scholarly journals GATA-3 is required for early T lineage progenitor development

2009 ◽  
Vol 206 (13) ◽  
pp. 2987-3000 ◽  
Author(s):  
Tomonori Hosoya ◽  
Takashi Kuroha ◽  
Takashi Moriguchi ◽  
Dustin Cummings ◽  
Ivan Maillard ◽  
...  
Keyword(s):  
T Cell ◽  
T Lymphocytes ◽  
Cell Cycle Progression ◽  
T Cell Development ◽  
Cell Development ◽  
Hematopoietic Progenitors ◽  
Cycle Progression ◽  
Hematopoietic Stem ◽  
Autonomous Development

Most T lymphocytes appear to arise from very rare early T lineage progenitors (ETPs) in the thymus, but the transcriptional programs that specify ETP generation are not completely known. The transcription factor GATA-3 is required for the development of T lymphocytes at multiple late differentiation steps as well as for the development of thymic natural killer cells. However, a role for GATA-3 before the double-negative (DN) 3 stage of T cell development has to date been obscured both by the developmental heterogeneity of DN1 thymocytes and the paucity of ETPs. We provide multiple lines of in vivo evidence through the analysis of T cell development in Gata3 hypomorphic mutant embryos, in irradiated mice reconstituted with Gata3 mutant hematopoietic cells, and in mice conditionally ablated for the Gata3 gene to show that GATA-3 is required for ETP generation. We further show that Gata3 loss does not affect hematopoietic stem cells or multipotent hematopoietic progenitors. Finally, we demonstrate that Gata3 mutant lymphoid progenitors exhibit neither increased apoptosis nor diminished cell-cycle progression. Thus, GATA-3 is required for the cell-autonomous development of the earliest characterized thymic T cell progenitors.

scholarly journals Concurrent visualization of trafficking, expansion, and activation of T lymphocytes and T-cell precursors in vivo

Blood ◽  
2010 ◽  
Vol 116 (11) ◽  
pp. e18-e25 ◽  
Author(s):  
Il-Kang Na ◽  
John C. Markley ◽  
Jennifer J. Tsai ◽  
Nury L. Yim ◽  
Bradley J. Beattie ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Lymphocytes ◽  
T Cell Activation ◽  
T Cell Development ◽  
Cell Development ◽  
Hematopoietic Stem ◽  
Wide Range ◽  
Nfat Activation

Abstract We have developed a dual bioluminescent reporter system allowing noninvasive, concomitant imaging of T-cell trafficking, expansion, and activation of nuclear factor of activated T cells (NFAT) in vivo. NFAT activation plays an important role in T-cell activation and T-cell development. Therefore we used this system to determine spatial-temporal activation patterns of (1) proliferating T lymphocytes during graft-versus-host disease (GVHD) and (2) T-cell precursors during T-cell development after allogeneic hematopoietic stem cell transplantation (HSCT). In the first days after HSCT, donor T cells migrated to the peripheral lymph nodes and the intestines, whereas the NFAT activation was dominant in the intestines, suggesting an important role for the intestines in the early stages of alloactivation during development of GVHD. After adoptive transfer of in vitro-derived T-cell receptor (TCR) H-Y transgenic T-cell precursors into B6 (H-2b) hosts of both sexes, NFAT signaling and development into CD4+ or CD8+ single-positive cells could only be detected in the thymus of female recipients indicating either absence of positive selection or prompt depletion of double-positive thymocytes in the male recipients. Because NFAT plays an important role in a wide range of cell types, our system could provide new insights into a variety of biologic processes.

scholarly journals CBFB-MYH11 hinders early T-cell development and induces massive cell death in the thymus

Blood ◽  
2006 ◽  
Vol 109 (8) ◽  
pp. 3432-3440 ◽  
Author(s):  
Ling Zhao ◽  
Jennifer L. Cannons ◽  
Stacie Anderson ◽  
Martha Kirby ◽  
Liping Xu ◽  
...  
Keyword(s):  
T Cell ◽  
Fusion Gene ◽  
T Cell Development ◽  
Cell Development ◽  
Compound Heterozygous ◽  
Thymocyte Development ◽  
Lymphoid Leukemia ◽  
Hematopoietic Stem ◽  
Fold Reduction

Abstract Recent studies suggest that the chromosome 16 inversion, associated with acute myeloid leukemia M4Eo, takes place in hematopoietic stem cells. If this is the case, it is of interest to know the effects of the resulting fusion gene, CBFB-MYH11, on other lineages. Here we studied T-cell development in mice expressing Cbfb-MYH11 and compared them with mice compound-heterozygous for a Cbfb null and a hypomorphic GFP knock-in allele (Cbfb−/GFP), which had severe Cbfb deficiency. We found a differentiation block at the DN1 stage of thymocyte development in Cbfb-MYH11 knock-in chimeras. In a conditional knock-in model in which Cbfb-MYH11 expression was activated by Lck-Cre, there was a 10-fold reduction in thymocyte numbers in adult thymus, resulting mainly from impaired survival of CD4+CD8+ thymocytes. Although Cbfb-MYH11 derepressed CD4 expression efficiently in reporter assays, such derepression was less pronounced in vivo. On the other hand, CD4 expression was derepressed and thymocyte development was blocked at DN1 and DN2 stages in E17.5 Cbfb−/GFP thymus, with a 20-fold reduction of total thymocyte numbers. Our data suggest that Cbfb-MYH11 suppressed Cbfb in several stages of T-cell development and provide a mechanism for CBFB-MYH11 association with myeloid but not lymphoid leukemia.

T-Cell Development Failure At β-Selection Checkpoint and TCRα/δ Locus Break Formation Associated with Chromosome 14 Translocation in Ataxia-Telangiectagia Mutated Deficient Mice

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 184-184
Author(s):  
Takeshi Isoda ◽  
Masatoshi Takagi ◽  
Jinhua Piao ◽  
Shun Nakagama ◽  
Masaki Sato ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
T Cell Development ◽  
Cell Development ◽  
Chromosome Break ◽  
Wild Type ◽  
Chromosome 14 ◽  
Hematopoietic Stem ◽  
T Cell Lymphopenia

Abstract Abstract 184FN2 Ataxia Telangiectagia (AT) is an autosomal recessive immunodeficiency, caused by mutation of ataxia telangiectagia mutated gene (ATM). ATM plays a crucial role for responding to DNA damages by extrinsic and intrinsic factors, and is a master regulator for maintaining DNA integrity. VDJ recombination and class switch recombination during lymphocyte maturation are the steps of intrinsic DNA damage response where ATM stabilizes DNA ends during recombination. ATM deficiency (ATM−/−) is known to predispose to T-cell lymphopenia and T-lineage lymphoma development. ATM−/− mouse has been shown to have a failure of T-cell development at the stage from double positive (DP) to single positive (SP) differentiation, which is due to a failure of T-cell receptor a (TCRa) recombination. Thymic lymphomas in ATM−/− mice have recently been shown to have a chromosome 14 translocation involving TCRd locus, suggesting that the first event for translocation arises during TCRd locus recombination at double negative (DN) stage. However, phenotypic features of T-cell development at DN phase and the timing of chromosome 14 translocation formation in ATM−/− are not fully elucidated. Here we demonstrate that T cells of ATM−/− mice show a failure at the transition from DN3a to DN3b at b and gd-selection checkpoints due to multiple TCR recombination failure in-vivo. Consistent with in-vivo developmental profiles of ATM−/− mice thymocytes, long term hematopoietic stem cells (LTR-HSCs) of ATM−/− mice cultured with OP9-DLL1 show a delay at b-selection checkpoint in chronological order. In this culture system, failures in gd-T-cell development are also observed in ATM−/− LTR-HSCs. Involvement of thymic stromas in the failure of this transition was ruled out by bone-marrow transplantation (BMT) of ATM−/− donor to WT recipient mice, where thymocytes reconstitution showed the same transition failure at b-selection checkpoint. Thymocytes in RAG2−/− mice are arrested at DN3 stage by a failure of cleavage of TCR genes, but the arrested thymocytes are known to progress to DP phase by anti-CD3e antibody stimulation. This experiment enables to analyze pre-TCR dependent differentiation signal machinery. Then anti-CD3e antibody was injected into RAG2−/−ATM−/− mouse and DN3 cells were shown to be led to DP phase, indicating that ATM itself is not involved in the differentiation program during DN to DP phase. These results suggested loss of ATM attenuates T cell differentiation at DN3a to DN3b transition due to inefficient TCRg, d and b locus recombination. Thus differentiation failure from DN3a to DN3b in ATM deficiency is presumably the primary cause of T cell lymphopenia at the stage prior to positive-selection. We next investigated when of the differentiation stages chromosome 14 translocation involving TCRa/d locus monitored. When the LTR-HSCs is cultured on the OP9-DLL1 cells with high-dose cytokine including 10 ng/ml of Flt3-L, IL-7 and SCF, differentiation of LTR-HSCs to T cells halt at DN2-3a phase before b-selection. Then, by reducing the Flt3-L and IL7 to 5 ng/ml and 1 ng/ml, respectively, the differentiation arrest is released and Tcell differentiation progresses from DN3a to DN3b. No detectable chromosome break at TCRad locus was observed at DN2-3a in wild type, while 5% of ATM−/− cells carried TCRad break, associated with chromosome 14 translocation in approximately 0.8 % of DN2-3a cells. After progression to DN3b-4 phase, TCRad locus break was still observed in AT cells at the frequency of 1%, and chromosome 14 translocations involving TCRad locus was observed in 12% of ATM−/− cells, which was in contrast to none in wild type cell. Mono- or bi-allelic TCRa/d breaks, chromosome 14 dicentric, and t (12:14) were also observed in minor population of ATM−/− cells. These results suggest that critical point for generation of chromosome 14 translocations involving TCRa/d locus lies at DN2-3a to 3b stages corresponding during b and gd selection checkpoint in ATM deficient thymocytes. Our findings revealed that developmental failure of T-cells in AT arises during b and gd–selection checkpoint, which leads to the breaks of TCRa/d locus and subsequent chromosome 14 translocation formation. Thus we propose T-lymphopenia and predisposition to T cell leukemia/lymphoma are tightly connected in ATM deficient condition. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Delta-like 4 is indispensable in thymic environment specific for T cell development

2008 ◽  
Vol 205 (11) ◽  
pp. 2507-2513 ◽  
Author(s):  
Katsuto Hozumi ◽  
Carolina Mailhos ◽  
Naoko Negishi ◽  
Ken-ichi Hirano ◽  
Takashi Yahata ◽  
...  
Keyword(s):  
T Cell ◽  
Notch Signaling ◽  
Cell Fate ◽  
T Cell Development ◽  
Cell Development ◽  
Thymic Epithelial Cells ◽  
Hematopoietic Progenitors ◽  
Double Positive

The thymic microenvironment is required for T cell development in vivo. However, in vitro studies have shown that when hematopoietic progenitors acquire Notch signaling via Delta-like (Dll)1 or Dll4, they differentiate into the T cell lineage in the absence of a thymic microenvironment. It is not clear, however, whether the thymus supports T cell development specifically by providing Notch signaling. To address this issue, we generated mice with a loxP-flanked allele of Dll4 and induced gene deletion specifically in thymic epithelial cells (TECs). In the thymus of mutant mice, the expression of Dll4 was abrogated on the epithelium, and the proportion of hematopoietic cells bearing the intracellular fragment of Notch1 (ICN1) was markedly decreased. Corresponding to this, CD4 CD8 double-positive or single-positive T cells were not detected in the thymus. Further analysis showed that the double-negative cell fraction was lacking T cell progenitors. The enforced expression of ICN1 in hematopoietic progenitors restored thymic T cell differentiation, even when the TECs were deficient in Dll4. These results indicate that the thymus-specific environment for determining T cell fate indispensably requires Dll4 expression to induce Notch signaling in the thymic immigrant cells.

scholarly journals Timely Controlled T Cell Receptor Expression of Genetically Engineered Precursor T Cells Requires Early Transgene Induction for Leukemia Control after Hematopoietic Stem Cell Transplantation

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 657-657
Author(s):  
Sayed Shahabuddin Hoseini ◽  
Martin Hapke ◽  
Jessica Herbst ◽  
Dirk Wedekind ◽  
Rolf Baumann ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Negative Selection ◽  
T Cell Development ◽  
Leukemia Cell ◽  
Cell Development ◽  
Receptor Expression ◽  
Hematopoietic Stem

Abstract BACKGROUND: The co-transplantation of hematopoietic stem cells (HS) with those that have been engineered to express tumor-reactive T cell receptors (TCRs) and differentiated into precursor T cells (preTs) may optimize tumor reduction. Since expression of potentially self-(tumor-) reactive TCRs will lead to negative selection upon thymic maturation, we investigated whether preTs forced to express a leukemia-reactive TCR under the control of a tetracycline-inducible promoter would allow timely controlled TCR expression thereby avoiding thymic negative selection. METHODS: Using lentiviral vectors, murine LSK cells were engineered to express a Tetracycline-inducible TCR directed against a surrogate leukemia antigen. TCR-transduced LSK cells were co-cultured on T cell development-supporting OP9-DL1 cells to produce preTs. Lethally-irradiated B6/NCrl recipients received syngeneic T cell-depleted bone marrow and 8 × 106 syngeneic or allogeneic (B10.A) TCR-engineered preTs. An otherwise lethal leukemia cell (C1498) challenge was given 28 days later. RESULTS: After in vivo maturation and gene induction up to 70% leukemia free survival was achieved in recipients of syngeneic TCR-transduced preTs (p<0.001) as shown in figure 1A. Importantly, transfer of allogeneic gene-manipulated preTs increased the survival of recipients (p<0.05) without inducing graft versus host disease (GVHD). Non-transduced preTs provided significantly lower leukemia protection being not significantly superior to the PBS controls. The progenies of engineered preTs gave rise to effector and central memory cells providing protection even after repeated leukemia challenge (Figure 1B and 1C). In vitro transduction and consecutive expansion of mature T cells required at least 40 × 106 cells/recipient to mediate similar anti-leukemia efficacy, risking the development of severe GVHD if of mismatched origin, and providing no long-term protection. Importantly, while transgene induction starting immediately after transplant forced CD8+ T cell development and was required to obtain a mature T cell subset of targeted specificity, late induction favored CD4 differentiation and failed to produce a leukemia-reactive population due to missing thymic positive selection. CONCLUSION: Co-transplanting TCR gene-engineered preTs is of high clinical relevance since small numbers of even mismatched HS can be transduced at a reasonable cost, expanded in vitro, stored if needed, and provide potent and long lasting leukemia protection. Figure 1 Figure 1. Co-transplantation of engineered preTs provides potent long-lasting anti-leukemia effects upon TCR-induction in vivo. (A) Lethally-irradiated B6 mice received syngeneic TCDBM cells and either non-transduced or TCR gene-transduced preTs. Doxycycline was given starting the day of transplantation. One month later, 1.2 x 106 C1498-OVA leukemia cells were injected via tail vein. Controls did not receive preTs. n = 10 to 15 per group. (B) Surviving mice of the co-transplantation experiments were re-challenged with C1498-OVA leukemia three months after the first challenge. Age matched non-transplanted mice were used as controls. Pooled data of two independent transplantations (n = 10) are shown. (C) 95 days after the second challenge, spleens of surviving animals were harvested (n = 4) and analyzed for the expression of T cell memory markers on the progenies of co-transplanted preTs. Disclosures No relevant conflicts of interest to declare.

Notch Repression by LRF Is Necessary for the Maintenance of Adult Hematopoietic Stem Cell Pool.

Blood ◽  
2010 ◽  
Vol 116 (21) ◽  
pp. 2633-2633
Author(s):  
Sung-UK Lee ◽  
Min Li ◽  
Manami Maeda ◽  
Nagisa Sakurai ◽  
Yuichi Ishikawa ◽  
...  
Keyword(s):  
Stem Cells ◽  
Bone Marrow ◽  
Stem Cell ◽  
T Cell ◽  
T Cell Development ◽  
Cell Development ◽  
Self Renewal ◽  
Hematopoietic Stem ◽  
Deficient Mice

Abstract Abstract 2633 Among the different stem cells, hematopoietic stem cells (HSCs) are one of the best studied and characterized stem cells. To maintain life-long hematopoiesis in the bone marrow (BM), signals governing the balance between self-renewal and differentiation are tightly regulated in HSC compartment. Notch signals are critical regulators of the lymphoid lineage fate, but their role in adult HSC function in the BM is currently under debate. LRF (Leukemia/Lymphoma Related Factor, also known as Zbtb7a/pokemon) is a transcription factor that acts as a proto-oncogene and plays a key role in lymphoid and erythroid development. Previously we reported that the pool of LT-HSCs, CD150+CD48−Flt3−Vcam-1+/&minus;IL7Rα−LSK (Lin−Sca-1+c-Kit+), was significantly reduced, while lymphoid-biased multi-potential progenitors (LMPPs: CD150−CD48+Flt3+Vcam-1+/&minus;IL7Rα−LSK) and common lymphoid progenitors (CLPs: Lin−CD150−CD48+Flt3+Vcam-1−IL7Rα+) were barely detectable in LRF deficient mice. This was due to excessive differentiation of HSC into aberrant CD4/CD8 DP (double positive) T cell development in the BM caused by high Notch activity, implicating LRF role on HSC maintenance. Both gene expression profile (GSEA and DAVID analysis) and Q-PCR results indicated that LRF deficient LT-HSCs had loss of stem cell signature; but gain of T cell signature and up-regulated Notch-target gene, Hes-1, without affecting mRNA expression of Notch (1-4) or related (DLL1, DLL4, Jagged-1) genes. To determine LRF function in HSCs, we performed in vivo and in vitro experiments: 1) 5-FU (5-fluorouracil, the chemotherapy agent) treated LRF deficient mice were not able to compensate for their loss of LT-HSCs; 2) multi-lineage defects were shown in second recipient mice transplanted with 1 million of LRF deficient bone marrow cells in serial bone marrow transplantation assays, suggesting that LRF deficient LT-HSCs had defect in self-renewal and 3) LRF deficient FL-HSCs (CD150+CD48−LSK cells) were cultured on OP9 cells expressing delta-like ligand (DLL1, DLL4 and Jagged1), and enhanced T cell differentiation was only observed when they were co-cultured with delta-expressing OP9 cells. Among the Notch family, these phenotypes were Notch1-dependent. In fact, Notch1flox/floxLRFflox/floxMx1-Cre+ mice demonstrated normal LT-HSC numbers and restored B cell development, and prolonged survival over LRFflox/floxMx1-Cre+ mice in sequential 5-FU treatment in vivo. To explore which Notch-ligand(s) in BM niche is responsible for aberrant T-cell development in LRF deficient mice as well, we treated wild-type and LRFflox/floxMx1-Cre+ with anti-DLL4 antibody twice per week for 3 weeks. DLL4 blockage in LRF deficient mice rescued B cell development and prevented the development of aberrant DP T-cell development in LRF deficient mice. To further elucidate the relationship between LRF and Notch in adult HSC function, we analyzed Notch protein expression levels in HSCs and performed in-depth analysis of HSC/progenitor (HSC, LMPP and CLP) compartments in wild-type and LRF knockout (KO). Interestingly, Notch1 proteins were differentially expressed in LT-HSCs and ~50 % of them were positive for Notch1, while Notch2 was abundantly expressed in LT-HSCs. Notch1 expressing LT-HSCs were in more active cell-cycle (S phase) and absent in LRF conditional knockout mice. It is most likely that Notch1 expressing LT-HSCs were continually differentiating toward T cells in the absence of LRF, as CD4+CD8+ T cells were evident in the BM 10 months after pIpC injection. Taken together, our data strongly indicate that LRF is indispensable for hematopoietic homeostasis by preventing the lymphoid-primed HSCs from Notch/Delta-mediated T-instructive signal in the BM niche. Currently we're investigating the functional significances of Notch1 expressing LT-HSCs in detail. Our studies help us to better understand the underlying mechanism for HSC fate decision (self-renewal v.s. differentiation) in stem cell biology and its therapeutic approach in regenerative medicine. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Identification of Two New HLA-A*1101-Restricted Tax Epitopes Recognized by Cytotoxic T Lymphocytes in an Adult T-Cell Leukemia Patient after Hematopoietic Stem Cell Transplantation

2005 ◽  
Vol 79 (15) ◽  
pp. 10088-10092 ◽  
Author(s):  
Nanae Harashima ◽  
Ryuji Tanosaki ◽  
Yukiko Shimizu ◽  
Kiyoshi Kurihara ◽  
Takao Masuda ◽  
...  
Keyword(s):  
Stem Cell ◽  
T Cell ◽  
T Lymphocytes ◽  
Hematopoietic Stem Cell Transplantation ◽  
Stem Cell Transplantation ◽  
Cytotoxic T Lymphocytes ◽  
Cell Leukemia ◽  
Hematopoietic Stem ◽  
Adult T Cell Leukemia

ABSTRACT We previously reported that Tax-specific CD8+ cytotoxic T lymphocytes (CTLs), directed to single epitopes restricted by HLA-A2 or A24, expanded in vitro and in vivo in peripheral blood mononuclear cells (PBMC) from some adult T-cell leukemia (ATL) patients after but not before allogeneic hematopoietic stem cell transplantation (HSCT). Here, we demonstrated similar Tax-specific CTL expansion in PBMC from another post-HSCT ATL patient without HLA-A2 or A24, whose CTLs equally recognized two newly identified epitopes, Tax88-96 and Tax272-280, restricted by HLA-A11, suggesting that these immunodominant Tax epitopes are present in the ATL patient in vivo.

3040 – IN VITRO RECAPITULATION OF MURINE T CELL DEVELOPMENT FROM SINGLE HEMATOPOIETIC STEM CELLS

2020 ◽  
Vol 88 ◽  
pp. S51
Author(s):  
Victoria Sun ◽  
Amelie Montel-Hagen ◽  
David Casero ◽  
Steven Tsai ◽  
Alexandre Zampieri ◽  
...  
Keyword(s):  
Stem Cells ◽  
T Cell ◽  
Hematopoietic Stem Cells ◽  
T Cell Development ◽  
Cell Development ◽  
Hematopoietic Stem

T-Cell Development from Hematopoietic Stem Cells

1998 ◽  
pp. 305-336 ◽  
Author(s):  
Koichi Akashi ◽  
Motonari Kondo ◽  
Annette M. Schlageter ◽  
Irving L. Weissman
Keyword(s):  
Stem Cells ◽  
T Cell ◽  
Hematopoietic Stem Cells ◽  
T Cell Development ◽  
Cell Development ◽  
Hematopoietic Stem

Antigen-presenting cells and T-lymphocytes homing to the thymus shape T cell development

2018 ◽  
Vol 204 ◽  
pp. 9-15 ◽  
Author(s):  
Jeremy Santamaria ◽  
Julie Darrigues ◽  
Joost P.M. van Meerwijk ◽  
Paola Romagnoli
Keyword(s):  
T Cell ◽  
T Lymphocytes ◽  
T Cell Development ◽  
Antigen Presenting Cells ◽  
Cell Development ◽  
Antigen Presenting
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