-cell receptor (BCR) signaling is a successful therapeutic strategy in mature B-cell malignancies. Precursor BCR (pre-BCR) signaling, which is critical during normal B lymphopoiesis, also plays an important role in pre-BCR + B cell acute lymphoblastic leukemia (B-ALL). Here, we investigated the activity and mechanism of action of the BTK inhibitor ibrutinib in preclinical models of B-ALL. Pre-BCR + ALL cells were exquisitely sensitive to ibrutinib at therapeutically relevant drug concentrations. In pre-BCR + ALL, ibrutinib thwarted autonomous and induced pre-BCR signaling, resulting in deactivation of PI3K/Akt signaling. Ibrutinib modulated the expression of pre-BCR regulators (PTPN6, CD22, CD72, and PKCβ) and substantially reduced BCL6 levels. Ibrutinib inhibited ALL cell migration toward CXCL12 and beneath marrow stromal cells and reduced CD44 expression. CRISPR-Cas9 gene editing revealed that both BTK and B lymphocyte kinase (BLK) are relevant targets of ibrutinib in pre-BCR + ALL. Consequently, in mouse xenograft models of pre-BCR + ALL, ibrutinib treatment significantly prolonged survival. Combination treatment of ibrutinib with dexamethasone or vincristine demonstrated synergistic activity against pre-BCR + ALL. These data corroborate ibrutinib as a promising targeted agent for pre-BCR + ALL and highlight the importance of ibrutinib effects on alternative kinase targets. © 2017 by The American Society of Hematology.

Ibrutinib inhibits pre-BCR+ B-cell acute lymphoblastic leukemia progression by targeting BTK and BLK

PubMed Central

Kim, Ekaterina; Hurtz, Christian; Koehrer, Stefan; Wang, Zhiqiang; Balasubramanian, Sriram; Chang, Betty Y.; Müschen, Markus; Davis, R. Eric

2017-01-01

Targeting B-cell receptor (BCR) signaling is a successful therapeutic strategy in mature B-cell malignancies. Precursor BCR (pre-BCR) signaling, which is critical during normal B lymphopoiesis, also plays an important role in pre-BCR+ B cell acute lymphoblastic leukemia (B-ALL). Here, we investigated the activity and mechanism of action of the BTK inhibitor ibrutinib in preclinical models of B-ALL. Pre-BCR+ ALL cells were exquisitely sensitive to ibrutinib at therapeutically relevant drug concentrations. In pre-BCR+ ALL, ibrutinib thwarted autonomous and induced pre-BCR signaling, resulting in deactivation of PI3K/Akt signaling. Ibrutinib modulated the expression of pre-BCR regulators (PTPN6, CD22, CD72, and PKCβ) and substantially reduced BCL6 levels. Ibrutinib inhibited ALL cell migration toward CXCL12 and beneath marrow stromal cells and reduced CD44 expression. CRISPR-Cas9 gene editing revealed that both BTK and B lymphocyte kinase (BLK) are relevant targets of ibrutinib in pre-BCR+ ALL. Consequently, in mouse xenograft models of pre-BCR+ ALL, ibrutinib treatment significantly prolonged survival. Combination treatment of ibrutinib with dexamethasone or vincristine demonstrated synergistic activity against pre-BCR+ ALL. These data corroborate ibrutinib as a promising targeted agent for pre-BCR+ ALL and highlight the importance of ibrutinib effects on alternative kinase targets. PMID:28031181

Regulation of T Cell Differentiation and Function by EZH2

PubMed Central

Karantanos, Theodoros; Christofides, Anthos; Bardhan, Kankana; Li, Lequn; Boussiotis, Vassiliki A.

2016-01-01

The enhancer of zeste homolog 2 (EZH2), one of the polycomb-group proteins, is the catalytic subunit of Polycomb-repressive complex 2 (PRC2) and induces the trimethylation of the histone H3 lysine 27 (H3K27me3) promoting epigenetic gene silencing. EZH2 contains a SET domain promoting the methyltransferase activity, while the three other protein components of PRC2, namely EED, SUZ12, and RpAp46/48, induce compaction of the chromatin permitting EZH2 enzymatic activity. Numerous studies highlight the role of this evolutionary conserved protein as a master regulator of differentiation in humans involved in the repression of the homeotic gene and the inactivation of X-chromosome. Through its effects in the epigenetic regulation of critical genes, EZH2 has been strongly linked to cell cycle progression, stem cell pluripotency, and cancer biology, being currently at the cutting edge of research. Most recently, EZH2 has been associated with hematopoietic stem cell proliferation and differentiation, thymopoiesis and lymphopoiesis. Several studies have evaluated the role of EZH2 in the regulation of T cell differentiation and plasticity as well as its implications in the development of autoimmune diseases and graft-versus-host disease (GVHD). The aim of this review is to summarize the current knowledge regarding the role of EZH2 in the regulation of the differentiation and function of T cells focusing on possible applications in various immune-mediated conditions, including autoimmune disorders and GVHD. PMID:27199994

The Endocytic Adaptor Eps15 Controls Marginal Zone B Cell Numbers

PubMed Central

Pozzi, Benedetta; Amodio, Stefania; Lucano, Caterina; Sciullo, Anna; Ronzoni, Simona; Castelletti, Daniela; Adler, Thure; Treise, Irina; Betsholtz, Ingrid Holmberg; Rathkolb, Birgit; Busch, Dirk H.; Wolf, Eckhard; Fuchs, Helmut; Gailus-Durner, Valérie; de Angelis, Martin Hrabě; Betsholtz, Christer; Casola, Stefano; Di Fiore, Pier Paolo; Offenhäuser, Nina

2012-01-01

Eps15 is an endocytic adaptor protein involved in clathrin and non-clathrin mediated endocytosis. In Caenorhabditis elegans and Drosophila melanogaster lack of Eps15 leads to defects in synaptic vesicle recycling and synapse formation. We generated Eps15-KO mice to investigate its function in mammals. Eps15-KO mice are born at the expected Mendelian ratio and are fertile. Using a large-scale phenotype screen covering more than 300 parameters correlated to human disease, we found that Eps15-KO mice did not show any sign of disease or neural deficits. Instead, altered blood parameters pointed to an immunological defect. By competitive bone marrow transplantation we demonstrated that Eps15-KO hematopoietic precursor cells were more efficient than the WT counterparts in repopulating B220+ bone marrow cells, CD19− thymocytes and splenic marginal zone (MZ) B cells. Eps15-KO mice showed a 2-fold increase in MZ B cell numbers when compared with controls. Using reverse bone marrow transplantation, we found that Eps15 regulates MZ B cell numbers in a cell autonomous manner. FACS analysis showed that although MZ B cells were increased in Eps15-KO mice, transitional and pre-MZ B cell numbers were unaffected. The increase in MZ B cell numbers in Eps15 KO mice was not dependent on altered BCR signaling or Notch activity. In conclusion, in mammals, the endocytic adaptor protein Eps15 is a regulator of B-cell lymphopoiesis. PMID:23226392

Antibody repertoire development in fetal and neonatal piglets. VI. B cell lymphogenesis occurs at multiple sites with differences in the frequency of in-frame rearrangements.

PubMed

Sinkora, Marek; Sun, Jishan; Sinkorová, Jana; Christenson, Ronald K; Ford, Steven P; Butler, John E

2003-02-15

B cell lymphogenesis in mammals occurs in various tissues during development but it is generally accepted that it operates by the same mechanism in all tissues. We show that in swine, the frequency of in-frame (IF) VDJ rearrangements differs among yolk sac, fetal liver, spleen, early thymus, bone marrow, and late thymus. All VDJ rearrangements recovered and analyzed on the 20th day of gestation (DG20) from the yolk sac were 100% IF. Those recovered at DG30 in the fetal liver were >90% IF, and this predominance of cells with apparently a single IF rearrangement continued in all organs until approximately DG45, which corresponds to the time when lymphopoiesis begins in the bone marrow. Thereafter, the proportion of IF rearrangements drops to approximately 71%, i.e., the value predicted whether VDJ rearrangement is random and both chromosomes were involved. Unlike other tissues, VDJs recovered from thymus after DG50 display a pattern suggesting no selection for IF rearrangements. Regardless of differences in the proportion of IF rearrangements, we observed no significant age- or tissue-dependent changes in CDR3 diversity, N region additions, or other characteristics of fetal VDJs during ontogeny. These findings indicate there are multiple sites of B cell lymphogenesis in fetal piglets and differences in the frequency of productive VDJ rearrangements at various sites. We propose the latter to result from differential selection or a developmentally dependent change in the intrinsic mechanism of VDJ rearrangement.

Homologs of CD83 from elasmobranch and teleost fish.

PubMed

Ohta, Yuko; Landis, Eric; Boulay, Thomas; Phillips, Ruth B; Collet, Bertrand; Secombes, Chris J; Flajnik, Martin F; Hansen, John D

2004-10-01

Dendritic cells are one of the most important cell types connecting innate and adaptive immunity, but very little is known about their evolutionary origins. To begin to study dendritic cells from lower vertebrates, we isolated and characterized CD83 from the nurse shark (Ginglymostoma cirratum (Gici)) and rainbow trout (Oncorhynchus mykiss (Onmy)). The open reading frames for Gici-CD83 (194 aa) and Onmy-CD83 (218 aa) display approximately 28-32% identity to mammalian CD83 with the presence of two conserved N-linked glycosylation sites. Identical with mammalian CD83 genes, Gici-CD83 is composed of five exons including conservation of phase for the splice sites. Mammalian CD83 genes contain a split Ig superfamily V domain that represents a unique sequence feature for CD83 genes, a feature conserved in both Gici- and Onmy-CD83. Gici-CD83 and Onmy-CD83 are not linked to the MHC, an attribute shared with mouse but not human CD83. Gici-CD83 is expressed rather ubiquitously with highest levels in the epigonal tissue, a primary site for lymphopoiesis in the nurse shark, whereas Onmy-CD83 mRNA expression largely paralleled that of MHC class II but at lower levels. Finally, Onmy-CD83 gene expression is up-regulated in virus-infected trout, and the promoter is responsive to trout IFN regulatory factor-1. These results suggest that the role of CD83, an adhesion molecule for cell-mediated immunity, has been conserved over 450 million years of vertebrate evolution.

Aberrantly Expressed OTX Homeobox Genes Deregulate B-Cell Differentiation in Hodgkin Lymphoma.

PubMed

Nagel, Stefan; Ehrentraut, Stefan; Meyer, Corinna; Kaufmann, Maren; Drexler, Hans G; MacLeod, Roderick A F

2015-01-01

In Hodgkin lymphoma (HL) we recently reported that deregulated homeobox gene MSX1 mediates repression of the B-cell specific transcription factor ZHX2. In this study we investigated regulation of MSX1 in this B-cell malignancy. Accordingly, we analyzed expression and function of OTX homeobox genes which activate MSX1 transcription during embryonal development in the neural plate border region. Our data demonstrate that OTX1 and OTX2 are aberrantly expressed in both HL patients and cell lines. Moreover, both OTX loci are targeted by genomic gains in overexpressing cell lines. Comparative expression profiling and subsequent pathway modulations in HL cell lines indicated that aberrantly enhanced FGF2-signalling activates the expression of OTX2. Downstream analyses of OTX2 demonstrated transcriptional activation of genes encoding transcription factors MSX1, FOXC1 and ZHX1. Interestingly, examination of the physiological expression profile of ZHX1 in normal hematopoietic cells revealed elevated levels in T-cells and reduced expression in B-cells, indicating a discriminatory role in lymphopoiesis. Furthermore, two OTX-negative HL cell lines overexpressed ZHX1 in correlation with genomic amplification of its locus at chromosomal band 8q24, supporting the oncogenic potential of this gene in HL. Taken together, our data demonstrate that deregulated homeobox genes MSX1 and OTX2 respectively impact transcriptional inhibition of (B-cell specific) ZHX2 and activation of (T-cell specific) ZHX1. Thus, we show how reactivation of a specific embryonal gene regulatory network promotes disturbed B-cell differentiation in HL.

Declining lymphoid progenitor fitness promotes aging-associated leukemogenesis.

PubMed

Henry, Curtis J; Marusyk, Andriy; Zaberezhnyy, Vadym; Adane, Biniam; DeGregori, James

2010-12-14

Aging is associated with the functional decline of cells, tissues, and organs. At the same time, age is the single most important prognostic factor in the development of most human cancers, including chronic myelogenous and acute lymphoblastic leukemias initiated by Bcr-Abl oncogenic translocations. Prevailing paradigms attribute the association between aging and cancers to the accumulation of oncogenic mutations over time, because the accrual of oncogenic events is thought to be the rate-limiting step in initiation and progression of cancers. Conversely, aging-associated functional decline caused by both cell-autonomous and non-cell-autonomous mechanisms is likely to reduce the fitness of stem and progenitor cell populations. This reduction in fitness should be conducive for increased selection of oncogenic mutations that can at least partially alleviate fitness defects, thereby promoting the initiation of cancers. We tested this hypothesis using mouse hematopoietic models. Our studies indicate that the dramatic decline in the fitness of aged B-lymphopoiesis coincides with altered receptor-associated kinase signaling. We further show that Bcr-Abl provides a much greater competitive advantage to old B-lymphoid progenitors compared with young progenitors, coinciding with restored kinase signaling pathways, and that this enhanced competitive advantage translates into increased promotion of Bcr-Abl-driven leukemias. Moreover, impairing IL-7-mediated signaling is sufficient to promote selection for Bcr-Abl-expressing B progenitors. These studies support an unappreciated causative link between aging and cancer: increased selection of oncogenic mutations as a result of age-dependent alterations of the fitness landscape.

Role of stromal cell-derived factor 1 (SDF1/CXCL12) in regulating anterior pituitary function.

PubMed

Barbieri, Federica; Bajetto, Adriana; Porcile, Carola; Pattarozzi, Alessandra; Schettini, Gennaro; Florio, Tullio

2007-03-01

Chemokines are key factors involved in the regulation of immune response, through the activation and control of leukocyte traffic, lymphopoiesis and immune surveillance. However, a large number of chemokines and their receptors are expressed in central nervous system (CNS) cells, either constitutively or induced by inflammatory stimuli, playing a role in many neuropathological processes. Stromal cell-derived factor 1 (SDF1) is a chemokine whose extra-immunological localization and functions have been extensively studied. SDF1 and its receptor CXCR4 were identified in both neurons and glia of many brain areas, including the hypothalamus, as well as at the pituitary level. Importantly, SDF1 and CXCR4 expression is increased in brain tumors in which their activity induced tumor cell proliferation and brain parenchyma invasion. Despite their localization, to date very few reports addressed the role of CXCR4 and SDF1 in the modulation of the hypothalamus/pituitary axis and their possible involvement in the development of pituitary adenomas. In this review, we discuss previous literature data on the role of chemokines in normal and adenomatous pituitary cells, focusing on recent data from our group showing that CXCR4 activation controls proliferation and both prolactin and GH release in the pituitary adenoma cell line GH4C1 through a complex network of intracellular signals. Thus, the SDF1/CXCR4 system together with other chemokinergic ligand-receptor pairs, may represent a novel regulatory pathway for pituitary function and, possibly, be involved in pituitary adenoma development. These lines of evidence suggest that the inhibition of chemokine receptors may represent a novel pharmacological target for the treatment of pituitary adenomas.

Human common acute lymphoblastic leukemia-derived cell lines are competent to recombine their T-cell receptor delta/alpha regions along a hierarchically ordered pathway.

PubMed

Hansen-Hagge, T E; Yokota, S; Reuter, H J; Schwarz, K; Bartram, C R

1992-11-01

Rearrangements of the T-cell receptor (TCR) delta locus are observed in the majority of human B-cell precursor acute lymphoblastic leukemias (ALL) with a striking predominance of V delta 2(D)D delta 3 recombinations in common ALL (cALL) patients. Recently, we and others showed that almost 20% of cALL cases are characterized by further recombination of V delta 2(D)D delta 3 segments to J alpha elements, thereby deleting the TCR delta locus in analogy to the delta Rec/psi J alpha pathway in differentiating alpha/beta-positive T cells. We report here that two human cALL-derived cell lines, REH and Nalm-6, are competent to recombine the TCR delta/alpha locus under standard tissue culture conditions. Analysis of different REH subclones obtained by limiting dilution of the initial culture showed a biased recombination of V delta 2D delta 3 to distinct J alpha elements. During prolonged tissue culture, a subclone acquired growth advantage and displaced parental cells as well as other subclones. Frequently, the DJ junctions of REH subclones contained extended stretches of palindromic sequences derived from modified D delta 3 coding elements. The other cell line, Nalm-6, started the TCR delta/alpha recombination with an unusual signal joint of a cryptic recombinase signal sequence (RSS) upstream of D delta 3 to the 3' RSS of D delta 3. The RSS dimer was subsequently rearranged in all investigated subclones to an identical J alpha element. Both cell lines might become valuable tools to unravel the complex regulation of TCR delta/alpha recombination pathways in malignant and normal lymphopoiesis.

Maternal Serum B-Cell Activating Factor Levels: Candidate Early Biomarker for Hypertensive Disorders of Pregnancy.

PubMed

Stohl, Hindi E; Lee, Richard H; Manetta, Joseph; Kikly, Kristine; Korst, Lisa M; Stohl, William

2017-11-01

Hypertensive disorders of pregnancy are a leading cause of maternal and perinatal morbidity and mortality. Early suppression of B-cell lymphopoiesis is necessary for a normal pregnancy. Dysregulation of factors critical to B-cell survival may result in pregnancy complications, including hypertension. In this prospective observational study at a single medical center, serum levels of BAFF (B-cell activating factor) were measured in pregnant participants at each trimester, at delivery, and postpartum and in nonpregnant controls at a single time point. Comparisons were made between nonpregnant and pregnant subjects and between time periods of pregnancy. First-trimester serum BAFF levels were further tested for association with hypertensive disorders of pregnancy. The study included 149 healthy pregnant women, 25 pregnant women with chronic hypertension, and 48 nonpregnant controls. Median first-trimester serum BAFF level (ng/mL) for healthy women (0.90) was lower than median serum BAFF levels for women with chronic hypertension (0.96; P =0.013) and controls (1.00; P =0.002). Serum BAFF levels steadily declined throughout pregnancy, with the median second-trimester level lower than the corresponding first-trimester level (0.77; P =0.003) and the median third-trimester level lower than the corresponding second-trimester level (0.72; P =0.025). The median first-trimester serum BAFF level was elevated in women who subsequently developed hypertension compared with women who remained normotensive (1.02 versus 0.85; P =0.012), with the area under the receiver operating characteristic curve being 0.709. First-trimester serum BAFF level may be an early and clinically useful predictor of hypertensive disorders of pregnancy. © 2017 American Heart Association, Inc.

Organotins Are Potent Activators of PPARγ and Adipocyte Differentiation in Bone Marrow Multipotent Mesenchymal Stromal Cells

PubMed Central

Yanik, Susan C.; Baker, Amelia H.; Mann, Koren K.; Schlezinger, Jennifer J.

2011-01-01

Adipocyte differentiation in bone marrow is potentially deleterious to both bone integrity and lymphopoiesis. Here, we examine the hypothesis that organotins, common environmental contaminants that are dual ligands for peroxisome proliferator–activated receptor (PPAR) γ and its heterodimerization partner retinoid X receptor (RXR), are potent activators of bone marrow adipogenesis. A C57Bl/6-derived bone marrow multipotent mesenchymal stromal cell (MSC) line, BMS2, was treated with rosiglitazone, a PPARγ agonist, bexarotene, an RXR agonist, or a series of organotins. Rosiglitazone and bexarotene potently activated adipocyte differentiation; however, bexarotene had a maximal efficacy of only 20% of that induced by rosiglitazone. Organotins (tributyltin [TBT], triphenyltin, and dibutyltin) also stimulated adipocyte differentiation (EC50 of 10–20nM) but with submaximal, structure-dependent efficacy. In coexposures, both bexarotene and TBT enhanced rosiglitazone-induced adipogenesis. To investigate the contribution of PPARγ to TBT-induced adipogenesis, we examined expression of PPARγ2, as well as its transcriptional target FABP4. TBT-induced PPARγ2 and FABP4 protein expression with an efficacy intermediate between rosiglitazone and bexarotene, similar to lipid accumulation. A PPARγ antagonist and PPARγ-specific small hairpin RNA suppressed TBT-induced differentiation, although to a lesser extent than rosiglitazone-induced differentiation, suggesting that TBT may engage alternate pathways. TBT and bexarotene, but not rosiglitazone, also induced the expression of TGM2 (an RXR target) and ABCA1 (a liver X receptor target). The results show that an environmental contaminant, acting with the same potency as a therapeutic drug, induces PPARγ-dependent adipocyte differentiation in bone marrow MSCs. Activation of multiple nuclear receptor pathways by organotins may have significant implications for bone physiology. PMID:21622945

Intestinal lymphangiectasia and thymic hypoplasia.

PubMed Central

Sorensen, R U; Halpin, T C; Abramowsky, C R; Hornick, D L; Miller, K M; Naylor, P; Incefy, G S

1985-01-01

We have evaluated the immunological abnormalities present in a 6 year old patient with primary intestinal and generalized lymphangiectasia confirmed by intestinal, lung and lymph node biopsies. Lymphocyte loss through the gut was confirmed by the detection of lymphocytes in her stool. An increased enteric protein loss was suggested by hypoproteinaemia, peripheral oedema, and a very short half-life for i.v. immune serum globulin (3 days). Lymphocyte subpopulation analysis revealed a selective loss of T lymphocytes, with a proportionally increased loss of the OKT4 positive helper/inducer subpopulation. Functionally, there was a decrease in proliferative responses to some mitogens and to allogeneic cells, and a lack of T cell help for in vitro B lymphocyte differentiation into immunoglobulin secreting cells. Natural killer function was normal. In this patient, a concomitant thymic deficiency was documented by failure to identify thymic tissue on a thymus biopsy and by an absence or decrease of the serum thymic factor (thymulin) and thymosin alpha 1. No compensatory lymphopoiesis was detected in the bone marrow. In an attempt to increase T lymphocyte development, the patient was treated with thymosin fraction 5. Daily treatment with this preparation resulted in a transient clinical improvement which could not be sustained on a weekly thymosin treatment schedule. However, lymphocyte numbers did not increase during this treatment. The findings in this patient support the notion that T lymphocytes are needed to stimulate thymic epithelium. In situations of excessive loss of long lived T lymphocytes a secondary thymic atrophy may occur and further contribute to the development of a deficiency in cell-mediated immunity. Images Fig. 1 Fig. 2 PMID:3971596

Aberrant activity of NKL homeobox gene NKX3-2 in a T-ALL subset

PubMed Central

Meyer, Corinna; Kaufmann, Maren; Zaborski, Margarete; MacLeod, Roderick A. F.; Drexler, Hans G.

2018-01-01

T-cell acute lymphoblastic leukemia (T-ALL) is a hematopoietic malignancy originating from T-cell progenitors in which differentiation is blocked at early stages. Physiological expression of specific NKL homeobox genes obeys a hematopoietic NKL-code implicated in the process of lymphopoiesis while in differentiated T-cells these genes are silenced. We propose that this developmental expression pattern underlies the observation that NKL homeobox genes are the most ubiquitous group of transcription factors deregulated in T-ALL, including TLX1, TLX3, NKX2-5 and NKX3-1. Here, we describe a novel member of the NKL homeobox gene subclass, NKX3-2 (BAPX1), which is aberrantly activated in 18% of pediatric T-ALL patients analyzed while being normally expressed in developing spleen. Identification of NKX3-2 expression in T-ALL cell line CCRF-CEM qualified these cells to model its deregulation and function in a leukemic context. Genomic and chromosomal analyses demonstrated normal configuration of the NKX3-2 locus at chromosome 4p15, thus excluding cytogenetic dysregulation. Comparative expression profiling analysis of NKX3-2 patient data revealed deregulated activity of BMP- and MAPK-signalling. These candidate pathways were experimentally confirmed to mediate aberrant NKX3-2 expression. We also show that homeobox gene SIX6, plus MIR17HG and GATA3 are downstream targets of NKX3-2 and plausibly contribute to the pathogenesis of this malignancy by suppressing T-cell differentiation. Finally, NKL homeobox gene NKX2-5 was activated by NKX3-2 in CCRF-CEM and by FOXG1 in PEER, representing mutually inhibitory activators of this translocated oncogene. Together, our findings reveal a novel oncogenic NKL homeobox gene subclass member which is aberrantly expressed in a large subset of T-ALL patients and participates in a deregulated gene network likely to arise in developing spleen. PMID:29746601

Deregulation of polycomb repressor complex 1 modifier AUTS2 in T-cell leukemia.

PubMed

Nagel, Stefan; Pommerenke, Claudia; Meyer, Corinna; Kaufmann, Maren; Drexler, Hans G; MacLeod, Roderick A F

2016-07-19

Recently, we identified deregulated expression of the B-cell specific transcription factor MEF2C in T-cell acute lymphoid leukemia (T-ALL). Here, we performed sequence analysis of a regulatory upstream section of MEF2C in T-ALL cell lines which, however, proved devoid of mutations. Unexpectedly, we found strong conservation between the regulatory upstream region of MEF2C (located at chromosomal band 5q14) and an intergenic stretch at 7q11 located between STAG3L4 and AUTS2, covering nearly 20 kb. While the non-coding gene STAG3L4 was inconspicuously expressed, AUTS2 was aberrantly upregulated in 6% of T-ALL patients (public dataset GSE42038) and in 3/24 T-ALL cell lines, two of which represented very immature differentiation stages. AUTS2 expression was higher in normal B-cells than in T-cells, indicating lineage-specific activity in lymphopoiesis. While excluding chromosomal aberrations, examinations of AUTS2 transcriptional regulation in T-ALL cells revealed activation by IL7-IL7R-STAT5-signalling and MEF2C. AUTS2 protein has been shown to interact with polycomb repressor complex 1 subtype 5 (PRC1.5), transforming this particular complex into an activator. Accordingly, expression profiling and functional analyses demonstrated that AUTS2 activated while PCGF5 repressed transcription of NKL homeobox gene MSX1 in T-ALL cells. Forced expression and pharmacological inhibition of EZH2 in addition to H3K27me3 analysis indicated that PRC2 repressed MSX1 as well. Taken together, we found that AUTS2 and MEF2C, despite lying on different chromosomes, share strikingly similar regulatory upstream regions and aberrant expression in T-ALL subsets. Our data implicate chromatin complexes PRC1/AUTS2 and PRC2 in a gene network in T-ALL regulating early lymphoid differentiation.

The development of primary and secondary lymphoid tissues in the nurse shark Ginglymostoma cirratum: B-cell zones precede dendritic cell immigration and T-cell zone formation during ontogeny of the spleen.

PubMed

Rumfelt, L L; McKinney, E C; Taylor, E; Flajnik, M F

2002-08-01

Secondary lymphoid tissue and immunoglobulin (Ig) production in mammals is not fully developed at birth, requiring time postnatally to attain all features required for adaptive immune responses. The immune system of newborn sharks - the oldest vertebrate group having adaptive immunity - also displays immature characteristics such as low serum IgM concentration and high levels of IgM1gj, an innate-like Ig. Primary and secondary lymphoid tissues in sharks and other cartilaginous fish were identified previously, but their cellular organization was not examined in detail. In this study of nurse shark lymphoid tissue, we demonstrate that the adult spleen contains well-defined, highly vascularized white pulp (WP) areas, composed of a central T-cell zone containing a major histocompatibility complex (MHC) class II+ dendritic cell (DC) network and a small number of Ig+ secretory cells, surrounded by smaller zones of surface Ig+ (sIg+) B cells. In neonates, splenic WPs are exclusively B-cell zones containing sIgM+-MHC class IIlow B cells; thus compartmentalized areas with T cells and DCs, as well as surface Ig novel antigen receptor (sIgNAR)-expressing B cells are absent at birth. Not until the pups are 5 months old do these WP areas become adult-like; concomitantly, sIgNAR+ B cells are readily detectable, indicating that this Ig class requires a 'mature immune-responsive environment'. The epigonal organ is the major site of neonatal B lymphopoiesis, based on the presence of developing B cells and recombination-activating gene 1 (RAG1)/terminal deoxynucleotidyl transferase (TdT) expression, indicative of antigen receptor rearrangement; such expression persists into adult life, whereas the spleen has negligible lymphopoietic activity. In adults but not neonates, many secretory B cells reside in the epigonal organ, suggesting, like in mammals, that B cells home to this primary lymphoid tissue after activation in other areas of the body.

Aberrant activity of NKL homeobox gene NKX3-2 in a T-ALL subset.

PubMed

Nagel, Stefan; Meyer, Corinna; Kaufmann, Maren; Zaborski, Margarete; MacLeod, Roderick A F; Drexler, Hans G

2018-01-01

T-cell acute lymphoblastic leukemia (T-ALL) is a hematopoietic malignancy originating from T-cell progenitors in which differentiation is blocked at early stages. Physiological expression of specific NKL homeobox genes obeys a hematopoietic NKL-code implicated in the process of lymphopoiesis while in differentiated T-cells these genes are silenced. We propose that this developmental expression pattern underlies the observation that NKL homeobox genes are the most ubiquitous group of transcription factors deregulated in T-ALL, including TLX1, TLX3, NKX2-5 and NKX3-1. Here, we describe a novel member of the NKL homeobox gene subclass, NKX3-2 (BAPX1), which is aberrantly activated in 18% of pediatric T-ALL patients analyzed while being normally expressed in developing spleen. Identification of NKX3-2 expression in T-ALL cell line CCRF-CEM qualified these cells to model its deregulation and function in a leukemic context. Genomic and chromosomal analyses demonstrated normal configuration of the NKX3-2 locus at chromosome 4p15, thus excluding cytogenetic dysregulation. Comparative expression profiling analysis of NKX3-2 patient data revealed deregulated activity of BMP- and MAPK-signalling. These candidate pathways were experimentally confirmed to mediate aberrant NKX3-2 expression. We also show that homeobox gene SIX6, plus MIR17HG and GATA3 are downstream targets of NKX3-2 and plausibly contribute to the pathogenesis of this malignancy by suppressing T-cell differentiation. Finally, NKL homeobox gene NKX2-5 was activated by NKX3-2 in CCRF-CEM and by FOXG1 in PEER, representing mutually inhibitory activators of this translocated oncogene. Together, our findings reveal a novel oncogenic NKL homeobox gene subclass member which is aberrantly expressed in a large subset of T-ALL patients and participates in a deregulated gene network likely to arise in developing spleen.

HOX gene expression in phenotypic and genotypic subgroups and low HOXA gene expression as an adverse prognostic factor in pediatric ALL.

PubMed

Starkova, Julia; Zamostna, Blanka; Mejstrikova, Ester; Krejci, Roman; Drabkin, Harry A; Trka, Jan

2010-12-01

HOX genes play an important role in both normal lymphopoiesis and leukemogenesis. However, HOX expression patterns in leukemia cells compared to normal lymphoid progenitors have not been systematically studied in acute lymphoblastic leukemia (ALL) subtypes. The RNA expression levels of HOXA, HOXB, and CDX1/2 genes were analyzed by qRT-PCR in a cohort of 61 diagnostic pediatric ALL samples and FACS-sorted subpopulations of normal lymphoid progenitors. The RNA expression of HOXA7-10, HOXA13, and HOXB2-4 genes was exclusively detected in leukemic cells and immature progenitors. The RNA expression of HOXB6 and CDX2 genes was exclusively detected in leukemic cells but not in B-lineage cells at any of the studied developmental stages. HOXA3-4, HOXA7, and HOXB3-4 genes were differentially expressed between BCP-ALL and T-ALL subgroups, and among genotypically defined MLL/AF4, TEL/AML1, BCR/ABL, hyperdiploid and normal karyotype subgroups. However, this differential expression did not define specific clusters in hierarchical cluster analysis. HOXA7 gene was low expressed at the RNA level in patients with hyperdiploid leukemia, whereas HOXB7 and CDX2 genes were low expressed in TEL/AML1-positive and BCR/ABL-positive cases, respectively. In contrast to previous findings in acute myeloid leukemia, high HOXA RNA expression was associated with an excellent prognosis in Cox's regression model (P = 0.03). In MLL/AF4-positive ALL, lower HOXA RNA expression correlated with the methylation status of their promoters. HOX gene RNA expression cannot discriminate leukemia subgroups or relative maturity of leukemic cells. However, HOXA RNA expression correlates with prognosis, and particular HOX genes are expressed in specific genotypically characterized subgroups.

NKL homeobox gene activities in hematopoietic stem cells, T-cell development and T-cell leukemia.

PubMed

Nagel, Stefan; Pommerenke, Claudia; Scherr, Michaela; Meyer, Corinna; Kaufmann, Maren; Battmer, Karin; MacLeod, Roderick A F; Drexler, Hans G

2017-01-01

T-cell acute lymphoblastic leukemia (T-ALL) cells represent developmentally arrested T-cell progenitors, subsets of which aberrantly express homeobox genes of the NKL subclass, including TLX1, TLX3, NKX2-1, NKX2-5, NKX3-1 and MSX1. Here, we analyzed the transcriptional landscape of all 48 members of the NKL homeobox gene subclass in CD34+ hematopoietic stem and progenitor cells (HSPCs) and during lymphopoiesis, identifying activities of nine particular genes. Four of these were expressed in HSPCs (HHEX, HLX1, NKX2-3 and NKX3-1) and three in common lymphoid progenitors (HHEX, HLX1 and MSX1). Interestingly, our data indicated downregulation of NKL homeobox gene transcripts in late progenitors and mature T-cells, a phenomenon which might explain the oncogenic impact of this group of genes in T-ALL. Using MSX1-expressing T-ALL cell lines as models, we showed that HHEX activates while HLX1, NKX2-3 and NKX3-1 repress MSX1 transcription, demonstrating the mutual regulation and differential activities of these homeobox genes. Analysis of a public T-ALL expression profiling data set comprising 117 patient samples identified 20 aberrantly activated members of the NKL subclass, extending the number of known NKL homeobox oncogene candidates. While 7/20 genes were also active during hematopoiesis, the remaining 13 showed ectopic expression. Finally, comparative analyses of T-ALL patient and cell line profiling data of NKL-positive and NKL-negative samples indicated absence of shared target genes but instead highlighted deregulation of apoptosis as common oncogenic effect. Taken together, we present a comprehensive survey of NKL homeobox genes in early hematopoiesis, T-cell development and T-ALL, showing that these genes generate an NKL-code for the diverse stages of lymphoid development which might be fundamental for regular differentiation.

Candida albicans induces selective development of macrophages and monocyte derived dendritic cells by a TLR2 dependent signalling.

PubMed

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

2011-01-01

As TLRs are expressed by haematopoietic stem and progenitor cells (HSPCs), these receptors may play a role in haematopoiesis in response to pathogens during infection. We have previously demonstrated that in in vitro defined conditions inactivated yeasts and hyphae of Candida albicans induce HSPCs proliferation and differentiation towards the myeloid lineage by a TLR2/MyD88 dependent pathway. In this work, we showed that C. albicans invasive infection with a low virulence strain results in a rapid expansion of HSPCs (identified as LKS cells: Lin(-) c-Kit(+) Sca-1(+) IL-7Rα(-)), that reach the maximum at day 3 post-infection. This in vivo expansion of LKS cells in TLR2(-/-) mice was delayed until day 7 post- infection. Candidiasis was, as expected, accompanied by an increase in granulopoiesis and decreased lymphopoiesis in the bone marrow. These changes were more pronounced in TLR2(-/-) mice correlating with their higher fungal burden. Accordingly, emigration of Ly6C(high) monocytes and neutrophils to spleen was increased in TLR2(-/-) mice, although the increase in macrophages and inflammatory macrophages was completely dependent on TLR2. Similarly, we detected for the first time, in the spleen of C. albicans infected control mice, a newly generated population of dendritic cells that have the phenotype of monocyte derived dendritic cells (moDCs) that were not generated in TLR2(-/-) infected mice. In addition, C. albicans signalling through TLR2/MyD88 and Dectin-1 promotes in vitro the differentiation of Lin(-) cells towards moDCs that secrete TNF-α and are able to kill the microorganism. Therefore, our results indicate that during infection C. albicans can directly stimulate progenitor cells through TLR2 and Dectin-1 to generate newly formed inflammatory macrophages and moDCs that may fulfill an essential role in defense mechanisms against the pathogen.

NKL homeobox gene activities in hematopoietic stem cells, T-cell development and T-cell leukemia

PubMed Central

Pommerenke, Claudia; Scherr, Michaela; Meyer, Corinna; Kaufmann, Maren; Battmer, Karin; MacLeod, Roderick A. F.; Drexler, Hans G.

2017-01-01

T-cell acute lymphoblastic leukemia (T-ALL) cells represent developmentally arrested T-cell progenitors, subsets of which aberrantly express homeobox genes of the NKL subclass, including TLX1, TLX3, NKX2-1, NKX2-5, NKX3-1 and MSX1. Here, we analyzed the transcriptional landscape of all 48 members of the NKL homeobox gene subclass in CD34+ hematopoietic stem and progenitor cells (HSPCs) and during lymphopoiesis, identifying activities of nine particular genes. Four of these were expressed in HSPCs (HHEX, HLX1, NKX2-3 and NKX3-1) and three in common lymphoid progenitors (HHEX, HLX1 and MSX1). Interestingly, our data indicated downregulation of NKL homeobox gene transcripts in late progenitors and mature T-cells, a phenomenon which might explain the oncogenic impact of this group of genes in T-ALL. Using MSX1-expressing T-ALL cell lines as models, we showed that HHEX activates while HLX1, NKX2-3 and NKX3-1 repress MSX1 transcription, demonstrating the mutual regulation and differential activities of these homeobox genes. Analysis of a public T-ALL expression profiling data set comprising 117 patient samples identified 20 aberrantly activated members of the NKL subclass, extending the number of known NKL homeobox oncogene candidates. While 7/20 genes were also active during hematopoiesis, the remaining 13 showed ectopic expression. Finally, comparative analyses of T-ALL patient and cell line profiling data of NKL-positive and NKL-negative samples indicated absence of shared target genes but instead highlighted deregulation of apoptosis as common oncogenic effect. Taken together, we present a comprehensive survey of NKL homeobox genes in early hematopoiesis, T-cell development and T-ALL, showing that these genes generate an NKL-code for the diverse stages of lymphoid development which might be fundamental for regular differentiation. PMID:28151996

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

Bone marrow stromal-B cell interactions in polycyclic aromatic hydrocarbon-induced pro/pre-B cell apoptosis.

PubMed

Allan, Lenka L; Mann, Koren K; Matulka, Raymond A; Ryu, Heui-Young; Schlezinger, Jennifer J; Sherr, David H

2003-12-01

Environmental polycyclic aromatic hydrocarbons (PAH) and related halogenated hydrocarbons are immunotoxic in a variety of systems. In a model system of B lymphopoiesis, PAH exposure rapidly induces apoptosis in CD43- pre-B and CD43+ pro/pre-B cells. Apoptosis induction by 7,12-dimethylbenzo[a]anthracene (DMBA) is dependent upon AhR+ bone marrow stromal cells and likely involves DMBA metabolism within the stromal cell. However, it is not known if PAH-treated stromal cells release free metabolites or soluble factors that may directly induce B cell death or if the effector death signal is delivered by stromal cell-B cell contact. Here, we demonstrate that supernatants from DMBA-treated bone marrow stromal cells contain an activity capable of inducing apoptosis in pro/pre-B cells cocultured with stromal cells. This activity (1) is not produced when stromal cells are cotreated with DMBA and alpha-naphthoflavone (alpha-NF), an aryl hydrocarbon receptor (AhR) and cytochrome P-450 inhibitor, (2) is > or = 50 kDa, (3) is trypsin and heat sensitive, and (4) is dependent on AhR+ stromal cells, which in turn deliver the effector death signal to pro/pre-B cells. The results (1) argue against a role for a soluble, stromal cell-derived cytokine as the effector of PAH-induced pro/pre-B cell death, (2) exclude the possibility of a free metabolite acting directly on AhR- pro/pre-B cell targets, and (3) suggest the elaboration by stromal cells of a relatively stable, DMBA metabolite-protein complex capable of acting on other stromal cells at some distance. Collectively, these studies suggest that, while stromal cell products, e.g., metabolite-protein complexes, may affect the function of distant stromal cells, the effector death signal delivered by stromal cells to bone marrow B cells is mediated by cell-cell contact.

Identification of BSAP (Pax-5) target genes in early B-cell development by loss- and gain-of-function experiments.

PubMed Central

Nutt, S L; Morrison, A M; Dörfler, P; Rolink, A; Busslinger, M

1998-01-01

The Pax-5 gene codes for the transcription factor BSAP which is essential for the progression of adult B lymphopoiesis beyond an early progenitor (pre-BI) cell stage. Although several genes have been proposed to be regulated by BSAP, CD19 is to date the only target gene which has been genetically confirmed to depend on this transcription factor for its expression. We have now taken advantage of cultured pre-BI cells of wild-type and Pax-5 mutant bone marrow to screen a large panel of B lymphoid genes for additional BSAP target genes. Four differentially expressed genes were shown to be under the direct control of BSAP, as their expression was rapidly regulated in Pax-5-deficient pre-BI cells by a hormone-inducible BSAP-estrogen receptor fusion protein. The genes coding for the B-cell receptor component Ig-alpha (mb-1) and the transcription factors N-myc and LEF-1 are positively regulated by BSAP, while the gene coding for the cell surface protein PD-1 is efficiently repressed. Distinct regulatory mechanisms of BSAP were revealed by reconstituting Pax-5-deficient pre-BI cells with full-length BSAP or a truncated form containing only the paired domain. IL-7 signalling was able to efficiently induce the N-myc gene only in the presence of full-length BSAP, while complete restoration of CD19 synthesis was critically dependent on the BSAP protein concentration. In contrast, the expression of the mb-1 and LEF-1 genes was already reconstituted by the paired domain polypeptide lacking any transactivation function, suggesting that the DNA-binding domain of BSAP is sufficient to recruit other transcription factors to the regulatory regions of these two genes. In conclusion, these loss- and gain-of-function experiments demonstrate that BSAP regulates four newly identified target genes as a transcriptional activator, repressor or docking protein depending on the specific regulatory sequence context. PMID:9545244

B cell signatures of BCWD-resistant and susceptible lines of rainbow trout: a shift towards more EBF-expressing progenitors and fewer mature B cells in resistant animals.

PubMed

Zwollo, Patty; Ray, Jocelyn C; Sestito, Michael; Kiernan, Elizabeth; Wiens, Gregory D; Kaattari, Steve; StJacques, Brittany; Epp, Lidia

2015-01-01

Bacterial cold water disease (BCWD) is a chronic disease of rainbow trout, and is caused by the Gram-negative bacterium Flavobacterium psychrophilum (Fp), a common aquaculture pathogen. The National Center for Cool and Cold Water Aquaculture has bred two genetic lines of rainbow trout: a line of Fp-resistant trout (ARS-Fp-R or R-line trout) and a line of susceptible trout (ARS-Fp-S, or S-line). Little is known about how phenotypic selection alters immune response parameters or how such changes relate to genetic disease resistance. Herein, we quantify interindividual variation in the distribution and abundance of B cell populations (B cell signatures) and examine differences between genetic lines of naive animals. There are limited trout-specific cell surface markers currently available to resolve B cell subpopulations and thus we developed an alternative approach based on detection of differentially expressed transcription factors and intracellular cytokines. B cell signatures were compared between R-line and S-line trout by flow cytometry using antibodies against transcription factors early B cell factor-1 (EBF1) and paired domain box protein Pax5, the pro-inflammatory cytokine IL-1β, and the immunoglobulin heavy chain mu. R-line trout had higher percentages of EBF(+) B myeloid/ progenitor and pre-B cells in PBL, anterior and posterior kidney tissues compared to S-line trout. The opposite pattern was detected in more mature B cell populations: R-line trout had lower percentages of both IgM(+) mature B cells and IgM-secreting cells in anterior kidney and PBL compared to S-line trout. In vitro LPS-activation studies of PBL and spleen cell cultures revealed no significant induction differences between R-line and S-line trout. Together, our findings suggest that selective resistance to BCWD may be associated with shifts in naive animal developmental lineage commitment that result in decreased B lymphopoiesis and increased myelopoiesis in BCWD resistant trout relative

Marrow Adipose Tissue Expansion Coincides with Insulin Resistance in MAGP1-Deficient Mice

PubMed Central

Walji, Tezin A.; Turecamo, Sarah E.; Sanchez, Alejandro Coca; Anthony, Bryan A.; Abou-Ezzi, Grazia; Scheller, Erica L.; Link, Daniel C.; Mecham, Robert P.; Craft, Clarissa S.

2016-01-01

Marrow adipose tissue (MAT) is an endocrine organ with the potential to influence skeletal remodeling and hematopoiesis. Pathologic MAT expansion has been studied in the context of severe metabolic challenge, including caloric restriction, high fat diet feeding, and leptin deficiency. However, the rapid change in peripheral fat and glucose metabolism associated with these models impedes our ability to examine which metabolic parameters precede or coincide with MAT expansion. Microfibril-associated glycoprotein-1 (MAGP1) is a matricellular protein that influences cellular processes by tethering signaling molecules to extracellular matrix structures. MAGP1-deficient (Mfap2−/−) mice display a progressive excess adiposity phenotype, which precedes insulin resistance and occurs without changes in caloric intake or ambulation. Mfap2−/− mice were, therefore, used as a model to associate parameters of metabolic disease, bone remodeling, and hematopoiesis with MAT expansion. Marrow adiposity was normal in Mfap2−/− mice until 6 months of age; however, by 10 months, marrow fat volume had increased fivefold relative to wild-type control at the same age. Increased gonadal fat pad mass and hyperglycemia were detectable in Mfap2−/− mice by 2 months, but peaked by 6 months. The development of insulin resistance coincided with MAT expansion. Longitudinal characterization of bone mass demonstrated a disconnection in MAT volume and bone volume. Specifically, Mfap2−/− mice had reduced trabecular bone volume by 2 months, but this phenotype did not progress with age or MAT expansion. Interestingly, MAT expansion in the 10-month-old Mfap2−/− mice was associated with modest alterations in basal hematopoiesis, including a shift from granulopoiesis to B lymphopoiesis. Together, these findings indicate MAT expansion is coincident with insulin resistance, but not excess peripheral adiposity or hyperglycemia in Mfap2−/− mice; and substantial MAT

c-Myb Coordinates Survival and the Expression of Genes That Are Critical for the Pre-BCR Checkpoint.

PubMed

Fahl, Shawn P; Daamen, Andrea R; Crittenden, Rowena B; Bender, Timothy P

2018-05-15

The c-Myb transcription factor is required for adult hematopoiesis, yet little is known about c-Myb function during lineage-specific differentiation due to the embryonic lethality of Myb -null mutations. We previously used tissue-specific inactivation of the murine Myb locus to demonstrate that c-Myb is required for differentiation to the pro-B cell stage, survival during the pro-B cell stage, and the pro-B to pre-B cell transition during B lymphopoiesis. However, few downstream mediators of c-Myb-regulated function have been identified. We demonstrate that c-Myb regulates the intrinsic survival of CD19 + pro-B cells in the absence of IL-7 by repressing expression of the proapoptotic proteins Bmf and Bim and that levels of Bmf and Bim mRNA are further repressed by IL-7 signaling in pro-B cells. c-Myb regulates two crucial components of the IL-7 signaling pathway: the IL-7Rα-chain and the negative regulator SOCS3 in CD19 + pro-B cells. Bypassing IL-7R signaling through constitutive activation of Stat5b largely rescues survival of c-Myb-deficient pro-B cells, whereas constitutively active Akt is much less effective. However, rescue of pro-B cell survival is not sufficient to rescue proliferation of pro-B cells or the pro-B to small pre-B cell transition, and we further demonstrate that c-Myb-deficient large pre-B cells are hypoproliferative. Analysis of genes crucial for the pre-BCR checkpoint demonstrates that, in addition to IL-7Rα, the genes encoding λ5, cyclin D3, and CXCR4 are downregulated in the absence of c-Myb, and λ5 is a direct c-Myb target. Thus, c-Myb coordinates survival with the expression of genes that are required during the pre-BCR checkpoint. Copyright © 2018 by The American Association of Immunologists, Inc.

Tissue breathing and topology of rats thymocytes surface under acute total γ-irradiation.

PubMed

Nikitina, I A; Gritsuk, A I

2017-12-01

Assessment of the effect of single total γ irradiation to the parameters of mitochondrial oxidation and the topology of the thymocyte surface. The study was performed in sexually mature white outbreeding male rats divided into three groups: two experimental and one control. The states of energy metabolism were determined by the rate of oxygen consumption by the thymus tissues on endogenous substrates at the presence of 2,4 dinitrophenol, uncoupler of a tissue breathing (TB) and oxidative phosphorylation (OP) after a single total γ irradiation at a dose of 1.0 Gy at 3, 10, 40 and 60 days. The topology of thymus cells was assessed using atomic force microscopy (AFM) and scanning electron microscopy (SEM). On the 3rd and 10th days after total gamma irradiation at a dose of 1.0 Gy, a significant decrease in respira tory activity was determined in thymus tissues on endogenous substrates. Simultaneously, on the 3rd day, pro nounced changes in the morphological parameters of thymocytes (height, volume, area of contact with the sub strate) and the topology of their surface were also observed. On the 10th day after irradiation, most of the morpho logical parameters of thymocytes, except for their volume, were characterized by restoration to normal. In the long term (on the 30th and 60th days after exposure), a gradual but not complete recovery of the respiratory activity of thymocytes was observed, accompanied by an increase in the degree of dissociation of TD and OP. The obtained data reflect and refine mechanisms of post radiation repair of lymphopoiesis, showing the presence of conjugated changes in the parameters of aerobic energy metabolism of thymocytes, morphology and topology of their surface. The synchronism of changes in the parameters under study is a reflection of the state of the cytoskeleton, the functional activity of which largely depends on the level and efficiency of mitochondrial oxidation. І. A. Nikitina, A. I. Gritsuk.

Candida albicans Induces Selective Development of Macrophages and Monocyte Derived Dendritic Cells by a TLR2 Dependent Signalling

PubMed Central

Yáñez, Alberto; Megías, Javier; O'Connor, José-Enrique; Gozalbo, Daniel; Gil, M. Luisa

2011-01-01

As TLRs are expressed by haematopoietic stem and progenitor cells (HSPCs), these receptors may play a role in haematopoiesis in response to pathogens during infection. We have previously demonstrated that in in vitro defined conditions inactivated yeasts and hyphae of Candida albicans induce HSPCs proliferation and differentiation towards the myeloid lineage by a TLR2/MyD88 dependent pathway. In this work, we showed that C. albicans invasive infection with a low virulence strain results in a rapid expansion of HSPCs (identified as LKS cells: Lin− c-Kit+ Sca-1+ IL-7Rα−), that reach the maximum at day 3 post-infection. This in vivo expansion of LKS cells in TLR2−/− mice was delayed until day 7 post- infection. Candidiasis was, as expected, accompanied by an increase in granulopoiesis and decreased lymphopoiesis in the bone marrow. These changes were more pronounced in TLR2−/− mice correlating with their higher fungal burden. Accordingly, emigration of Ly6Chigh monocytes and neutrophils to spleen was increased in TLR2−/− mice, although the increase in macrophages and inflammatory macrophages was completely dependent on TLR2. Similarly, we detected for the first time, in the spleen of C. albicans infected control mice, a newly generated population of dendritic cells that have the phenotype of monocyte derived dendritic cells (moDCs) that were not generated in TLR2−/− infected mice. In addition, C. albicans signalling through TLR2/MyD88 and Dectin-1 promotes in vitro the differentiation of Lin− cells towards moDCs that secrete TNF-α and are able to kill the microorganism. Therefore, our results indicate that during infection C. albicans can directly stimulate progenitor cells through TLR2 and Dectin-1 to generate newly formed inflammatory macrophages and moDCs that may fulfill an essential role in defense mechanisms against the pathogen. PMID:21935459

Growing B Lymphocytes in a Three-Dimensional Culture System

NASA Technical Reports Server (NTRS)

Wu, J. H. David; Bottaro, Andrea

2010-01-01

within 3D cultures that have been modified to foster lymphopoiesis retain an immunophenotype that closely recapitulates cells in fresh bone marrow harvests. The 3D culture system has been found to be capable of supporting long-lived (8 weeks) populations of B and T lymphocytes from peripheral lymphoid organs, in the absence of activation signals, to an extent not achievable by conventional culture techniques. Interestingly, it has been found that 3D-culture B cells display a phenotype that has characteristics of both B1a and B2 cells. These promising preliminary observations suggest that the 3D culture system could be used with success in the study of peripheral-B-lymphocyte biology and in the development of biotechnological techniques and processes.