Distinctive Roles of Stem Cell Factor and IL-7 in T Cell Development.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3156-3156
Author(s):  
Hongfang Wang ◽  
L. Jeanne Pierce ◽  
Gerald J. Spangrude
Keyword(s):  
Stem Cell ◽  
Cell Differentiation ◽  
T Cell ◽  
Notch Signaling ◽  
Stem Cell Factor ◽  
Critical Role ◽  
T Cell Development ◽  
Cell Development ◽  
T Cell Differentiation ◽  
Lineage Differentiation

Abstract Notch signaling plays a critical role in T lineage commitment during lymphoid differentiation. However, Notch signaling alone is not sufficient to support T cell development through the CD4/CD8 double positive (DP) stage in vitro. We here report distinct effects of several cytokines on T cell differentiation in the OP9-DL1 cell culture model. Our studies show that Flt3 ligand enhances the proliferation of progenitors but has no obvious effect on differentiation. In contrast, stem cell factor (SCF) favors the proliferation of CD4/CD8 double negative (DN) lymphoid progenitors and inhibits differentiation to the DP stage in a dose-dependent manner. Differentiation of the NK lineage is promoted under these conditions. Conversely, blocking the function of SCF that is expressed endogenously by OP9-DL1 cells inhibits proliferation of lymphoid progenitors and accelerates T lineage differentiation. IL-7 is necessary for differentiation from the DP to the CD8 single positive (SP) stage, and is also required for γδ T lineage development. We also find a dosage effect of IL-7 during T cell development. OP9 and OP9-DL1 stromal cells produce endogenous levels of IL-7 that are sufficient to support B and DP T cell differentiation. However, the amount of endogenous IL-7 is not sufficient to support T cell differentiation from the DP to the SP stage. Addition of exogenous IL-7 (1–10 ng/ml) to the cultures promotes SP differentiation, while blocking endogenous IL-7 with anti-IL-7 antibody inhibits both B and T cell development. We conclude that activation through the Notch pathway is sufficient to suppress B lineage differentiation and thereby promote T lineage commitment, but is not sufficient to promote the subsequent stages of T cell development. SCF promotes expansion and directs NK lineage differentiation at the expense of T cell development, while IL-7 provides both proliferation as well as T lineage differentiation signals. T cell development from the DN to the DP stage requires a low amount of IL-7, while differentiation from the DP to the SP stage requires a higher level of IL-7. The balance between the effects mediated by these cytokines, along with Notch signaling, plays a critical role in regulating development of the T and NK lineages.

Notch Signaling in Hematopoietic Precursor Cells Maintains the Expression of Genes Required for Stem Cell Self-Renewal and Promotes the Expression of Genes Associated with T cell Differentiation.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 1699-1699
Author(s):  
Keizo Kato ◽  
Barbara Varnum-Finney ◽  
Irwin D. Bernstein
Keyword(s):  
Stem Cell ◽  
Cell Differentiation ◽  
T Cell ◽  
Notch Signaling ◽  
T Cell Differentiation ◽  
The Self ◽  
Self Renewal ◽  
Notch Ligand ◽  
Expression Of Genes ◽  
Hematopoietic Precursor

Abstract Notch Signaling in Hematopoietic Precursor Cells Maintains the Expression of Genes Required for Stem Cell Self-renewal and Promotes the Expression of Genes Associated with T cell Differentiation. Keizo Kato, Barbara Varnum-Finney, Irwin D. Bernstein Clinical Research Division, Fred Hutchinson Cancer Research Center, Seattle, WA, USA We have previously shown that Notch signaling promotes the self-renewal of hematopoietic precursors, including short-term repopulating cells, and induces early T-cell differentiation. Here we evaluate gene expression in murine Lin-Sca-1+c-kit+ Hoechst side population (SKSP) bone marrow cells during culture with the immobilized Notch ligand, Delta1ext-IgG, consisting of the extracellular domain of Delta1 fused to the Fc domain of human-IgG1 for 28 days with SCF, IL-6, IL-11 and Flt-3-ligand. We analyzed hematopoietic stem cell (HSC) associated genes, including polycomb genes, Bmi-1 and Rae28, required for HSC self-renewal, and Rex-1 required for embryonic stem cell self-renewal, together with early T-lymphoid differentiation associated genes such as GATA-3, pre-Ta and CD3e, by semi-quantitative or real-time RT-PCR. After culture for 7 or 14 days with Delta1ext-IgG, the expression of Bmi-1, Rae28 and Rex-1 was greater in cultures containing Delta1ext-IgG compared to those without. Bmi-1, Rae28 and Rex-1 were likely not direct targets of Notch signaling since the expression in SKSP cells was equivalent after 3 hours culture in the presence or absence of Notch ligand, whereas rapid up-regulation of the direct Notch target Hes1 was observed 3 hours after incubation with Delta1ext-IgG. Expression of GATA-3, pre-Ta and CD3e was induced by Notch signaling since their expression was seen by 7-14 days with but not without Delta1ext-IgG. Furthermore, the expression of these genes was dependent on Notch signaling since removal of cells from Notch ligand after culture for 28 days led to a rapid reduction of Hes1 expression within 3 hours, and a slower reduction in genes associated with self-renewal observed after 2 days. Our results suggest that Notch signaling regulates the self-renewal of hematopoietic precursors by maintaining the expression of genes known to be required for stem cell self-renewal, while also promoting the expression of T-cell differentiation-associated genes.

scholarly journals B cell adaptor for PI3-kinase (BCAP) modulates CD8+ effector and memory T cell differentiation

2018 ◽  
Vol 215 (9) ◽  
pp. 2429-2443 ◽  
Author(s):  
Mark D. Singh ◽  
Minjian Ni ◽  
Jenna M. Sullivan ◽  
Jessica A. Hamerman ◽  
Daniel J. Campbell
Keyword(s):  
T Cells ◽  
Cell Differentiation ◽  
T Cell ◽  
B Cell ◽  
Cd8 T Cells ◽  
T Cell Development ◽  
Cell Development ◽  
T Cell Differentiation ◽  
Pi3k Signaling ◽  
Memory T Cell

CD8+ T cells respond to signals via the T cell receptor (TCR), costimulatory molecules, and immunoregulatory cytokines by developing into diverse populations of effector and memory cells. The relative strength of phosphoinositide 3-kinase (PI3K) signaling early in the T cell response can dramatically influence downstream effector and memory T cell differentiation. We show that initial PI3K signaling during T cell activation results in up-regulation of the signaling scaffold B cell adaptor for PI3K (BCAP), which further potentiates PI3K signaling and promotes the accumulation of CD8+ T cells with a terminally differentiated effector phenotype. Accordingly, BCAP-deficient CD8+ T cells have attenuated clonal expansion and altered effector and memory T cell development following infection with Listeria monocytogenes. Thus, induction of BCAP serves as a positive feedback circuit to enhance PI3K signaling in activated CD8+ T cells, thereby acting as a molecular checkpoint regulating effector and memory T cell development.

Dysregulation of T-Cell Development in Adrenal Glucocorticoid-Deprived Rats

2009 ◽  
Vol 234 (9) ◽  
pp. 1067-1074 ◽  
Author(s):  
Zorica Stojić-Vukanić ◽  
Aleksandra Rauški ◽  
Duško Kosec ◽  
Katarina Radojević ◽  
Ivan Pilipović ◽  
...  
Keyword(s):  
Cell Differentiation ◽  
T Cell ◽  
T Cell Development ◽  
Cell Development ◽  
T Cell Differentiation ◽  
Male Rats ◽  
Double Positive ◽  
Cell Repertoire ◽  
Single Positive ◽  
The Impact

A number of different experimental approaches have been used to elucidate the impact of basal levels of adrenal gland-derived glucocorticoids (GCs) on T cell development, and thereby T cell-mediated immune responses. However, the relevance of the adrenal GCs to T cell development is still far from clear. This study was undertaken to explore the relevance of basal levels of GCs to T cell differentiation/maturation. Eight days post-adrenalectomy in adult male rats the thymocyte yield, apoptotic and proliferative rate and the relationship amongst major thymocyte subsets, as defined by TCRαβ/CD4/CD8 expression, were examined using flow cytometry. Adrenal GC deprivation decreased thymocyte apoptosis and altered the kinetics of T cell differentiation/maturation. In the adrenalectomized rats there was increased thymic hypercellularity and an over-representation of the CD4+CD8+ double positive (DP) TCRαβlow cells entering selection, as well as increased numbers of their DP TCRαβ− immediate precursors. These changes were accompanied with under-representation of the postselected DP TCRαβhigh and the most mature CD4−CD8+ and, particularly, CD4+CD8− single positive (SP) TCRαβhigh cells. This data suggests that withdrawal of adrenal GCs produces alterations in the thymocyte selection processes, possibly affecting the diversity of functional T cell repertoire and generation of potentially self-reactive cells as indicated by the reduced proportion and number of CD4−CD8− double negative TCRαβhigh cells. In addition, it indicates that GCs influence the post-selection maturation of thymocytes and plays a regulatory role in controlling the ratio of mature CD4+CD8−/CD4−CD8+ SP TCRαβhigh cells.

scholarly journals Artificial thymic organoids represent a reliable tool to study T-cell differentiation in patients with severe T-cell lymphopenia

2020 ◽  
Vol 4 (12) ◽  
pp. 2611-2616 ◽  
Author(s):  
Marita Bosticardo ◽  
Francesca Pala ◽  
Enrica Calzoni ◽  
Ottavia M. Delmonte ◽  
Kerry Dobbs ◽  
...  
Keyword(s):  
Cell Differentiation ◽  
T Cell ◽  
T Cell Development ◽  
Cell Development ◽  
T Cell Differentiation ◽  
Cd8 Cells ◽  
Cd34 Cells ◽  
T Cell Lymphopenia ◽  
Stromal Cell Line

Abstract The study of early T-cell development in humans is challenging because of limited availability of thymic samples and the limitations of in vitro T-cell differentiation assays. We used an artificial thymic organoid (ATO) platform generated by aggregating a DLL4-expressing stromal cell line (MS5-hDLL4) with CD34+ cells isolated from bone marrow or mobilized peripheral blood to study T-cell development from CD34+ cells of patients carrying hematopoietic intrinsic or thymic defects that cause T-cell lymphopenia. We found that AK2 deficiency is associated with decreased cell viability and an early block in T-cell development. We observed a similar defect in a patient carrying a null IL2RG mutation. In contrast, CD34+ cells from a patient carrying a missense IL2RG mutation reached full T-cell maturation, although cell numbers were significantly lower than in controls. CD34+ cells from patients carrying RAG mutations were able to differentiate to CD4+CD8+ cells, but not to CD3+TCRαβ+ cells. Finally, normal T-cell differentiation was observed in a patient with complete DiGeorge syndrome, consistent with the extra-hematopoietic nature of the defect. The ATO system may help determine whether T-cell deficiency reflects hematopoietic or thymic intrinsic abnormalities and define the exact stage at which T-cell differentiation is blocked.

An early decrease in Notch activation is required for human TCR-αβ lineage differentiation at the expense of TCR-γδ T cells

Blood ◽  
2009 ◽  
Vol 113 (13) ◽  
pp. 2988-2998 ◽  
Author(s):  
Inge Van de Walle ◽  
Greet De Smet ◽  
Magda De Smedt ◽  
Bart Vandekerckhove ◽  
Georges Leclercq ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Notch Signaling ◽  
Target Genes ◽  
T Cell Development ◽  
Γδ T Cells ◽  
Cell Development ◽  
Lineage Differentiation ◽  
Human T Cell ◽  
Notch Activation

Abstract Although well characterized in the mouse, the role of Notch signaling in the human T-cell receptor αβ (TCR-αβ) versus TCR-γδ lineage decision is still unclear. Although it is clear in the mouse that TCR-γδ development is less Notch dependent compared with TCR-αβ differentiation, retroviral overexpression studies in human have suggested an opposing role for Notch during human T-cell development. Using the OP9-coculture system, we demonstrate that changes in Notch activation are differentially required during human T-cell development. High Notch activation promotes the generation of T-lineage precursors and γδ T cells but inhibits differentiation toward the αβ lineage. Reducing the amount of Notch activation rescues αβ-lineage differentiation, also at the single-cell level. Gene expression analysis suggests that this is mediated by differential sensitivities of Notch target genes in response to changes in Notch activation. High Notch activity increases DTX1, NRARP, and RUNX3 expression, genes that are down-regulated during αβ-lineage differentiation. Furthermore, increased interleukin-7 levels cannot compensate for the Notch dependent TCR-γδ development. Our results reveal stage-dependent molecular changes in Notch signaling that are critical for normal human T-cell development and reveal fundamental molecular differences between mouse and human.

scholarly journals Analysis of Thymocyte Development Reveals That the Gtpase Rhoa Is a Positive Regulator of T Cell Receptor Responses in Vivo

2001 ◽  
Vol 194 (7) ◽  
pp. 903-914 ◽  
Author(s):  
Isabelle Corre ◽  
Manuel Gomez ◽  
Susina Vielkind ◽  
Doreen A. Cantrell
Keyword(s):  
T Cells ◽  
Cell Differentiation ◽  
T Cell ◽  
T Cell Receptor ◽  
T Cell Development ◽  
Cell Receptor ◽  
Cell Development ◽  
T Cell Differentiation ◽  
Thymocyte Development

Loss of function of the guanine nucleotide binding protein RhoA blocks pre-T cell differentiation and survival indicating that this GTPase is a critical signaling molecule during early thymocyte development. Previous work has shown that the Rho family GTPase Rac-1 can initiate changes in actin dynamics necessary and sufficient for pre-T cell development. The present data now show that Rac-1 actions in pre-T cells require Rho function but that RhoA cannot substitute for Rac-1 and induce the actin cytoskeletal changes necessary for pre-T cell development. Activation of Rho is thus not sufficient to induce pre-T cell differentiation or survival in the absence of the pre-T cell receptor (TCR). The failure of RhoA activation to impact on pre-TCR–mediated signaling was in marked contrast to its actions on T cell responses mediated by the mature TCR α/β complex. Cells expressing active RhoA were thus hyperresponsive in the context of TCR-induced proliferation in vitro and in vivo showed augmented positive selection of thymocytes expressing defined TCR complexes. This reveals that RhoA function is not only important for pre-T cells but also plays a role in determining the fate of mature T cells.

scholarly journals Distinct roles of IL-7 and stem cell factor in the OP9-DL1 T-cell differentiation culture system

2006 ◽  
Vol 34 (12) ◽  
pp. 1730-1740 ◽  
Author(s):  
Hongfang Wang ◽  
L. Jeanne Pierce ◽  
Gerald J. Spangrude
Keyword(s):  
Stem Cell ◽  
Cell Differentiation ◽  
T Cell ◽  
Stem Cell Factor ◽  
Culture System ◽  
T Cell Differentiation

Constitutive Expression of MEF2C, LYL1, or LMO2 in CD34+ HSCs Blocks the in Vitro Differentiation before the T-Cell Commitment Point and Could be Oncogenic in Early T-Cell Progenitor ALL

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 2423-2423
Author(s):  
Kirsten Canté-Barrett ◽  
Rui D Mendes ◽  
Wilco K Smits ◽  
Rob Pieters ◽  
Jules PP Meijerink
Keyword(s):  
Gene Expression ◽  
T Cells ◽  
Cell Differentiation ◽  
T Cell ◽  
T Cell Development ◽  
Cell Development ◽  
T Cell Differentiation ◽  
Cell Commitment

Abstract Background: T-cell development in the thymus is a complex process that depends on sequential transcriptional and epigenetic events that induce T-cell lineage commitment and simultaneously suppress alternative cell fates. In T-cell acute lymphoblastic leukemia (T-ALL), aberrantly expressed oncogenes result in the arrest of developing thymocytes, which can lead to the acquisition of secondary mutations, uncontrolled proliferation and disease progression. MEF2C is often expressed as a result of chromosomal rearrangements in immature, early T-cell progenitor ALL (ETP-ALL), but is also expressed in normal thymocyte progenitors before T-cell commitment (in the ETP stage). As the only hematopoietic lineage, thymocytes that have passed the T-cell commitment checkpoint (as well as mature T-cells) do no longer express MEF2C. Aims: We aimed to investigate the effect of constitutive MEF2C expression on early T-cell development. OP9-DL1 co-cultures have been most useful for mimicking in vitro T-cell development starting with hematopoietic stem cells (HSCs) derived from human cord blood or bone marrow. We also aimed to investigate the impact of MEF2C in comparison to LYL1 and LMO2; two T-ALL oncogenes also highly expressed at the ETP stage. Methods: We have utilized the OP9-DL1 in vitro co-culture system to gradually differentiate CD34+ HSCs from umbilical cord blood into the T-cell lineage. HSCs in this co-culture will recapitulate in vivo T-cell development as measured by incremental acquisition of surface markers CD7, CD5, CD1a, and reach the CD4, CD8 double-positive (DP) stage. We generated gene expression profiles of 11 subsequent in vitro stages of differentiation to help us match them to in vivo development stages. We investigated in vitro T-cell differentiation of HSCs after lentiviral transduction with MEF2C or control vectors, as well as with other transcriptional regulators LYL1 and LMO2 that are expressed at the ETP stage. Results: The major change in gene expression of subsequent early T-cell differentiation stages defines two distinct T-cell differentiation clusters that correlate with in vivo pre- and post-T-cell commitment profiles. We found that T-cell commitment occurs in CD7+ CD5+ cells before the acquisition of CD1a surface expression. Expression of control vectors in HSCs does not affect the in vitro T-cell differentiation, but MEF2C expression blocks differentiation into the direction of T-cells as measured by the failure of most cells to acquire CD7 as the first marker. Instead, with increased passage number cells gradually lose CD34 expression and eventually disappear from the co-culture. Similar effects were observed for the expression of LYL1 and LMO2; LYL1 expression arrests the cells at the most immature CD7+ ETP stage and prevents the transition towards CD7+ CD5+ cells, whereas LMO2 expressing cells reach the CD7+ CD5+ stage but fail to acquire CD1a as a marker of T-cell commitment. Summary/Conclusion: The gene expression profiles of 11 human in vitro T-cell differentiation subsets has enabled us to pinpoint T-cell commitment to a stage in which cells have acquired CD7 and CD5, just prior to the acquisition of CD1a. MEF2C, LYL1, and LMO2, expressed in ETP-ALL as well as in normal thymocyte progenitors, do not allow the transition to T-cell commitment when constitutively expressed. These proteins each result in the arrest of in vitro differentiating T-cells at different ETP stages, all before the T-cell commitment as marked by CD1a expression. Constitutive expression of MEF2C, LYL1, or LMO2 in very early thymocyte progenitors is incompatible with development into and beyond the T-cell commitment checkpoint and these proteins could therefore play important roles in the pathogenesis of ETP-ALL. Disclosures No relevant conflicts of interest to declare.

scholarly journals A role for proapoptotic Bax and Bak in T-cell differentiation and transformation

Blood ◽  
2010 ◽  
Vol 116 (24) ◽  
pp. 5237-5246 ◽  
Author(s):  
Subhrajit Biswas ◽  
Qiong Shi ◽  
Lauren Matise ◽  
Susan Cleveland ◽  
Utpal Dave ◽  
...  
Keyword(s):  
T Cells ◽  
Cell Differentiation ◽  
T Cell ◽  
Progenitor Cells ◽  
T Cell Development ◽  
B Cell Lymphoma ◽  
Cell Development ◽  
T Cell Differentiation ◽  
Intrinsic Pathway ◽  
Deficient Mice

Abstract Proapoptotic Bax and Bak are the key B-cell lymphoma-2 family members mediating apoptosis through the intrinsic pathway. Cells doubly deficient for Bax and Bak are profoundly resistant to apoptotic stimuli originating from multiple stimuli. Here we describe mice in which Bax and Bak have been deleted specifically in T-cells using Lck-Cre. In these T cell–specific BaxBak-deficient mice, early T-cell progenitors accumulate in the thymus, with relative depletion of more mature T cells. In addition, bone marrow progenitor cells fail to progress to the double positive stage when cultured on OP9 stromal cells expressing the Notch ligand Delta-like 1, consistent with a critical role for Bax and Bak in early T-cell development. Over time, T cell–specific BaxBak-deficient mice progress to an aggressive T-cell lymphoblastic leukemia/lymphoma. Interestingly, quantitative real-time polymerase chain reaction analysis of BaxBak-deficient T-cell lymphomas does not display amplification of the Notch signal transduction pathway, commonly activated in T-cell leukemia in both mouse and man. Bax and Bak, key regulators of the intrinsic pathway of apoptosis, are thus required to prevent T-cell malignancy, and for normal T-cell differentiation, regulating early T-cell development at the stage of early T-lineage progenitor cells.

scholarly journals From Murine to Human Nude/SCID: The Thymus, T-Cell Development and the Missing Link

2012 ◽  
Vol 2012 ◽  
pp. 1-12 ◽  
Author(s):  
Rosa Romano ◽  
Loredana Palamaro ◽  
Anna Fusco ◽  
Leucio Iannace ◽  
Stefano Maio ◽  
...  
Keyword(s):  
Cell Differentiation ◽  
T Cell ◽  
T Cell Development ◽  
Cell Development ◽  
Lymphoid Organ ◽  
T Cell Differentiation ◽  
Developmentally Regulated ◽  
Molecular Alterations ◽  
Stromal Component ◽  
Increased Susceptibility

Primary immunodeficiencies (PIDs) are disorders of the immune system, which lead to increased susceptibility to infections. T-cell defects, which may affect T-cell development/function, are approximately 11% of reported PIDs. The pathogenic mechanisms are related to molecular alterations not only of genes selectively expressed in hematopoietic cells but also of the stromal component of the thymus that represents the primary lymphoid organ for T-cell differentiation. With this regard, the prototype of athymic disorders due to abnormal stroma is the Nude/SCID syndrome, first described in mice in 1966. In man, the DiGeorge Syndrome (DGS) has long been considered the human prototype of a severe T-cell differentiation defect. More recently, the human equivalent of the murine Nude/SCID has been described, contributing to unravel important issues of the T-cell ontogeny in humans. Both mice and human diseases are due to alterations of the FOXN1, a developmentally regulated transcription factor selectively expressed in skin and thymic epithelia.

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