Analysis of TCR, pTα, and RAG-1 in T-acute lymphoblastic leukemias improves understanding of early human T-lymphoid lineage commitment

Blood ◽  
2003 ◽  
Vol 101 (7) ◽  
pp. 2693-2703 ◽  
Author(s):  
Vahid Asnafi ◽  
Kheira Beldjord ◽  
Emmanuelle Boulanger ◽  
Béatrice Comba ◽  
Patricia Le Tutour ◽  
...  

T-acute lymphoblastic leukemias (T-ALLs) derive from human T-lymphoid precursors arrested at various early stages of development. Correlation of phenotype and T-cell receptor (TCR) status with RAG-1 and pTα transcription in 114 T-ALLs demonstrated that they largely reflect physiologic T-lymphoid development. Half the TCRαβ lineage T-ALLs expressed a pre-TCR, as evidenced by RAG-1, pTα, and cTCRβ expression, absence of TCRδ deletion, and a sCD3−, CD1a+, CD4/8 double-positive (DP) phenotype, in keeping with a population undergoing β selection. Most TCRγδ T-ALLs were pTα, terminal deoxynucleotidyl transferase (TdT), and RAG-1lo/neg, double-negative/single-positive (DN/SP), and demonstrated only TCRβ DJ rearrangement, whereas 40% were pTα, TdT, and RAG-1 positive, DP, and demonstrated TCRβ V(D)J rearrangement, with cTCRβ expression in proportion. As such they may correspond to TCRαβ lineage precursors selected by TCRγδ expression, to early γδ cells recently derived from a pTα+ common αβ/γδ precursor, or to a lineage-deregulated αβ/γδ intermediate. Approximately 30% of T-ALLs were sCD3/cTCRβ− and corresponded to nonrestricted thymic precursors because they expressed non–T-restricted markers such as CD34, CD13, CD33, and CD56 and were predominantly DN, CD1a, pTα, and RAG-1 low/negative, despite immature TCRδ and TCRγ rearrangements. TCR gene configuration identified progressive T-lymphoid restriction. T-ALLs, therefore, provide homogeneous expansions of minor human lymphoid precursor populations that can aid in the understanding of healthy human T-cell development.

Blood ◽  
1999 ◽  
Vol 93 (9) ◽  
pp. 3033-3043 ◽  
Author(s):  
Bianca Blom ◽  
Martie C.M. Verschuren ◽  
Mirjam H.M. Heemskerk ◽  
Arjen Q. Bakker ◽  
Ellen J. van Gastel-Mol ◽  
...  

Abstract Recent studies have identified several populations of progenitor cells in the human thymus. The hematopoietic precursor activity of these populations has been determined. The most primitive human thymocytes express high levels of CD34 and lack CD1a. These cells acquire CD1a and differentiate into CD4+CD8+ through CD3−CD4+CD8− and CD3−CD4+CD8+β− intermediate populations. The status of gene rearrangements in the various TCR loci, in particular of TCRδ and TCRγ, has not been analyzed in detail. In the present study we have determined the status of TCR gene rearrangements of early human postnatal thymocyte subpopulations by Southern blot analysis. Our results indicate that TCRδ rearrangements initiate in CD34+CD1a− cells preceding those in the TCRγ and TCRβ loci that commence in CD34+CD1a+ cells. Furthermore, we have examined at which cellular stage TCRβ selection occurs in humans. We analyzed expression of cytoplasmic TCRβ and cell-surface CD3 on thymocytes that lack a mature TCRβ. In addition, we overexpressed a constitutive-active mutant of p56lckF505 by retrovirus-mediated gene transfer in sequential stages of T-cell development and analyzed the effect in a fetal thymic organ culture system. Evidence is presented that TCRβ selection in humans is initiated at the transition of the CD3−CD4+CD8− into the CD4+CD8+β− stage.


1992 ◽  
Vol 2 (2) ◽  
pp. 95-101 ◽  
Author(s):  
Cees de Heer ◽  
Bernard de Geus ◽  
Henk-Jan Schuurma ◽  
Henk Van Loveren ◽  
Jan Rozing

T-cell receptor (TCR)ß-chain usage and expression of the CD3, CD4, and CD8 differentiation antigens were analyzed in 14 spontaneous AKR lymphomas. Lymphoma cells massively infiltrated and/or proliferated in the organs analyzed (thymus, spleen, and mesenteric lymph nodes), giving rise to a loss of organ structure. One lymphoma occurred only in the thymus, and failed to express CD3, CD4, and CD8. All other lymphomas expressed the CD3/TCR complex. With respect to CD4 and CD8 expression, the lymphomas were either double-negative (DN), double-positive (DP), or single-positive (SP). The frequency of DP (CD4+8+) lymphomas was low compared to the frequency of DP thymocytes in a normal AKR thymus. A substantial heterogeneity was seen in the intensity of CD4 and CD8 expression among various lymphomas, which was independent of the level of CD3 expression. Considering TCR Vßgene family usage, 2 out of 14 lymphomas expressed Vß6. Normally, Vß6+thymocytes are deleted from the thymocyte pool at the immature DP stage of T-cell development in AKR mice. These data support the hypothesis that the lymphocytes in the immature DP stage of T-cell development are susceptible to the induction of AKR lymphomagenesis. The presence of Vß6+lymphoma cells indicates that the lymphomagenesis is accompanied by a defective clonal deletion of cells expressing a possible autoreactive TCR.


2015 ◽  
Vol 112 (25) ◽  
pp. 7773-7778 ◽  
Author(s):  
Hyung-Ok Lee ◽  
Xiao He ◽  
Jayati Mookerjee-Basu ◽  
Dai Zhongping ◽  
Xiang Hua ◽  
...  

The transcription factor T-helper-inducing POZ/Krueppel-like factor (ThPOK, encoded by the Zbtb7b gene) plays widespread and critical roles in T-cell development, particularly as the master regulator of CD4 commitment. Here we show that mice expressing a constitutive T-cell–specific ThPOK transgene (ThPOKconst mice) develop thymic lymphomas. These tumors resemble human T-cell acute lymphoblastic leukemia (T-ALL), in that they predominantly exhibit activating Notch1 mutations. Lymphomagenesis is prevented if thymocyte development is arrested at the DN3 stage by recombination-activating gene (RAG) deficiency, but restored by introduction of a T-cell receptor (TCR) transgene or by a single injection of anti-αβTCR antibody into ThPOKconst RAG-deficient mice, which promotes development to the CD4+8+ (DP) stage. Hence, TCR signals and/or traversal of the DN (double negative) > DP (double positive) checkpoint are required for ThPOK-mediated lymphomagenesis. These results demonstrate a novel link between ThPOK, TCR signaling, and lymphomagenesis. Finally, we present evidence that ectopic ThPOK expression gives rise to a preleukemic and self-perpetuating DN4 lymphoma precursor population. Our results collectively define a novel role for ThPOK as an oncogene and precisely map the stage in thymopoiesis susceptible to ThPOK-dependent tumor initiation.


Blood ◽  
1999 ◽  
Vol 93 (9) ◽  
pp. 3033-3043 ◽  
Author(s):  
Bianca Blom ◽  
Martie C.M. Verschuren ◽  
Mirjam H.M. Heemskerk ◽  
Arjen Q. Bakker ◽  
Ellen J. van Gastel-Mol ◽  
...  

Recent studies have identified several populations of progenitor cells in the human thymus. The hematopoietic precursor activity of these populations has been determined. The most primitive human thymocytes express high levels of CD34 and lack CD1a. These cells acquire CD1a and differentiate into CD4+CD8+ through CD3−CD4+CD8− and CD3−CD4+CD8+β− intermediate populations. The status of gene rearrangements in the various TCR loci, in particular of TCRδ and TCRγ, has not been analyzed in detail. In the present study we have determined the status of TCR gene rearrangements of early human postnatal thymocyte subpopulations by Southern blot analysis. Our results indicate that TCRδ rearrangements initiate in CD34+CD1a− cells preceding those in the TCRγ and TCRβ loci that commence in CD34+CD1a+ cells. Furthermore, we have examined at which cellular stage TCRβ selection occurs in humans. We analyzed expression of cytoplasmic TCRβ and cell-surface CD3 on thymocytes that lack a mature TCRβ. In addition, we overexpressed a constitutive-active mutant of p56lckF505 by retrovirus-mediated gene transfer in sequential stages of T-cell development and analyzed the effect in a fetal thymic organ culture system. Evidence is presented that TCRβ selection in humans is initiated at the transition of the CD3−CD4+CD8− into the CD4+CD8+β− stage.


Blood ◽  
2011 ◽  
Vol 117 (26) ◽  
pp. 7053-7062 ◽  
Author(s):  
Margherita Ghisi ◽  
Alberto Corradin ◽  
Katia Basso ◽  
Chiara Frasson ◽  
Valentina Serafin ◽  
...  

Abstract Ontogenesis of T cells in the thymus is a complex process whose molecular control is poorly understood. The present study investigated microRNAs involved in human thymocyte differentiation by comparing the microRNA expression profiles of thymocytes at the double-positive, single-positive CD4+ and single-positive CD8+ maturation stages. Microarray analysis showed that each thymocyte population displays a distinct microRNA expression profile that reflects their developmental relationships. Moreover, analysis of small-RNA libraries generated from human unsorted and double-positive thymocytes and from mature peripheral CD4+ and CD8+ T lymphocytes, together with the microarray data, indicated a trend toward up-regulation of microRNA expression during T-cell maturation after the double-positive stage and revealed a group of microRNAs regulated during normal T-cell development, including miR-150, which is strongly up-regulated as maturation progresses. We showed that miR-150 targets NOTCH3, a member of the Notch receptor family that plays important roles both in T-cell differentiation and leukemogenesis. Forced expression of miR-150 reduces NOTCH3 levels in T-cell lines and has adverse effects on their proliferation and survival. Overall, these findings suggest that control of the Notch pathway through miR-150 may have an important impact on T-cell development and physiology.


2005 ◽  
Vol 202 (1) ◽  
pp. 111-121 ◽  
Author(s):  
Troy A. Baldwin ◽  
Michelle M. Sandau ◽  
Stephen C. Jameson ◽  
Kristin A. Hogquist

Sequential rearrangement of the T cell receptor for antigen (TCR) β and α chains is a hallmark of thymocyte development. This temporal control is lost in TCR transgenics because the α chain is expressed prematurely at the CD4−CD8− double negative (DN) stage. To test the importance of this, we expressed the HYα chain at the physiological CD4+CD8+ double positive (DP) stage. The reduced DP and increased DN cellularity typically seen in TCR transgenics was not observed when the α chain was expressed at the appropriate stage. Surprisingly, antigen-driven selection events were also altered. In male mice, thymocyte deletion now occurred at the single positive or medullary stage. In addition, no expansion of CD8αα intestinal intraepithelial lymphocytes (IELs) was observed, despite the fact that HY transgenics have been used to model IEL development. Collectively, these data establish the importance of proper timing of TCR expression in thymic development and selection and emphasize the need to use models that most accurately reflect the physiologic process.


1992 ◽  
Vol 176 (6) ◽  
pp. 1619-1624 ◽  
Author(s):  
B A Vandekerckhove ◽  
R Baccala ◽  
D Jones ◽  
D H Kono ◽  
A N Theofilopoulos ◽  
...  

Implantation of pieces of human fetal liver and thymus into SCID mice results in the development of a human thymus-like organ, in which sustained lymphopoiesis is reproducibly observed. In this model, T cell development can be experimentally manipulated. To study the influence of thymic selection on the development of the human T cell repertoire, the T cell receptor (TCR) V beta gene repertoire of double-positive (CD4+CD8+) and single-positive (CD4+CD8- and CD4-CD8+) T cells was analyzed in the SCID-hu thymus using a multiprobe ribonuclease protection assay. TCR diversity in double-positive SCID-hu thymocytes was found to be comparable with that present in the thymus of the fetal liver donor, did not change with time, and was independent of the origin of the thymus donor. Thymic selection in SCID-hu thymus induces changes in V beta usage by the single-positive CD4+ or CD8+ T cells comparable with those previously reported for single-positive cells present in a normal human thymus. Finally, significant differences were observed in the V beta usage by CD4 or CD8 single-positive T cells that matured from genetically identical stem cells in different thymic environments. Collectively, these data suggest: first, that the generation of TCR diversity at the double-positive stage is determined by the genotype of the stem cells; and second, that polymorphic determinants expressed by thymic epithelium measurably influence the V beta repertoire of mature single-positive T cells.


2015 ◽  
Vol 35 (22) ◽  
pp. 3854-3865 ◽  
Author(s):  
Kristy R. Stengel ◽  
Yue Zhao ◽  
Nicholas J. Klus ◽  
Jonathan F. Kaiser ◽  
Laura E. Gordy ◽  
...  

Hdac3 is a key target for Hdac inhibitors that are efficacious in cutaneous T cell lymphoma. Moreover, the regulation of chromatin structure is critical as thymocytes transition from an immature cell with open chromatin to a mature T cell with tightly condensed chromatin. To define the phenotypes controlled by Hdac3 during T cell development, we conditionally deletedHdac3using theLck-Cretransgene. This strategy inactivatedHdac3in the double-negative stages of thymocyte development and caused a significant impairment at the CD8 immature single-positive (ISP) stage and the CD4/CD8 double-positive stage, with few mature CD4+or CD8+single-positive cells being produced. WhenHdac3−/−mice were crossed withBcl-xL-,Bcl2-, orTCRβ-expressing transgenic mice, a modest level of complementation was found. However, when the null mice were crossed with mice expressing a fully rearranged T cell receptor αβ transgene, normal levels of CD4 single-positive cells were produced. Thus, Hdac3 is required for the efficient transit from double-negative stage 4 through positive selection.


2018 ◽  
Vol 46 (4) ◽  
pp. 441-449
Author(s):  
Sowmya Angusamy ◽  
Tamer Mansour ◽  
Mohammed Abdulmageed ◽  
Rachel Han ◽  
Brian C. Schutte ◽  
...  

Abstract Background: The adaptive immune system of neonates is relatively underdeveloped. The thymus is an essential organ for adaptive T cell development and might be affected during the natural course of oxygen induced lung injury. The effect of prolonged hyperoxia on the thymus, thymocyte and T cell development, and its proliferation has not been studied extensively. Methods: Neonatal mice were exposed to 85% oxygen (hyperoxia) or room air (normoxia) up to 28 days. Flow cytometry using surface markers were used to assay for thymocyte development and proliferation. Results: Mice exposed to prolonged hyperoxia had evidence of lung injury associated alveolar simplification, a significantly lower mean weight, smaller thymic size, lower mean thymocyte count and higher percentage of apoptotic thymocytes. T cells subpopulation in the thymus showed a significant reduction in the count and proliferation of double positive and double negative T cells. There was a significant reduction in the count and proliferation of single positive CD4+ and CD8+ T cells. Conclusions: Prolonged hyperoxia in neonatal mice adversely affected thymic size, thymocyte count and altered the distribution of T cells sub-populations. These results are consistent with the hypothesis that prolonged hyperoxia causes defective development of T cells in the thymus.


Blood ◽  
1998 ◽  
Vol 92 (2) ◽  
pp. 383-393 ◽  
Author(s):  
David J. Izon ◽  
Sofia Rozenfeld ◽  
Stephen T. Fong ◽  
László Kömüves ◽  
Corey Largman ◽  
...  

Abstract Hox homeobox genes play a crucial role in specifying the embryonic body pattern. However, a role for Hox genes in T-cell development has not been explored. The Hoxa-9 gene is expressed in normal adult and fetal thymuses. Fetal thymuses of mice homozygous for an interruption of the Hoxa-9 gene are one eighth normal size and have a 25-fold decrease in the number of primitive thymocytes expressing the interleukin-2 receptor (IL-2R, CD25). Progression to the double positive (CD4+CD8+) stage is dramatically retarded in fetal thymic organ cultures. This aberrant development is associated with decreased amounts of intracellular CD3 and T-cell receptor β (TCRβ) and reduced surface expression of IL-7R and E-cadherin. Mutant thymocytes show a significant increase in apoptotic cell death and premature downregulation of bcl-2 expression. A similar phenotype is seen in primitive thymocytes from adult Hoxa-9−/− mice and from mice transplanted with Hoxa-9−/−marrow. Hoxa-9 appears to play a previously unsuspected role in T-cell ontogeny by modulating cell survival of early thymocytes and by regulating their subsequent differentiation.


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