scholarly journals Normal T-Cell Development and Immune Functions in TRIM-Deficient Mice

2006 ◽  
Vol 26 (9) ◽  
pp. 3639-3648 ◽  
Author(s):  
Uwe Kölsch ◽  
Börge Arndt ◽  
Dirk Reinhold ◽  
Jonathan A. Lindquist ◽  
Nicole Jüling ◽  
...  
Keyword(s):  
Immune Response ◽  
T Cells ◽  
T Cell ◽  
T Cell Development ◽  
Cell Development ◽  
T Cell Subsets ◽  
Immune Functions ◽  
Deficient Mice

ABSTRACT The transmembrane adaptor molecule TRIM is strongly expressed within thymus and in peripheral CD4+ T cells. Previous studies suggested that TRIM is an integral component of the T-cell receptor (TCR)/CD3 complex and might be involved in regulating TCR cycling. To elucidate the in vivo function of TRIM, we generated TRIM-deficient mice by homologous recombination. TRIM−/− mice develop normally and are healthy and fertile. However, the animals show a mild reduction in body weight that appears to be due to a decrease in the size and/or cellularity of many organs. The morphology and anatomy of nonlymphoid as well as primary and secondary lymphoid organs is normal. The frequency of thymocyte and peripheral T-cell subsets does not differ from control littermates. In addition, a detailed analysis of lymphocyte development revealed that TRIM is not required for either positive or negative selection. Although TRIM−/− CD4+ T cells showed an augmented phosphorylation of the serine/threonine kinase Akt, the in vitro characterization of peripheral T cells indicated that proliferation, survival, activation-induced cell death, migration, adhesion, TCR internalization and recycling, TCR-mediated calcium fluxes, tyrosine phosphorylation, and mitogen-activated protein family kinase activation are not affected in the absence of TRIM. Similarly, the in vivo immune response to T-dependent and T-independent antigens as well as the clinical course of experimental autoimmune encephalomyelitis, a complex Th1-mediated autoimmune model, is comparable to that of wild-type animals. Collectively, these results demonstrate that TRIM is dispensable for T-cell development and peripheral immune functions. The lack of an evident phenotype could indicate that TRIM shares redundant functions with other transmembrane adaptors involved in regulating the immune response.

Differential use of FasL- and perforin-mediated cytolytic mechanisms by T-cell subsets involved in graft-versus-myeloid leukemia responses

Blood ◽  
2000 ◽  
Vol 96 (3) ◽  
pp. 1047-1055 ◽  
Author(s):  
Michael H. Hsieh ◽  
Robert Korngold
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Myeloid Leukemia ◽  
T Cell Subsets ◽  
Leukemia Cell Line ◽  
Deficient Mice

In graft-versus-leukemia (GVL) responses, the cellular subsets and effector mechanisms responsible for cytotoxicity against leukemic cells in vivo remain poorly characterized. A murine model of syngeneic GVL that features CD4+ and CD8+T-cell responses against the MMB3.19 myeloid leukemia cell line has been previously described. MMB3.19 expresses high levels of functional Fas and tumor necrosis factor (TNF) receptors that do not transduce proapoptotic signals. Through the use of perforin- and Fas ligand (FasL)-deficient mice, it was demonstrated that CD4+ T cells mediate anti-MMB3.19 effects in vivo primarily through the use of FasL and secondarily through perforin mechanisms. Conversely, CD8+ T cells induce GVL effects primarily through the use of perforin and minimally through FasL mechanisms. Although the in vivo observations of CD8+ T cells were reflective of their in vitro cytotoxic T lymphocyte (CTL) activity, for CD4+ T cells, in vitro responses were dominated by the perforin pathway. In addition, the diminished capacity of T cells from perforin- and FasL-deficient mice to lyse MMB3.19 target cells appeared directly related to their deficient cytotoxic functions rather than to defects in activation because these cells were fully capable of mounting proliferative responses to the tumor cells. These findings demonstrate that GVL responses of T-cell subsets can involve preferential use of different cytotoxic mechanisms. In particular, these findings identify a role for both FasL-employing CD4+CTLs and the more novel perforin-utilizing CD4+ T-cell subset in responses against a myeloid leukemia.

Differential use of FasL- and perforin-mediated cytolytic mechanisms by T-cell subsets involved in graft-versus-myeloid leukemia responses

Blood ◽  
2000 ◽  
Vol 96 (3) ◽  
pp. 1047-1055 ◽  
Author(s):  
Michael H. Hsieh ◽  
Robert Korngold
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Cd4 T Cells ◽  
Myeloid Leukemia ◽  
T Cell Subsets ◽  
Leukemia Cell Line ◽  
Deficient Mice

Abstract In graft-versus-leukemia (GVL) responses, the cellular subsets and effector mechanisms responsible for cytotoxicity against leukemic cells in vivo remain poorly characterized. A murine model of syngeneic GVL that features CD4+ and CD8+T-cell responses against the MMB3.19 myeloid leukemia cell line has been previously described. MMB3.19 expresses high levels of functional Fas and tumor necrosis factor (TNF) receptors that do not transduce proapoptotic signals. Through the use of perforin- and Fas ligand (FasL)-deficient mice, it was demonstrated that CD4+ T cells mediate anti-MMB3.19 effects in vivo primarily through the use of FasL and secondarily through perforin mechanisms. Conversely, CD8+ T cells induce GVL effects primarily through the use of perforin and minimally through FasL mechanisms. Although the in vivo observations of CD8+ T cells were reflective of their in vitro cytotoxic T lymphocyte (CTL) activity, for CD4+ T cells, in vitro responses were dominated by the perforin pathway. In addition, the diminished capacity of T cells from perforin- and FasL-deficient mice to lyse MMB3.19 target cells appeared directly related to their deficient cytotoxic functions rather than to defects in activation because these cells were fully capable of mounting proliferative responses to the tumor cells. These findings demonstrate that GVL responses of T-cell subsets can involve preferential use of different cytotoxic mechanisms. In particular, these findings identify a role for both FasL-employing CD4+CTLs and the more novel perforin-utilizing CD4+ T-cell subset in responses against a myeloid leukemia.

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.

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.

Kras Is Critical for CD8 T Cell Antiviral Function

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 284-284
Author(s):  
Yuhong Chen ◽  
Yongwei Zheng ◽  
Xiaona You ◽  
Gang Xin ◽  
Mei Yu ◽  
...  
Keyword(s):  
Immune Response ◽  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
T Cell Development ◽  
Cd8 T Cell ◽  
Cell Immune Response ◽  
Deficient Mice ◽  
Lcmv Infection

Abstract Small GTPases regulate multiple signaling pathways and individual Ras member can have distinct biological function. To overcome embryonic lethality of Kras-deficient mice, we generated and examined mice with hematopoietic- and T cell-specific deletion of Kras. In VavCreKrasfl/fl mice with hematopoietic deletion of Kras, thymic T-cell development was normal based on the presence of normal populations of total, CD4- CD8-, CD4+ CD8+, CD4+ and CD8+ thymocytes. The populations of splenic CD4+ and CD8+ T cells were also comparable between VavCreKrasfl/fl relative to control mice. In addition, no consistent defects in the 3 H-thymidine incorporation rate of Kras-deficient splenic CD4+ or CD8+ T cells in response to anti-CD3 or anti-CD3 plus IL-2 was detected. Nonetheless, we studied the effect of Kras deficiency on CD8 T-cell immune response to acute infection of the Armstrong strain of lymphocytic choriomeningitis virus (LCMV). Sub-lethally irradiated Rag1-deficient mice transplanted with bone marrow (BM) cells from VavCreKrasfl/fl or control mice were subjected to LCMV infection. Infection-induced expansion of CD8 T cells and generation of LCMV epitope gp33-specific CD8 T cells were markedly reduced in the recipients that received the BM from VavCreKrasfl/fl relative to control mice. Following in vitro stimulation with the LCMV epitope gp33, the induction of IFNg-expressing CD8 T cells from LCMV-infected recipients that received the BM from VavCreKrasfl/fl mice was dramatically reduced. Further, BM chimeric mice with CD8 T cell-specific deficiency generated by transplantation of lethally irradiated CD8 T cell-depleted CD45.1 congenic mice with a mixture of BM cells from VavCreKrasfl/fl mice and BM cells from CD8 T cell-deficient mice exhibited an impaired CD8 T-cell immune response to LCMV infection. Lastly, we examined the role of Kras in TCR signaling. The level of total TCR-activated Ras (Ras-GTP) was markedly reduced in Kras-deficient relative to control CD8 T cells. Importantly, TCR-induced ERK1/2 activation was impaired in Kras-deficient relative to control CD8 T cells. Consistently, TCR-induced activation of Raf-1 and MEK1/2 was markedly reduced in mutant CD8 T cells. However, TCR-induced JNK and p38 activation as well as Ca2+ flux were normal in Kras-deficient CD8 T cells. Of note, TCR-induced activation of Ca2+ flux, JNK and p38 as well as ERK1/2, MEK1/2 and Raf1 was normal in Kras-deficient relative to control CD4 cells. Taken together, these data demonstrate that Kras is dispensable for T cell development or TCR-induced proliferation of CD4 or CD8 T cells in vitro, but regulates TCR-induced activation of the Raf-1/MEK/ERK pathway in CD8 but not CD4 T cells and intrinsically controls CD8 T-cell immune response to viral infection. Disclosures No relevant conflicts of interest to declare.

696 Brentuximab vedotin, a CD30-directed antibody-drug conjugate, selectively depletes activated Tregs in vitro and in vivo

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A738-A738
Author(s):  
Bryan Grogan ◽  
Reice James ◽  
Michelle Ulrich ◽  
Shyra Gardai ◽  
Ryan Heiser ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Efflux Pump ◽  
Apoptotic Cell ◽  
T Cell Subsets ◽  
Memory Cells ◽  
Antibody Drug Conjugate ◽  
Selective Depletion

BackgroundRegulatory T cells (Tregs) play an important role in maintaining immune homeostasis, preventing excessive inflammation in normal tissues. In cancer, Tregs hamper anti-tumor immunosurveillance and facilitate immune evasion. Selective targeting of intratumoral Tregs is a potentially promising treatment approach. Orthogonal evaluation of tumor-infiltrating lymphocytes (TILs) in solid tumors in mice and humans have identified CCR8, and several tumor necrosis family receptors (TNFRs), including TNFSFR8 (CD30), as receptors differentially upregulated on intratumoral Tregs compared to normal tissue Tregs and other intratumoral T cells, making these intriguing therapeutic targets.Brentuximab vedotin (BV) is approved for classical Hodgkin lymphoma (cHL) across multiple lines of therapy including frontline use in stage III/IV cHL in combination with doxorubicin, vinblastine, and dacarbazine. BV is also approved for certain CD30-expressing T-cell lymphomas. BV is comprised of a CD30-directed monoclonal antibody conjugated to the highly potent microtubule-disrupting agent monomethyl auristatin E (MMAE).The activity of BV in lymphomas is thought to primarily result from tumor directed intracellular MMAE release, leading to mitotic arrest and apoptotic cell death.The role CD30 plays in normal immune function is unclear, with both costimulatory and proapoptotic roles described. CD30 is transiently upregulated following activation of memory T cells and expression has been linked to highly activated/suppressive IRF4+ effector Tregs.MethodsHere we evaluated the activity of BV on CD30-expressing T cell subsets in vitro and in vivo.ResultsTreatment of enriched T cell subsets with clinically relevant concentrations of BV drove selective depletion of CD30-expressing Tregs > CD30-expressingCD4+ T memory cells, with minimal effects on CD30-expressing CD8+ T memory cells. In a humanized xeno-GVHD model, treatment with BV selectively depleted Tregs resulting in accelerated wasting and robust T cell expansion. The observed differential activity on Tregs is likely attributable to significant increases in CD30 expression and reduced efflux pump activity relative to other T cell subsets. Interestingly, blockade of CD25 signaling prevents CD30 expression on T cell subsets without impacting proliferation, suggesting a link between CD25, the high affinity IL-2 receptor, and CD30 expression.ConclusionsTogether, these data suggest that BV may have an immunomodulatory effect through selective depletion of highly suppressive CD30-expressing Tregs.AcknowledgementsThe authors would like to thank Michael Harrison, PharmD for their assistance in abstract preparation.Ethics ApprovalAnimals studies were approved by and conducted in accordance with Seattle Genetics Institutional Care and Use Committee protocol #SGE-024.

scholarly journals Inactivation of c-Cbl Reverses Neonatal Lethality and T Cell Developmental Arrest of SLP-76–deficient Mice

2004 ◽  
Vol 200 (1) ◽  
pp. 25-34 ◽  
Author(s):  
Y. Jeffrey Chiang ◽  
Connie L. Sommers ◽  
Martha S. Jordan ◽  
Hua Gu ◽  
Lawrence E. Samelson ◽  
...  
Keyword(s):  
Signal Transduction ◽  
T Cells ◽  
T Cell ◽  
T Cell Development ◽  
Adaptor Protein ◽  
Premature Death ◽  
Cell Receptor ◽  
Cell Development ◽  
Neonatal Lethality ◽  
Deficient Mice

c-Cbl is an adaptor protein that negatively regulates signal transduction events involved in thymic-positive selection. To further characterize the function of c-Cbl in T cell development, we analyzed the effect of c-Cbl inactivation in mice deficient in the scaffolding molecule SLP-76. SLP-76–deficient mice show a high frequency of neonatal lethality; and in surviving mice, T cell development is blocked at the DN3 stage. Inactivation of c-cbl completely reversed the neonatal lethality seen in SLP-76–deficient mice and partially reversed the T cell development arrest in these mice. SLP-76−/− Cbl−/− mice exhibited marked expansion of polarized T helper type (Th)1 and Th2 cell peripheral CD4+ T cells, lymphoid infiltrates of parenchymal organs, and premature death. This rescue of T cell development is T cell receptor dependent because it does not occur in recombination activating gene 2−/− SLP-76−/− Cbl−/− triple knockout mice. Analysis of the signal transduction properties of SLP-76−/− Cbl−/− T cells reveals a novel SLP-76– and linker for activation of T cells–independent pathway of extracellular signal–regulated kinase activation, which is normally down-regulated by c-Cbl.

scholarly journals Induction of unresponsiveness and impaired T cell expansion by staphylococcal enterotoxin B in CD28-deficient mice.

1996 ◽  
Vol 183 (6) ◽  
pp. 2481-2488 ◽  
Author(s):  
H W Mittrücker ◽  
A Shahinian ◽  
D Bouchard ◽  
T M Kündig ◽  
T W Mak
Keyword(s):  
T Cells ◽  
T Cell ◽  
Staphylococcal Enterotoxin ◽  
Staphylococcal Enterotoxin B ◽  
Rapid Induction ◽  
Cd28 Costimulation ◽  
Enterotoxin B ◽  
Deficient Mice

We used CD28-deficient mice to analyze the importance of CD28 costimulation for the response against Staphylococcal enterotoxin B (SEB) in vivo. CD28 was necessary for the strong expansion of V beta 8+ T cells, but not for deletion. The lack of expansion was not due to a failure of SEB to activate V beta 8+ T cells, as V beta 8+ T cells from both CD28-/- and CD28+/+ mice showed similar phenotypic changes within the first 24 h after SEB injection and cell cycle analysis showed that an equal percentage of V beta 8+ T cells started to proliferate. However, the phenotype and the state of proliferation of V beta 8+ T cells was different at later time points. Furthermore, in CD28-/- mice injection with SEB led to rapid induction of unresponsiveness in SEB responsive T cells, indicated by a drastic reduction of proliferation after secondary SEB stimulation in vitro. Unresponsiveness could also be demonstrated in vivo, as CD28-/- mice produced only marginal amounts of TNF alpha after rechallenge with SEB. In addition CD28-/- mice were protected against a lethal toxic shock induced by a second injection with SEB. Our results indicate that CD28 costimulation is crucial for the T cell-mediated toxicity of SEB and demonstrate that T cell stimulation in the absence of CD28 costimulation induces unresponsiveness in vivo.

Functional and Molecular Analysis of Maturation of Thymocytes in Bcl10 or Malt1 Deficient Mice.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3323-3323
Author(s):  
Philipp J. Jost ◽  
Uta Ferch ◽  
Stephanie Weiss ◽  
Stephanie Leeder ◽  
Olaf Gross ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Transgenic Animals ◽  
Adaptor Proteins ◽  
Cell Receptor ◽  
Single Positive ◽  
T Cell Selection ◽  
Deficient Mice

Abstract Development of immature T cells in the thymus requires signals through the clonotypic T cell receptor (TCR). Thymocytes expressing a functionally inactive or autoreactive TCR are deleted via apoptosis (negative selection). Thymocytes expressing a functionally active but not autoreactive TCR are selected through inhibition of cell death (positive selection). Deregulation of this process is likely to result in autoimmunity or lymphomagenesis of T cells. The intracellular mechanisms by which the balance between TCR-dependent survival and apoptosis are regulated are largely unknown. A central regulator of survival and apoptosis in the immune system is the transcription factor NF-κB. Activation of NF-κB in mature T-cells requires the adaptor proteins Bcl10 and Malt1. Using gene-targeted mice deficient for Bcl10 or Malt1, we show that Bcl10 and Malt1 are also required for TCR-induced NF-κB activation in immature T cells. Furthermore, to elucidate the process of T cell selection within the thymus, we have crossed Bcl10 or Malt1 deficient mice into mice with genetic backgrounds expressing defined TCR transgenes. Using specific peptide agonists of these TCR transgenes, we show that neither in vivo nor in vitro development into single positive (SP) CD4 or CD8 positive T cells is altered in Bcl10 or Malt1 deficient mice. Absolute numbers and ratio of SP T cells found within the thymus or in peripheral lymphnodes of transgenic animals are normal. These findings indicate that Bcl10 and Malt1 activate NF-κB in thymocytes but are dispensable for maturation of immature T cells in this model system.

The in Vitro and In Vivo Oncogenic Activity of Homodimeric Mutant of Interleukin-7 Receptor a Chain (IL7Rα) Highlights the Significance of the IL7Rα/Jak1 Pathway in T-Cell Acute Lymphoblastic Leukemia

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 73-73
Author(s):  
Kazuaki Yokoyama ◽  
Nozomi Yokoyama ◽  
Kiyoko Izawa ◽  
Ai Kotani ◽  
Ratanakanit Harnprasopwat ◽  
...  
Keyword(s):  
T Cell ◽  
Lymphoblastic Leukemia ◽  
T Cell Development ◽  
Cell Development ◽  
Gamma Delta ◽  
Interleukin 7 ◽  
Signaling Axis ◽  
A Chain

Abstract Abstract 73 Interleukin-7 (IL7) is essential for T cell development and homeostasis. Dysregulation of signals that control normal T-cell development has been implicated in the onset of T-cell acute lymphoblastic leukemia (T-ALL). By analogy to activating mutations in the Notch pathways, we hypothesized that any mutations in the IL7 signaling axis might also contribute to T-ALL. Direct sequencing of human IL7 receptor a chain (hIL7RA) gene in a panel of 16 T-ALL cell lines identified two types of mutations in two different cell lines. One was an insertion mutation of 4 amino acids (LSRC) in the transmembrane region (INS, Fig.1A) from DND-41, a gamma-delta TCR+ T-ALL cell line, and the other was a truncated, loss-of-function, mutation in the cytoplasmic region from MOLT-4. We demonstrated that hIL7RA-INS mutant spontaneously formed a homodimer and constitutively activated downstream signals including Stat family members (1, 3 and 5), Akt and Erk via Jak1, but not Jak3. Next, we investigated oncogenic activity of hIL7RA-INS in primary hematopoietic progenitor cells. To this aim, lin− E.14 Balb/c fetal liver (FL) cells were retrovirally transduced with hIL7RA-INS in parallel with hIL7RA-wild type (WT), and then tested for their cytokine dependence in vitro. As expected, only hIL7RA-INS-transduced lin−FL cells showed abrogation of cytokine dependence. hIL7RA-transduced lin−FL cells were also transplanted into lethally irradiated syngeneic mice. Within 7–9 weeks after transplantation of lin−FL cells transduced with hIL7RA-INS, but not with hIL7RA-WT, recipient mice developed well-tolerated myelo- and lymphoproliferative disorders, characterized by marked leukocytosis, systemic lymphadenopathy and splenomegaly (Fig.1B). Notably, concomitant increase in hIL7RA+gamma-delta TCR+ T cells and decrease in B cells were observed in peripheral blood (Fig.1C). Histological examination of bone marrow, spleen and liver specimens from diseased mice revealed moderate to severe myeloid hyperplasia, disrupted splenic architecture by disseminated mature myeloid cells and infiltration of both myeloid and mononuclear cells into hepatic parenchyma, respectively. In addition, recipient mice for hIL7RA-INS-transduced lin−FL cells frequently manifested ruffled fur as well as mononuclear cell infiltration into salivary gland and pericardium, suggesting an autoimmune-like disorder. However, during median follow-up of 11 weeks, these recipient mice did not develop either overt leukemia or lymphoma, indicating that additional transforming events are required for evolution to aggressive hematological malignancies. These in vivo findings highlighted the possibility that aberrant signals via IL7RA in hematopoietic stem/progenitor cells might preferentially stimulate myelopoiesis over lymphopoiesis, and also confirmed the essential role of IL7RA in gamma-delta TCR+ T cell development, previously shown by IL7RA-knockout mice. Taken together, we speculated that dysregulated IL7RA signaling axis might be involved in the onset of T-ALL, especially with gamma-delta TCR+ phenotype. Finally, the present study, together with the recent report (JEM 208:901, 2011), emphasizes the significance of the sequential Notch-IL7RA pathways in the pathogenesis of T-ALL as well as the dominant role of the IL7RA/Jak1 axis in IL7 proliferative signal. Disclosures: No relevant conflicts of interest to declare.

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