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Sample records for double positive thymocytes

  1. An Essential Role for Nuclear Factor κB in Promoting Double Positive Thymocyte Apoptosis

    PubMed Central

    Hettmann, Thore; DiDonato, Joseph; Karin, Michael; Leiden, Jeffrey M.

    1999-01-01

    To examine the role of nuclear factor (NF)-κB in T cell development and activation in vivo, we produced transgenic mice that express a superinhibitory mutant form of inhibitor κB-α (IκB-αA32/36) under the control of the T cell–specific CD2 promoter and enhancer (mutant [m]IκB-α mice). Thymocyte development proceeded normally in the mIκB-α mice. However, the numbers of peripheral CD8+ T cells were significantly reduced in these animals. The mIκB-α thymocytes displayed a marked proliferative defect and significant reductions in interleukin (IL)-2, IL-3, and granulocyte/macrophage colony-stimulating factor production after cross-linking of the T cell antigen receptor. Perhaps more unexpectedly, double positive (CD4+CD8+; DP) thymocytes from the mIκB-α mice were resistant to α-CD3–mediated apoptosis in vivo. In contrast, they remained sensitive to apoptosis induced by γ-irradiation. Apoptosis of wild-type DP thymocytes after in vivo administration of α-CD3 mAb was preceded by a significant reduction in the level of expression of the antiapoptotic gene, bcl-xL. In contrast, the DP mIκB-α thymocytes maintained high level expression of bcl-xL after α-CD3 treatment. Taken together, these results demonstrated important roles for NF-κB in both inducible cytokine expression and T cell proliferation after TCR engagement. In addition, NF-κB is required for the α-CD3–mediated apoptosis of DP thymocytes through a pathway that involves the regulation of the antiapoptotic gene, bcl-xL. PMID:9874571

  2. The frequency of double-positive thymocytes expressing an alphabeta TCR clonotype regulates peripheral CD4 T cell compartment homeostasis.

    PubMed

    Reed, Amy J; Zarrabi, Yasaman; Perate, Alison L; Jeganathan, Arjun; Naji, Ali; Noorchashm, Hooman

    2005-11-01

    The present study aimed to determine whether the frequency of double positive (DP) thymocytes expressing alphabeta T-cell receptor (TCR) clonotypes at the time of selection regulates peripheral CD4 T-cell compartment size. Scid recipients were inoculated with various ratios of TCR Calpha(0/0) and wild-type bone marrow (BM) stem cells. Increasing the frequency of TCR Calpha(0/0) thymocytes at steady-state introduced a graded decrease in the maturation probability of the total DP thymocyte pool. At 12-14 weeks following BM inoculation, the frequency of TCR Calpha(0/0) DP thymocytes was inversely correlated with that of CD4 single positive (SP) thymocytes. Notwithstanding, a decreased frequency of wild-type DP thymocytes led to a marked increase in their transit efficiency from the DP to SP compartments. The frequency-dependent increase in thymocyte transit efficiency was associated with a CD4 SP cell surface phenotype indicative of increased antigenic experience. Importantly, the frequency of DP thymocytes capable of expressing TCR clonotypes dictated the steady-state size of the peripheral CD4 T cell compartment and its potential for homeostatic proliferation. Collectively, these results indicate that the efficiency of DP to CD4 SP transit is a frequency dependent process, which determines (1) the steady-state size of the peripheral T cell compartment and (2) the threshold for homeostatic expansion of peripheral CD4 T cells.

  3. Cooperation of Gata3, c-Myc and Notch in malignant transformation of double positive thymocytes.

    PubMed

    van Hamburg, Jan Piet; de Bruijn, Marjolein J W; Dingjan, Gemma M; Beverloo, H Berna; Diepstraten, Hans; Ling, Kam-Wing; Hendriks, Rudi W

    2008-06-01

    Gata transcription factors are critical regulators of proliferation and differentiation implicated in various human cancers, but specific genes activated by Gata proteins remain to be identified. We previously reported that enforced expression of Gata3 during T cell development in CD2-Gata3 transgenic mice induced CD4(+)CD8(+) double-positive (DP) T cell lymphoma. Here, we show that the presence of the DO11.10 T-cell receptor transgene, which directs DP cells towards the CD4 lineage, resulted in enhanced lymphoma development and a dramatic increase in thymocyte cell size in CD2-Gata3 transgenic mice. CD2-Gata3 DP cells expressed high levels of the proto-oncogene c-Myc but the Notch1 signaling pathway, which is known to induce c-Myc, was not activated. Gene expression profiling showed that in CD2-Gata3 lymphoma cells transcription of c-Myc and its target genes was further increased. A substantial fraction of CD2-Gata3 lymphomas had trisomy of chromosome 15, leading to an increased c-Myc gene dose. Interestingly, most lymphomas showed high expression of the Notch targets Deltex1 and Hes1, often due to activating Notch1 PEST domain mutations. Therefore, we conclude that enforced Gata3 expression converts DP thymocytes into a pre-malignant state, characterized by high c-Myc expression, whereby subsequent induction of Notch1 signaling cooperates to establish malignant transformation. The finding that Gata3 regulates c-Myc expression levels, in a direct or indirect fashion, may explain the parallel phenotypes of mice with overexpression or deficiency of either of the two transcription factors.

  4. Over-expression of Runx1 transcription factor impairs the development of thymocytes from the double-negative to double-positive stages.

    PubMed

    Wong, Won F; Nakazato, Megumi; Watanabe, Toshio; Kohu, Kazuyoshi; Ogata, Takehiro; Yoshida, Naomi; Sotomaru, Yusuke; Ito, Mamoru; Araki, Kimi; Telfer, Janice; Fukumoto, Manabu; Suzuki, Daisuke; Sato, Takehito; Hozumi, Katsuto; Habu, Sonoko; Satake, Masanobu

    2010-06-01

    Runx1 transcription factor is highly expressed at a CD4/CD8-double-negative (DN) stage of thymocyte development but is down-regulated when cells proceed to the double-positive (DP) stage. In the present study, we examined whether the down-regulation of Runx1 is necessary for thymocyte differentiation from the DN to DP stage. When Runx1 was artificially over-expressed in thymocytes by Lck-driven Cre, the DN3 population was unaffected, as exemplified by proper pre-T-cell receptor expression, whereas the DN4 population was perturbed as shown by the decrease in the CD27(hi) sub-fraction. In parallel, the growth rate of DN4 cells was reduced by half, as measured by bromodeoxyuridine incorporation. These events impaired the transition of DN4 cells to the DP stage, resulting in the drastic reduction of the number of DP thymocytes. The Runx1 gene has two promoters, a proximal and a distal promoter; and, in thymocytes, endogenous Runx1 was mainly transcribed from the distal promoter. Interestingly, only distal, but not proximal, Runx1 over-expression exhibited an inhibitory effect on thymocyte differentiation, suggesting that the distal Runx1 protein may fulfil a unique function. Our collective results indicate that production of the distal Runx1 protein must be adequately down-regulated for thymocytes to transit from the DN to the DP stage, a critical step in the massive expansion of the T-cell lineage.

  5. The frequency of double-positive thymocytes expressing an αβ TCR clonotype regulates peripheral CD4 T cell compartment homeostasis

    PubMed Central

    Reed, Amy J; Zarrabi, Yasaman; Perate, Alison L; Jeganathan, Arjun; Naji, Ali; Noorchashm, Hooman

    2005-01-01

    The present study aimed to determine whether the frequency of double positive (DP) thymocytes expressing αβ T-cell receptor (TCR) clonotypes at the time of selection regulates peripheral CD4 T-cell compartment size. Scid recipients were inoculated with various ratios of TCR Cα0/0 and wild-type bone marrow (BM) stem cells. Increasing the frequency of TCR Cα0/0 thymocytes at steady-state introduced a graded decrease in the maturation probability of the total DP thymocyte pool. At 12–14 weeks following BM inoculation, the frequency of TCR Cα0/0 DP thymocytes was inversely correlated with that of CD4 single positive (SP) thymocytes. Notwithstanding, a decreased frequency of wild-type DP thymocytes led to a marked increase in their transit efficiency from the DP to SP compartments. The frequency-dependent increase in thymocyte transit efficiency was associated with a CD4 SP cell surface phenotype indicative of increased antigenic experience. Importantly, the frequency of DP thymocytes capable of expressing TCR clonotypes dictated the steady-state size of the peripheral CD4 T cell compartment and its potential for homeostatic proliferation. Collectively, these results indicate that the efficiency of DP to CD4 SP transit is a frequency dependent process, which determines (1) the steady-state size of the peripheral T cell compartment and (2) the threshold for homeostatic expansion of peripheral CD4 T cells. PMID:16236130

  6. β-Catenin stabilization stalls the transition from double-positive to single-positive stage and predisposes thymocytes to malignant transformation

    PubMed Central

    Guo, Zhuyan; Dose, Marei; Kovalovsky, Damian; Chang, Rui; O'Neil, Jennifer; Look, A. Thomas; von Boehmer, Harald; Khazaie, Khashayarsha

    2007-01-01

    Activation of β-catenin has been causatively linked to the etiology of colon cancer. Conditional stabilization of this molecule in pro-T cells promotes thymocyte development without the requirement for pre-TCR signaling. We show here that activated β-catenin stalls the developmental transition from the double-positive (DP) to the single-positive (SP) thymocyte stage and predisposes DP thymocytes to transformation. β-Catenin–induced thymic lymphomas have a leukemic arrest at the early DP stage. Lymphomagenesis requires Rag activity, which peaks at this developmental stage, as well as additional secondary genetic events. A consistent secondary event is the transcriptional up-regulation of c-Myc, whose activity is required for transformation because its conditional ablation abrogates lymphomagenesis. In contrast, the expression of Notch receptors as well as targets is reduced in DP thymocytes with stabilized β-catenin and remains low in the lymphomas, indicating that Notch activation is not required or selected for in β-catenin–induced lymphomas. Thus, β-catenin activation may provide a mechanism for the induction of T-cell–acute lymphoblastic leukemia (T-ALL) that does not depend on Notch activation. PMID:17317856

  7. Low glucocorticoid receptor (GR), high Dig2 and low Bcl-2 expression in double positive thymocytes of BALB/c mice indicates their endogenous glucocorticoid hormone exposure.

    PubMed

    Boldizsár, Ferenc; Pálinkás, László; Czömpöly, Tamás; Bartis, Domokos; Németh, Péter; Berki, Timea

    2006-01-01

    Several studies have shown that of the four major thymocyte subsets, the CD4/CD8 double positive (DP) thymocytes are the most sensitive to in vivo glucocorticoid hormone (GC)-induced apoptosis. Our aim was to analyse fine molecular differences among thymocyte subgroups that could underlie this phenomenon. Therefore, we characterised the glucocorticoid hormone receptor (GR) expression of thymocyte subgroups both at the mRNA and protein levels by real-time PCR and flow cytometry, and correlated these features to their apoptotic sensitivity. We also investigated the time-dependent effects of the GC agonist dexamethasone (DX) with or without GC antagonist (RU486) treatments on GR mRNA/protein expression. We also analysed the expression of two apoptosis-related gene products: dexamethasone-induced gene 2 (Dig2) mRNA and Bcl-2 protein. We found that DN thymocytes had the highest GR expression, followed by CD8 single positive (SP), CD4 SP and DP thymocytes in 4-week-old BALB/c mice, both at the mRNA and protein levels, respectively. In DP cells, the Dig2 expression was significantly higher, while the Bcl-2 expression was significantly lower than in DN, CD4 SP and CD8 SP thymocytes. Single high dose DX treatment caused time-dependent depletion of DP thymocytes due to their higher apoptosis rate, which could not be abolished with RU486 pretreatment. After a single high dose DX treatment, there was a transient, significant increase of the GR mRNA and protein level of unsorted thymocytes after 8 and 16 h, followed by a significant decrease at 24 h, respectively. The time-dependent GR expression changes after DX administration could not be inhibited by the GC antagonist RU486. Twenty-four hours after exposure to high dose DX the DN, CD4 SP and CD8 SP cells showed a significant decrease of GR mRNA and protein expression, whereas the DP thymocytes, showed no significant alteration of GR mRNA or protein expression. The kinetical analysis of GR expression and apoptotic marker

  8. Notch induces human T-cell receptor γδ+ thymocytes to differentiate along a parallel, highly proliferative and bipotent CD4 CD8 double-positive pathway.

    PubMed

    Van Coppernolle, S; Vanhee, S; Verstichel, G; Snauwaert, S; van der Spek, A; Velghe, I; Sinnesael, M; Heemskerk, M H; Taghon, T; Leclercq, G; Plum, J; Langerak, A W; Kerre, T; Vandekerckhove, B

    2012-01-01

    In wild-type mice, T-cell receptor (TCR) γδ(+) cells differentiate along a CD4 CD8 double-negative (DN) pathway whereas TCRαβ(+) cells differentiate along the double-positive (DP) pathway. In the human postnatal thymus (PNT), DN, DP and single-positive (SP) TCRγδ(+) populations are present. Here, the precursor-progeny relationship of the various PNT TCRγδ(+) populations was studied and the role of the DP TCRγδ(+) population during T-cell differentiation was elucidated. We demonstrate that human TCRγδ(+) cells differentiate along two pathways downstream from an immature CD1(+) DN TCRγδ(+) precursor: a Notch-independent DN pathway generating mature DN and CD8αα SP TCRγδ(+) cells, and a Notch-dependent, highly proliferative DP pathway generating immature CD4 SP and subsequently DP TCRγδ(+) populations. DP TCRγδ(+) cells are actively rearranging the TCRα locus, and differentiate to TCR(-) DP cells, to CD8αβ SP TCRγδ(+) cells and to TCRαβ(+) cells. Finally, we show that the γδ subset of T-cell acute lymphoblastic leukemias (T-ALL) consists mainly of CD4 SP or DP phenotypes carrying significantly more activating Notch mutations than DN T-ALL. The latter suggests that activating Notch mutations in TCRγδ(+) thymocytes induce proliferation and differentiation along the DP pathway in vivo.

  9. Protein kinase D regulates positive selection of CD4+ thymocytes through phosphorylation of SHP-1

    PubMed Central

    Ishikawa, Eri; Kosako, Hidetaka; Yasuda, Tomoharu; Ohmuraya, Masaki; Araki, Kimi; Kurosaki, Tomohiro; Saito, Takashi; Yamasaki, Sho

    2016-01-01

    Thymic selection shapes an appropriate T cell antigen receptor (TCR) repertoire during T cell development. Here, we show that a serine/threonine kinase, protein kinase D (PKD), is crucial for thymocyte positive selection. In T cell-specific PKD-deficient (PKD2/PKD3 double-deficient) mice, the generation of CD4 single positive thymocytes is abrogated. This defect is likely caused by attenuated TCR signalling during positive selection and incomplete CD4 lineage specification in PKD-deficient thymocytes; however, TCR-proximal tyrosine phosphorylation is not affected. PKD is activated in CD4+CD8+ double positive (DP) thymocytes on stimulation with positively selecting peptides. By phosphoproteomic analysis, we identify SH2-containing protein tyrosine phosphatase-1 (SHP-1) as a direct substrate of PKD. Substitution of wild-type SHP-1 by phosphorylation-defective mutant (SHP-1S557A) impairs generation of CD4+ thymocytes. These results suggest that the PKD–SHP-1 axis positively regulates TCR signalling to promote CD4+ T cell development. PMID:27670070

  10. Gasp, a Grb2-associating protein, is critical for positive selection of thymocytes

    PubMed Central

    Patrick, Michael S.; Oda, Hiroyo; Hayakawa, Kunihiro; Sato, Yoshinori; Eshima, Koji; Kirikae, Teruo; Iemura, Shun-ichiro; Shirai, Mutsunori; Abe, Takaya; Natsume, Tohru; Sasazuki, Takehiko; Suzuki, Harumi

    2009-01-01

    T cells develop in the thymus through positive and negative selection, which are responsible for shaping the T cell receptor (TCR) repertoire. To elucidate the molecular mechanisms involved in selection remains an area of intense interest. Here, we identified and characterized a gene product Gasp (Grb2-associating protein, also called Themis) that is critically required for positive selection. Gasp is a cytosolic protein with no known functional motifs that is expressed only in T cells, especially immature CD4/CD8 double positive (DP) thymocytes. In the absence of Gasp, differentiation of both CD4 and CD8 single positive cells in the thymus was severely inhibited, whereas all other TCR-induced events such as β-selection, negative selection, peripheral activation, and homeostatic proliferation were unaffected. We found that Gasp constitutively associates with Grb2 via its N-terminal Src homology 3 domain, suggesting that Gasp acts as a thymocyte-specific adaptor for Grb2 or regulates Ras signaling in DP thymocytes. Collectively, we have described a gene called Gasp that is critical for positive selection. PMID:19805304

  11. Involvement of p21ras distinguishes positive and negative selection in thymocytes.

    PubMed Central

    Swan, K A; Alberola-Ila, J; Gross, J A; Appleby, M W; Forbush, K A; Thomas, J F; Perlmutter, R M

    1995-01-01

    Small molecular weight GTP binding proteins of the ras family have been implicated in signal transduction from the T cell antigen receptor (TCR). To test the importance of p21ras in the control of thymocyte development, we generated mice expressing a dominant-negative p21ras protein (H-rasN17) in T lineage cells under the control of the lck proximal promoter. Proliferation of thymocytes from lck-H-rasN17 mice in response to TCR stimulation was nearly completely blocked, confirming the importance of p21ras in mediating TCR-derived signals in mature CD4+8- or CD8+4- thymocytes. In contrast, some TCR-derived signals proceeded unimpaired in the CD4+8+ thymocytes of mice expressing dominant-negative p21ras. Analysis of thymocyte development in mice made doubly transgenic for the H-Y-specific TCR and lck-H-rasN17 demonstrated that antigen-specific negative selection occurs normally in the presence of p21H-rasN17. Superantigen-induced negative selection in vivo also proceeded unhindered in H-rasN17 thymocytes. In contrast, positive selection of thymocytes in the H-Y mice was severely compromised by the presence of p21H-rasN17. These observations demonstrate that positive and negative selection, two conceptually antithetical consequences of TCR stimulation, are biochemically distinguishable. Images PMID:7835338

  12. TNF activation of NF-κB is essential for development of single-positive thymocytes

    PubMed Central

    Ley, Steven C.

    2016-01-01

    NF-κB activation has been implicated at multiple stages of thymic development of T cells, during which it is thought to mediate developmental signals originating from the T cell receptor (TCR). However, the Card11–Bcl10–Malt1 (CBM) complex that is essential for TCR activation of NF-κB in peripheral T cells is not required for thymocyte development. It has remained unclear whether the TCR activates NF-κB independent of the CBM complex in thymocyte development or whether another NF-κB activating receptor is involved. In the present study, we generated mice in which T cells lacked expression of both catalytic subunits of the inhibitor of κB kinase (IKK) complex, IKK1 and IKK2, to investigate this question. Although early stages of T cell development were unperturbed, maturation of CD4 and CD8 single-positive (SP) thymocytes was blocked in mice lacking IKK1/2 in the T cell lineage. We found that IKK1/2-deficient thymocytes were specifically sensitized to TNF-induced cell death in vitro. Furthermore, the block in thymocyte development in IKK1/2-deficient mice could be rescued by blocking TNF with anti-TNF mAb or by ablation of TNFRI expression. These experiments reveal an essential role for TNF activation of NF-κB to promote the survival and development of single positive T cells in the thymus. PMID:27432943

  13. Positive selection of transgenic receptor-bearing thymocytes by Kb antigen is altered by Kb mutations that involve peptide binding.

    PubMed Central

    Sha, W C; Nelson, C A; Newberry, R D; Pullen, J K; Pease, L R; Russell, J H; Loh, D Y

    1990-01-01

    A specific interaction between the class I major histocompatibility complex molecule Kb and thymocytes expressing the antigen receptor from the cytolytic T lymphocyte 2C enhances maturation of T cells of the CD8 lineage in transgenic mice. By analyzing transgenic mice backcrossed to Kbm mutant strains of mice, we have identified five bm mutations of the Kb antigen-encoding gene that alter the positive selection of thymocytes induced by Kb antigen. Compared with Kb, Kbm10 and Kbm1 did not induce significant maturation of 2C T-cell receptor-bearing thymocytes, and Kbm8 antigen positively selected for transgenic thymocytes only weakly. Altering residue 77 of Kb molecule from aspartic acid to serine made Kbm3 and Kbm11 allogeneic targets for the 2C antigen receptor and caused deletion of transgenic thymocytes. This deletion spared T cells that expressed low levels of CD8, a result differing from the total deletion of CD8-bearing T cells seen in mice that expressed the original target alloantigen Ld. This evidence indicates that (i) self-peptides bound to thymic major histocompatibility complex molecules can influence the positive selection of thymocytes and (ii) thymocytes with apparently weak interaction with self-major histocompatibility complex antigens can escape clonal deletion. PMID:2117275

  14. CD45-null transgenic mice reveal a positive regulatory role for CD45 in early thymocyte development, in the selection of CD4+CD8+ thymocytes, and B cell maturation

    PubMed Central

    1996-01-01

    The CD45 transmembrane glycoprotein has been shown to be a protein phosphotyrosine phosphatase and to be important in signal transduction in T and B lymphocytes. We have employed gene targeting to create a strain of transgenic mice that completely lacks expression of all isoforms of CD45. The spleens from CD45-null mice contain approximately twice the number of B cells and one fifth the number of T cells found in normal controls. The increase in B cell numbers is due to the specific expansion of two B cell subpopulations that express high levels of immunoglobulin (IgM) staining. T cell development is significantly inhibited in CD45-null animals at two distinct stages. The efficiency of the development of CD4-CD8- thymocytes into CD4+ CD8+ thymocytes is reduced by twofold, subsequently the frequency of successful maturation of the double positive population into mature, single positive thymocytes is reduced by a further four- to fivefold. In addition, we demonstrate that CD45-null thymocytes are severely impaired in their apoptotic response to cross-linking signals via T cell receptor (TCR) in fetal thymic organ culture. In contrast, apoptosis can be induced normally in CD45-null thymocytes by non-TCR- mediated signals. Since both positive and negative selection require signals through the TCR complex, these findings suggest that CD45 is an important regulator of signal transduction via the TCR complex at multiple stages of T cell development. CD45 is absolutely required for the transmission of mitogenic signals via IgM and IgD. By contrast, CD45-null B cells proliferate as well as wild-type cells to CD40- mediated signals. The proliferation of B cells in response to CD38 cross-linking is significantly reduced but not abolished by the CD45- null mutation. We conclude that CD45 is not required at any stage during the generation of mature peripheral B cells, however its loss reveals a previously unrecognized role for CD45 in the regulation of certain subpopulations of

  15. HDAC3 Is Required for the Downregulation of RORγt during Thymocyte Positive Selection.

    PubMed

    Philips, Rachael L; Chen, Meibo W; McWilliams, Douglas C; Belmonte, Paul J; Constans, Megan M; Shapiro, Virginia Smith

    2016-07-15

    To generate functional peripheral T cells, proper gene regulation during T cell development is critical. In this study, we found that histone deacetylase (HDAC) 3 is required for T cell development. T cell development in CD2-icre HDAC3 conditional knockout (cKO) mice (HDAC3-cKO) was blocked at positive selection, resulting in few CD4 and CD8 T cells, and it could not be rescued by a TCR transgene. These single-positive thymocytes failed to upregulate Bcl-2, leading to increased apoptosis. HDAC3-cKO mice failed to downregulate retinoic acid-related orphan receptor (ROR) γt during positive selection, similar to the block in positive selection in RORγt transgenic mice. In the absence of HDAC3, the RORC promoter was hyperacetylated. In the periphery, the few CD4 T cells present were skewed toward RORγt(+) IL-17-producing Th17 cells, leading to inflammatory bowel disease. Positive selection of CD8 single-positive thymocytes was restored in RORγt-KO Bcl-xL transgenic HDAC3-cKO mice, demonstrating that HDAC3 is required at positive selection to downregulate RORγt.

  16. JL1, a novel differentiation antigen of human cortical thymocyte

    PubMed Central

    1993-01-01

    Expression of a novel thymocyte differentiation antigen, JL1, defined by a monoclonal antibody (mAb) developed against human thymocytes showed a specificity for stage II double positive (CD4+CD8+) human cortical thymocytes. This antigen was not expressed at detectable levels on medullary thymocytes, mature peripheral leukocytes, bone marrow cells or on other types of tissues elsewhere in the human body. Immunohistologic analysis revealed that JL1 had a clear pattern of distribution on cortical thymocytes. Immunoprecipitation of 125I- labeled cell lysates from human thymocytes and Molt-4 leukemic cell line with anti-JL1 mAb yielded a 120-130-kD single chain glycoprotein. When immunoprecipitation of cell lysate was done after endoglycosidase F treatment, JL1 antigen was still detected by antibody but the band showed a reduction in apparent molecular mass of approximately 5 kD. This suggests that, although JL1 molecule contains carbohydrate group, this does not form a critical part of the antigenic determinant for anti-JL1 antibody. JL1 antigen appears to be the first double positive, stage-specific differentiation antigen of human thymocyte reported so far. This antigen would be a useful marker for lymphoblastic malignancy of stage II thymocyte origin and it may be involved in the thymocyte education process. PMID:8376947

  17. Yin Yang 1 Promotes Thymocyte Survival by Downregulating p53.

    PubMed

    Chen, Liang; Foreman, Daniel P; Sant'Angelo, Derek B; Krangel, Michael S

    2016-03-15

    Yin Yang 1 (YY1) is a zinc finger protein that functions as a transcriptional activator or repressor and participates in multiple biological processes, including development and tumorigenesis. To investigate the role of YY1 in developing T cells, we used mouse models that depleted YY1 at two distinct stages of thymocyte development. When YY1 was depleted in CD4(-)CD8(-) double-negative thymocytes, development to the CD4(+)CD8(+) double-positive stage was impaired, due to increased apoptosis that prevented expansion of post-β-selection thymocytes. When YY1 was depleted in double-positive thymocytes, they underwent increased cell-autonomous apoptosis in vitro and displayed a shorter lifespan in vivo, as judged by their ability to undergo secondary Vα-to-Jα recombination. Mechanistically, we found that the increased apoptosis in YY1-deficient thymocytes was attributed to overexpression of p53, because concurrent loss of p53 completely rescued the developmental defects of YY1-deficient thymocytes. These results indicated that YY1 functions as a critical regulator of thymocyte survival and that it does so by suppressing the expression of p53.

  18. Growth hormone in the presence of laminin modulates interaction of human thymic epithelial cells and thymocytes in vitro.

    PubMed

    Lins, Marvin Paulo; de Araújo Vieira, Larissa Fernanda; Rosa, Alfredo Aurélio Marinho; Smaniotto, Salete

    2016-09-02

    Several evidences indicate that hormones and neuropeptides function as immunomodulators. Among these, growth hormone (GH) is known to act on the thymic microenvironment, supporting its role in thymocyte differentiation. The aim of this study was to evaluate the effect of GH on human thymocytes and thymic epithelial cells (TEC) in the presence of laminin. GH increased thymocyte adhesion on BSA-coated and further on laminin-coated surfaces. The number of migrating cells in laminin-coated membrane was higher in GH-treated thymocyte group. In both results, VLA-6 expression on thymocytes was constant. Also, treatment with GH enhanced laminin production by TEC after 24 h in culture. However, VLA-6 integrin expression on TEC remained unchanged. Finally, TEC/thymocyte co-culture model demonstrated that GH elevated absolute number of double-negative (CD4(-)CD8(-)) and single-positive CD4(+) and CD8(+) thymocytes. A decrease in cell number was noted in double-positive (CD4(+)CD8(+)) thymocytes. The results of this study demonstrate that GH is capable of enhancing the migratory capacity of human thymocytes in the presence of laminin and promotes modulation of thymocyte subsets after co-culture with TEC.

  19. Altered bone marrow lymphopoiesis and interleukin-6-dependent inhibition of thymocyte differentiation contribute to thymic atrophy during Trypanosoma cruzi infection.

    PubMed

    Carbajosa, Sofía; Gea, Susana; Chillón-Marinas, Carlos; Poveda, Cristina; Maza, Mª Carmen; Fresno, Manuel; Gironès, Núria

    2017-01-28

    Thymic atrophy occurs during infection being associated with apoptosis of double positive (DP) and premature exit of DP and double negative (DN) thymocytes. We observed for the first time that a significant bone marrow aplasia and a decrease in common lymphoid progenitors (CLPs) preceded thymic alterations in mice infected with Trypanosoma cruzi. In addition, depletion of the DN2 stage was previous to the DN1, indicating an alteration in the differentiation from DN1 to DN2 thymocytes. Interestingly, infected mice deficient in IL-6 expression showed higher numbers of DP and CD4+ thymocytes than wild type infected mice, while presenting similar percentages of DN1 thymocytes. Moreover, the drop in late differentiation stages of DN thymocytes was partially abrogated in comparison with wild type littermates. Thus, our results suggest that thymic atrophy involves a drop in CLPs production in bone marrow and IL-6-dependent and independent mechanisms that inhibits the differentiation of DN thymocytes.

  20. Age-related synthesis of glucocorticoids in thymocytes

    SciTech Connect

    Qiao Shengjun Chen Liying; Okret, Sam; Jondal, Mikael

    2008-10-01

    Glucocorticoids (GCs) are primarily synthesized in the adrenal glands but an ectopic production has also been reported in the brain, the gastrointestinal tract and in thymic epithelial cells (TEC). Here we show that thymocytes express genes encoding for all enzymes required for de novo GC synthesis and produce the hormone as demonstrated by both a GC specific reporter assay and a corticosterone specific ELISA assay. Interestingly, GC synthesis is detectable in cells from young mice (4 weeks) and thereafter increases during aging (14-22 weeks) together with an increased gene expression of the rate-limiting enzymes StAR and CYP11A1. Hormone production occurred at a thymocyte differentiation stage characterized by being double positive for the CD4 and CD8 surface markers but was found to be unrelated to CD69 expression, a marker for thymocytes undergoing positive selection. No GC synthesis was found in resting or anti-CD3 activated CD4 and CD8 positive T cells isolated from the spleen. Thymocyte-derived GC had an anti-proliferative effect on a GR-transfected cell line and induced apoptosis in thymocytes. The age- and differentiation stage-related GC synthesis in thymocytes may play a role in the involution process that the thymus gland undergoes.

  1. CBAP promotes thymocyte negative selection by facilitating T-cell receptor proximal signaling

    PubMed Central

    Ho, K-C; Chiang, Y-J; Lai, A C-Y; Liao, N-S; Chang, Y-J; Yang-Yen, H-F; Yen, J J-Y

    2014-01-01

    T-cell receptor (TCR)-transduced signaling is critical to thymocyte development at the CD4/CD8 double-positive stage, but the molecules involved in this process are not yet fully characterized. We previously demonstrated that GM-CSF/IL-3/IL-5 receptor common β-chain-associated protein (CBAP) modulates ZAP70-mediated T-cell migration and adhesion. On the basis of the high expression of CBAP during thymocyte development, we investigated the function of CBAP in thymocyte development using a CBAP knockout mouse. CBAP-deficient mice showed normal early thymocyte development and positive selection. In contrast, several negative selection models (including TCR transgene, superantigen staphylococcal enterotoxin B, and anti-CD3 antibody treatment) revealed an attenuation of TCR-induced thymocyte deletion in CBAP knockout mice. This phenotype correlated with a reduced accumulation of BIM upon TCR crosslinking in CBAP-deficient thymocytes. Loss of CBAP led to reduced TCR-induced phosphorylation of proteins involved in both proximal and distal signaling events, including ZAP70, LAT, PLCγ1, and JNK1/2. Moreover, TCR-induced association of LAT signalosome components was reduced in CBAP-deficient thymocytes. Our data demonstrate that CBAP is a novel component in the TCR signaling pathway and modulates thymocyte apoptosis during negative selection. PMID:25393474

  2. Progressive Changes in CXCR4 Expression That Define Thymocyte Positive Selection Are Dispensable For Both Innate and Conventional αβT-cell Development.

    PubMed

    Lucas, Beth; White, Andrea J; Parnell, Sonia M; Henley, Peter M; Jenkinson, William E; Anderson, Graham

    2017-07-11

    The ordered migration of immature thymocytes through thymic microenvironments generates both adaptive MHC restricted αβT-cells and innate CD1d-restricted iNKT-cells. While several chemokine receptors and ligands control multiple stages of this process, their involvement during early thymocyte development often precludes direct analysis of potential roles during later developmental stages. For example, because of early lethality of CXCR4(-/-) mice, and stage-specific requirements for CXCR4 in thymus colonisation and pre-TCR mediated selection, its role in thymic positive selection is unclear. Here we have examined CXCR4-CXCL12 interactions during the maturation of CD4(+)CD8(+) thymocytes, including downstream stages of iNKT and αβT-cell development. We show CXCL12 expression is a common feature of cortical thymic epithelial cells, indicating widespread availability throughout the cortex. Moreover, CXCR4 expression by CD4(+)CD8(+) pre-selection thymocytes is progressively downregulated following both MHC and CD1d-restricted thymic selection events. However, using CD4(Cre)-mediated deletion to bypass its involvement in CD4(-)CD8(-) thymocyte development, we show CXCR4 is dispensable for the maintenance and intrathymic positioning of CD4(+)CD8(+) thymocytes, and their ability to generate mature αβT-cells and CD1d-restricted iNKT-cells. Collectively, our data define dynamic changes in CXCR4 expression as a marker for intrathymic selection events, and show its role in T-cell development is restricted to pre-CD4(+)CD8(+) stages.

  3. In Silico Modeling of Itk Activation Kinetics in Thymocytes Suggests Competing Positive and Negative IP4 Mediated Feedbacks Increase Robustness

    PubMed Central

    Mukherjee, Sayak; Rigaud, Stephanie; Seok, Sang-Cheol; Fu, Guo; Prochenka, Agnieszka; Dworkin, Michael; Gascoigne, Nicholas R. J.; Vieland, Veronica J.; Sauer, Karsten; Das, Jayajit

    2013-01-01

    The inositol-phosphate messenger inositol(1,3,4,5)tetrakisphosphate (IP4) is essential for thymocyte positive selection by regulating plasma-membrane association of the protein tyrosine kinase Itk downstream of the T cell receptor (TCR). IP4 can act as a soluble analog of the phosphoinositide 3-kinase (PI3K) membrane lipid product phosphatidylinositol(3,4,5)trisphosphate (PIP3). PIP3 recruits signaling proteins such as Itk to cellular membranes by binding to PH and other domains. In thymocytes, low-dose IP4 binding to the Itk PH domain surprisingly promoted and high-dose IP4 inhibited PIP3 binding of Itk PH domains. However, the mechanisms that underlie the regulation of membrane recruitment of Itk by IP4 and PIP3 remain unclear. The distinct Itk PH domain ability to oligomerize is consistent with a cooperative-allosteric mode of IP4 action. However, other possibilities cannot be ruled out due to difficulties in quantitatively measuring the interactions between Itk, IP4 and PIP3, and in generating non-oligomerizing Itk PH domain mutants. This has hindered a full mechanistic understanding of how IP4 controls Itk function. By combining experimentally measured kinetics of PLCγ1 phosphorylation by Itk with in silico modeling of multiple Itk signaling circuits and a maximum entropy (MaxEnt) based computational approach, we show that those in silico models which are most robust against variations of protein and lipid expression levels and kinetic rates at the single cell level share a cooperative-allosteric mode of Itk regulation by IP4 involving oligomeric Itk PH domains at the plasma membrane. This identifies MaxEnt as an excellent tool for quantifying robustness for complex TCR signaling circuits and provides testable predictions to further elucidate a controversial mechanism of PIP3 signaling. PMID:24066087

  4. Serious Infection Risk and Immune Recovery after Double Unit Cord Blood Transplantation without Anti-thymocyte Globulin

    PubMed Central

    Sauter, Craig; Abboud, Michelle; Jia, Xiaoyu; Heller, Glenn; Gonzales, Anne-Marie; Lubin, Marissa; Hawke, Rebecca; Perales, Miguel-Angel; van den Brink, Marcel R.; Giralt, Sergio; Papanicolaou, Genovefa; Scaradavou, Andromachi; Small, Trudy N.; Barker, Juliet N.

    2011-01-01

    Factors contributing to infection risk following cord blood transplantation (CBT) include the use of anti-thymocyte globulin (ATG), prolonged neutropenia, and failure to transfer immunity. To potentially reduce the infection risk we have investigated double unit CBT without ATG, and have evaluated the nature of serious infections in the first year after CBT using this approach. Seventy-two predominantly adult patients were transplanted for hematologic malignancies; 52 patients received myeloablative and 20 had non-myeloablative conditioning. The peak incidence of bacterial infections, fungal infections, or bacterial/ fungal pneumonias was in the first 30 days post-transplant and affected 32%, 14%, and 10% of patients, respectively. Three such infections contributed to early mortality. The peak incidence of viral infections was 31-60 days post-transplant affecting 30% of patients. Cytomegalovirus (CMV) was the most common viral infection. CMV infections prior to day 120 (n = 23) had no relationship with graft-versus-host disease (GVHD), whereas CMV infections after day 120 (n = 5), and all Epstein-Barr virus viremia (EBV, n = 5) and adeno-viral enteritis (n = 2) occurred exclusively in the context of GVHD therapy or corticosteroid use for another indication. Viral infections had the highest lethality: 2 were a direct cause of death and 3 contributed to death. Patients exhibited steady immune recovery achieving a median CD3+4+ T-cell count > 200 cells/microL by day 120, and after day 120 there were no infection-related deaths. Our results suggest that double unit CBT without ATG is associated with prompt T-cell recovery, and unlike CBT incorporating ATG, infection is rarely a primary cause of death. However, CBT without ATG is associated with a significant risk of GVHD and serious infections remain a burden, especially in the setting of GVHD. New strategies are required to further reduce infectious complications after CBT and will require earlier neutrophil recovery

  5. Thymic education curtailed: defective immune responses in nude rats reconstituted with immature thymocyte subsets.

    PubMed

    Yang, C P; Bell, E B

    1994-04-01

    We have studied the ability of thymocyte subsets from allotype marked donors to populate athymic nude rats with T cells and to restore immune responsiveness. Following adoptive transfer, CD4-CD8- double-negative (DN) thymocytes (lymphoid precursor cells) or the CD4+CD8+ double-positive (DP) subset (intermediate thymocytes) or CD4+CD8- single-positive (CD4 SP) cells (mature thymocytes) each generated a permanent population of CD4+ progeny in syngeneic nude recipients. DN and DP thymocytes also produced small numbers of CD8+ cells; there was no evidence of a CD4-CD8- or CD4+CD8+ donor cell population. CD4 SP thymocytes conferred T cell functions [graft-versus-host (GVH) responses, allograft rejection and thymus-dependent antibody responses] on nude rats that were almost indistinguishable from those conferred by mature peripheral recirculating CD4 T cells. Transfer of DP thymocytes extended the life-span of the immunoincompetent nudes and produced CD4+ progeny with near normal GVH responsiveness. However, DP-derived CD4+ cells were deficient at inducing allograft rejection and provided little or no help for antibody synthesis. The CD4+ progeny of DN thymocytes did not prolong the survival of nude recipients, gave reduced GVH reactivity, showed almost no capacity to initiate skin allograft rejection and failed to help B cells produce antibody. The results suggest that intrathymic development proceeds stepwise; each stage is accompanied by acquisition of additional properties that are reflected by T cell responses in the periphery. Thymic education does not become complete until the SP stage is reached when thymocytes become fully independent of the thymic microenvironment.

  6. A threshold level of NFATc1 activity facilitates thymocyte differentiation and opposes notch-driven leukaemia development

    PubMed Central

    Klein-Hessling, Stefan; Rudolf, Ronald; Muhammad, Khalid; Knobeloch, Klaus-Peter; Maqbool, Muhammad Ahmad; Cauchy, Pierre; Andrau, Jean-Christophe; Avots, Andris; Talora, Claudio; Ellenrieder, Volker; Screpanti, Isabella; Serfling, Edgar; Patra, Amiya Kumar

    2016-01-01

    NFATc1 plays a critical role in double-negative thymocyte survival and differentiation. However, the signals that regulate Nfatc1 expression are incompletely characterized. Here we show a developmental stage-specific differential expression pattern of Nfatc1 driven by the distal (P1) or proximal (P2) promoters in thymocytes. Whereas, preTCR-negative thymocytes exhibit only P2 promoter-derived Nfatc1β expression, preTCR-positive thymocytes express both Nfatc1β and P1 promoter-derived Nfatc1α transcripts. Inducing NFATc1α activity from P1 promoter in preTCR-negative thymocytes, in addition to the NFATc1β from P2 promoter impairs thymocyte development resulting in severe T-cell lymphopenia. In addition, we show that NFATc1 activity suppresses the B-lineage potential of immature thymocytes, and consolidates their differentiation to T cells. Further, in the pTCR-positive DN3 cells, a threshold level of NFATc1 activity is vital in facilitating T-cell differentiation and to prevent Notch3-induced T-acute lymphoblastic leukaemia. Altogether, our results show NFATc1 activity is crucial in determining the T-cell fate of thymocytes. PMID:27312418

  7. Mutation of the phospholipase C-γ1–binding site of LAT affects both positive and negative thymocyte selection

    PubMed Central

    Sommers, Connie L.; Lee, Jan; Steiner, Kevin L.; Gurson, Jordan M.; DePersis, Corinne L.; El-Khoury, Dalal; Fuller, Claudette L.; Shores, Elizabeth W.; Love, Paul E.; Samelson, Lawrence E.

    2005-01-01

    Linker for activation of T cells (LAT) is a scaffolding adaptor protein that is critical for T cell development and function. A mutation of LAT (Y136F) that disrupts phospholipase C-γ1 activation and subsequent calcium influx causes a partial block in T cell development and leads to a severe lymphoproliferative disease in homozygous knock-in mice. One possible contribution to the fatal disease of LAT Y136F knock-in mice could be from autoreactive T cells generated in these mice because of altered thymocyte selection. To examine the impact of the LAT Y136F mutation on thymocyte positive and negative selection, we bred this mutation onto the HY T cell receptor (TCR) transgenic, recombination activating gene-2 knockout background. Female mice with this genotype showed a severe defect in positive selection, whereas male mice exhibited a phenotype resembling positive selection (i.e., development and survival of CD8hi HY TCR-specific T cells) instead of negative selection. These results support the hypothesis that in non-TCR transgenic, LAT Y136F knock-in mice, altered thymocyte selection leads to the survival and proliferation of autoreactive T cells that would otherwise be negatively selected in the thymus. PMID:15795236

  8. Altered thymocyte and T cell development in neonatal mice with hyperoxia-induced lung injury.

    PubMed

    Angusamy, Sowmya; Mansour, Tamer; Abdulmageed, Mohammed; Han, Rachel; Schutte, Brian C; LaPres, John; Harkema, Jack R; Omar, Said A

    2017-08-19

    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. 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. 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. 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.

  9. Constitutive activation of integrin alpha 4 beta 1 defines a unique stage of human thymocyte development

    PubMed Central

    1994-01-01

    Our understanding of thymocyte development and of the positive and negative selection events involved in shaping the repertoire of mature T lymphocytes has been greatly facilitated by the use of transgenic and gene knockout animals. Much less is known about the factors that control the homing and population of the thymus by T cell precursors and the subsequent migration of developing thymocytes through the thymic architecture. As the integrins represent a candidate group of cell surface receptors that may regulate thymocyte development, we have analyzed the expression and function of alpha 4 beta 1 and alpha 5 beta 1 on human thymocytes. A major portion of double positive (CD4+ CD8+) human thymocytes express alpha 4 beta 1 in a constitutively active form and adhere to fibronectin and vascular cell adhesion molecule 1. alpha 4 beta 1 expression is similar on adherent and nonadherent populations, thus, activity reflects the receptor state and not simple expression. The adherent cells are immature, expressing high levels of CD4/CD8 and low levels of CD3 and CD69. In contrast, nonadherent cells possess the phenotype of thymocytes after positive selection, expressing intermediate levels of CD4 and/or CD8 and high levels of CD3 and CD69. The adherent population fails to respond to activation with anti-CD3 and fibronectin, whereas nonadherents exhibit an alpha 5 beta 1- dependent proliferation. Differential regulation of alpha 4 beta 1 and alpha 5 beta 1 receptors may provide a mechanism controlling cellular traffic, differentiation, and positive selection of thymocytes. PMID:8163937

  10. Pre-TCR signaling and CD8 gene bivalent chromatin resolution during thymocyte development.

    PubMed

    Harker, Nicola; Garefalaki, Anna; Menzel, Ursula; Ktistaki, Eleni; Naito, Taku; Georgopoulos, Katia; Kioussis, Dimitris

    2011-06-01

    The CD8 gene is silent in CD4(-)CD8(-) double-negative thymocytes, expressed in CD4(+)CD8(+) double-positive cells, and silenced in cells committing to the CD4(+) single-positive (SP) lineage, remaining active in the CD8(+) SP lineage. In this study, we show that the chromatin of the CD8 locus is remodeled in C57BL/6 and B6/J Rag1(-/-) MOM double-negative thymocytes as indicated by DNaseI hypersensitivity and widespread bivalent chromatin marks. Pre-TCR signaling coincides with chromatin bivalency resolution into monovalent activating modifications in double-positive and CD8 SP cells. Shortly after commitment to CD4 SP cell lineage, monovalent repressive characteristics and chromatin inaccessibility are established. Differential binding of Ikaros, NuRD, and heterochromatin protein 1α on the locus during these processes may participate in the complex regulation of CD8.

  11. Malnutrition alters the rates of apoptosis in splenocytes and thymocyte subpopulations of rats

    PubMed Central

    Ortiz, R; Cortés, L; Cortés, E; Medina, H

    2009-01-01

    Malnutrition continues to be a major public health problem throughout the developing world. Nutritional deficiencies may be the most common cause of secondary immunodeficiency states in humans. It has been suggested that nutritional imbalances can induce apoptosis in a variety of cell types. The purpose of this study was to examine the effect of severe malnutrition on cell subsets and the frequency of spontaneous and/or dexamethasone-induced cell death in vivo in the thymus and spleen from severely malnourished, lactating rats. Apoptosis frequency was estimated by flow cytometry using annexin-V and terminal transferase-mediated dUTP nick-end labelling assay assays. The results obtained in the present study indicate that malnutrition is associated with a significant increase of spontaneously apoptotic cells in the thymus (9·8-fold) and spleen (2·4-fold). Increase in apoptosis was associated largely with CD4+CD8+ double-positive thymocytes. Unexpectedly, similar frequencies of spontaneous apoptosis of these cells were found in both well-nourished and malnourished rats. In contrast, consistent increases in the apoptosis of CD4−CD8− double-negative thymocytes were observed in malnourished rats. In addition, single-positive CD8+ and single-positive CD4+ thymocytes had higher frequencies of apoptosis in malnourished rats. The frequency of total dexamethasone-induced apoptosis was found to be similar in both groups of animals. Nevertheless, in malnourished dexamethasone-treated animals, the percentage of apoptotic double-negative thymocytes was significantly higher than in well-nourished animals, while the rate of apoptosis was lower among double-positive cells. In general, the thymus appears more sensitive to the effects of malnutrition and dexamethasone than the spleen. Furthermore, double-negative thymocytes appear to be the most affected. PMID:19076833

  12. Malnutrition alters the rates of apoptosis in splenocytes and thymocyte subpopulations of rats.

    PubMed

    Ortiz, R; Cortés, L; Cortés, E; Medina, H

    2009-01-01

    Malnutrition continues to be a major public health problem throughout the developing world. Nutritional deficiencies may be the most common cause of secondary immunodeficiency states in humans. It has been suggested that nutritional imbalances can induce apoptosis in a variety of cell types. The purpose of this study was to examine the effect of severe malnutrition on cell subsets and the frequency of spontaneous and/or dexamethasone-induced cell death in vivo in the thymus and spleen from severely malnourished, lactating rats. Apoptosis frequency was estimated by flow cytometry using annexin-V and terminal transferase-mediated dUTP nick-end labelling assay assays. The results obtained in the present study indicate that malnutrition is associated with a significant increase of spontaneously apoptotic cells in the thymus (9.8-fold) and spleen (2.4-fold). Increase in apoptosis was associated largely with CD4(+)CD8(+) double-positive thymocytes. Unexpectedly, similar frequencies of spontaneous apoptosis of these cells were found in both well-nourished and malnourished rats. In contrast, consistent increases in the apoptosis of CD4(-)CD8(-) double-negative thymocytes were observed in malnourished rats. In addition, single-positive CD8(+) and single-positive CD4(+) thymocytes had higher frequencies of apoptosis in malnourished rats. The frequency of total dexamethasone-induced apoptosis was found to be similar in both groups of animals. Nevertheless, in malnourished dexamethasone-treated animals, the percentage of apoptotic double-negative thymocytes was significantly higher than in well-nourished animals, while the rate of apoptosis was lower among double-positive cells. In general, the thymus appears more sensitive to the effects of malnutrition and dexamethasone than the spleen. Furthermore, double-negative thymocytes appear to be the most affected.

  13. False Position, Double False Position and Cramer's Rule

    ERIC Educational Resources Information Center

    Boman, Eugene

    2009-01-01

    We state and prove the methods of False Position (Regula Falsa) and Double False Position (Regula Duorum Falsorum). The history of both is traced from ancient Egypt and China through the work of Fibonacci, ending with a connection between Double False Position and Cramer's Rule.

  14. False Position, Double False Position and Cramer's Rule

    ERIC Educational Resources Information Center

    Boman, Eugene

    2009-01-01

    We state and prove the methods of False Position (Regula Falsa) and Double False Position (Regula Duorum Falsorum). The history of both is traced from ancient Egypt and China through the work of Fibonacci, ending with a connection between Double False Position and Cramer's Rule.

  15. Apoptosis of nur77/N10-transgenic thymocytes involves the Fas/Fas ligand pathway.

    PubMed Central

    Weih, F; Ryseck, R P; Chen, L; Bravo, R

    1996-01-01

    The orphan nuclear receptor Nur77/N10 has recently been demonstrated to be involved in apoptosis of T cell hybridomas. We report here that chronic expression of Nur77/N10 in thymocytes of transgenic mice results in a dramatic reduction of CD4+CD8+ double-positive as well as CD4+CD8- and CD4-CD8+ single-positive cell populations due to an early onset of apoptosis. CD4-CD8- double-negative and CD25+ precursor cells, however, are unaffected. Moreover, nur77/N10-transgenic thymocytes show increased expression of Fas ligand (FasL), while the levels of the Fas receptor (Fas) are not increased. The mouse spontaneous mutant gld (generalized lymphoproliferative disease) carries a point mutation in the extracellular domain of the FasL gene that abolishes the ability of FasL to bind to Fas. Thymuses from nur77/N10-transgenic mice on a gld/gld background have increased cellularity and an almost normal profile of thymocyte subpopulations. Our results demonstrate that one pathway of apoptosis triggered by Nur77/N10 in double-positive thymocytes occurs through the upregulation of FasL expression resulting in increased signaling through Fas. Images Fig. 1 Fig. 2 Fig. 4 PMID:8643610

  16. Ternary complex factors SAP-1 and Elk-1, but not net, are functionally equivalent in thymocyte development.

    PubMed

    Costello, Patrick; Nicolas, Robert; Willoughby, Jane; Wasylyk, Bohdan; Nordheim, Alfred; Treisman, Richard

    2010-07-15

    The ternary complex factors (TCFs; SAP-1, Elk-1, and Net) are serum response factor cofactors that share many functional properties and are coexpressed in many tissues. SAP-1, the predominant thymus TCF, is required for thymocyte positive selection. In this study, we assessed whether the different TCFs are functionally equivalent. Elk-1 deletion, but not the hypomorphic Net(delta) mutation, exacerbated the SAP-1 positive selection phenotype, but triply deficient thymocytes were no more defective than SAP-1(-/-) Elk-1(-/-) cells. Inactivation of the other TCFs did not affect SAP-1-independent processes, including beta-selection, regulatory T cell selection, and negative selection, although reduced marginal zone B cells were observed in SAP-1(-/-) Elk-1(-/-) animals. Ectopic expression of Elk-1, but not Net, rescued positive selection of SAP-1(-/-) thymocytes; thus, SAP-1 and Elk-1 are functionally equivalent in this system, and the SAP-1 null selection phenotype reflects only its high expression in the thymus. Array analysis of TCR-stimulated double-positive cells identified SAP-1-dependent inducible genes whose transcription was further impaired in SAP-1(-/-) Elk-1(-/-) cells; thus, these genes, which include Egr-1 and Egr-2, represent candidate mediators of positive selection. Chromatin immunoprecipitation revealed subtly different promoter targeting between the different TCFs. Ectopic expression of Egr-1 restored positive selection in SAP-1 null thymocytes, establishing it (and possibly other Egr family members) as the major effector for ERK-SAP-1 signaling in thymocyte positive selection.

  17. Loss of Gαi proteins impairs thymocyte development, disrupts T-cell trafficking, and leads to an expanded population of splenic CD4(+)PD-1(+)CXCR5(+/-) T-cells.

    PubMed

    Hwang, Il-Young; Harrison, Kathleen; Park, Chung; Kehrl, John H

    2017-06-23

    Thymocyte and T cell trafficking relies on signals initiated by G-protein coupled receptors. To address the importance of the G-proteins Gαi2 and Gαi3 in thymocyte and T cell function, we developed several mouse models. Gαi2 deficiency in hematopoietic progenitors led to a small thymus, a double negative (DN)1/DN2 thymocyte transition block, and an accumulation of mature single positive (SP) thymocytes. Loss at the double positive (DP) stage of thymocyte development caused an increase in mature cells within the thymus. In both models an abnormal distribution of memory and naïve CD4 T cells occurred, and peripheral CD4 and CD8 T cells had reduced chemoattractant responses. The loss of Gαi3 had no discernable impact, however the lack of both G-proteins commencing at the DP stage caused a severe T cell phenotype. These mice lacked a thymic medullary region, exhibited thymocyte retention, had a peripheral T cell deficiency, and lacked T cell chemoattractant responses. Yet a noteworthy population of CD4(+)PD-1(+)CXCR5(+/-) cells resided in the spleen of these mice likely due to a loss of regulatory T cell function. Our results delineate a role for Gαi2 in early thymocyte development and for Gαi2/3 in multiple aspects of T cell biology.

  18. Phenylhydroquinone induces loss of thymocytes through cell cycle arrest and apoptosis elevation in p53-dependent pathway.

    PubMed

    Nakata, Yuichiro; Nishi, Kosuke; Nishimoto, Sogo; Sugahara, Takuya

    2013-01-01

    ortho-Phenylphenol has been employed in post-harvest treatment of citrus fruits. Although o-phenylphenol has been reported to cause carcinomas in the urinary tract in rats, toxicity to the immune organs is still unknown. Herein, we report that administration of o-phenylphenol induces thymic atrophy and loss of thymocytes in female BALB/c mice. The influence seems to result from inhibition of the thymocyte development, because increased and decreased populations of the CD4⁻ CD8⁻ double-negative and CD4⁺ CD8⁺ double-positive thymocytes were observed in the o-phenylphenol-administered mice, respectively. ortho-Phenylphenol is metabolized to phenylhydroquinone by cytochrome P450 monooxygenases. Phenylhydroquinone made cell cycle of thymocytes to be arrested through reduced expression of the genes associated with G₂/M phase and through phosphorylation of p53 at Ser15. Phosphorylation of p53 at Ser15 was upregulated by activation of not only ATR but also Erk1/2 and p38, leading to increase of apoptosis. Gene expression of cytochrome P450 1A1 (CYP1A1) was promoted in thymocytes from the o-phenylphenol-administered mice. Overall, our results suggest that o-phenylphenol induces CYP1A1 expression and is metabolized into phenylhydroquinone by the expressed CYP1A1 in thymocytes. The produced phenylhydroquinone in turn induces inhibition of thymocyte development through cell cycle arrest and apoptosis in the p53-dependent pathway.

  19. Dynamics of ATP-induced Calcium Signaling in Single Mouse Thymocytes

    PubMed Central

    Ross, Paul E.; Ehring, George R.; Cahalan, Michael D.

    1997-01-01

    Extracellular ATP (ATPo) elicits a robust change in the concentration of intracellular Ca2+ ([Ca2+]i) in fura-2–loaded mouse thymocytes. Most thymocytes (60%) exposed to ATPo exhibited a biphasic rise in [Ca2+]i; [Ca2+]i rose slowly at first to a mean value of 260 nM after 163 s and then increased rapidly to a peak level of 735 nM. In many cells, a declining plateau, which lasted for more than 10 min, followed the crest in [Ca2+]i. Experiments performed in the absence of extracellular [Ca2+]o abolished the rise in thymocyte [Ca2+]i, indicating that Ca2+ influx, rather than the release of stored Ca2+, is stimulated by ATPo. ATPo- mediated Ca2+ influx was potentiated as the [Mg2+]o was reduced, confirming that ATP4− is the active agonist form. In the absence of Mg2+o, 3′-O-(4-benzoyl)benzoyl-ATP (BzATP) proved to be the most effective agonist of those tested. The rank order of potency for adenine nucleotides was BzATP4−>ATP4−>MgATP2−>ADP3−, suggesting purinoreceptors of the P2X7/P2Z class mediate the ATPo response. Phenotyping experiments illustrate that both immature (CD4−CD8−, CD4+CD8+) and mature (CD4+CD8−, CD4−CD8+) thymocyte populations respond to ATP. Further separation of the double-positive population by size revealed that the ATPo-mediated [Ca2+]i response was much more pronounced in large (actively dividing) than in small (terminally differentiated) CD4+CD8+ thymocytes. We conclude that thymocytes vary in sensitivity to ATPo depending upon the degree of maturation and suggest that ATPo may be involved in processes that control cellular differentiation within the thymus. PMID:9281578

  20. Interferon-regulatory factors during development of CD4 and CD8 thymocytes.

    PubMed Central

    Simon, A K; Desrois, M; Schmitt-Verhulst, A M

    1997-01-01

    Selection events in the thymus occur at the double-positive CD4+ CD8+ (DP) developmental stage leading either to further differentiation of the CD4+ and CD8+ lineages or to deletion. The interferon-regulatory factor IRF-1 has been implicated in signalling for T-cell death and also in CD8+ thymic differentiation. IRF-1 is an activator and IRF-2 a repressor of gene transcription regulated by type 1 interferons (IFN). To evaluate the role of IRF-1 and IRF-2 in the differentiation of CD4 and CD8 thymocytes, we analysed their DNA-binding activity before and after antigenic stimulation at different stages of thymic development and in peripheral T cells. Unseparated, double-positive and single-positive thymocytes as well as peripheral T lymphocytes from mice transgenic (tg) for a T-cell receptor (TCR), restricted either by major histocompatibility complex class I or class II, were stimulated by their nominal antigen. Our results demonstrate that the DNA-binding activity of IRF-2 and, weakly, that of IRF-1 are inducible in total thymocytes in response to antigen. There is no induction of IRF-1/IRF-2 binding activity at the double-positive stage of thymic development in the MHC class II-restricted model whereas in the MHC class I-restricted model IRF-1/IRF-2 activity is induced weakly. At the single-positive stage, antigen induces the IRF-1/IRF-2 DNA binding in both CD4+ and CD8+ thymocytes, but not in mature lymphocytes from the periphery. This pattern of expression suggests that IRF-1/IRF-2 binding activities resulting from antigen stimulation are developmentally regulated. No evidence for a selective role of IRF-1 in the development of the CD8+ lineage was found, however. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:9301521

  1. A Double-Blind, Double-Dummy, Flexible-Design Randomized Multicenter Trial: Early Safety of Single- Versus Divided-Dose Rabbit Anti-Thymocyte Globulin Induction in Renal Transplantation.

    PubMed

    Stevens, R B; Wrenshall, L E; Miles, C D; Farney, A C; Jie, T; Sandoz, J P; Rigley, T H; Osama Gaber, A

    2016-06-01

    A previous nonblinded, randomized, single-center renal transplantation trial of single-dose rabbit anti-thymocyte globulin induction (SD-rATG) showed improved efficacy compared with conventional divided-dose (DD-rATG) administration. The present multicenter, double-blind/double-dummy STAT trial (Single dose vs. Traditional Administration of Thymoglobulin) evaluated SD-rATG versus DD-rATG induction for noninferiority in early (7-day) safety and tolerability. Ninety-five patients (randomized 1:1) received 6 mg/kg SD-rATG or 1.5 mg/kg/dose DD-rATG, with tacrolimus-mycophenolate maintenance immunosuppression. The primary end point was a composite of fever, hypoxia, hypotension, cardiac complications, and delayed graft function. Secondary end points included 12-month patient survival, graft survival, and rejection. Target enrollment was 165 patients with an interim analysis scheduled after 80 patients. Interim analysis showed primary end point noninferiority of SD-rATG induction (p = 0.6), and a conditional probability of <1.73% of continued enrollment producing a significant difference (futility analysis), leading to early trial termination. Final analysis (95 patients) showed no differences in occurrence of primary end point events (p = 0.58) or patients with no, one, or more than one event (p = 0.81), or rejection, graft, or patient survival (p = 0.78, 0.47, and 0.35, respectively). In this rigorously blinded trial in adult renal transplantation, we have shown SD-rATG induction to be noninferior to DD-rATG induction in early tolerability and equivalent in 12-month safety. (Clinical Trials.gov #NCT00906204.).

  2. CAML is a p56Lck-interacting protein that is required for thymocyte development.

    PubMed

    Tran, David D; Edgar, Contessa E; Heckman, Karin L; Sutor, Shari L; Huntoon, Catherine J; van Deursen, Jan; McKean, David L; Bram, Richard J

    2005-08-01

    Calcium modulating cyclophilin ligand (CAML) is a ubiquitously expressed protein implicated in T cell signaling, although its mechanism and physiologic role in the immune system are unknown. We show here that CAML is essential for peripheral T cell development. Inactivation of CAML in mouse thymocytes lowered the numbers of double-positive and single-positive thymocytes, concomitant with reduced positive and enhanced negative selection. We found that CAML interacts with p56Lck and appears to regulate subcellular localization of the kinase in both resting and T cell receptor (TCR)-stimulated cells. CAML-deficient cells displayed enhanced p56lck and ZAP-70 phosphorylation and increased IL2 production and cell death after TCR stimulation, suggesting that CAML may act as a negative regulator of p56lck. Our data establish a novel role for CAML as an essential mediator of T cell survival during thymopoiesis and indicate that its loss deregulates p56Lck signaling.

  3. Selective manipulation of the human T-cell receptor repertoire expressed by thymocytes in organ culture.

    PubMed Central

    Merkenschlager, M; Fisher, A G

    1992-01-01

    A recently described organ culture system for human thymocytes is shown to support the generation of a diverse T-cell receptor repertoire in vitro: thymocytes of the alpha beta lineage, including representatives of the V beta families 5.2/5.3, 6.7, and 8, accounted for the majority of T-cell receptor-positive cells throughout a 3-week culture period. Thymocytes bearing gamma delta receptors were also identified, particularly among the CD4 CD8 double-negative subset. The T-cell receptor repertoire expressed in organ culture responded to experimental manipulation with staphylococcal enterotoxins. Staphylococcal enterotoxin D (a powerful activator of human peripheral T cells expressing V beta 5.2/5.3 receptors) caused a marked reduction of V beta 5.2/5.3 expression, as determined with the V beta-specific antibody 42/1C1. Evidence is presented that this loss of V beta 5.2/5.3 expression resulted from the selective deletion of activated thymocytes by apoptosis, in concert with T-cell receptor modulation. These effects of staphylococcal enterotoxin D were specific (since staphylococcal enterotoxin E did not influence V beta 5.2/5.3 expression) and V beta-selective (since expression of V beta 6.7 remained unaffected by staphylococcal enterotoxin D). On the basis of these observations, we suggest that thymic organ culture provides a powerful approach to study the generation of the human T-cell repertoire. Images PMID:1584760

  4. Coreceptor gene imprinting governs thymocyte lineage fate

    PubMed Central

    Adoro, Stanley; McCaughtry, Thomas; Erman, Batu; Alag, Amala; Van Laethem, François; Park, Jung-Hyun; Tai, Xuguang; Kimura, Motoko; Wang, Lie; Grinberg, Alex; Kubo, Masato; Bosselut, Remy; Love, Paul; Singer, Alfred

    2012-01-01

    Immature thymocytes are bipotential cells that are signalled during positive selection to become either helper- or cytotoxic-lineage T cells. By tracking expression of lineage determining transcription factors during positive selection, we now report that the Cd8 coreceptor gene locus co-opts any coreceptor protein encoded within it to induce thymocytes to express the cytotoxic-lineage factor Runx3 and to adopt the cytotoxic-lineage fate, findings we refer to as ‘coreceptor gene imprinting'. Specifically, encoding CD4 proteins in the endogenous Cd8 gene locus caused major histocompatibility complex class II-specific thymocytes to express Runx3 during positive selection and to differentiate into CD4+ cytotoxic-lineage T cells. Our findings further indicate that coreceptor gene imprinting derives from the dynamic regulation of specific cis Cd8 gene enhancer elements by positive selection signals in the thymus. Thus, for coreceptor-dependent thymocytes, lineage fate is determined by Cd4 and Cd8 coreceptor gene loci and not by the specificity of T-cell antigen receptor/coreceptor signalling. This study identifies coreceptor gene imprinting as a critical determinant of lineage fate determination in the thymus. PMID:22036949

  5. Positive and negative selection of the T cell repertoire: what thymocytes see and don't see

    PubMed Central

    Klein, Ludger; Kyewski, Bruno; Allen, Paul M.; Hogquist, Kristin A.

    2016-01-01

    The fate of developing T cells is specified by interactions of their antigen receptor with self-peptide/MHC complexes displayed by thymic antigen presenting cells (APCs). Various thymic APCs subsets are strategically positioned in particular thymic microenvironments and orchestrate the selection of a functional and self-tolerant T cell repertoire. Here, we will review the different strategies that these APCs employ to sample and process self-antigens and thereby generate partly unique, ‘idiosyncratic’ peptide/MHC ligandomes. We will discuss how the particular composition of these APC-subset-specific peptide/MHC ligandomes not only shapes the T cell repertoire in the thymus, but may also indelibly imprint the behavior of mature T cells in the periphery. PMID:24830344

  6. The onset of Fas expression parallels the acquisition of CD8 and CD4 in fetal and adult alpha beta thymocytes.

    PubMed

    Andjelić, S; Drappa, J; Lacy, E; Elkon, K B; Nikolić-Zugić, J

    1994-01-01

    Fas is an apoptosis-related cell surface molecule whose defective transcription results in the lpr defect and autoimmunity. Recent analysis of Fas mRNA and protein expression in normal mice showed high expression in the thymus, on activated T cells, and on 5-10% of peripheral T cells. To investigate the role of Fas in the thymus, we analyzed its expression in fetal and adult thymocyte subsets. Fas was not expressed on fetal nor adult CD8-CD4- (double-negative, DN) T cell precursors. The earliest precursors that expressed low levels of FAS were the immediate precursors of DP thymocytes that bear the CD44-CD25-CD8loCD4loTCRlo phenotype. Other DN cells that expressed Fas appeared to be either non-T cells or mature alpha beta + DN thymocytes. The onset of Fas expression followed the onset of expression of CD8 and CD4 and Fas expression reached its peak in CD8+CD4+ double-positive (DP) thymocytes. Both single-positive (SP) subsets were largely Fas+ (CD8 SP < CD4 SP) but expressed lower levels of Fas than DP cells. However, a majority (> 60%) of the most mature HSA(lo) SP cells (2-5% of all SP thymocytes) were Fas- and the remainder of the HSA(lo) SP cells was Fas(lo). We observed two main differences between Fas expression on fetal versus adult thymocytes. First, up to 90% of fetal gamma delta + DN cells expressed high levels of Fas, in contrast to the very low expression (< 7% Fas+ cells) among adult gamma delta + thymocytes. Second, whereas virtually all adult DP cells were Fas+, up to 75% of fetal day 16 DP cells were Fas-.(ABSTRACT TRUNCATED AT 250 WORDS)

  7. CD6 modulates thymocyte selection and peripheral T cell homeostasis

    PubMed Central

    Consuegra-Fernández, Marta; Girard, Laura; Aranda, Fernando; Martínez, Vanesa-Gabriela; Sarukhan, Adelaida; Malissen, Marie

    2016-01-01

    The CD6 glycoprotein is a lymphocyte surface receptor putatively involved in T cell development and activation. CD6 facilitates adhesion between T cells and antigen-presenting cells through its interaction with CD166/ALCAM (activated leukocyte cell adhesion molecule), and physically associates with the T cell receptor (TCR) at the center of the immunological synapse. However, its precise role during thymocyte development and peripheral T cell immune responses remains to be defined. Here, we analyze the in vivo consequences of CD6 deficiency. CD6−/− thymi showed a reduction in both CD4+ and CD8+ single-positive subsets, and double-positive thymocytes exhibited increased Ca2+ mobilization to TCR cross-linking in vitro. Bone marrow chimera experiments revealed a T cell–autonomous selective disadvantage of CD6−/− T cells during development. The analysis of TCR-transgenic mice (OT-I and Marilyn) confirmed that abnormal T cell selection events occur in the absence of CD6. CD6−/− mice displayed increased frequencies of antigen-experienced peripheral T cells generated under certain levels of TCR signal strength or co-stimulation, such as effector/memory (CD4+TEM and CD8+TCM) and regulatory (T reg) T cells. The suppressive activity of CD6−/− T reg cells was diminished, and CD6−/− mice presented an exacerbated autoimmune response to collagen. Collectively, these data indicate that CD6 modulates the threshold for thymocyte selection and the generation and/or function of several peripheral T cell subpopulations, including T reg cells. PMID:27377588

  8. CCR9 AND CCR7 are overexpressed in CD4(-) CD8(-) thymocytes of myasthenia gravis patients.

    PubMed

    Li, Qianru; Liu, Pingping; Xuan, Xiaoyan; Zhang, Junfeng; Zhang, Yun; Zhu, Zhengkun; Gao, Feng; Zhang, Qingyong; Du, Ying

    2017-01-01

    Chemokine CC motif receptors 9 and 7 (CCR9 and CCR7) play a major role in the migration of T-cell precursors to the thymus to initiate T thymopoiesis. However, their role in development of T-cells in myasthenia gravis (MG) patients has not been fully elucidated. Expression and distribution of CCR9(+) and CCR7(+) cells were detected by flow cytometry and immunofluorescence. Real-time polymerase chain reaction was used to check the adhesion molecules on CD4(-) CD8(-) double-negative (DN) thymocytes. CCR9 and CCR7 expression by DN thymocytes increased in the MG thymus; the levels of CCR9, CCR7, interleukin-7R mRNA increased, and CXCR4 levels decreased compared with levels in the non-MG thymus. More CCR7 and CCR9 double-positive (DP) thymocytes were gathered near the subcapsular region in MG thymus. Enhanced expression of CCR9 and CCR7 may complicate the differentiation of DP thymocytes from the DN stage in MG thymus. Muscle Nerve, 2016 Muscle Nerve 55: 84-90, 2017. © 2015 Wiley Periodicals, Inc.

  9. Interleukin-7 co-ordinates proliferation, differentiation and Tcra recombination during thymocyte β-selection

    PubMed Central

    Boudil, Amine; Matei, Irina R.; Shih, Han-Yu; Bogdanoski, Goce; Yuan, Julie S.; Chang, Stephen G.; Montpellier, Bertrand; Kowalski, Paul E.; Voisin, Veronique; Bashir, Shaheena; Bader, Gary D.; Krangel, Michael S.; Guidos, Cynthia J.

    2015-01-01

    pre-T cell receptor (TCR) and Notch signaling induce transient self-renewal or “β-selection” of TCRβ+ CD4 CD8 double-negative-3 (DN3) and DN4 progenitors that differentiate into CD4 CD8 double positive (DP) thymocytes which then rearrange Tcra. Interleukin-7 (IL-7) promotes Bcl2-dependent survival of TCRβ− DN thymocytes, but IL-7 functions during β-selection remain unclear. Here, we show that IL-7 signals TCRβ+ DN3 and DN4 thymocytes to upregulate genes involved in cell growth and represses Bcl6. Accordingly, IL-7-deficient DN4 cells lacked trophic receptors and did not proliferate, but rearranged Tcra prematurely and differentiated rapidly. Bcl6 deletion, but not BCL2 over-expression, partially restored DN4 self-renewal in the absence of IL-7. Thus, IL-7 critically collaborates with pre-TCR and Notch signaling to coordinate proliferation, differentiation and Tcra recombination during β-selection. PMID:25729925

  10. Commitment of Immature CD4+8+ Thymocytes to the CD4 Lineage Requires CD3 Signaling but Does Not Require Expression of Clonotypic T Cell Receptor (TCR) Chains

    PubMed Central

    Suzuki, Harumi; Shinkai, Yoichi; Granger, Lawrence G.; Alt, Frederick W.; Love, Paul E.; Singer, Alfred

    1997-01-01

    As a consequence of positive selection in the thymus, immature CD4+8+ double-positive, [DP] thymocytes selectively terminate synthesis of one coreceptor molecule and, as a result, differentiate into either CD4+ or CD8+ T cells. The decision by individual DP thymocytes to terminate synthesis of one or the other coreceptor molecule is referred to as lineage commitment. Previously, we reported that the intrathymic signals that induced commitment to the CD4 versus CD8 T cell lineages were markedly asymmetric. Notably, CD8 commitment appeared to require lineage-specific signals, whereas CD4 commitment appeared to occur in the absence of lineage-specific signals by default. Consequently, it was unclear whether CD4 commitment, as revealed by selective termination of CD8 coreceptor synthesis, occurred in all DP thymocytes, or whether CD4 commitment occurred only in T cell receptor (TCR)–CD3-signaled DP thymocytes. Here, we report that selective termination of CD8 coreceptor synthesis does not occur in DP thymocytes spontaneously. Rather, CD4 commitment in DP thymocytes requires signals transduced by either CD3 or ζ chains, which can signal CD4 commitment even in the absence of clonotypic TCR chains. PMID:9206993

  11. Differential susceptibility and maturation of thymocyte subsets during Salmonella Typhimurium infection: insights on the roles of glucocorticoids and Interferon-gamma

    PubMed Central

    Majumdar, Shamik; Deobagkar-Lele, Mukta; Adiga, Vasista; Raghavan, Abinaya; Wadhwa, Nitin; Ahmed, Syed Moiz; Rananaware, Supriya Rajendra; Chakraborty, Subhashish; Joy, Omana; Nandi, Dipankar

    2017-01-01

    The thymus is known to atrophy during infections; however, a systematic study of changes in thymocyte subpopulations has not been performed. This aspect was investigated, using multi-color flow cytometry, during oral infection of mice with Salmonella Typhimurium (S. Typhimurium). The major highlights are: First, a block in the developmental pathway of CD4−CD8− double negative (DN) thymocytes is observed. Second, CD4+CD8+ double positive (DP) thymocytes, mainly in the DP1 (CD5loCD3lo) and DP2 (CD5hiCD3int), but not DP3 (CD5intCD3hi), subsets are reduced. Third, single positive (SP) thymocytes are more resistant to depletion but their maturation is delayed, leading to accumulation of CD24hiCD3hi SP. Kinetic studies during infection demonstrated differences in sensitivity of thymic subpopulations: Immature single positive (ISP) > DP1, DP2 > DN3, DN4 > DN2 > CD4+ > CD8+. Upon infection, glucocorticoids (GC), inflammatory cytokines, e.g. Ifnγ, etc are induced, which enhance thymocyte death. Treatment with RU486, the GC receptor antagonist, increases the survival of most thymic subsets during infection. Studies with Ifnγ−/− mice demonstrated that endogenous Ifnγ produced during infection enhances the depletion of DN2-DN4 subsets, promotes the accumulation of DP3 and delays the maturation of SP thymocytes. The implications of these observations on host cellular responses during infections are discussed. PMID:28091621

  12. Effects of mistletoe extract on murine thymocytes in vivo and on glucocorticoid-induced cell count reduction.

    PubMed

    Hajtó, Tibor; Berki, Timea; Pálinkás, László; Boldizsár, Ferenc; Németh, Péter

    2006-02-01

    Mistletoe extracts are widely used in cancer patients due to their cytostatic and immunomodulatory effects. Essential components include mistletoe lectins which act as biomodulators with proinflammatory and apoptosisinducing effects. This study investigates the acute and longterm effects of standardized mistletoe extract (Iscador(R) M spec 5 mg) on thymocyte subpopulations and peripheral T-cells using a murine (Balb/c) model. Using cell surface CD4/CD8 staining and flow cytometry, we followed the changes in CD4-CD8- double-negative (DN), CD4(+)CD8(+) double-positive (DP) and CD4(+) or CD8(+) single-positive (SP) T-cells 24 h after single or repeated injections of 3 different dilutions (1:12, 1:60, 1:300) corresponding to 2.1, 0.42 and 0.08 mg/kg of Iscador. Thymocyte apoptosis was detected by flow cytometry using Annexin V and propidium iodide. 24 h after a single injection of the 2 lower doses, the number of DN thymocytes increased significantly with an enhanced ratio of apoptotic cells. Following administration of the lowest dose, in peripheral blood the CD4(+)/CD8(+) ratio was elevated. In the long-term trial, Balb/c mice were treated twice a week with 3 different doses of Iscador +/- 20 mg/kg of dexamethasone (DX), resulting in significantly enhanced DN thymocytes and elevated levels of apoptotic cells after treatment with the 2 lower doses. Iscador also inhibited the DX-induced reduction in the thymic DN cell count, as well as the DX-induced decrease in the CD4(+)/CD8(+) ratio and CD4(+) in the peripheral blood. Our results suggest that standardized mistletoe extract modulates proliferation and apoptosis of thymocytes in a dose-dependent manner and may act lymphoprotective during DX treatment.

  13. Thymic exosomes promote the final maturation of thymocytes

    PubMed Central

    Lundberg, Vanja; Berglund, Martin; Skogberg, Gabriel; Lindgren, Susanne; Lundqvist, Christina; Gudmundsdottir, Judith; Thörn, Karolina; Telemo, Esbjörn; Ekwall, Olov

    2016-01-01

    Extensive knowledge has been gained the last years concerning mechanisms underlying the selection of single positive thymocytes in the thymic medulla. Less is known regarding other important processes in the thymic medulla such as the regulation of late stage thymocyte maturation. We have previously reported that exosomes are abundant in the thymus with a phenotype that indicates an epithelial cell origin and immunoregulatory properties. In this study we use an in vitro system to investigate the effects of thymic exosomes on the maturation of single positive thymocytes as well as effects on nTreg formation. We show that thymic exosomes promote the maturation of single positive CD4+CD25− cells into mature thymocytes with S1P1+Qa2+ and CCR7+Qa2+ phenotypes. Furthermore, we show that thymic exosomes reduce the formation of CD4+CD25+FoxP3+ thymocytes and that these exosome effects are independent of dendritic cell co-stimulation but require intact exosomal RNA content and surface proteins. An efficient direct uptake of exosomes by both thymocytes and thymic DC’s is also demonstrated. In conclusion, this study demonstrates that exosomes may represent a new route of communication within the thymus. PMID:27824109

  14. Preselection Thymocytes Are More Sensitive to T Cell Receptor Stimulation Than Mature T Cells

    PubMed Central

    Davey, Gayle M.; Schober, Sonya L.; Endrizzi, Bart T.; Dutcher, Angela K.; Jameson, Stephen C.; Hogquist, Kristin A.

    1998-01-01

    During T cell development, thymocytes which are tolerant to self-peptides but reactive to foreign peptides are selected. The current model for thymocyte selection proposes that self-peptide–major histocompatibility complex (MHC) complexes that bind the T cell receptor with low affinity will promote positive selection while those with high affinity will result in negative selection. Upon thymocyte maturation, such low affinity self-peptide–MHC ligands no longer provoke a response, but foreign peptides can incidentally be high affinity ligands and can therefore stimulate T cells. For this model to work, thymocytes must be more sensitive to ligand than mature T cells. Contrary to this expectation, several groups have shown that thymocytes are less responsive than mature T cells to anti-T cell receptor for antigen (TCR)/CD3 mAb stimulation. Additionally, the lower TCR levels on thymocytes, compared with T cells, would potentially correlate with decreased thymocyte sensitivity. Here we compared preselection thymocytes and mature T cells for early activation events in response to peptide–MHC ligands. Remarkably, the preselection thymocytes were more responsive than mature T cells when stimulated with low affinity peptide variants, while both populations responded equally well to the antigenic peptide. This directly demonstrates the increased sensitivity of thymocytes compared with T cells for TCR engagement by peptide–MHC complexes. PMID:9815264

  15. T3 expression by human thymocytes in culture.

    PubMed Central

    Aiello, F B; Musiani, P; Maggiano, N; Larocca, L M; Piantelli, M

    1985-01-01

    By panning procedures employing T6 and T3 monoclonal antibody, human thymocytes were fractionated into two subpopulations depleted of T6- or T3-positive (T6+, T3+) cells. Unfractionated thymocytes and T6- and T3-depleted subpopulations were separately cultured for 48 h in RPMI 1640 medium with 10% FCS or in HB 101 serum-free medium. Determining the phenotype of unfractionated thymocytes at various time intervals, a time-dependent increase of T3+ cells was observed. An inverse relationship was found between the percentage of T3+ cells and the T6 and peanut agglutinin (PNA) reactive thymocytes. When the surface antigen expression in the T3-depleted population (greater than 95% T6+ and PNA+ cells) was analysed, a strong increase of T3+ cells and a complementary reduction of T6+ and PNA+ cells was evidenced. During that time the surface phenotype of the T6-depleted population (greater than 80% T3+ cells) showed the same trend of differentiation, as the other thymocyte preparations. These results indicate that a conspicuous fraction of human thymocytes and particularly of those characterized by a cortical phenotype (PNA+ and T6+ cells), are able to express mature T-cell antigens when cultured in vitro in the absence of the thymic microenvironment influence. However, the in vitro acquisition of a mature phenotype is not accompanied by a parallel achievement of the capacity to respond to mitogens such as PHA or T3 monoclonal antibody. PMID:3876184

  16. The DNA damage- and transcription-associated protein paxip1 controls thymocyte development and emigration.

    PubMed

    Callen, Elsa; Faryabi, Robert B; Luckey, Megan; Hao, Bingtao; Daniel, Jeremy A; Yang, Wenjing; Sun, Hong-Wei; Dressler, Greg; Peng, Weiqun; Chi, Hongbo; Ge, Kai; Krangel, Michael S; Park, Jung-Hyun; Nussenzweig, André

    2012-12-14

    Histone 3 lysine 4 trimethylation (H3K4me3) is associated with promoters of active genes and found at hot spots for DNA recombination. Here we have shown that PAXIP1 (also known as PTIP), a protein associated with MLL3 and MLL4 methyltransferase and the DNA damage response, regulates RAG-mediated cleavage and repair during V(D)J recombination in CD4(+) CD8(+) DP thymocytes. Loss of PAXIP1 in developing thymocytes diminished Jα H3K4me3 and germline transcription, suppressed double strand break formation at 3' Jα segments, but resulted in accumulation of unresolved T cell receptor α-chain gene (Tcra) breaks. Moreover, PAXIP1 was essential for release of mature single positive (SP) αβ T cells from the thymus through transcriptional activation of sphingosine-1-phosphate receptor S1pr1 as well as for natural killer T cell development. Thus, in addition to maintaining genome integrity during Tcra rearrangements, PAXIP1 controls distinct transcriptional programs during DP differentiation necessary for Tcra locus accessibility, licensing mature thymocytes for trafficking and natural killer T cell development.

  17. From pre-DP, post-DP, SP4, and SP8 Thymocyte Cell Counts to a Dynamical Model of Cortical and Medullary Selection

    PubMed Central

    Sawicka, Maria; Stritesky, Gretta L.; Reynolds, Joseph; Abourashchi, Niloufar; Lythe, Grant; Molina-París, Carmen; Hogquist, Kristin A.

    2014-01-01

    Cells of the mature αβ T cell repertoire arise from the development in the thymus of bone marrow precursors (thymocytes). αβ T cell maturation is characterized by the expression of thousands of copies of identical αβ T cell receptors and the CD4 and/or CD8 co-receptors on the surface of thymocytes. The maturation stages of a thymocyte are: (1) double negative (DN) (TCR−, CD4− and CD8−), (2) double positive (DP) (TCR+, CD4+ and CD8+), and (3) single positive (SP) (TCR+, CD4+ or CD8+). Thymic antigen presenting cells provide the appropriate micro-architecture for the maturation of thymocytes, which “sense” the signaling environment via their randomly generated TCRs. Thymic development is characterized by (i) an extremely low success rate, and (ii) the selection of a functional and self-tolerant T cell repertoire. In this paper, we combine recent experimental data and mathematical modeling to study the selection events that take place in the thymus after the DN stage. The stable steady state of the model for the pre-DP, post-DP, and SP populations is identified with the experimentally measured cell counts from 5.5- to 17-week-old mice. We make use of residence times in the cortex and the medulla for the different populations, as well as recently reported asymmetric death rates for CD4 and CD8 SP thymocytes. We estimate that 65.8% of pre-DP thymocytes undergo death by neglect. In the post-DP compartment, 91.7% undergo death by negative selection, 4.7% become CD4 SP, and 3.6% become CD8 SP. Death by negative selection in the medulla removes 8.6% of CD4 SP and 32.1% of CD8 SP thymocytes. Approximately 46.3% of CD4 SP and 27% of CD8 SP thymocytes divide before dying or exiting the thymus. PMID:24592261

  18. Lunatic fringe enhances competition for delta-like Notch ligands but does not overcome defective pre-TCR signaling during thymocyte beta-selection in vivo.

    PubMed

    Visan, Ioana; Yuan, Julie S; Liu, Ying; Stanley, Pamela; Guidos, Cynthia J

    2010-10-15

    Notch1 activation by Delta-like (DL) Notch ligands is essential to induce T cell commitment and to suppress B cell development from thymus-seeding progenitors. Thymus-seeding progenitor competition for DL4 is critically regulated by Lunatic Fringe (Lfng), which glycosylates epidermal growth factor repeats in the Notch1 extracellular domain to enhance binding avidity for DL ligands. Notch1 activation is also essential for the process of β-selection, which drives TCRβ(+) CD4/CD8 double-negative 3 (DN3) precursors to proliferate and generate a large pool of CD4/CD8 double-positive thymocytes. We have used several genetic approaches to determine the importance of Lfng-Notch1 interactions in regulating competition of preselection and postselection DN3 thymocytes for DL ligands in vivo. Surprisingly, although Lfng overexpression enhanced DL4 binding by preselection DN3a thymocytes, it did not confer them with a competitive advantage in mixed chimeras. In contrast, Lfng overexpression enhanced competition of post-β-selection DN3b precursors for DL ligands. Lfng modification of O-fucose in the Notch1 ligand-binding domain contributed to but was not solely responsible for the developmental effects of Lfng overexpression. Although previous studies have suggested that pre-TCR-deficient DN3 thymocytes compete poorly for DL ligands, Lfng overexpression did not fully restore double-positive thymocyte pools from DN3b cells with pre-TCR signaling defects. Thus, pre-TCR and Notch signaling have largely nonoverlapping functions in β-selection. Collectively, our data reveal that Lfng enhances DN3b precursor competition for intrathymic DL ligands to maximize Notch-induced clonal expansion during the earliest stage of β-selection.

  19. Investigation of the effect of mistletoe (Viscum album L.) extract Iscador on the proliferation and apoptosis of murine thymocytes.

    PubMed

    Hajtò, Tibor; Berki, Timea; Pàlinkàs, Làszlò; Boldizsàr, Ferenc; Németh, Péter

    2006-06-01

    Mistletoe (Viscum album L.) extracts (ME) have been shown to exhibit a bell-shaped curve of immunological efficacy and mistletoe lectins (MLs) were found to play an important role in this phenomenon. The aim of present in vivo study was to investigate the acute- and long-term effect of a standardized ME (Iscador M special) on thymocyte subpopulations and peripheral T cells using a murine (Balb/c) model. In thymus CD4-CD8- double negative (DN), CD4+CD8+ double positive (DP), CD4+ or CD8+ single positive T cells were determined 24 h after a single injection or following a long-term treatment (twice a week for 4 weeks) with three different dilutions of ME which are corresponding to 4.5 ng/ kg, 22.5 ng/kg and 112.5 ng/kg doses of MLs. The apoptosis of the thymocytes was also tested by flow cytometry using Annexin V and propidium iodide. 24 h after a single injection of ME only the lowest dose caused in the blood samples an elevated CD4+/CD8+ ratio and in thymus an enhanced proliferation of DN thymocytes indicating a similar bell-shaped curve of immunological efficacy. After a treatment for four weeks these responses were less intensive indicating that none of the three doses are immunologically optimal. Surprisingly, both in the acute and in the long-term trial only the lower doses induced significant enhancements in the ratio of apoptotic thymocytes. In addition, ME inhibited the dexamethasone (DX)-induced reduction of DN cell count in thymus, as well as the DX-induced decrease of CD4+/CD8+ ratio and CD4+ cell level in peripheral blood. These in vivo results suggest that investigation of thymocytes in vivo can be helpful in the immunological dose-finding since standardized ME is able to modulate the proliferation and apoptosis of thymocytes with a bell-shaped curve of efficacy. In addition, ME may act lymphoprotectively during DX treatment.

  20. Increased Susceptibility of  Thymocytes to Apoptosis in Mice Lacking AIM, a Novel Murine Macrophage-derived Soluble Factor Belonging to the Scavenger Receptor Cysteine-rich Domain Superfamily

    PubMed Central

    Miyazaki, Toru; Hirokami, Yumiko; Matsuhashi, Nobuyuki; Takatsuka, Hisakazu; Naito, Makoto

    1999-01-01

    Apoptosis of cells must be regulated both positively and negatively in response to a variety of stimuli in the body. Various environmental stresses are known to initiate apoptosis via differential signal transduction cascades. However, induction of signals that may inhibit apoptosis is poorly understood, although a number of intracellular molecules that mediate inhibition of apoptosis have been identified. Here we present a novel murine macrophage-specific 54-kD secreted protein which inhibits apoptosis (termed AIM, for apoptosis inhibitor expressed by macrophages). AIM belongs to the macrophage scavenger receptor cysteine-rich domain superfamily (SRCR-SF), members of which share a highly homologous conserved cysteine-rich domain. In AIM-deficient mice, the thymocyte numbers were diminished to half those in wild-type mice, and CD4/CD8 double-positive (DP) thymocytes were strikingly more susceptible to apoptosis induced by both dexamethasone and irradiation in vivo. Recombinant AIM protein significantly inhibited cell death of DP thymocytes in response to a variety of stimuli in vitro. These results indicate that in the thymus, AIM functions in trans to induce resistance to apoptosis within DP cells, and thus supports the viability of DP thymocytes before thymic selection. PMID:9892623

  1. THEMIS, a new T cell specific protein important for late thymocyte development

    PubMed Central

    Lesourne, Renaud; Uehara, Shoji; Lee, Jan; Song, Ki-Duk; Li, LiQi; Pinkhasov, Julia; Zhang, Yongqing; Weng, Nan-Ping; Wildt, Kathryn F.; Wang, Lie; Bosselut, Remy; Love, Paul E.

    2010-01-01

    During positive selection, thymocytes transition through a stage during which T cell receptor (TCR) signaling controls CD4 versus CD8 lineage choice and subsequent maturation. Here, we describe a new T cell specific protein, THEMIS, that performs a distinct function during this stage. In Themis-/- mice, thymocyte selection was impaired and the number of transitional CD4+CD8int thymocytes as well as CD4 and CD8 single positive thymocytes was decreased. Remarkably, although no overt TCR-proximal signaling deficiencies were detected, Themis-/-CD4+CD8int thymocytes exhibited developmental defects consistent with attenuated signaling that were reversible by increased TCR stimulation. These results identify THEMIS as a critical component of the T cell developmental program and suggest that THEMIS functions to sustain and/or integrate signals required for proper lineage commitment and maturation. PMID:19597498

  2. Contributions of the T Cell Receptor–associated CD3γ–ITAM to Thymocyte Selection

    PubMed Central

    Haks, Mariëlle C.; Pépin, Elsa; van den Brakel, Jeroen H.N.; Smeele, Sigrid A.A.; Belkowski, Stanley M.; Kessels, Helmut W.H.G.; Krimpenfort, Paul; Kruisbeek, Ada M.

    2002-01-01

    The immunoreceptor tyrosine-based activation motifs (ITAMs) in the CD3 chains associated with the T cell receptor (TCR) are crucial for TCR signaling. To probe the role of the CD3γ–ITAM in T cell development, we created knock-in mice in which the CD3γ chain of the TCR complex is replaced by a mutant signaling-deficient CD3γ chain, lacking the CD3γ–ITAM. This mutation results in considerable impairment in positive selection in the polyclonal TCR repertoire. When CD3γ–ΔITAM mice are crossed to mice expressing transgenic F5 TCRs, their thymocytes are completely unable to perform positive selection in vivo in response to intrathymic ligands. Also, the in vitro positive selection response of double-positive (DP) thymocytes with F5–CD3γ–ΔITAM mutant receptors to their agonist ligand and many of its variants is severely impaired or abrogated. Yet, the binding and dissociation constants of agonist ligands for the F5 receptor are not affected by the CD3γ–ΔITAM mutation. Furthermore, DP thymocytes with mutant receptors can respond to agonist ligand with normal antigen sensitivity and to normal levels, as shown by their ability to induce CD69 up-regulation, TCR down-regulation, negative selection, and ZAP70 and c-Jun NH2-terminal kinase activation. In sharp contrast, induction of extracellular signal-regulated kinase (ERK) activation and linker for activation of T cells (LAT) phosphorylation are severely impaired in these cells. Together, these findings underscore that intrinsic properties of the TCR–CD3 complex regulate selection at the DP checkpoint. More importantly, this analysis provides the first direct genetic evidence for a role of the CD3γ–ITAM in TCR-driven thymocyte selection. PMID:12093866

  3. Expression of T cell receptors by thymocytes: in situ staining and biochemical analysis.

    PubMed Central

    Cristanti, A; Colantoni, A; Snodgrass, R; von Boehmer, H

    1986-01-01

    We have examined the in situ expression of T cell receptor (TCR) V beta 8 protein in murine thymus during ontogeny using the monoclonal antibody F23.1. Positive cells were first detected at day 15 of gestation (0.6%). By day 16 the frequency of positive cells increased dramatically (4.18%). From day 16 to day 17 positive cells doubled (8.17%). The first clusters of F23.1 positive cells were seen at day 17. In the cortex, positive cells decreased from 14% in the newborn mice to 9.8% in 8-week-old mice, whereas in the medulla the frequency remained unchanged at 20%. The antibody F23.1, as well as an antiserum raised against the constant region of the beta chain, immunoprecipitated receptor dimers from highly purified Lyt2+, L3T4+ thymocytes and from two thymic lymphomas of cortical phenotype which express full size alpha and beta mRNA. The receptor dimer could not be precipitated from Lyt2-, L3T4- thymocytes. The results are discussed with regard to intrathymic T cell repertoire selection. Images Fig. 1. Fig. 2. Fig. 3. Fig. 4. Fig. 5. PMID:2431899

  4. Infection of human thymocytes by Epstein-Barr virus.

    PubMed

    Watry, D; Hedrick, J A; Siervo, S; Rhodes, G; Lamberti, J J; Lambris, J D; Tsoukas, C D

    1991-04-01

    The Epstein-Barr Virus (EBV) causes infectious mononucleosis, and has been strongly associated with certain human cancers. The virus is thought to exclusively bind to B lymphocytes and epithelial cells via receptors (CR2/CD21) that also interact with fragments of the third component of complement (C3). Recent evidence, however, has challenged this belief. We have used two-color immunofluorescence analysis using biotin-conjugated EBV and streptavidin-phycoerythrin along with fluorescein-conjugated anti-T cell antibodies and demonstrated that CD1-positive, CD3-dull (immature) human thymocytes express functional EBV receptors. In four replicate experiments, the binding of EBV to thymocytes ranged between 8 and 18%. This interaction is specific as evidenced by inhibition with nonconjugated virus, anti-CR2 antibodies, aggregated C3, and an antibody to the gp350 viral glycoprotein that the virus uses to bind to CR2. EBV can infect the thymocytes as evaluated by the presence of episomal EBV-DNA in thymocytes that had been incubated with the virus as short as 12 days or as long as 6 weeks. Episomal DNA analysis was performed by Southern blotting with a EBV-DNA probe that hybridizes to the first internal reiteration of the viral DNA. The presence of the EBV genome is also supported by the detection of EBV nuclear antigen 1 in infected thymocytes, assessed by Western blotting with EBV-immune sera. The EBV infection is specific as determined by blocking experiments using anti-CR2 and anti-gp350 antibodies. Finally, virus infection of thymocytes can act synergistically along with interleukin 2 and induce a lymphokine-dependent cellular proliferation. In view of previously reported cases of EBV-positive human T cell lymphomas, the possibility is raised that EBV may be involved in cancers of T lymphocytes that have not been previously appreciated.

  5. Role for Ets-2Thr-72 Transcription Factor in Stage-specific Thymocyte Development and Survival

    PubMed Central

    Fisher, Ian B.; Ostrowski, Mike; Muthusamy, Natarajan

    2012-01-01

    Interference of Ras signaling deregulates thymocyte development in mouse models. However, the role of Ets-2, a transcription factor that is phosphorylated on a critical threonine residue (Thr-72) by the Ras/MAPK pathway in thymocyte development, has not been defined. Transgenic mice overexpressing a phosphomutant Ets-2 (T72A) in the thymus displayed reduced thymus size associated with a 60–80% reduction in thymocyte populations. The transgenic mice exhibited a 20-fold increase in a c-Kit+ CD4+ CD8+ CD3− population and a 5-fold increase in a unique CD5low population associated with a partial developmental block at the DN2-DN3 stage of thymocytes. Transgenic thymocytes exhibited increased apoptosis, and overexpression of Bcl-2 rescued the hypocellularity and associated thymocyte developmental block in double transgenic mice. The observed defects in these mice are not dependent on Ets-1 expression. These studies implicate for the first time a stage-specific Ets-1-independent regulatory role for Ets-2 in early thymocyte development and survival. PMID:22128184

  6. 2,3,7,8-Tetrachlorodibenzo-p-dioxin induces premature activation of the KLF2 regulon during thymocyte development.

    PubMed

    McMillan, Brian J; McMillan, Susanne N; Glover, Ed; Bradfield, Christopher A

    2007-04-27

    The environmental pollutant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, dioxin) causes numerous and diverse toxic events via activation of the aryl hydrocarbon receptor, including atrophy of the thymus. Exposure to TCDD induces acute thymocyte cell loss, which occurs concomitantly with proliferation arrest and premature emigration of triple negative (TN; CD4(-), CD8(-), CD3(-)) T cell progenitors. In this report, we demonstrate that TCDD exposure results in dysregulation of KLF2 (Kruppel-like factor 2) expression in developing thymocytes. The Klf2 gene encodes an Sp1-like zinc finger transcription factor that functions as a central regulator of T lymphocyte proliferation and trafficking. During normal thymocyte development, KLF2 is expressed exclusively in CD4 and CD8 single positive T cells and promotes a nonproliferative, promigratory phenotype. In mice exposed to TCDD, however, the Klf2 gene is prematurely expressed in TN thymocytes. Administration of a 100 microg/kg dose of TCDD results in a approximately 15-fold induction of KLF2 as early as the TN2 (CD44(+), CD25(+)) stage of development and immediately precedes acute cell loss in the TN3, TN4, and double positive (CD4(+), CD8(+)) cell stages. Induction of KLF2 occurs within 12 h of TCDD exposure and is fully dependent on expression of the aryl hydrocarbon receptor. In addition, TCDD exposure alters the expression of several factors comprising the KLF2 regulon, including Edg1/S1P(1), beta(7) integrin, CD52, Cdkn2d (cyclin-dependent kinase inhibitor 2D), s100a4, and IL10R alpha. These findings indicate that the pollutant TCDD interferes with early thymopoeisis via ectopic expression of the KLF2 regulon.

  7. Mushroom acidic glycosphingolipid induction of cytokine secretion from murine T cells and proliferation of NK1.1 {alpha}/{beta} TCR-double positive cells in vitro

    SciTech Connect

    Nozaki, Hirofumi; Itonori, Saki; Sugita, Mutsumi; Nakamura, Kimihide; Ohba, Kiyoshi; Suzuki, Akemi; Kushi, Yasunori

    2008-08-29

    Interferon (IFN)-{gamma} and interleukin (IL)-4 regulate many types of immune responses. Here we report that acidic glycosphingolipids (AGLs) of Hypsizigus marmoreus and Pleurotus eryngii induced secretion of IFN- {gamma} and IL-4 from T cells in a CD11c-positive cell-dependent manner similar to that of {alpha}-galactosylceramide ({alpha}-GalCer) and isoglobotriaosylceramide (iGb3), although activated T cells by AGLs showed less secretion of cytokine than those activated by {alpha}-GalCer. In addition, stimulation of these mushroom AGLs induced proliferation of NK1.1 {alpha}/{beta} TCR-double positive cells in splenocytes. Administration of a mixture of {alpha}-GalCer and AGLs affected the stimulation of {alpha}-GalCer and generally induced a subtle Th1 bias for splenocytes but induced an extreme Th2 bias for thymocytes. These results suggested that edible mushroom AGLs contribute to immunomodulation.

  8. Washington Double Star Catalog Cross Index (1950 position sort)

    NASA Technical Reports Server (NTRS)

    1993-01-01

    A machine-readable version of the Washington Catalog of Visual Double Stars (WDS) was prepared in 1984 on the basis of a data file that was collected and maintained for more than a century by a succession of double-star observers. Although this catalog is being continually updated, a new copy for distribution is not expected to be available for a few years. The WDS contains DM numbers, but many of these are listed only in the notes, which makes it difficult to search for double-star information, except by position. Hence, a cross index that provides complete DM identifications is desirable, and it appears useful to add HD numbers for systems in that catalog. Aitken Double Star (ADS) numbers were retained from the WDS, but no attempt was made to correct these except for obvious errors.

  9. IL-23 Promotes TCR-mediated Negative Selection of Thymocytes through the Upregulation of IL-23 Receptor and RORγt

    PubMed Central

    Li, Hao; Hsu, Hui-Chen; Wu, Qi; Yang, PingAr; Li, Jun; Luo, Bao; Oukka, Mohamed; Steele, Claude H.; Cua, Daniel J; Grizzle, William E.; Mountz, John D.

    2014-01-01

    Summary Transient thymic involution is frequently found during inflammation, yet the mode of action of inflammatory cytokines is not well defined. Here we report that interleukin-23 (IL-23) production by the thymic dendritic cells (DCs) promotes apoptosis of the CD4hiCD8hi double positive (DP) thymocytes. A deficiency in IL-23 signaling interferes with negative selection in the male Db/H-Y T-cell receptor (TCR) transgenic mice. IL-23 plus TCR signaling results in significant up-regulation of IL-23 receptor (IL-23R) expressed predominantly on CD4hiCD8hiCD3+αβTCR+ DP thymocytes, and leads to RORγt dependent apoptosis. These results extend the action of IL-23 beyond its peripheral effects to a unique role in TCR mediated negative selection including elimination of natural T regulatory cells in the thymus. PMID:25001511

  10. Clonally Expanding Thymocytes Having Lineage Capability in Gamma-Ray-Induced Mouse Atrophic Thymus

    SciTech Connect

    Yamamoto, Takashi; Morita, Shin-ichi; Go, Rieka; Obata, Miki; Katsuragi, Yoshinori; Fujita, Yukari; Maeda, Yoshitaka; Yokoyama, Minesuke; Aoyagi, Yutaka; Ichikawa, Hitoshi; Mishima, Yukio; Kominami, Ryo

    2010-05-01

    Purpose: To characterize, in the setting of gamma-ray-induced atrophic thymus, probable prelymphoma cells showing clonal growth and changes in signaling, including DNA damage checkpoint. Methods and Materials: A total of 111 and 45 mouse atrophic thymuses at 40 and 80 days, respectively, after gamma-irradiation were analyzed with polymerase chain reaction for D-J rearrangements at the TCRbeta locus, flow cytometry for cell cycle, and Western blotting for the activation of DNA damage checkpoints. Results: Limited D-J rearrangement patterns distinct from normal thymus were detected at high frequencies (43 of 111 for 40-day thymus and 21 of 45 for 80-day thymus). Those clonally expanded thymocytes mostly consisted of CD4{sup +}CD8{sup +} double-positive cells, indicating the retention of lineage capability. They exhibited pausing at a late G1 phase of cell cycle progression but did not show the activation of DNA damage checkpoints such as gammaH2AX, Chk1/2, or p53. Of interest is that 17 of the 52 thymuses showing normal D-J rearrangement patterns at 40 days after irradiation showed allelic loss at the Bcl11b tumor suppressor locus, also indicating clonal expansion. Conclusion: The thymocytes of clonal growth detected resemble human chronic myeloid leukemia in possessing self-renewal and lineage capability, and therefore they can be a candidate of the lymphoma-initiating cells.

  11. Themis controls thymocyte selection through regulation of T cell receptor-mediated signaling

    PubMed Central

    Fu, Guo; Vallée, Sébastien; Rybakin, Vasily; McGuire, Marielena V.; Ampudia, Jeanette; Brockmeyer, Claudia; Salek, Mogjiborahman; Fallen, Paul R.; Hoerter, John A.H.; Munshi, Anil; Huang, Yina H.; Hu, Jianfang; Fox, Howard S.; Sauer, Karsten; Acuto, Oreste; Gascoigne, Nicholas R.J.

    2009-01-01

    Themis (Thymocyte expressed molecule involved in selection), a member of a family of proteins with unknown functions, is highly conserved among vertebrates. Here we found that Themis is expressed in high amounts in thymocytes between the pre-T cell receptor (TCR) and positive selection checkpoints, and in low amounts in mature T cells. Themis-deficient thymocytes exhibit defective positive selection, which results in reduced numbers of mature thymocytes. Negative selection is also impaired in Themis-deficient mice. A higher percentage of Themis-deficient T cells exhibit CD4+CD25+Foxp3+ regulatory and CD62LloCD44hi memory phenotypes than in wild-type mice. Supporting a role for Themis in TCR signaling, this protein is phosphorylated quickly after TCR stimulation, and is needed for optimal TCR-driven Ca2+ mobilization and Erk activation. PMID:19597499

  12. FMC46, a cell protrusion-associated leukocyte adhesion molecule-1 epitope on human lymphocytes and thymocytes.

    PubMed

    Pilarski, L M; Turley, E A; Shaw, A R; Gallatin, W M; Laderoute, M P; Gillitzer, R; Beckman, I G; Zola, H

    1991-07-01

    In this report, we describe a 76-kDa glycoprotein recognized by mAb FMC46 that, by virtue of its concentration on cell protrusions involved in motility, may be important in lymphoid cell locomotion. FMC46 detects an epitope of the leukocyte adhesion molecule-1 (LAM-1), a member of the selecting family (LAM-1, Endothelial Leukocyte Adhesion Molecular-1 (ELAM-1), and Granule Membrane Protein-140 (GMP-140), that is expressed on LAM-1-transfected cell lines, is a glycosylation epitope based on its loss after culture in tunicamycin, and is closely related to the LAM-1.2 epitope. FMC46 is expressed at high density on the majority of CD45RA+ and CD45RO+ peripheral blood T cells (60 to 70%) and on a subset of thymocytes that includes the multinegative CD3- CD4- CD8- progenitor cells (100% FMC46hi) and the CD45R0- presumptive thymic generative lineage (70% FMC46hi). It appears at reduced density and frequency on CD45RA- thymocytes (50% FMC46lo), comprised mainly of death-committed thymocytes. Among thymic subsets defined by expression of CD4 and/or CD8, FMC46 is expressed at high density predominantly on a subset of single-positive cells and not on double-positive cells. These results suggest a fundamental role for LAM-1 in thymic development, with a high density preferentially expressed on cells involved in thymic generative processes and a low density on cells progressing to intrathymic death. A major subset of peripheral blood B cells and thymic B cells also express FMC46. Immunohistochemistry on frozen sections indicated strong staining in splenic follicles and around blood vessels, staining of the thymic medulla and subcapsular areas, and staining of the mantle zone of germinal centers of the lymph node. FMC46+ lymphocytes accumulated along high endothelial venules in the lymph node. On locomoting multinegative thymocytes, FMC46 is concentrated on the leading tip of extended processes, on pseudopods, and on ruffles, unlike the distribution of either CD44 or TQ1 (LAM 1

  13. Huge positive magnetoresistance in antiferromagnetic double perovskite metals

    NASA Astrophysics Data System (ADS)

    Nand Singh, Viveka; Majumdar, Pinaki

    2014-07-01

    Metals with large positive magnetoresistance are rare. We demonstrate that antiferromagnetic metallic states, as have been predicted for the double perovskites, are excellent candidates for huge positive magnetoresistance. An applied field suppresses long range antiferromagnetic order leading to a state with short range antiferromagnetic correlations and strong electronic scattering. The field induced resistance ratio can be more than tenfold, at moderate field, in a structurally ordered system, and continues to be almost twofold even in systems with ˜25% antisite disorder. Although our explicit demonstration is in the context of a two- dimensional spin-fermion model of the double perovskites, the mechanism we uncover is far more general, complementary to the colossal negative magnetoresistance process, and would operate in other local moment metals that show a field driven suppression of non-ferromagnetic order.

  14. Huge positive magnetoresistance in antiferromagnetic double perovskite metals.

    PubMed

    Singh, Viveka Nand; Majumdar, Pinaki

    2014-07-23

    Metals with large positive magnetoresistance are rare. We demonstrate that antiferromagnetic metallic states, as have been predicted for the double perovskites, are excellent candidates for huge positive magnetoresistance. An applied field suppresses long range antiferromagnetic order leading to a state with short range antiferromagnetic correlations and strong electronic scattering. The field induced resistance ratio can be more than tenfold, at moderate field, in a structurally ordered system, and continues to be almost twofold even in systems with ∼ 25% antisite disorder. Although our explicit demonstration is in the context of a two- dimensional spin-fermion model of the double perovskites, the mechanism we uncover is far more general, complementary to the colossal negative magnetoresistance process, and would operate in other local moment metals that show a field driven suppression of non-ferromagnetic order.

  15. [Apoptosis and thymocyte development (epithelial cells as inducers of thymocyte apoptosis)].

    PubMed

    Iarilin, A A; Bulanova, E G; Sharova, N I; Budagian, V M

    1998-01-01

    Apoptosis, together with proliferation, is a main factor of selection of the clones of developing T-lymphocytes: the clones not supported by positive selection are subject to apoptosis and apoptosis accounts for discarding of potentially autoaggressive clones, i.e., for negative selection in the thymus and peripheral lymphoid tissue. Realization of apoptosis at different stages of the development of T-lymphocytes depends to a varying extent on Fas, Bcl-2, p53, and other regulators. The dendritic cells are the main cell type, the contact with determines apoptosis of T-lymphocytes. A possible role of the epithelial cells was shown in few models (on murine cells) and was not practically studied. We obtained a line of epithelial cells of the human thymus cells HTSC, cocultivation with which induces apoptosis of immature thymocytes and blood T-cells activated by mitogens. Development of apoptosis is suppressed by inhibitors of protein and RNA synthesis, chelators Ca2+, ions Zn2+, and factors destroying the cytoskeleton components. In this model, interaction of pairs of molecules CD4-HLA class II and LFA-1-ICAM-1. When in contact with the HTSC cells, the thymocytes of mice mutant for Fas-receptor (line MRL.lpr) are subject to apoptosis, but when this receptor is present, it affects the development of apoptosis.

  16. Double-positive Goodpasture's syndrome with concomitant active pulmonary tuberculosis.

    PubMed

    Kashif, Waqar; Yaqub, Sonia; Mahmood, Syed Faisal; Patel, Junaid

    2013-07-01

    Anti-glomerular basement membrane (anti-GBM) disease usually presents as rapidly progressive glomerulonephritis, and, when accompanied with pulmonary hemorrhage, it is called Goodpasture's syndrome. Anti-neutrophilic cytoplasmic antibodies (ANCA) may co-exist with anti-GBM antibodies. In most of these "double positive" cases, ANCA is specific for myeloperoxidase (p-ANCA). We report a rare case of a critically ill patient c-ANCA-associated double-positive Goodpasture's syndrome with concomitant tuberculosis that was successfully treated with immunosuppression, plasmapheresis and anti-tuberculous therapy (ATT). A 32-year-old gentleman with a 15 pack-year smoking history presented with massive hemoptysis, respiratory failure and oliguria. Laboratory investigation revealed anemia, elevated creatinine and active urinary sediment. Chest X-ray revealed bilateral pulmonary infiltrates. Broad-spectrum antibiotics and intravenous corticosteroids were started. Bronchoscopy showed alveolar hemorrhage and smears from bronchial lavage from both lungs were positive for acid fast bacillus (AFB). Vasculitis work-up revealed high titers of c-ANCA and anti-GBM antibodies. Kidney biopsy revealed crescents in >50% glomeruli on light microscopy. Immunofluorescence showed linear deposition of IgG and C3. The patient received pulse methylprednisone for three days followed by oral prednisone and ATT. In addition, he also underwent nine sessions of plasmapheresis. Oral Cyclophosphamide was added on Day 10. The patient showed remarkable recovery as his lung fields cleared and his kidney function got stabilized. Cyclophosphamide was continued for three months and then switched to azathioprine. At six months, the creatinine is 1.2 mg/dL, with minimal proteinuria and a normal chest X-ray. To the best of our knowledge, this is the only reported case of double-positive Goodpasture's syndrome (c-ANCA and anti GBM) with active tuberculosis treated successfully.

  17. The orphan adapter protein SLY1 as a novel anti-apoptotic protein required for thymocyte development

    PubMed Central

    Reis, Bernhard; Pfeffer, Klaus; Beer-Hammer, Sandra

    2009-01-01

    Background SH3 containing Lymphocyte Protein (SLY1) is a putative adapter protein exclusively expressed in lymphocytes which is involved in antigen receptor induced activation. We previously have generated SLY1Δ/Δ mice harbouring a partial deletion in the N-terminal region of SLY1 which revealed profound immunological defects in T and B cell functions. Results In this study, T cell development in SLY1-/- and SLY1Δ/Δ mice was analysed ex vivo and upon cultivation with the bone marrow stromal cell line OP9. SLY1-deficient thymocytes were compromised in inducing nutrient receptor expression and ribosomal protein S6 phosphorylation, indicating a defect in mTOR complex activation. Furthermore, SLY1 was identified as a novel anti-apoptotic protein required for developmental progression of T cell precursors to the CD4+CD8+ double-positive stage by protecting from premature programmed cell death initiation in developing CD4-CD8- double-negative thymocytes. In addition, SLY1 phosphorylation was differentially regulated upon Notch ligand-mediated stimulation and expression of the preTCR. Conclusion Thus, our results suggest a non-redundant role for SLY1 in integrating signals from both receptors in early T cell progenitors in the thymus. PMID:19604361

  18. CD5-CK2 Signaling Modulates Erk Activation and Thymocyte Survival

    PubMed Central

    Mier-Aguilar, Carlos A.; Cashman, Kevin S.; Raman, Chander; Soldevila, Gloria

    2016-01-01

    CD5 is well recognized for its importance in thymic selection. Although this property of CD5 has been attributed to its ITIM-domain dependent regulation of TCR-signal strength, the mechanism has not been established. A second major signaling domain within the cytoplasmic tail of CD5 is a CK2 binding/activation domain (CD5-CK2BD). Using a gene-targeted mouse in which the CD5-CK2BD is selectively ablated (CD5-ΔCK2BD), we determined that loss of function of CD5-CK2 signaling in a MHC-II selecting TCR transgenic (OT-II) mouse resulted in decrease in double positive (DP) thymocytes, which correlated with enhanced apoptosis. Remarkably, DP cells expressing high levels of CD5 and CD69 and single positive (CD4+SP) thymocytes were increased in CD5-ΔCK2BD mice indicating that CD5-CK2 signaling regulates positive selection and promotes survival. Consistent with this possibility, we determined that the activation and nuclear localization of ERK as well as apoptosis was greater in thymic populations from OTII CD5-ΔCK2BD mice than OTII CD5-WT mice following injection of OVA323-339-peptide. The mobilization of Ca2+, an early event of TCR activation, was not altered by the loss of CD5-CK2 signaling. Collectively, these data demonstrate that the CD5-CK2 signaling axis regulates positive selection by modulating activation of ERK and promoting survival independent of proximal TCR signals. PMID:28030587

  19. Synthesis and apparent secretion of prothymosin alpha by different subpopulations of calf and rat thymocytes.

    PubMed Central

    Franco, F J; Diaz, C; Barcia, M; Arias, P; Gomez-Marquez, J; Soriano, F; Mendez, E; Freire, M

    1989-01-01

    Prothymosin alpha (ProT alpha), a polypeptide containing the sequence of thymosin alpha 1 (T alpha 1) at its NH2-terminus, has been isolated from calf thymocytes in a concentration in the order of that found in the whole thymic gland. As deduced from the analysis of their tryptic peptides, calf ProT alpha differs from the rat polypeptide at least in an alanine residue replacing valine at position 92. Thymocytes cultured in a radioactive medium exhibit an important secretory activity, ProT alpha being one of the products synthesized and exported to the culture medium. Large and small thymocyte subpopulations from calf and rat differ in their capacity to synthesize ProT alpha. The polypeptide is produced in a major concentration by large thymocytes. However, all the calf and rat thymocyte subpopulations show a similar capacity to secrete ProT alpha, the amount of the newly synthesized polypeptide recovered from cell culture supernatants being 80-90% of that found in thymocyte extracts. Images Figure 1 PMID:2787781

  20. Membranes as Sensitive Targets in Thymocyte Apoptosis

    DTIC Science & Technology

    1993-01-01

    water soluble analogue function. Apoptotic death is, on the other hand, a of vitamin E and inhibitor of membrane damage , inhibits DNA fragmentation in...participate in the process, or how their damage irradiated cells. This suggests that trolox irreversibly inhibits a results in cell death. The most...of Ca2" in irradiated thymocytes was more than twice precedes cell death (Ramakrishnan and Catravas that of unirradiated thymocytes. Membrane damage

  1. CAML regulates Bim-dependent thymocyte death

    PubMed Central

    Edgar, Contessa E.; Lindquist, Lonn D.; McKean, David L.; Strasser, Andreas; Bram, Richard J.

    2010-01-01

    Appropriate control of apoptosis during T lymphocyte differentiation is critical for destruction of T cells bearing potentially autoreactive or useless immuno-receptors, and for survival of those T cells bearing antigen receptors that may recognize foreign proteins. Despite the well-established importance of thymocyte survival, the exact signals regulating thymocyte apoptosis have not been fully elucidated. Here, we show that thymocytes lacking the endoplasmic reticulum (ER) protein CAML failed to undergo normal T cell development and exhibited dramatically increased rates of apoptosis. In vitro, CAML-deficient thymocytes accumulated high levels of reactive oxygen species (ROS) and underwent abnormally accelerated death in response to several cytotoxic stimuli, including treatment with etoposide, cytokine deprivation or Fas ligation. Although neither p53 deletion nor loss of Fas rescued the survival and continued development of CAML-deficient thymocytes, removal of the pro-apoptotic BH3-only Bcl-2 family member Bim significantly restored their survival. This work reveals CAML to be a critically important regulator of ROS- and Bim-dependent thymocyte death. PMID:20300112

  2. Fringe positions in double-exposure speckle photography.

    PubMed

    Hinsch, K

    1989-12-15

    Double-exposure records in speckle photography or particle image velocimetry are often evaluated by analysis of the system of Young's diffraction fringes. Fringe spacing, necessary to calculate the displacement, is determined from the positions of fringe maxima or minima. These, however, are influenced by the diffraction halo function and by fringe visibility. A generalized theory of the effects is presented, including position dependent visibility and fringe phase. Evaluations are given for disk-shaped particle images in particle image velocimetry, and for coherent and incoherent speckle photography. Fringe shifts are determined numerically for commonly encountered values of fringe density and visibility thus presenting a basis for rapid assessment of accuracy in metrological experiments.

  3. Proteasome activities decrease during dexamethasone-induced apoptosis of thymocytes.

    PubMed

    Beyette, J; Mason, G G; Murray, R Z; Cohen, G M; Rivett, A J

    1998-06-01

    The induction of apoptosis in thymocytes by the glucocorticoid dexamethasone was used as a model system to investigate whether there are changes in 20 S and 26 S proteasome activities during apoptosis. We observed that thymocytes contain high concentrations of proteasomes and that following treatment with dexamethasone, cell extracts showed a decrease in proteasome chymotrypsin-like activity which correlated with the degree of apoptosis observed. The decrease in chymotrypsin-like activity of 20 S and 26S proteasomes was still apparent after these complexes had been partially purified from apoptotic thymocyte extracts and was therefore not due to competition resulting from a general increase in protein turnover. The trypsin-like and peptidylglutamylpeptide hydrolase activities of proteasome complexes were also observed to decrease during apoptosis, but these decreases were reversed by the inhibition of apoptosis by the caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp(OMe)-fluoromethylketone. However, the chymotrypsin-like activity of proteasomes decreased further in the presence of the apoptosis inhibitor. Val-Ala-Asp-fluoromethylketone was found to inhibit the chymotrypsin- and trypsin-like activity of 26 S proteasomes in vitro. The decrease in proteasome activities in apoptosis did not appear to be due to a decrease in the concentration of total cellular proteasomes. Thus, the early decreases in 20 S and 26 S proteasome activities during apoptosis appear to be due to a down-regulation of their proteolytic activities and not to a decrease in their protein concentration. These data suggest that proteasomes may be responsible, in thymocytes, for the turnover of a protein that functions as a positive regulator of apoptosis.

  4. Presenilin-dependent γ-secretase activity modulates thymocyte development

    PubMed Central

    Doerfler, Petra; Shearman, Mark S.; Perlmutter, Roger M.

    2001-01-01

    In neuronal cells, presenilin-dependent γ-secretase activity cleaves amyloid precursor proteins to release Aβ peptides, and also catalyzes the release of the intracellular domain of the transmembrane receptor Notch. Accumulation of aberrant Aβ peptides appears to be causally related to Alzheimer's disease. Inhibition of Aβ peptide production is therefore a potential target for therapeutic intervention. Notch proteins play an important role in cell fate determination in many different organisms and at different stages of development, for example in mammalian T cell development. We therefore addressed whether structurally diverse γ-secretase inhibitors impair Notch function by studying thymocyte development in murine fetal thymic organ cultures. Here we show that high concentrations of the most potent inhibitors blocked thymocyte development at the most immature stage. In contrast, lower concentrations or less potent inhibitors impaired differentiation at a later stage, most notably suppressing the development of CD8 single-positive T cells. These phenotypes are consistent with an impairment of Notch signaling by γ-secretase inhibitors and define a strict Notch dose dependence of consecutive stages during thymocyte development. PMID:11470902

  5. Mask image position correction for double patterning lithography

    NASA Astrophysics Data System (ADS)

    Saito, Masato; Itoh, Masamitsu; Ikenaga, Osamu; Ishigo, Kazutaka

    2008-05-01

    Application of double patterning technique has been discussed for lithography of HP 3X nm device generation. In this case, overlay budget for lithography becomes so hard that it is difficult to achieve it with only improvement of photomask's position accuracy. One of the factors of overlay error will be induced by distortion of photomask after chucking on the mask stage of exposure tool, because photomasks are bended by the force of vacuum chucking. Recently, mask flatness prediction technique was developed. This technique is simulating the surface shape of mask when it is on the mask stage by using the flatness data of free-standing state blank and the information of mask chucking stage. To use this predicted flatness data, it is possible to predict a pattern position error after exposed and it is possible to correct it on the photomask. A blank supplier developed the flatness data transfer system to mask vender. Every blanks are distinguished individually by 2D barcode mark on blank which including serial number. The flatness data of each blank is linked with this serial number, and mask vender can use this serial number as a key code to mask flatness data. We developed mask image position correction system by using 2D barcode mark linked to predicted flatness data, and position accuracy assurance system for these masks. And with these systems, we made some masks actually.

  6. Effect of dietary argan oil on fatty acid composition, proliferation, and phospholipase D activity of rat thymocytes.

    PubMed

    Benzaria, Amal; Meskini, Nadia; Dubois, Madeleine; Croset, Martine; Némoz, Georges; Lagarde, Michel; Prigent, Annie-France

    2006-06-01

    Argan oil is receiving increasing attention due to its potential health benefits in the prevention of cardiovascular risk, but no information to date is available about its possible effect on immune cells and functions. To address this issue male rats were fed one of five diets that contained fish oil, argan oil, olive oil, coconut oil, or sunflower oil for 4 wk. The fatty acid composition of plasma and thymocyte lipids was then analyzed in relation to the mitogen-induced proliferation and phospholipase D (PLD) activity of thymocytes. The 18:2omega-6 proportion in thymocyte phospholipids from rats fed argan oil was significantly lower than that observed in phospholipids from rats fed sunflower oil and fish oil but higher than that found in the olive oil and coconut oil groups. Further, a significant positive linear relation was found between thymocyte proliferation and the 18:2omega-6 proportion in thymocyte phospholipids, whatever the diet. The proliferation response of thymocytes to mitogenic activation was also inversely correlated to PLD activity measured in intact thymocytes. Subsequent western blotting experiments indicated that the diet-induced variations in PLD activity mainly reflected variations in the expression of PLD2 protein. On the whole, the present study shows that the effects of argan oil on immune cells are very similar to those of olive oil, and that, as a consequence, argan oil can be used as a balanced dietary supply without marked adverse effects on immune cell function.

  7. Positive cooperative regulation of double binding sites for human acetylcholinesterase.

    PubMed

    Liu, Hao; Ye, Wei; Chen, Hai-Feng

    2016-10-25

    Acetylcholinesterase is a potent enzyme that regulates neurotransmission by rapidly hydrolyzing the neurotransmitter acetylcholine in synapses of the nervous system. As drug target of anti-AD, it has catalytic and peripheral anionic sites. However, the regulation relation between these two sites is unclear. Therefore, we constructed dynamics fluctuation network based on all-atom molecular dynamics simulations to reveal the regulation mechanism. The results suggest that the correlation network in double-site system (hAChE/TZ5) is distinctly different from that in the free state and single-site systems (hAChE/huprine and hAChE/1YL). The community network analysis indicates that the information freely transfers from the peripheral anionic site to the catalytic active site in hAChE/TZ5. Furthermore, the binding free energy between the inhibitor and hAChE for hAChE/TZ5 is significantly lower than of either hAChE/huprine or hAChE/1YL. Thus, a hypothesis of 'positive cooperative regulation' is proposed for the regulation of double binding sites and further confirmed by the weakening and mutation community analyses. Finally, one possible cooperative regulation pathway of W86-TZ5-W286 was identified based on the shortest path algorithm and was confirmed by the network perturbation analysis. Interestingly, the regulation pathway for single-site systems is significantly different from that of dual-site system. The process targeting on the shortest pathway can better regulate the hydrolyzing the neurotransmitter acetylcholine and significantly inhibit the aggregation of Aβ amyloid.

  8. Neuroendocrine control of T cell development in mammals: role of growth hormone in modulating thymocyte migration.

    PubMed

    Savino, Wilson

    2007-09-01

    The thymus gland is a primary lymphoid organ, in which bone-marrow-derived T cell precursors undergo differentiation, eventually leading to migration of positively selected cells to the peripheral lymphoid organs. This differentiation occurs along with cell migration in the context of the thymic microenvironment, a three-dimensional network formed by epithelial cells, macrophages, dendritic cells, fibroblasts and extracellular matrix components. A series of data clearly shows that growth hormone (GH) pleiotropically modulates thymic functions. For example, GH upregulates proliferation of thymocytes and thymic epithelial cells. Accordingly, GH-transgenic mice, as well as animals and humans treated with exogenous GH, exhibit an enhanced cellularity in the organ. Growth hormone stimulates the secretion of thymic hormones, cytokines and chemokines by the thymic microenvironment, as well as the production of extracellular matrix proteins, leading to an increase in thymocyte migratory responses and intrathymic traffic of developing T cells. In addition, GH stimulates the in vivo export of thymocytes from the organ, as ascertained by studies with intrathymic injection of GH in normal mice and with GH-transgenic mice. Moreover, since GH is produced by thymocytes and thymic epithelial cells, which express GH receptors, we should consider that, in addition to the classic endocrine pathway, the GH control of the thymus may include an autocrine/paracrine pathway. Finally, since GH promotes a replenishment of the thymus and an increase of thymocyte export, it could be envisioned as a potential adjuvant therapeutic agent in the treatment of immunodeficiencies associated with thymic atrophy.

  9. Critical role of SP thymocyte motility in regulation of thymic output in neonatal Aire-/- mice.

    PubMed

    Jin, Rong; Aili, Abudureyimujiang; Wang, Yuqing; Wu, Jia; Sun, Xiuyuan; Zhang, Yu; Ge, Qing

    2017-01-03

    Autoimmune regulator (Aire) is essential in the perinatal period to prevent the multiorgan autoimmunity. Here we show that Aire-regulated single positive thymocyte trafficking in neonatal period is critical for thymic egress. Reduced thymic emigration was found in Aire-/- mice during neonatal period, leading to enhanced homeostatic expansion of peripheral T cells as early as 2 weeks of age. In neonatal Aire-/- mice, thymic expression of CCR7 ligands were dramatically reduced, resulting in decreased thymocyte motility and thymocyte emigration. This reduction of thymic egress in Aire-/- mice was alleviated beyond 3 weeks of age by an early upregulation of S1P1 signaling. As the numbers and quality of thymic emigrants are essential for the establishment and maintenance of peripheral tolerance, the reduced thymic emigration during neonatal period may deteriorate autoimmunity caused by the emigration of autoreactive T cells.

  10. Ontogeny of fetal CD8lo4lo thymocytes: expression of CD44, CD25 and early expression of TcR alpha mRNA.

    PubMed

    Andjelić, S; Jain, N; Nikolić-Zugić, J

    1993-09-01

    CD8lo4lo cells are the immediate precursors of immature CD8hi4loTcRlo, CD8lo4hiTcRlo and CD8hi4hiTcRlo double-positive (DP) thymocytes in the adult murine thymus. These cells are the first subset in the adult thymus to express accessory CD8 and CD4 molecules, to rearrange the T cell receptor (TcR) alpha chain genes and to express the TcR alpha beta heterodimer at low levels at the surface. Here, we investigate the fetal ontogeny of CD8lo4lo cells. We detect these cells on day 15 of fetal development. They dominate the thymus on day 15.5, to become progressively less prominent thereafter. An important characteristic of fetal CD8lo4lo cells is the early expression of TcR alpha mRNA (on fetal day 15, 36-48 h earlier than reported previously). Our results also suggest, but do not prove, that the receptor may be expressed on the surface as early as day 15.5. Fetal CD8lo4lo cells, like their adult counterparts, become DP in vitro. However, early fetal CD8lo4lo thymocytes express both CD44 and CD25--unlike the adult subset--and that links them to their putative precursors, fetal CD44+CD25+ double-negative cells. This finding underscores the difference between adult and fetal thymocytes in turnover of membrane molecules and/or the kinetics of progression through phenotypic stages.

  11. Cutting Edge: CD3 ITAM Diversity Is Required for Optimal TCR Signaling and Thymocyte Development.

    PubMed

    Bettini, Matthew L; Chou, Po-Chein; Guy, Clifford S; Lee, Thomas; Vignali, Kate M; Vignali, Dario A A

    2017-09-01

    For the αβ or γδTCR chains to integrate extracellular stimuli into the appropriate intracellular cellular response, they must use the 10 ITAMs found within the CD3 subunits (CD3γε, CD3δε, and ζζ) of the TCR signaling complex. However, it remains unclear whether each specific ITAM sequence of the individual subunit (γεδζ) is required for thymocyte development or whether any particular CD3 ITAM motif is sufficient. In this article, we show that mice utilizing a single ITAM sequence (γ, ε, δ, ζa, ζb, or ζc) at each of the 10 ITAM locations exhibit a substantial reduction in thymic cellularity and limited CD4(-)CD8(-) (double-negative) to CD4(+)CD8(+) (double-positive) maturation because of low TCR expression and signaling. Together, the data suggest that ITAM sequence diversity is required for optimal TCR signal transduction and subsequent T cell maturation. Copyright © 2017 by The American Association of Immunologists, Inc.

  12. Scavenger receptor BI and HDL regulate thymocyte apoptosis in sepsis

    PubMed Central

    Guo, Ling; Zheng, Zhong; Ai, Junting; Howatt, Deborah A.; Mittelstadt, Paul R.; Thacker, Seth; Daugherty, Alan; Ashwell, Jonathan D.; Remaley, Alan T.; Li, Xiang-An

    2014-01-01

    Objective Thymocyte apoptosis is a major event in sepsis; however, how this process is regulated remains poorly understood. Approach and Results Septic stress induces glucocorticoids (GC) production which triggers thymocyte apoptosis. Here, we used scavenger receptor BI (SR-BI) null mice, which are completely deficient in inducible GC (iGC) in sepsis, to investigate the regulation of thymocyte apoptosis in sepsis. Cecal ligation and puncture (CLP) induced profound thymocyte apoptosis in SR-BI+/+ mice, but no thymocyte apoptosis in SR-BI−/− mice due to lack of iGC. Unexpectedly, supplementation of GC only partly restored thymocyte apoptosis in SR-BI−/− mice. We demonstrated that HDL is a critical modulator for thymocyte apoptosis. SR-BI+/+ HDL significantly enhanced GC-induced thymocyte apoptosis but SR-BI−/− HDL had no such activity. Further study revealed that SR-BI+/+ HDL modulates GC-induced thymocyte apoptosis via promoting glucocorticoid receptor translocation, but SR-BI−/− HDL loses such regulatory activity. To understand why SR-BI−/− HDL loses its regulatory activity, we analyzed HDL cholesterol contents. There was 3-fold enrichment of unesterified cholesterol in SR-BI−/− HDL compared with SR-BI+/+ HDL. Normalization of unesterified cholesterol in SR-BI−/− HDL by probucol administration or LCAT expression restored GC-induced thymocyte apoptosis, and incorporating unesterified cholesterol into SR-BI+/+ HDL rendered SR-BI+/+ HDL dysfunctional. Using lckCre-GRfl/fl mice in whom thymocytes lack CLP-induced thymocyte apoptosis, we showed that lckCre-GRfl/fl mice were significantly more susceptible to CLP-induced septic death than GRfl/fl control mice, suggesting that GC-induced thymocyte apoptosis is required for protection against sepsis. Conclusions The findings in this study reveal a novel regulatory mechanism of thymocyte apoptosis in sepsis by SR-BI and HDL. PMID:24603680

  13. CD28 in thymocyte development and peripheral T cell activation in mice exposed to suspended particulate matter

    SciTech Connect

    Drela, Nadzieja . E-mail: ndrela@biol.uw.edu.pl; Zesko, Izabela; Jakubowska, Martyna; Biernacka, Marzena

    2006-09-01

    The CD28:B7 signaling pathway is very important for the activity of mature peripheral T lymphocytes and thymocyte development. The proper development of thymocytes into mature single positive CD4{sup +}and CD8{sup +} T cells is crucial for almost all immune functions. In naturally occurring conditions, T cells maturation in the thymus is influenced by environmental agents. The expression of CD28 and the distribution of CD28{sup low/high} thymocytes have been examined at various stages of thymocyte development in BALB/c mice exposed to air-suspended particulate matter (ASM). Acute exposure to ASM resulted in the decrease of CD28 expression in the total thymocyte population. The increase of the percentage of CD28{sup low} and the decrease of CD28{sup high} thymocytes were observed, which may account for the acceleration of thymocyte development under the conditions of elevated risk resulting from the exposure of animals to environmental xenobiotics. ASM exposure resulted in the increase of the level of proliferation of lymph node T cells induced by anti-CD3 and anti-CD28 monoclonal antibodies activation despite normal expression of CD28 molecule. In contrast, the level of proliferation of spleen T cells was lowered or normal dependently of the concentration of stimuli used for activation. Results of these studies demonstrate that acute exposure of mice to ASM can result in the progression of two contrasting processes in the immune system: upregulation of thymocyte development, which contributes to the maintenance of peripheral T cell pool, and over-activation of lymph node lymphocytes, which may lead to uncontrolled immunostimulation.

  14. Fli-1 regulates the DN2 to DN3 thymocyte transition and promotes γδ T-cell commitment by enhancing TCR signal strength.

    PubMed

    Smeets, Monique F M A; Wiest, David L; Izon, David J

    2014-09-01

    Friend leukemia integration 1 (Fli-1) is a member of the Ets transcription factor family and is expressed during T-cell development; however, the role Fli-1 plays in early T-cell differentiation has not been elucidated. In this report, we demonstrate that in mouse, Fli-1 overexpression retards the CD4(-) CD8(-) double-negative (DN) to CD4(+) CD8(+) double-positive (DP) transition by deregulating normal DN thymocyte development. Specifically, Fli-1 expression moderates the DN2 and DN3 developmental transitions. We further show that Fli-1 overexpression partially mimics strong TCR signals in developing DN thymocytes and thereby enhances γδ T-cell development. Conversely, Fli-1 knockdown by small hairpin RNA reverses the lineage bias from γδ T cells and directs DN cells to the αβ lineage by attenuating TCR signaling. Therefore, Fli-1 plays a critical role in both the DN2 to DN3 transition and αβ/γδ lineage commitment.

  15. A selective plasmin inhibitor, trans-aminomethylcyclohexanecarbonyl-L-(O-picolyl)tyrosine-octylamide (YO-2), induces thymocyte apoptosis.

    PubMed

    Lee, Eibai; Enomoto, Riyo; Takemura, Kazu; Tsuda, Yuko; Okada, Yoshio

    2002-04-01

    The treatment of rat thymocytes with YO-2, a novel inhibitor of plasmin, resulted in an increase in DNA fragmentation. DNA fragmentation was also induced by another YO compounds such as YO-0, -3, -4 and -5. These YO compounds are the inhibitor of plasmin activity. On the other hand, YO-1, -6 and -8 that hardly inhibit plasmin activity had no effect on DNA fragmentation. Analysis of fragmented DNA from thymocytes treated with YO-2 by agarose gel electrophoresis revealed that the compound caused internucleosomal DNA fragmentation. In addition, judging from a laser scanning microscopy, annexin V-positive and propidium iodide-negative cells were increased by the treatment of the cells with the compound. Moreover, chromatin condensation was observed in thymocytes treated with the compound. These results demonstrated that YO-2 induces thymocyte apoptosis. There seemed to be some correlation between the apoptosis induced by YO compounds and their plasmin inhibitory effect. However, because the other protease inhibitors including pepstatin A, leupeptin, AEBSF, DFP and E-64-d did not affect DNA fragmentation, YO compounds are likely to have unique mechanism on plasmin or to show the effect on the other plasmin-like proteases. The plasmin inhibitory activity may have an important role in YO-2-induced apoptosis. Furthermore, the stimulations of caspase-8, -9 and -3-like activities were observed in thymocytes treated with YO-2. These results suggest that YO-2 induces thymocyte apoptosis via activation of caspase cascade.

  16. Temporal Expression of Bim Limits the Development of Agonist-Selected Thymocytes and Skews Their TCRβ Repertoire

    PubMed Central

    Li, Kun-Po; Fahnrich, Anke; Roy, Eron; Cuda, Carla M.; Grimes, H. Leighton; Perlman, Harris R.; Kalies, Kathrin; Hildeman, David A.

    2017-01-01

    CD8αα TCRαβ+ intestinal intraepithelial lymphocytes play a critical role in promoting intestinal homeostasis, although mechanisms controlling their development and peripheral homeostasis remain unclear. In this study, we examined the spatiotemporal role of Bim in the thymic selection of CD8αα precursors and the fate of these cells in the periphery. We found that T cell–specific expression of Bim during early/cortical, but not late/medullary, thymic development controls the agonist selection of CD8αα precursors and limits their private TCRβ repertoire. During this process, agonist-selected double-positive cells lose CD4/8 coreceptor expression and masquerade as double-negative (DN) TCRαβhi thymocytes. Although these DN thymocytes fail to re-express coreceptors after OP9-DL1 culture, they eventually mature and accumulate in the spleen where TCR and IL-15/STAT5 signaling promotes their conversion to CD8αα cells and their expression of gut-homing receptors. Adoptive transfer of splenic DN cells gives rise to CD8αα cells in the gut, establishing their precursor relationship in vivo. Interestingly, Bim does not restrict the IL-15–driven maturation of CD8αα cells that is critical for intestinal homeostasis. Thus, we found a temporal and tissue-specific role for Bim in limiting thymic agonist selection of CD8αα precursors and their TCRβ repertoire, but not in the maintenance of CD8αα intraepithelial lymphocytes in the intestine. PMID:27852740

  17. Early Thymocyte Development Is Regulated by Modulation of E2a Protein Activity

    PubMed Central

    Engel, Isaac; Johns, Carol; Bain, Gretchen; Rivera, Richard R.; Murre, Cornelis

    2001-01-01

    The E2A gene encodes the E47 and E12 basic helix-loop-helix (bHLH) transcription factors. T cell development in E2A-deficient mice is partially arrested before lineage commitment. Here we demonstrate that E47 expression becomes uniformly high at the point at which thymocytes begin to commit towards the T cell lineage. E47 protein levels remain high until the double positive developmental stage, at which point they drop to relatively moderate levels, and are further downregulated upon transition to the single positive stage. However, stimuli that mimic pre-T cell receptor (TCR) signaling in committed T cell precursors inhibit E47 DNA-binding activity and induce the bHLH inhibitor Id3 through a mitogen-activated protein kinase kinase–dependent pathway. Consistent with these observations, a deficiency in E2A proteins completely abrogates the developmental block observed in mice with defects in TCR rearrangement. Thus E2A proteins are necessary for both initiating T cell differentiation and inhibiting development in the absence of pre-TCR expression. Mechanistically, these data link pre-TCR mediated signaling and E2A downstream target genes into a common pathway. PMID:11560990

  18. In vitro proliferation and cloning of CD3- CD16+ cells from human thymocyte precursors

    PubMed Central

    1991-01-01

    Purified CD3-4- thymocytes were obtained by depletion of CD3+ and CD4+ cells from fresh thymocyte suspensions. 5-15% of these cells were found to express CD16 antigen, while other natural killer (NK) cell markers were virtually absent. Double fluorescence analysis revealed that 20- 40% of thymic CD16+ cells coexpressed CD1, while approximately half were cyCD3+. When cultured in the presence of peripheral blood lymphocytes and H9 leukemia cell line as a source of irradiated feeder cells and interleukin 2 (IL-2), CD3-4- thymocytes underwent extensive proliferation. In addition, after 1-2 wk of culture, 30-50% of these cells were found to express CD16 surface antigen. Cloning under limiting dilution conditions of either CD3-4- or CD3-4-16- thymocytes in the presence of irradiated H9 cells resulted in large proportions (approximately 50%) of CD16+ clones. On the basis of the expression of surface CD16 and/or cyCD3 antigen, clones could be grouped in the following subsets: CD16+ cyCD3+; CD16+ cyCD3-; CD16- cyCD3+; and CD16- cyCD3-. All clones expressed CD56 surface antigen, displayed a strong cytolytic activity against NK sensitive (K562) and NK-resistant (M14) target cells, and produced IFN-gamma and tumor necrosis factor, but not IL-2. Similar to peripheral NK cells, thymic CD16+ cells expressed transcripts for CD16 and for CD3 epsilon (Biassoni, R., S. Ferrini, I. Prigione, A. Moretta, and E.O. Long, 1988. J. Immunol. 140:1685.) and zeta chains (Anderson, P., M. Caligiuri, J. Ritz, and S.F. Schlossman. 1989. Nature [Lond.]. 341:159). Therefore, it appears that cells that are phenotypically and functionally similar to CD3- CD16+ NK cells may arise from immature thymocytes. PMID:1711562

  19. Pak2 is required for actin cytoskeleton remodeling, TCR signaling, and normal thymocyte development and maturation

    PubMed Central

    Phee, Hyewon; Au-Yeung, Byron B; Pryshchep, Olga; O'Hagan, Kyle Leonard; Fairbairn, Stephanie Grace; Radu, Maria; Kosoff, Rachelle; Mollenauer, Marianne; Cheng, Debra; Chernoff, Jonathan; Weiss, Arthur

    2014-01-01

    The molecular mechanisms that govern thymocyte development and maturation are incompletely understood. The P21-activated kinase 2 (Pak2) is an effector for the Rho family GTPases Rac and Cdc42 that regulate actin cytoskeletal remodeling, but its role in the immune system remains poorly understood. In this study, we show that T-cell specific deletion of Pak2 gene in mice resulted in severe T cell lymphopenia accompanied by marked defects in development, maturation, and egress of thymocytes. Pak2 was required for pre-TCR β-selection and positive selection. Surprisingly, Pak2 deficiency in CD4 single positive thymocytes prevented functional maturation and reduced expression of S1P1 and KLF2. Mechanistically, Pak2 is required for actin cytoskeletal remodeling triggered by TCR. Failure to induce proper actin cytoskeletal remodeling impaired PLCγ1 and Erk1/2 signaling in the absence of Pak2, uncovering the critical function of Pak2 as an essential regulator that governs the actin cytoskeleton-dependent signaling to ensure normal thymocyte development and maturation. DOI: http://dx.doi.org/10.7554/eLife.02270.001 PMID:24843022

  20. Aire deficiency results in decreased expression of CCR4 and CCR7 ligands and in delayed migration of CD4+ thymocytes1

    PubMed Central

    Laan, Martti; Kisand, Kai; Kont, Vivian; Möll, Kaidi; Tserel, Liina; Scott, Hamish S.; Peterson, Pärt

    2009-01-01

    Autoimmune regulator (Aire) has been viewed as a central player in the induction of tolerance. This study examines, whether Aire can modulate the production of the thymic chemokines involved in cortico-medullary migration and thus play a role in intrathymic thymocyte migration and maturation. Aire deficiency resulted in reduced gene expression and protein levels of the CCR4 and CCR7 ligands in whole thymi of mice, as determined by quantitative PCR analysis and ELISA. The expression of the CCR4 ligands coincided with Aire expression in the CD80high medullary epithelial cells (mTECs), whereas the expression of the CCR7 ligands was detected in other cell populations. Also, the expression pattern of the CCR4 and CCR7 ligands follows that of Aire during postnatal but not during embryonic development. In vitro, overexpression of Aire resulted in an upregulation of selected CCR4 and CCR7 ligands, which induced selective migration of double-positive (DP) and single positive (SP) CD4+ cells. In vivo, Aire deficiency resulted in a diminished emigration of mature CD4+ T-cells from the thymi of 5-day-old mice In conclusion, Aire regulates the production of CCR4 and CCR7 ligands in mTECs and alters the coordinated maturation and migration of thymocytes. These results suggest a novel mechanism behind the Aire-dependent induction of central tolerance. PMID:19923453

  1. Selective inhibition of immature CD4-CD8+ thymocyte proliferation, but not differentiation, by the thymus atrophy-inducing compound di-n-butyltin dichloride.

    PubMed Central

    Pieters, R H; Bol, M; Ariëns, T; Punt, P; Seinen, W; Bloksma, N; Penninks, A H

    1994-01-01

    Effects of the thymus atrophy-inducing organotin compound di-n-butyltin dichloride (DBTC) on the differentiation and proliferation of immature rat thymocyte subsets were studied in vivo and in vitro. Incubation of freshly isolated CD4-CD8- or immature CD4-CD8+ (characterized as CD4-CD53-) thymocytes with 10(-7) M DBTC for 18 hr did not affect cell recovery or their ability to differentiate to CD4-CD8+ cells and CD4+CD8+ or to CD4+CD8+ cells, respectively. The same treatment decreased the spontaneous as well as the phytohaemagglutinin (PHA)-induced proliferation in both subsets. However, the inhibition of proliferation by DBTC of immature CD4-CD8+, but not of CD4-CD8- thymocytes, appeared to increase with their growth rate. Data show that differentiation of immature thymocytes can proceed independently of proliferation and that DBTC causes thymus atrophy by selectively inhibiting the proliferation of immature CD4-CD8+ thymocytes. Administration to rats of DBTC via the diet for 14 days resulted in an initial decrease of thymoblast number by day 2, followed by a decrease in the total number of thymocytes by day 4. Total thymocyte numbers were lowest on day 7 and did not significantly change thereafter. CD4/CD8 thymocyte subset distributions were similar to controls on day 4, but on day 7 of feeding a marked reduction of the percentage of CD4+CD8+ thymocytes and consequently an increase of the percentages of the three other CD4/CD8 subsets were found. Thereafter, the CD4/CD8 subset distribution recovered, reaching near control values on day 14, despite the very low numbers of thymoblasts and of total thymocytes at that time. Data together indicate that DBTC reduces the production of CD4+CD8+ and mature single-positive thymocytes by selectively inhibiting immature CD4-CD8+ thymocyte proliferation but without affecting the differentiation capacity of these cells. This suggests that thymocyte proliferation and differentiation are separately regulated processes. PMID

  2. Targeted loss of SHP1 in murine thymocytes dampens TCR signaling late in selection.

    PubMed

    Martinez, Ryan J; Morris, Anna B; Neeld, Dennis K; Evavold, Brian D

    2016-09-01

    SHP1 is a tyrosine phosphatase critical to proximal regulation of TCR signaling. Here, analysis of CD4-Cre SHP1(fl/fl) conditional knockout thymocytes using CD53, TCRβ, CD69, CD4, and CD8α expression demonstrates the importance of SHP1 in the survival of post selection (CD53(+) ), single-positive thymocytes. Using Ca(2+) flux to assess the intensity of TCR signaling demonstrated that SHP1 dampens the signal strength of these same mature, postselection thymocytes. Consistent with its dampening effect, TCR signal strength was also probed functionally using peptides that can mediate selection of the OT-I TCR, to reveal increased negative selection mediated by lower-affinity ligand in the absence of SHP1. Our data show that SHP1 is required for the survival of mature thymocytes and the generation of the functional T-cell repertoire, as its absence leads to a reduction in the numbers of CD4(+) and CD8(+) naïve T cells in the peripheral lymphoid compartments.

  3. Characterization of thymic atrophy and the mechanism of thymocyte depletion after in vivo exposure to a mixture of herbicides.

    PubMed

    de la Rosa, Patricia; Barnett, John B; Schafer, Rosana

    2005-01-22

    3,4-Dichloropropionanilide (propanil) and 2,4-dichlorophenoxyacetic acid (2,4-D) are two commonly used herbicides that are marketed as a chemical mixture. It was hypothesized that the interaction between these two herbicides, when administered as a mixture, would result in a greater effect on the immune system than the individual components of the mixture. The present study demonstrates in a murine model that a mixture of propanil and 2,4-D, when compared to single herbicide exposures, exacerbates decreases in thymocyte populations 2 d postexposure and inhibits the repopulation of T-cells in the thymus 7 d postexposure. Exposure to 150 mg herbicide/kg body weight of propanil or 2,4-D alone had no effect on thymus weight. In contrast, decreases in the ratio of thymus weight to body weight (TW:BW) occurred 2 d after treatment with the mixture of 150 mg propanil/kg body weight + 150 mg 2,4-D/kg body weight (150/150). Thymic atrophy was associated with a decrease in the double-positive thymocyte population (CD4+CD8+) and correlated with sera corticosterone levels from 600 to 1000 pg/ml. Therefore, the hypothesis was tested that glucocorticoids, induced after exposure to herbicides, were responsible for the thymic atrophy and depletion of thymocytes. However, similar levels of corticosterone were induced after exposure to 50, 100, or 150 mg propanil/kg body weight, and 50/50 or 100/100 mixture treatments, doses that did not produce thymic atrophy or cell loss. In addition, RU 486, a glucocorticoid receptor blocker, only partially abrogated the thymic atrophy in mice exposed to the 150/150 mixture of herbicides. These results suggest that glucocorticoids are only partially responsible for herbicide-induced thymic atrophy. This study demonstrates that the effects of exposure to a mixture of chemicals cannot always be predicted based on single exposure data and emphasizes the importance of mixture-based studies.

  4. Evidence that FTY720 induces rat thymocyte apoptosis.

    PubMed

    Isoyama, Naohito; Takai, Kimio; Tsuchida, Masahiro; Matsumura, Masafumi; Naito, Katsusuke

    2006-04-01

    FTY720, a novel immunomodulator with the potential to improve immunosuppressive therapy after organ transplantation, is currently under clinical investigation. FTY720 drastically decreases blood lymphocytes, especially T cells, accelerating lymphocyte homing to secondary lymphoid organs. However, its immunosuppressive effects remain unknown. We investigated these effects in rat thymocytes. Rats were intramuscularly injected with 10mg/kg/day FTY720 or saline for 7days. Thymuses were removed on days 0, 1, 3, 5, 7 and 14 after treatment. Three-color analysis was performed with a flow cytofluorometer. Apoptotic nuclei in the tissue sections were identified by TUNEL. Genomic DNA was then extracted and samples were electrophoresed on 2.0% agarose gel. FTY720 reduced the total number of thymocytes and, with time, significantly reduced the percentage of CD4+8+ TCRalphabeta(negative/low) thymocytes. Light microscopy of thymuses of FTY720-treated rats revealed obvious reductions in the size of the cortical region. TUNEL analysis showed that FTY720 induced thymocyte apoptosis in the cortical region. Furthermore, DNA fragmentation was observed in thymocytes treated with FTY720, indicating thymocyte apoptosis. FTY720 reduced the number of CD4+8+ thymocytes before TCRalphabeta expression resulting in impaired thymocyte differentiation and maturation. This might be an immunosuppressive effect of FTY720.

  5. The kinetics of T cell antigen receptor expression by subgroups of CD4+8+ thymocytes: delineation of CD4+8+3(2+) thymocytes as post- selection intermediates leading to mature T cells

    PubMed Central

    1991-01-01

    Cortical thymocytes from adult mice, separated on the basis of coexpression of CD4 and CD8 or of binding of high levels of peanut agglutinin (PNA), were subdivided according to the level of expression of the T cell receptor (TCR)-CD3 complex. The incidence of dividing cells in the resultant subpopulations was determined by DNA staining. Precursor-product relationships and the timing of TCR-CD3 acquisition were studied using continuous in vivo [3H]TdR labeling and radioautography. The extent of intrathymic selection for TCR specificity in the subpopulations was determined from the incidence of cells bearing V beta 6 or V beta 17a in different mouse strains. The majority of dividing CD4+8+ blast cells expressed extremely low levels of TCR-CD3, indicating that TCR expression and specificity selection generally occurred after division ceased. The [3H]TdR-labeling studies indicated that postdivision TCR expression was rapid, and that those nondividing cortical thymocytes which had not expressed significant levels of TCR by day 1, remained extremely low or negative for their entire 3.6-d lifespan. Small cortical thymocytes which expressed moderate levels of TCR-CD3, were predominantly an unselected population with a lifespan of 3.8 d. A small subgroup of CD4+8+ PNA+ cortical thymocytes expressing high levels of TCR-CD3 was identified as a nondividing intermediate between the small cortical thymocytes expressing moderate levels of TCR and mature medullary thymocytes. These intermediates showed a 1-d lag in [3H]TdR labeling, then a 3.4-d transit time. The cell flux through this intermediate subpopulation was approximately 10(6) cells/d, similar to the rate of turnover of mature thymocytes; thus, although only 3-4% of thymocytes progressed to this intermediate state, once reaching it most then progressed to full maturity. In accordance with this, the incidence of the V beta selection markers within the intermediate subpopulation indicated that both positive and negative

  6. Investigation of positive streamers by double pulse experiments

    NASA Astrophysics Data System (ADS)

    Nijdam, Sander; Takahashi, Eiichi; Markosyan, Aram H.; Ebert, Ute

    2013-09-01

    Streamer discharges are influenced by background ionization and other effects of previous discharges. We have studied the influence of repeating positive streamer discharges by applying two subsequent high voltage pulses with a variable interval (200 ns to 40 ms) between them. The discharges are studied with two ICCD cameras that image the discharge during either the first or the second voltage pulse. Experiments have been performed in a 103 mm point-plane gap at a pressure of 133 mbar in artificial air, pure nitrogen and pure argon. We have found a range of phenomena that depend on the inter-pulse time Δt . For small Δt , (below 1 μs for air and nitrogen and below 15 μs for argon) the streamers just continue their old paths. At larger Δt the conductivity has decreased too much for such continuation. However, parts of the old paths do glow up again like secondary streamers. At still larger Δt (roughly above 2.5 μs for air and 30 μs for nitrogen) new channels appear. At first they avoid the entire area of the previous discharge; next they follow the edges of the old channels; then they start to follow the old channels exactly and finally (Δt > 1 ms) they become fully independent of the old paths. This work was supported by JSPS KAKENHI Grant Number 24560249 as well as under FY2012 Researcher Exchange Program between the Japan Society for the Promotion of Science and The Netherlands Organisation for Scientific Research.

  7. Critical role of SP thymocyte motility in regulation of thymic output in neonatal Aire−/− mice

    PubMed Central

    Jin, Rong; Aili, Abudureyimujiang; Wang, Yuqing; Wu, Jia; Sun, Xiuyuan; Zhang, Yu; Ge, Qing

    2017-01-01

    Autoimmune regulator (Aire) is essential in the perinatal period to prevent the multiorgan autoimmunity. Here we show that Aire-regulated single positive thymocyte trafficking in neonatal period is critical for thymic egress. Reduced thymic emigration was found in Aire−/− mice during neonatal period, leading to enhanced homeostatic expansion of peripheral T cells as early as 2 weeks of age. In neonatal Aire−/− mice, thymic expression of CCR7 ligands were dramatically reduced, resulting in decreased thymocyte motility and thymocyte emigration. This reduction of thymic egress in Aire−/− mice was alleviated beyond 3 weeks of age by an early upregulation of S1P1 signaling. As the numbers and quality of thymic emigrants are essential for the establishment and maintenance of peripheral tolerance, the reduced thymic emigration during neonatal period may deteriorate autoimmunity caused by the emigration of autoreactive T cells. PMID:27965471

  8. Biotin deficiency blocks thymocyte maturation, accelerates thymus involution, and decreases nose-rump length in mice.

    PubMed

    Báez-Saldaña, Armida; Ortega, Enrique

    2004-08-01

    Biotin deficiency in experimental animals causes low body weight as well as several phenomena suggestive of an altered immune system. We reported previously that chronic biotin deficiency in mice decreases body weight and alters the number and proportion of lymphocyte subpopulations in the spleen. To further characterize the effects of biotin deficiency, we studied in detail the maturation of thymocytes and the status of biotin in the thymus, as well as the body length of biotin-deficient mice. Male Balb/cAnN mice were fed for up to 20 wk either standard control diet, a biotin-deficient diet, or a biotin-sufficient diet. At different times, nose-rump length, weight of the thymus, spleen and liver, total number of cells in the spleen and thymus, pyruvate carboxylase (PC) and propionyl CoA carboxylase (PCC) activity in thymus cells, and the proportion of distinct thymocyte subsets were determined. These variables did not differ between mice fed the control and biotin-sufficient diets. In contrast, biotin-deficient mice differed from biotin-sufficient mice in all of the analyzed variables. PC and PCC specific activities of thymocytes of mice fed the biotin-depleting diet decreased during the first 4 wk by 84.5%. The maturation of thymocytes in biotin-deficient mice was arrested at the double-negative stage. Our results suggest that biotin deficiency in mice causes an accelerated involution of the thymus and decreases nose-rump length, but these effects do not correlate in magnitude or in temporality with the sharp decrease in the activity of the biotin-dependent carboxylases. As such, the possibility that the aforementioned effects are not related directly to the prosthetic function of biotin should be considered.

  9. Peanut agglutinin (PNA)-binding properties of murine thymocyte subpopulation.

    PubMed Central

    Dumont, F; Nardelli, J

    1979-01-01

    Surface receptors for peanut agglutinin (PNA), a lectin with D-galactose specificity, were detected on mouse thymocytes using fluorescence microscopy. Depending on mouse strain, 69-85% of unseparated thymocytes could thus be characterized as PNA+. Electrophoretic fractionation of thymocytes from normal or immunosuppressive drug-treated donors revealed an inverse relationship between PNA-binding properties and cell electrophoretic mobility (EPM). Thus, all thymocytes recovered in the lowest EPM fractions were strongly PNA+ whereas those in the highest EPM fractions were in the majority PNA-. Most of the cells collected in the intermediate EPM range were PNA+ but staining with the fluoresceinated lectin appeared weaker than for the low EPM thymocytes. Reciprocal experiments in which thymocytes were separated by PNA-mediated aggregation into fractions with different affinities for the lectin and then subjected to physical analysis, definitely established that PNA+ cells are of lower EPM than PNA- cells and that these two cell types also differ in size distribution. These data show that the four physical subpopulations of thymocytes previously described present distinctive PNA-binding properties: Th1 and Th2 cells can be classified as strongly PNA+, Th3 cells as less intensely PNA+, and Th4 cells as mostly PNA-. Images Figure 1 PMID:313899

  10. Condylar position control during maxillary surgery: the condylar positioning appliance and three-dimensional double splint method.

    PubMed

    Schwestka, R; Engelke, D; Kubein-Meesenburg, D

    1990-01-01

    A new method for positioning the maxilla and condyle after Le Fort I osteotomy maintains the patient's vertical dimension (ie, the relation of the mandible to the skull above the osteotomy plane) in the preoperative and postoperative positions during both cast surgery and actual surgery. During surgery the condylar positioning appliance is fixed to the anterolateral zygoma and the lateral cortex of the mandibular ramus bilaterally to orient the mandible in centric relation. The condylar positioning appliance is used with the three-dimensional double splint method. Two prefabricated splints enable three-dimensional positioning of the maxilla in the fixed mandibular position during surgery. Postoperatively, the mandible can be rotated into the new centric occlusion.

  11. The double universal joint wrist on a manipulator: Solution of inverse position kinematics and singularity analysis

    NASA Technical Reports Server (NTRS)

    Williams, Robert L., III

    1992-01-01

    This paper presents three methods to solve the inverse position kinematics position problem of the double universal joint attached to a manipulator: (1) an analytical solution for two specific cases; (2) an approximate closed form solution based on ignoring the wrist offset; and (3) an iterative method which repeats closed form position and orientation calculations until the solution is achieved. Several manipulators are used to demonstrate the solution methods: cartesian, cylindrical, spherical, and an anthropomorphic articulated arm, based on the Flight Telerobotic Servicer (FTS) arm. A singularity analysis is presented for the double universal joint wrist attached to the above manipulator arms. While the double universal joint wrist standing alone is singularity-free in orientation, the singularity analysis indicates the presence of coupled position/orientation singularities of the spherical and articulated manipulators with the wrist. The cartesian and cylindrical manipulators with the double universal joint wrist were found to be singularity-free. The methods of this paper can be implemented in a real-time controller for manipulators with the double universal joint wrist. Such mechanically dextrous systems could be used in telerobotic and industrial applications, but further work is required to avoid the singularities.

  12. Proliferation of thymocytes in relation to T-cell receptor beta-chain expression.

    PubMed

    Parkin, I G; Owen, J J; Jenkinson, E J

    1988-05-01

    During proliferation and differentiation of maturing thymocytes, T-cell receptor beta-chain products are first expressed in the cytoplasm. Only subsequently are they expressed on the cell surface, presumably as part of the alpha beta/CD3 receptor complex. This study uses double immunofluorescence labelling to identify these cytoplasmic and surface phases separately in relationship to cell-cycle parameters. The use of a mitotic arrest agent and tritiated thymidine autoradiography both show that cells with cytoplasmic beta-chains are in cell cycle, whereas cells with surface beta-chains are cycling slowly, if at all.

  13. A Cellular Automata-Based Mathematical Model for Thymocyte Development

    PubMed Central

    Souza-e-Silva, Hallan; Savino, Wilson; Feijóo, Raúl A.; Vasconcelos, Ana Tereza Ribeiro

    2009-01-01

    Intrathymic T cell development is an important process necessary for the normal formation of cell-mediated immune responses. Importantly, such a process depends on interactions of developing thymocytes with cellular and extracellular elements of the thymic microenvironment. Additionally, it includes a series of oriented and tunely regulated migration events, ultimately allowing mature cells to cross endothelial barriers and leave the organ. Herein we built a cellular automata-based mathematical model for thymocyte migration and development. The rules comprised in this model take into account the main stages of thymocyte development, two-dimensional sections of the normal thymic microenvironmental network, as well as the chemokines involved in intrathymic cell migration. Parameters of our computer simulations with further adjusted to results derived from previous experimental data using sub-lethally irradiated mice, in which thymus recovery can be evaluated. The model fitted with the increasing numbers of each CD4/CD8-defined thymocyte subset. It was further validated since it fitted with the times of permanence experimentally ascertained in each CD4/CD8-defined differentiation stage. Importantly, correlations using the whole mean volume of young normal adult mice revealed that the numbers of cells generated in silico with the mathematical model fall within the range of total thymocyte numbers seen in these animals. Furthermore, simulations made with a human thymic epithelial network using the same mathematical model generated similar profiles for temporal evolution of thymocyte developmental stages. Lastly, we provided in silico evidence that the thymus architecture is important in the thymocyte development, since changes in the epithelial network result in different theoretical profiles for T cell development/migration. This model likely can be used to predict thymocyte evolution following therapeutic strategies designed for recovery of the thymus in diseases

  14. Peptides Regulate Cortical Thymocytes Differentiation, Proliferation, and Apoptosis

    PubMed Central

    Khavinson, V. Kh.; Polyakova, V. O.; Linkova, N. S.; Dudkov, A. V.; Kvetnoy, I. M.

    2011-01-01

    The processes of differentiation, proliferation, and apoptosis were studied in a cell culture of human cortical thymocytes under the influence of short peptides T-32 (Glu-Asp-Ala) and T-38 (Lys-Glu-Asp). Peptides T-32 and T-38 amplified cortical thymocytes differentiation towards regulatory T cells, increased their proliferative activity, and decreased the level of apoptosis. Moreover, peptides under study stimulated proliferative and antiapoptotic activity of the mature regulatory T cells. PMID:22312461

  15. Laminin-Mediated Interactions in Thymocyte Migration and Development

    PubMed Central

    Savino, Wilson; Mendes-da-Cruz, Daniella Arêas; Golbert, Daiane Cristina Ferreira; Riederer, Ingo; Cotta-de-Almeida, Vinicius

    2015-01-01

    Intrathymic T-cell differentiation is a key process for the development and maintenance of cell-mediated immunity, and occurs concomitantly to highly regulated migratory events. We have proposed a multivectorial model for describing intrathymic thymocyte migration. One of the individual vectors comprises interactions mediated by laminins (LMs), a heterotrimeric protein family of the extracellular matrix. Several LMs are expressed in the thymus, being produced by microenvironmental cells, particularly thymic epithelial cells (TECs). Also, thymocytes and epithelial cells express integrin-type LM receptors. Functionally, it has been reported that the dy/dy mutant mouse (lacking the LM isoform 211) exhibits defective thymocyte differentiation. Several data show haptotactic effects of LMs upon thymocytes, as well as their adhesion on TECs; both effects being prevented by anti-LM or anti-LM receptor antibodies. Interestingly, LM synergizes with chemokines to enhance thymocyte migration, whereas classe-3 semaphorins and B ephrins, which exhibit chemorepulsive effects in the thymus, downregulate LM-mediated migratory responses of thymocytes. More recently, we showed that knocking down the ITGA6 gene (which encodes the α6 integrin chain of LM receptors) in human TECs modulates a large number of cell migration-related genes and results in changes of adhesion pattern of thymocytes onto the thymic epithelium. Overall, LM-mediated interactions can be placed at the cross-road of the multivectorial process of thymocyte migration, with a direct influence per se, as well as by modulating other molecular interactions associated with the intrathymic-trafficking events. PMID:26635793

  16. Increased TCR signal strength in DN thymocytes promotes development of gut TCRαβ((+))CD8αα((+)) intraepithelial lymphocytes.

    PubMed

    Grandjean, Capucine L; Sumaria, Nital; Martin, Stefania; Pennington, Daniel J

    2017-09-06

    CD4((+))CD8((+)) "double positive" (DP) thymocytes differentiate into diverse αβ T cell sub-types using mechanistically distinct programs. For example, conventional αβ T cells develop from DP cells after partial-agonist T cell receptor (TCR) interactions with self-peptide/MHC, whereas unconventional αβ T cells, such as TCRαβ((+))CD8αα((+)) intraepithelial lymphocytes (IELs), require full-agonist TCR interactions. Despite this, DP cells appear homogeneous, and it remains unclear how distinct TCR signalling instructs distinct developmental outcomes. Moreover, whether TCR signals at earlier stages of development, for example in CD4((-))CD8((-)) double negative (DN) cells, impact on later fate decisions is presently unknown. Here, we assess four strains of mice that display altered TCR signal strength in DN cells, which correlates with altered generation of unconventional TCRαβ((+))CD8αα((+)) IELs. FVB/n mice (compared to C57BL/6 animals) and mice with altered preTCRα (pTα) expression, both displayed weaker TCR signalling in DN cells, an inefficient DN-to-DP transition, and reduced contribution of TCRαβ((+))CD8αα((+)) IELs to gut epithelium. Conversely, TCRαβ((+))CD8αα((+)) IEL development was favoured in mice with increased TCR signal strength in DN cells. Collectively, these data suggest TCR signal strength in DN cells directly impacts on subsequent DP cell differentiation, fundamentally altering the potential of thymocyte progenitors to adopt conventional versus unconventional T cell fates.

  17. GRB2-Mediated Recruitment of THEMIS to LAT Is Essential for Thymocyte Development

    PubMed Central

    Paster, Wolfgang; Brockmeyer, Claudia; Fu, Guo; Simister, Philip C.; de Wet, Ben; Martinez-Riaño, Ana; Hoerter, John A. H.; Feller, Stephan M.; Wülfing, Christoph; Gascoigne, Nicholas R. J.

    2013-01-01

    Thymocyte-expressed molecule involved in selection (THEMIS) is a recently identified regulator of thymocyte positive selection. THEMIS’s mechanism of action is unknown, and whether it has a role in TCR-proximal signaling is controversial. In this article, we show that THEMIS and the adapter molecule growth factor receptor–bound protein 2 (GRB2) associate constitutively through binding of a conserved PxRPxK motif within the proline-rich region 1 of THEMIS to the C-terminal SH3-domain of GRB2. This association is indispensable for THEMIS recruitment to the immunological synapse via the transmembrane adapter linker for activation of T cells (LAT) and for THEMIS phosphorylation by Lck and ZAP-70. Two major sites of tyrosine phosphorylation were mapped to a YY-motif close to proline-rich region 1. The YY-motif was crucial for GRB2 binding, suggesting that this region of THEMIS might control local phosphorylation-dependent conformational changes important for THEMIS function. Finally, THEMIS binding to GRB2 was required for thymocyte development. Our data firmly assign THEMIS to the TCR-proximal signaling cascade as a participant in the LAT signalosome and suggest that the THEMIS–GRB2 complex might be involved in shaping the nature of Ras signaling, thereby governing thymic selection. PMID:23460737

  18. The conveyor belt hypothesis for thymocyte migration: participation of adhesion and de-adhesion molecules.

    PubMed

    Villa-Verde, D M; Calado, T C; Ocampo, J S; Silva-Monteiro, E; Savino, W

    1999-05-01

    Thymocyte differentiation is the process by which bone marrow-derived precursors enter the thymus, proliferate, rearrange the genes and express the corresponding T cell receptors, and undergo positive and/or negative selection, ultimately yielding mature T cells that will represent the so-called T cell repertoire. This process occurs in the context of cell migration, whose cellular and molecular basis is still poorly understood. Kinetic studies favor the idea that these cells leave the organ in an ordered pattern, as if they were moving on a conveyor belt. We have recently proposed that extracellular matrix glycoproteins, such as fibronectin, laminin and type IV collagen, among others, produced by non-lymphoid cells both in the cortex and in the medulla, would constitute a macromolecular arrangement allowing differentiating thymocytes to migrate. Here we discuss the participation of both molecules with adhesive and de-adhesive properties in the intrathymic T cell migration. Functional experiments demonstrated that galectin-3, a soluble beta-galactoside-binding lectin secreted by thymic microenvironmental cells, is a likely candidate for de-adhesion proteins by decreasing thymocyte interaction with the thymic microenvironment.

  19. Prenatal cadmium exposure dysregulates sonic hedgehog and Wnt/beta-catenin signaling in the thymus resulting in altered thymocyte development

    SciTech Connect

    Hanson, Miranda L.; Brundage, Kathleen M.; Schafer, Rosana; Tou, Janet C.; Barnett, John B.

    2010-01-15

    Cadmium (Cd) is both an environmental pollutant and a component of cigarette smoke. Although evidence demonstrates that adult exposure to Cd causes changes in the immune system, there are limited reports in the literature of immunomodulatory effects of prenatal exposure to Cd. The sonic hedgehog (Shh) and Wnt/beta-catenin pathways are required for thymocyte maturation. Several studies have demonstrated that Cd exposure affects these pathways in different organ systems. This study was designed to investigate the effect of prenatal Cd exposure on thymocyte development, and to determine if these effects were linked to dysregulation of Shh and Wnt/beta-catenin pathways. Pregnant C57Bl/6 mice were exposed to an environmentally relevant dose (10 ppm) of Cd throughout pregnancy and effects on the thymus were assessed on the day of birth. Thymocyte phenotype was determined by flow cytometry. A Gli:luciferase reporter cell line was used to measure Shh signaling. Transcription of target genes and translation of key components of both signaling pathways were assessed using real-time RT-PCR and western blot, respectively. Prenatal Cd exposure increased the number of CD4{sup +} cells and a subpopulation of double-negative cells (DN; CD4{sup -}CD8{sup -}), DN4 (CD44{sup -}CD25{sup -}). Shh and Wnt/beta-catenin signaling were both decreased in the thymus. Target genes of Shh (Patched1 and Gli1) and Wnt/beta-catenin (c-fos, and c-myc) were affected differentially among thymocyte subpopulations. These findings suggest that prenatal exposure to Cd dysregulates two signaling pathways in the thymus, resulting in altered thymocyte development.

  20. THP-1 macrophage lipid accumulation unaffected by fatty acid double bond geometric or positional configuration

    USDA-ARS?s Scientific Manuscript database

    Dietary fatty acid type alters atherosclerotic lesion progression and macrophage lipid accumulation. Incompletely elucidated are the mechanisms by which fatty acids differing in double-bond geometric or positional configuration alter arterial lipid accumulation. The objective of this study was to ev...

  1. Altered Expression of Galectin-3 Induces Cortical Thymocyte Depletion and Premature Exit of Immature Thymocytes during Trypanosoma cruzi Infection

    PubMed Central

    Silva-Monteiro, Elizangela; Reis Lorenzato, Luciana; Kenji Nihei, Oscar; Junqueira, Mara; Rabinovich, Gabriel Adrián; Hsu, Daniel Kaiyuan; Liu, Fu-Tong; Savino, Wilson; Chammas, Roger; Villa-Verde, Déa Maria Serra

    2007-01-01

    During acute infection with Trypanosoma cruzi, the causative agent of Chagas’ disease, the thymus undergoes intense atrophy followed by a premature escape of CD4+CD8+ immature cortical thymocytes. Here we report a pivotal role for the endogenous lectin galectin-3 in accelerating death of thymocytes and migration of these cells away from the thymus after T. cruzi infection. We observed a pronounced increase in galectin-3 expression that paralleled the extensive depletion of CD4+CD8+ immature thymocytes after infection. In vitro, recombinant galectin-3 induced increased levels of death in cortical immature thymocytes. Consistent with the role of galectin-3 in promoting cell death, thymuses from gal-3−/− mice did not show cortical thymocyte depletion after parasite infection in vivo. In addition, galectin-3 accelerated laminin-driven CD4+CD8+ thymocyte migration in vitro and in vivo induced exportation of CD4+CD8+ cells from the thymus to the peripheral compartment. Our findings provide evidence of a novel role for galectin-3 in the regulation of thymus physiology and identify a potential mechanism based on protein-glycan interactions in thymic atrophy associated with acute T. cruzi infection. PMID:17255323

  2. Altered expression of galectin-3 induces cortical thymocyte depletion and premature exit of immature thymocytes during Trypanosoma cruzi infection.

    PubMed

    Silva-Monteiro, Elizangela; Reis Lorenzato, Luciana; Kenji Nihei, Oscar; Junqueira, Mara; Rabinovich, Gabriel Adrián; Hsu, Daniel Kaiyuan; Liu, Fu-Tong; Savino, Wilson; Chammas, Roger; Villa-Verde, Déa Maria Serra

    2007-02-01

    During acute infection with Trypanosoma cruzi, the causative agent of Chagas' disease, the thymus undergoes intense atrophy followed by a premature escape of CD4+CD8+ immature cortical thymocytes. Here we report a pivotal role for the endogenous lectin galectin-3 in accelerating death of thymocytes and migration of these cells away from the thymus after T. cruzi infection. We observed a pronounced increase in galectin-3 expression that paralleled the extensive depletion of CD4+CD8+ immature thymocytes after infection. In vitro, recombinant galectin-3 induced increased levels of death in cortical immature thymocytes. Consistent with the role of galectin-3 in promoting cell death, thymuses from gal-3-/- mice did not show cortical thymocyte depletion after parasite infection in vivo. In addition, galectin-3 accelerated laminin-driven CD4+CD8+ thymocyte migration in vitro and in vivo induced exportation of CD4+CD8+ cells from the thymus to the peripheral compartment. Our findings provide evidence of a novel role for galectin-3 in the regulation of thymus physiology and identify a potential mechanism based on protein-glycan interactions in thymic atrophy associated with acute T. cruzi infection.

  3. Large amplitude double layers in a positively charged dusty plasma with nonthermal electrons

    SciTech Connect

    Djebli, M.; Marif, H.

    2009-06-15

    A pseudopotential approach is used to investigate large amplitude dust-acoustic solitary structures for a plasma composed of positively charged dust, cold electrons, and nonthermal hot electrons. Numerical investigation for an adiabatic situation is conducted to examine the existence region of the wave. The negative potential of the double layers is found to be dependent on nonthermal parameters, Mach number, and electrons temperature. A range of the nonthermal parameters values exists for which two possible double layers for the same plasma mix at different Mach numbers and with significant different amplitudes. The present model is used to investigate localized structures in the lower-altitude Earth's ionosphere.

  4. [Effects of dithiocarbamates on the mitotic activity of thymocytes].

    PubMed

    Hennighausen, G; Kröning, G

    1988-01-01

    The effects of dithiocarbamates on the mitotic activity of thymocytes were studied in vitro and in vivo. The mitotic activity was determined by counting of the metaphases blocked by colchicine. Using thymocytes of mice the mitotic activity in short-time cultures was inhibited (50%) by the following concentrations of the substances under study (EC50-values in mol/l): tetraethylthiuramdisulfide; (1; disulfiram) 4.2.10(-9); diethyldithiocarbamate (2) 3.2.10(-8), Cu-2, 2.6.10(-8), Zn-dimethyldithiocarbamate (3; Ziram) 1.9.10(-8), Zn-ethylene-bis-dithiocarbamate (4; Zineb) 2.3.10(-6). Using rat and guinea-pig thymocytes the ED50- values for 1 and 2 were found in the same concentration range as compared with thymocytes of mice. There was no impact on the vitality of cells by the mitosis inhibiting concentrations. In vivo the mitotic activity of thymocytes in mice was decreased only after application of high doses (500 mg/kg i.p.) of 2. After depletion of reduced glutathione (GSH) by the pretreatment of mice with diethylmaleate the effects of 2 were increased in vivo. In vitro GSH (10(-4) mol/l) decreased the mitosis inhibiting activity of 2 in mice thymocytes and the inhibition of the SH-enzyme ecto-ATPase in rat thymocytes. It is supposed that the inhibition of mitosis is due to the reaction of dithiocarbamates with SH-groups of microtubules and other functionally important proteines. The formation of tiyl radicals from dithiocarbamates may play a certain role in redox processes which can be influenced by GSH.

  5. Role of Extracellular Matrix-Mediated Interactions in Thymocyte Migration

    PubMed Central

    Dalmau, Sérgio Ranto; Dealmeida, Vinícius Cotta

    2000-01-01

    Cell adhesion, migration, differentiation and survival or death is amongst a large spectrum of biological responses that can be elicited by ligation of extracellular matrix components to their corresponding receptors. As regards the physiology of the thymus, cell migration is a crucial event in the general process of T cell differentiation. Studies on the intrathymic distribution of ECM components revealed that fibronectin, laminin and type IV collagen, are not restrictedly located at typical basement membrane sites, also forming a thick network in the medullary region of the thymic lobules, whereas very thin ECM fibers are found within the cortex. These ECM components are essentially produced by thymic microenvironmental cells, which also drive thymocyte differentiation. Signals triggered by ECM are conveyed into thymocytes or microenvironmental cells through specific membrane receptors, and most of them belong to the integrin type, such as the VLA-3, VLA-4, VLA-5 and VLA-6. In vitro studies revealed that adhesion of thymocytes to thymic microenvironmental cells is mediated by extracellular matrix. Such an adhesion is preferentially done by immature thymocytes. Importantly, ECM-mediated interactions also govern the entrance and exit of thymocytes in the lymphoepithelial complexes named thymic nurse cells. Lastly, pathological conditions, including infectious and autoimmune diseases, in which changes of ECM ligands and receptors are observed, course with alterations in thymocyte migration and death. In conclusion, the fact that ECM can modulate traffic, differentiation, death and survival of normal thymocytes adds clues for understanding how ECM-mediated interactions behave in the thymus, not only in normal, but also in pathological conditions. PMID:11097218

  6. Human αβ and γδ Thymocyte Development: TCR Gene Rearrangements, Intracellular TCRβ Expression and γδ Developmental Potential – Differences between Men and Mice1,2

    PubMed Central

    Joachims, Michelle L.; Chain, Jennifer L.; Hooker, Scott W.; Knott-Craig, Christopher J.; Thompson, Linda F.

    2006-01-01

    To evaluate the role of the TCR in the αβ/γδ lineage choice during human thymocyte development, molecular analyses of the TCRβ locus in γδ cells and the TCRγ and δ loci in αβ cells were undertaken. TCRβ variable gene segments remained largely in germline configuration in γδ cells, indicating that commitment to the γδ lineage occurred prior to complete TCRβ rearrangements in most cases. The few TCRβ rearrangements detected were primarily out-of-frame, suggesting that productive TCRβ rearrangements diverted cells away from the γδ lineage. In contrast, in αβ cells, the TCRγ locus was almost completely rearranged with a random productivity profile; the TCRδ locus contained primarily non-productive rearrangements. Productive γ rearrangements were, however, depleted compared to pre-selected cells. Productive TCRγ and δ rearrangements rarely occurred in the same cell, suggesting that αβ cells developed from cells unable to produce a functional γδ TCR. Intracellular TCRβ expression correlated with the up regulation of CD4 and concomitant down regulation of CD34, and plateaued at the early double positive stage. Surprisingly, however, some early double positive thymocytes retained γδ potential in culture. We present a model for human thymopoiesis which includes γδ development as a default pathway, an instructional role for the TCR in the αβ/γδ lineage choice, and a prolonged developmental window for β-selection and γδ lineage commitment. Aspects that differ from the mouse are the status of TCR gene rearrangements at the non-expressed loci, the timing of β-selection, and maintenance of γδ potential through the early double positive stage of development. PMID:16424183

  7. A silencer-proximal intronic region is required for sustained CD4 expression in post-selection thymocytes

    PubMed Central

    Henson, David M.; Chou, Chun; Sakurai, Nagisa; Egawa, Takeshi

    2014-01-01

    It has been proposed that differential kinetics of CD4/CD8 co-receptors regulate fate choice of selected thymocytes. Sustained signals by interaction between MHC class II and TCR/CD4 is required for commitment to the CD4 helper lineage. While prematurely terminated MHC-TCR/CD4 interaction in transgenic mouse models results in lineage redirection, it is unclear if CD4 expression is actively maintained by endogenous cis-elements to facilitate prolonged signaling under physiological conditions. Here we show that sustained CD4 expression in post-selection thymocytes requires an intronic sequence containing an uncharacterized DNase I hypersensitivity site (DHS) located 3’ to the silencer. Despite normal CD4 expression before selection, thymocytes lacking a 1.5 kb sequence in intron 1 including the 0.4 kb silencer and the DHS, but not the 0.4 kb silencer alone, failed to maintain CD4 expression upon positive selection and are redirected to the CD8 lineage following MHC class II-restricted selection. Furthermore, CpG dinucleotides adjacent to the DHS are hypermethylated in CD8+ T cells. These results indicate that the 1.5 kb cis-element is required in post-selection thymocytes for helper lineage commitment, presumably mediating the maintenance of CD4 expression, and suggest that inactivation of the cis-element by DNA methylation may contribute to epigenetic Cd4 silencing. PMID:24729613

  8. Aberrant Development of Thymocytes in Mice Lacking Laminin-2

    PubMed Central

    Magner, William J.; Chang, Andrew C.; Owens, Jennie; Hong, M-J. P.; Brooks, Andrew

    2000-01-01

    In previous in vitro studies, we proposed a role for the extracellular matrix component, laminin- 2, and its integrin receptor, VLA-6, in thymocyte development. The characterization of two dystrophic mouse strains with different defects in laminin-2 allowed us to examine this proposal in vivo. Mice deficient in laminin-2, dy/dy, show a significant reduction in thymus size and number of thymocytes compared to normal littermates. These mice also exhibited apparent alterations of thymic architecture. Examination of the CD4/CD8 populations in dy/dy thymi showed large relative increases in the DN (CD4-CD8-) and SP (CD4+CD8-, CD4-CD8+) populations and a significant decrease in the DP (CD4+CD8+) population. Further examination of the DN population for CD44 and CD25 expression showed a remarkable decrease in the more mature pre-T cell populations. Analysis of apoptosis in situ, and by flow cytometry, in dy/dy thymi revealed a significant increase in apoptotic DN thymocytes in the capsule and subcapsular regions. Interestingly, thymocyte development appeared to proceed normally in dystrophic mice expressing a mutant form of laminin-2, dy2J, as well as, in fetal and neonatal dy/dy mice. We propose that laminin-2 plays an active role in thymocyte development by delivering cell survival and differentiation signals at specific stages of development in young adult mice. PMID:11097211

  9. Large amplitude dust-acoustic double layers in non-thermal plasmas with positive and negative dust

    NASA Astrophysics Data System (ADS)

    Maharaj, S. K.; Bharuthram, R.; Singh, S. V.; Pillay, S. R.; Lakhina, G. S.

    2011-11-01

    The existence of large amplitude double layers in a plasma composed of cold negative dust, adiabatic positive dust, non-thermal ions and Boltzmann electrons is investigated using the Sagdeev pseudopotential technique. Both positive potential and negative potential double layers are found to be supported by the model. The variation of the maximum amplitudes of the double layers and corresponding Mach numbers are examined as a function of various plasma parameters. In particular, we investigate to what extent ion non-thermal effects are required for positive potential double layers to occur.

  10. Large amplitude dust-acoustic double layers in non-thermal plasmas with positive and negative dust

    SciTech Connect

    Maharaj, S. K.; Bharuthram, R.; Singh, S. V.; Lakhina, G. S.; Pillay, S. R.

    2011-11-29

    The existence of large amplitude double layers in a plasma composed of cold negative dust, adiabatic positive dust, non-thermal ions and Boltzmann electrons is investigated using the Sagdeev pseudopotential technique. Both positive potential and negative potential double layers are found to be supported by the model. The variation of the maximum amplitudes of the double layers and corresponding Mach numbers are examined as a function of various plasma parameters. In particular, we investigate to what extent ion non-thermal effects are required for positive potential double layers to occur.

  11. Molecular dynamics study of lipid bilayers modeling the plasma membranes of normal murine thymocytes and leukemic GRSL cells.

    PubMed

    Andoh, Yoshimichi; Okazaki, Susumu; Ueoka, Ryuichi

    2013-04-01

    Molecular dynamics (MD) calculations for the plasma membranes of normal murine thymocytes and thymus-derived leukemic GRSL cells in water have been performed under physiological isothermal-isobaric conditions (310.15K and 1 atm) to investigate changes in membrane properties induced by canceration. The model membranes used in our calculations for normal and leukemic thymocytes comprised 23 and 25 kinds of lipids, respectively, including phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidylinositol, sphingomyelin, lysophospholipids, and cholesterol. The mole fractions of the lipids adopted here were based on previously published experimental values. Our calculations clearly showed that the membrane area was increased in leukemic cells, and that the isothermal area compressibility of the leukemic plasma membranes was double that of normal cells. The calculated membranes of leukemic cells were thus considerably bulkier and softer in the lateral direction compared with those of normal cells. The tilt angle of the cholesterol and the conformation of the phospholipid fatty acid tails both showed a lower level of order in leukemic cell membranes compared with normal cell membranes. The lateral radial distribution function of the lipids also showed a more disordered structure in leukemic cell membranes than in normal cell membranes. These observations all show that, for the present thymocytes, the lateral structure of the membrane is considerably disordered by canceration. Furthermore, the calculated lateral self-diffusion coefficient of the lipid molecules in leukemic cell membranes was almost double that in normal cell membranes. The calculated rotational and wobbling autocorrelation functions also indicated that the molecular motion of the lipids was enhanced in leukemic cell membranes. Thus, here we have demonstrated that the membranes of thymocyte leukemic cells are more disordered and more fluid than normal cell membranes. Copyright © 2013

  12. Using ambient ozone for assignment of double bond position in unsaturated lipids.

    PubMed

    Ellis, Shane R; Hughes, Jessica R; Mitchell, Todd W; in het Panhuis, Marc; Blanksby, Stephen J

    2012-03-07

    Unsaturated lipids deposited onto a range of materials are observed to react with the low concentrations of ozone present in normal laboratory air. Parent lipids and ozonolysis cleavage products are both detected directly from surfaces by desorption electrospray ionisation mass spectrometry (DESI-MS) with the resulting mass spectra providing clear evidence of the double bond position within these molecules. This serendipitous process has been coupled with thin-layer chromatography (TLC) to provide a simple but powerful approach for the detailed structural elucidation of lipids present in complex biological extracts. Lipid extracts from human lens were deposited onto normal phase TLC plates and then developed to separate components according to lipid class. Exposure of the developed plates to laboratory air for ca. 1 h prior to DESI-MS analysis gave rise to ozonolysis products allowing for the unambiguous identification of double bond positions in even low abundant, unsaturated lipids. In particular, the co-localization of intact unsaturated lactosylceramides (LacCer) with products from their oxidative cleavage provide the first evidence for the presence of three isomeric LacCer (d18:0/24:1) species in the ocular lens lipidome, i.e., variants with double bonds at the n-9, n-7 and n-5 positions.

  13. Test of local position invariance using a double-cavity laser system

    SciTech Connect

    Agachev, A. R.; Belov, I. Yu.; Bochkarev, V. V.; Daishev, R. A.; Mavrin, S. V.; Murzakhanov, Z. G.; Skochilov, A. F. Chugunov, Yu. P.; Shindyaev, O. P.

    2010-01-15

    The results of testing local position invariance, which is a constituent of the Einstein equivalence principle, in a 'null' gravitational redshift experiment are reported. The processing of the experimental data collected during the five-month operation of a double-c avity laser system, where one cavity operates in the free generation mode and the frequency of the second cavity is stabilized with the nonlinear ultranarrow absorption resonance of the methane molecule, has confirmed the universality of the gravitational redshift law at a level of 0.9%. This result almost doubly improves the best existing accuracy (1.7%) of testing local position invariance for clocks of different physical natures.

  14. Calcineurin sets the bandwidth for discrimination of signals during thymocyte development

    PubMed Central

    Gallo, Elena M.; Winslow, Monte M.; Canté-Barrett, Kirsten; Radermacher, Amy N.; Ho, Lena; McGinnis, Lisa; Iritani, Brian; Neilson, Joel R.; Crabtree, Gerald R.

    2017-01-01

    At critical times in development, cells are able to convert graded signals into discrete developmental outcomes; however, the mechanisms involved are poorly understood. During thymocyte development, cell fate is determined by signals originating from the α β T-cell receptor. Low-affinity/avidity interactions between the T-cell receptor and peptide–MHC complexes direct differentiation to the single-positive stage (positive selection), whereas high-affinity/avidity interactions induce death by apoptosis (negative selection)1,2. Here we show that mice deficient in both calcineurin and nuclear factor of activated T cells (NFAT)c2/c3 lack a population of preselection thymocytes with enhanced ability to activate the mitogen-activated protein kinase (Raf–MEK–ERK) pathway, and fail to undergo positive selection. This defect can be partially rescued with constitutively active Raf, indicating that calcineurin controls MAPK signalling. Analysis of mice deficient in both Bim (which is required for negative selection) and calcineurin revealed that calcineurin-induced ERK (extracellular signal-regulated kinase) sensitization is required for differentiation in response to ‘weak’ positive selecting signals but not in response to ‘strong’ negative selecting signals (which normally induce apoptosis). These results indicate that early calcineurin/NFAT signalling produces a developmental period of ERK hypersensitivity, allowing very weak signals to induce positive selection. This mechanism might be generally useful in the discrimination of graded signals that induce different cell fates. PMID:18046413

  15. GENETIC CORRELATION OF A MOUSE LIGHT CHAIN VARIABLE REGION MARKER WITH A THYMOCYTE SURFACE ANTIGEN

    PubMed Central

    Gottlieb, Paul D.

    1974-01-01

    The inbred and congenic strain distribution of the IH-peptide marker in the variable region of mouse immunoglobulin light chains has been compared with other known genetic markers. A positive correlation was noted between the IH-peptide marker and expression of the Ly-3.1 thymocyte cell surface antigen. This suggests that the locus responsible for IH-peptide expression is genetically linked to the Ly-2 and Ly-3 loci in linkage group XI on chromsome 6 of the mouse. PMID:4547628

  16. Double hysteresis loops and large negative and positive electrocaloric effects in tetragonal ferroelectrics.

    PubMed

    Wu, Hong-Hui; Zhu, Jiaming; Zhang, Tong-Yi

    2015-10-07

    Phase field modelling and thermodynamic analysis are employed to investigate depolarization and compression induced large negative and positive electrocaloric effects (ECEs) in ferroelectric tetragonal crystalline nanoparticles. The results show that double-hysteresis loops of polarization versus electric field dominate at temperatures below the Curie temperature of the ferroelectric material, when the mechanical compression exceeds a critical value. In addition to the mechanism of pseudo-first-order phase transition (PFOPT), the double-hysteresis loops are also caused by the abrupt rise of macroscopic polarization from the abc phase to the c phase or the sudden fall of macroscopic polarization from the c phase to the abc phase when the temperature increases. This phenomenon is called the electric-field-induced-pseudo-phase transition (EFIPPT) in the present study. Similar to the two types of PFOPTs, the two types of EFIPPTs cause large negative and positive ECEs, respectively, and give the maximum absolute values of negative and positive adiabatic temperature change (ATC ΔT). The temperature associated with the maximum absolute value of negative ATC ΔT is lower than that associated with the maximum positive ATC ΔT. Both maximum absolute values of ATC ΔTs change with the variation in the magnitude of an applied electric field and depend greatly on the compression intensity.

  17. Absence of equifinality of hand position in a double-step unloading task.

    PubMed

    Norouzi-Gheidari, Nahid; Archambault, Philippe

    2010-08-01

    Equifinality, during arm reaching movements, relates to the capacity of the neuromuscular system to attain the same final position in the presence or absence of transient perturbations. There have been several controversies regarding equifinality in the literature. A brief elastic perturbation, applied during a fast arm movement or just before its initiation, typically does not affect final arm position. On the other hand, several experiments have shown that velocity-dependent perturbations, such as Coriolis force or negative damping, while transient in nature, have a significant effect on final arm position when compared to unperturbed movements. In this study, an unloading paradigm was used to study the role of reflexes with respect to equifinality. The effects on final arm position of suddenly decreasing a static load maintained by fourteen subjects were analyzed. Subjects maintained an initial load produced by a double-joint manipulandum moving in the horizontal plane. The load was suddenly decreased, either in one or in two successive steps with different time intervals, resulting in a rapid reflex-mediated change in arm position. Unloading led to short-latency changes in the activity of shoulder and elbow muscles and significant variations in tonic activity. It was found that the final hand position was shorter for double- versus single-step unloading if the time between two successive changes in load was greater than 100 ms. With a shorter time interval, the final hand positions were the same. This difference in final hand positions was inversely proportional to the hand velocity at the time of the second change in load. Further, agonist/antagonist co-activation increased in double-step unloading. Thus, the change in both the load and the movement velocity may influence the magnitude of the unloading reflex. This may be indicative of a dependence of stretch reflexes on velocity. Perturbation may cause a reflex-mediated increase in joint stiffness, which

  18. Increased CD4+/CD8+ Double-Positive T Cells in Chronic Chagasic Patients

    PubMed Central

    Giraldo, Nicolas A.; Bolaños, Natalia I.; Cuellar, Adriana; Guzman, Fanny; Uribe, Ana Maria; Bedoya, Astrid; Olaya, Natalia; Cucunubá, Zulma M.; Roa, Nubia; Rosas, Fernando; Velasco, Víctor; Puerta, Concepción J.; González, John M.

    2011-01-01

    Background CD4+/CD8+ double positive (DP) T cells have been described in healthy individuals as well as in patients with autoimmune and chronic infectious diseases. In chronic viral infections, this cell subset has effector memory phenotype and displays antigen specificity. No previous studies of double positive T cells in parasite infections have been carried out. Methodology/Principal Findings Seventeen chronic chagasic patients (7 asymptomatic and 10 symptomatic) and 24 non-infected donors, including 12 healthy and 12 with non-chagasic cardiomyopathy donors were analyzed. Peripheral blood was stained for CD3, CD4, CD8, HLA-DR and CD38, and lymphocytes for intracellular perforin. Antigen specificity was assessed using HLA*A2 tetramers loaded with T. cruzi K1 or influenza virus epitopes. Surface expression of CD107 and intracellular IFN-γ production were determined in K1-specific DP T cells from 11 chagasic donors. Heart tissue from a chronic chagasic patient was stained for both CD8 and CD4 by immunochemistry. Chagasic patients showed higher frequencies of DP T cells (2.1%±0.9) compared with healthy (1.1%±0.5) and non-chagasic cardiomyopathy (1.2%±0.4) donors. DP T cells from Chagasic patients also expressed more HLA-DR, CD38 and perforin and had higher frequencies of T. cruzi K1-specific cells. IFN-γ production in K1-specific cells was higher in asymptomatic patients after polyclonal stimulation, while these cells tended to degranulate more in symptomatic donors. Immunochemistry revealed that double positive T cells infiltrate the cardiac tissue of a chagasic donor. Conclusions Chagasic patients have higher percentages of circulating double positive T cells expressing activation markers, potential effector molecules and greater class I antigenic specificity against T. cruzi. Although K1 tetramer positive DP T cell produced little IFN-γ, they displayed degranulation activity that was increased in symptomatic patients. Moreover, K1-specific DP T cells can

  19. Position resolution of a double junction superconductive detector based on a single material

    NASA Astrophysics Data System (ADS)

    Samedov, V. V.

    2008-02-01

    The Naples group from Istituto Nazionale di Fisica Nucleare presented the results of theoretical investigations of a new class of superconductive radiation detectors - double junction superconductive detector based on a single material [1]. In such detectors, the absorption of energy occurs in a long superconductive strip while two superconductive tunnel junctions positioned at the ends of the strip provide the readout of the signals. The main peculiarity of this type of detectors is that they are based on a single superconducting material, i.e., without trapping layers at the ends of the strip. In this paper, general approach to the position resolution of this type of detectors has been attempted. The formula for the position resolution is derived. It is shown that the application of the aluminium for the absorber may be the best possible way not only due to the small gap energy, but also mainly for STJ fabrication technology based on the aluminium oxide tunnel barrier.

  20. CHANGES IN POLARIZATION POSITION ANGLE ACROSS THE ECLIPSE IN THE DOUBLE PULSAR SYSTEM

    SciTech Connect

    Yuen, R.; Manchester, R. N.; Burgay, M.; Camilo, F.; Kramer, M.; Melrose, D. B.; Stairs, I. H.

    2012-06-20

    We investigate the changes in polarization position angle in radiation from pulsar A around the eclipse in the Double Pulsar system PSR J0737-3039A/B at the 20 cm and 50 cm wavelengths using the Parkes 64 m telescope. The changes are {approx}2{sigma} during and shortly after the eclipse at 20 cm but less significant at 50 cm. We show that the changes in position angle during the eclipse can be modeled by differential synchrotron absorption in the eclipse regions. Position angle changes after the eclipse are interpreted as Faraday rotation in the magnetotail of pulsar B. Implied charge densities are consistent with the Goldreich-Julian density, suggesting that the particle energies in the magnetotail are mildly relativistic.

  1. Influence of Preoperative Musculotendinous Junction Position on Rotator Cuff Healing After Double-Row Repair.

    PubMed

    Tashjian, Robert Z; Erickson, Gregory A; Robins, Richard J; Zhang, Yue; Burks, Robert T; Greis, Patrick E

    2017-06-01

    The primary purpose of this study was to determine the effect of the preoperative position of the musculotendinous junction (MTJ) on rotator cuff healing after double-row arthroscopic rotator cuff repair. A secondary purpose was to evaluate how tendon length and MTJ position change when the rotator cuff heals. Preoperative and postoperative magnetic resonance imaging (MRI) scans of 42 patients undergoing arthroscopic double-row rotator cuff repair were reviewed. Patients undergoing repairs with other constructs or receiving augmented repairs (platelet-rich fibrin matrix) who had postoperative MRI scans were excluded. Preoperative MRI scans were evaluated for anteroposterior tear size, tendon retraction, tendon length, muscle quality, and MTJ position with respect to the glenoid in the coronal plane. The position of the MTJ was referenced off the glenoid face as either lateral or medial. Postoperative MRI scans were evaluated for healing, tendon length, and MTJ position. Of 42 tears, 36 (86%) healed, with 27 of 31 small to medium tears (87%) and 9 of 11 large to massive tears (82%) healing. Healing occurred in 94% of tears that had a preoperative MTJ lateral to the face of the glenoid but only 56% of tears that had a preoperative MTJ medial to the glenoid face (P = .0135). The measured tendon length increased an average of 14.4 mm in patients whose tears healed compared with shortening by 6.4 mm in patients with tears that did not heal (P < .001). The MTJ lateralized an average of 6.1 mm in patients whose tears healed compared with medializing 1.9 mm in patients whose tears did not heal (P = .026). The overall follow-up period of the study was from April 2005 to September 2014 (113 months). The preoperative MTJ position is predictive of postoperative healing after double-row rotator cuff repair. The position of the MTJ with respect to the glenoid face is a reliable, identifiable marker on MRI scans that can be predictive of healing. Level IV, retrospective

  2. Transcriptional control of transglutaminase 2 expression in mouse apoptotic thymocytes.

    PubMed

    Sándor, Katalin; Daniel, Bence; Kiss, Bea; Kovács, Fruzsina; Szondy, Zsuzsa

    2016-08-01

    Transglutaminase 2 (TGM2) is a ubiquitously expressed multifunctional protein, which participates in various biological processes including thymocyte apoptosis. As a result, the transcriptional regulation of the gene is complex and must depend on the cell type. Previous studies from our laboratory have shown that in dying thymocytes the expression of Tgm2 is induced by external signals derived from engulfing macrophages, such as retinoids, transforming growth factor (TGF)-β and adenosine, the latter triggering the adenylate cyclase signaling pathway. The existence of TGF-β and retinoid responsive elements in the promoter region of Tgm2 has already been reported, but the intergenic regulatory elements participating in the regulation of Tgm2 have not yet been identified. Here we used publicly available results from DNase I hypersensitivity analysis followed by deep sequencing and chromatin immunoprecipitation followed by deep sequencing against CCCTC-binding factor (CTCF), H3K4me3, H3K4me1 and H3K27ac to map a putative regulatory element set for Tgm2 in thymocytes. By measuring eRNA expressions of these putative enhancers in retinoid, rTGF-β or dibutiryl cAMP-exposed thymocytes we determined which of them are functional. By applying ChIP-qPCR against SMAD4, retinoic acid receptor, retinoid X receptor, cAMP response element binding protein, P300 and H3K27ac under the same conditions, we identified two enhancers of Tgm2, which seem to act as integrators of the TGF-β, retinoid and adenylate cyclase signaling pathways in dying thymocytes. Our study describes a novel strategy to identify and characterize the signal-specific functional enhancer set of a gene by integrating genome-wide datasets and measuring the production of enhancer specific RNA molecules.

  3. A rehabilitation training system with double-CCD camera and automatic spatial positioning technique

    NASA Astrophysics Data System (ADS)

    Lin, Chern-Sheng; Wei, Tzu-Chi; Lu, An-Tsung; Hung, San-Shan; Chen, Wei-Lung; Chang, Chia-Chang

    2011-03-01

    This study aimed to develop a computer game for machine vision integrated rehabilitation training system. The main function of the system is to allow users to conduct hand grasp-and-place movement through machine vision integration. Images are captured by a double-CCD camera, and then positioned on a large screen. After defining the right, left, upper, and lower boundaries of the captured images, an automatic spatial positioning technique is employed to obtain their correlation functions, and lookup tables are defined for cameras. This system can provide rehabilitation courses and games that allow users to exercise grasp-and-place movements, in order to improve their upper limb movement control, trigger trunk control, and balance training.

  4. Positive reactions to placebo in children undergoing double-blind, placebo-controlled food challenge.

    PubMed

    Ahrens, B; Niggemann, B; Wahn, U; Beyer, K

    2014-04-01

    The gold standard in the diagnosis of food allergy is the double-blind, placebo-controlled oral food challenge (DBPCFC). During this challenge, patients receive the allergenic food and placebo on separate randomized days, while being monitored for clinical reactions. Interestingly, some reactions are assessed as positive although the patients had received placebo. The aim of our study was to analyze incidence and characteristics of positive placebo reactions during DBPCFCs. In food-allergic children, we retrospectively analyzed positive placebo reactions in DBPCFCs in 740 placebo challenges in our department. Individual characteristics were compared, such as age or IgE levels, as well as clinical symptoms. Of all placebo challenges, 2.8% (21 of 740) were assessed as positive. Young children (age ≤ 1.5 years) had more (P = 0.047) positive placebo challenges (4.0%) compared to older children (age > 1.5 years; 1.5%). Children with positive placebo challenges had higher levels of total IgE (median 201 kU/L) compared to negatively classified children (median 110 kU/L). In children with positive placebo reactions, skin symptoms were observed significantly more often, with a worsening of atopic eczema (AE) as the most reported symptom. Placebo reactions in DBPCFC are not common. Worsening of AE is the most frequent clinical reaction associated with positive placebo challenges, and young children (age ≤ 1.5 years) seem to be affected more often. Therefore - contrary to current recommendations - DBPCFC tests should be considered in infants and young children, especially those with a history of AE. © 2014 John Wiley & Sons Ltd.

  5. Determination of Double Bond Positions and Geometry of Methyl Linoleate Isomers with Dimethyl Disulfide Adducts by GC/MS.

    PubMed

    Shibamoto, Shigeaki; Murata, Tasuku; Yamamoto, Kouhei

    2016-09-01

    The dimethyl disulfide (DMDS) adduct method is one of the convenient and effective methods for determining double bond positions of unsaturated fatty acid methyl esters (FAME) except conjugated FAME. When analyzed using gas chromatography/electron ionization-mass spectrometry (GC/EI-MS), unsaturated FAME with DMDS added to the double bonds yields high intensity MS spectra of characteristic ions. The MS spectra of characteristic ions can then be used to easily confirm double bond positions. Here we explore the GC/EI-MS analysis of the DMDS adducts of methyl linoleate geometrical isomers isolated by high performance liquid chromatography (HPLC) with a silver nitrate column. For C18:2-c9, c12 and C18:2-t9, t12, DMDS randomly formed adducts with double bonds at either carbon 9-10 or carbon 12-13, but not both at the same time due to steric hindrance. For C18:2-c9, t12 and C18:2-t9, c12, however, DMDS only formed adducts with the double bond in the cis configuration. Consequently, when analyzing fatty acids with methylene interrupted double bonds, with one double bond in the cis and one in the trans configuration, double bond positions cannot be completely confirmed.

  6. Separation of human thymocytes at different stages of maturation by centrifugation on a discontinuous gradient of colloidal silica gel.

    PubMed

    Goust, J M; Perry, L R

    1981-06-01

    Separation of human intrathymic cells on a discontinuous gradient of colloidal silica gel (Percoll) yielded four layers. The first (density 1.054 +/- 0.002 g/ml) contained stromal cells and a few thymocytes positive for terminal deoxynucleotidyl transferase (Tdt), most of which were bound to large Tdt-negative non-T cells. The second layer (1.069+/- 0.003 g/ml) contained large Tdt-negative thymocytes. The third and forth layers (1.075 +/-0.004 and 1.085 +/- 0.003 g/ml, respectively) contained smaller T cells, more than 95% of which were Tdt-positive. Functional studies revealed that cells from the first layer had a high level of spontaneous [3H]thymidine uptake but did not respond to lectins; the second layer responded to PHA, ConA, and allogeneic stimuli; and the third and fourth layers did not respond to lectin stimulation. Addition of cells from the first layer to the other layers at a 1 : 10 ratio significantly increased the mitogenic responses of the cells from the second layer, but not of those from the third or fourth layer. These results suggest that, as in mice and rats, low-density intrathymic thymocytes in humans represent more mature T cells, the percentage of which increases with age.

  7. Differential roles for Bim and Nur77 in thymocyte clonal deletion induced by ubiquitous self-antigen.

    PubMed

    Hu, Qian Nancy; Baldwin, Troy A

    2015-03-15

    Negative selection, primarily mediated through clonal deletion of self-reactive thymocytes, is critical for establishing self-tolerance and preventing autoimmunity. Recent studies suggest that the molecular mechanisms of negative selection differ depending on the thymic compartment and developmental stage at which thymocytes are deleted. Using the physiological HY(cd4) TCR transgenic model of negative selection against ubiquitous self-antigen, we previously found that one of the principal mediators implicated in clonal deletion, Bim, is required for caspase-3 activation but is ultimately dispensable for negative selection. On the basis of these data, we hypothesized that Nur77, another molecule thought to be a key mediator of clonal deletion, could be responsible for Bim-independent deletion. Despite comparable Nur77 induction in thymocytes during negative selection, Bim deficiency resulted in an accumulation of high-affinity-signaled thymocytes as well as impairment in caspase-mediated and caspase-independent cell death. Although these data suggested that Bim may be required for Nur77-mediated cell death, we found that transgenic Nur77 expression was sufficient to induce apoptosis independently of Bim. However, transgenic Nur77-induced apoptosis was significantly inhibited in the context of TCR signaling, suggesting that endogenous Nur77 could be similarly regulated during negative selection. Although Nur77 deficiency alone did not alter positive or negative selection, combined deficiency in Bim and Nur77 impaired clonal deletion efficiency and significantly increased positive selection efficiency. Collectively, these data shed light on the different roles for Bim and Nur77 during ubiquitous Ag-mediated clonal deletion and highlight potential differences from their reported roles in tissue-restricted Ag-mediated clonal deletion.

  8. Proteasomes play an essential role in thymocyte apoptosis.

    PubMed Central

    Grimm, L M; Goldberg, A L; Poirier, G G; Schwartz, L M; Osborne, B A

    1996-01-01

    Cell death in many different organisms requires the activation of proteolytic cascades involving cytosolic proteases. Here we describe a novel requirement in thymocyte cell death for the 20S proteasome, a highly conserved multicatalytic protease found in all eukaryotes. Specific inhibitors of proteasome function blocked cell death induced by ionizing radiation, glucocorticoids or phorbol ester. In addition to inhibiting apoptosis, these signals prevented the cleavage of poly(ADP-ribose) polymerase that accompanies many cell deaths. Since overall rates of protein degradation were not altered significantly during cell death in thymocytes, these results suggest that the proteasome may either degrade regulatory protein(s) that normally inhibit the apoptotic pathway or may proteolytically activate protein(s) than promote cell death. Images PMID:8670888

  9. Newly developed double neural network concept for reliable fast plasma position control

    NASA Astrophysics Data System (ADS)

    Jeon, Young-Mu; Na, Yong-Su; Kim, Myung-Rak; Hwang, Y. S.

    2001-01-01

    Neural network is considered as a parameter estimation tool in plasma controls for next generation tokamak such as ITER. The neural network has been reported to be so accurate and fast for plasma equilibrium identification that it may be applied to the control of complex tokamak plasmas. For this application, the reliability of the conventional neural network needs to be improved. In this study, a new idea of double neural network is developed to achieve this. The new idea has been applied to simple plasma position identification of KSTAR tokamak for feasibility test. Characteristics of the concept show higher reliability and fault tolerance even in severe faulty conditions, which may make neural network applicable to plasma control reliably and widely in future tokamaks.

  10. Positional calibrations of the germanium double sided strip detectors for the Compton spectrometer and imager

    NASA Astrophysics Data System (ADS)

    Lowell, A.; Boggs, S.; Chiu, J. L.; Kierans, C.; McBride, S.; Tseng, C. H.; Zoglauer, A.; Amman, M.; Chang, H. K.; Jean, P.; Lin, C. H.; Sleator, C.; Tomsick, J.; von Ballmoos, P.; Yang, C. Y.

    2016-08-01

    The Compton Spectrometer and Imager (COSI) is a medium energy gamma ray (0.2 - 10 MeV) imager designed to observe high-energy processes in the universe from a high altitude balloon platform. At its core, COSI is comprised of twelve high purity germanium double sided strip detectors which measure particle interaction energies and locations with high precision. This manuscript focuses on the positional calibrations of the COSI detectors. The interaction depth in a detector is inferred from the charge collection time difference between the two sides of the detector. We outline our previous approach to this depth calibration and also describe a new approach we have recently developed. Two dimensional localization of interactions along the faces of the detector (x and y) is straightforward, as the location of the triggering strips is simply used. However, we describe a possible technique to improve the x/y position resolution beyond the detector strip pitch of 2 mm. With the current positional calibrations, COSI achieves an angular resolution of 5.6 +/- 0.1 degrees at 662 keV, close to our expectations from simulations.

  11. Position effects influencing intrachromosomal repair of a double-strand break in budding yeast.

    PubMed

    Wang, Ruoxi W; Lee, Cheng-Sheng; Haber, James E

    2017-01-01

    Repair of a double-strand break (DSB) by an ectopic homologous donor sequence is subject to the three-dimensional arrangement of chromosomes in the nucleus of haploid budding yeast. The data for interchromosomal recombination suggest that searching for homology is accomplished by a random collision process, strongly influenced by the contact probability of the donor and recipient sequences. Here we explore how recombination occurs on the same chromosome and whether there are additional constraints imposed on repair. Specifically, we examined how intrachromosomal repair is affected by the location of the donor sequence along the 813-kb chromosome 2 (Chr2), with a site-specific DSB created on the right arm (position 625 kb). Repair correlates well with contact frequencies determined by chromosome conformation capture-based studies (r = 0.85). Moreover, there is a profound constraint imposed by the anchoring of the centromere (CEN2, position 238 kb) to the spindle pole body. Sequences at the same distance on either side of CEN2 are equivalently constrained in recombining with a DSB located more distally on one arm, suggesting that sequences on the opposite arm from the DSB are not otherwise constrained in their interaction with the DSB. The centromere constraint can be partially relieved by inducing transcription through the centromere to inactivate CEN2 tethering. In diploid cells, repair of a DSB via its allelic donor is strongly influenced by the presence and the position of an ectopic intrachromosomal donor.

  12. Correlation between Femoral Guidewire Position and Tunnel Communication in Double Bundle Anterior Cruciate Ligament Reconstruction

    PubMed Central

    Lee, Sang Hyuk; Choi, Jun Young; Kim, Dong Hee; Kang, Bun Jung; Nam, Dae Cheol; Yoon, Hong Kwon

    2014-01-01

    Purpose The object of this study was to determine the shortest possible distances of antero-medial (AM) and postero-lateral (PL) guide wire tunnel positions required to prevent femoral bone tunnel communication in double-bundle anterior cruciate ligament (ACL) reconstruction using human cadaver knees. Materials and Methods The centers of femoral AM and PL bundles of 16 cadaveric knees were drilled with guide wires and the distances of guide wires, were measured upon entrance into the bone. Femoral tunnel drilling was performed using transportal technique. The diameters of AM and PL graft were 8 mm and 6 mm, respectively. CT scans were taken on each knee, and 3-dimensional models were constructed to identify the femoral tunnel position and to create AM and PL tunnel virtual cylinders. Thickness of the bone bridge between the two tunnels was measured. Results In four out of six specimens, in which the guide wires were placed at less than or equal to 9 mm, communication was noted. In specimens with guide wires placed at distances greater than or equal to 10 mm, communication was not noted. The two groups showed a statistically significant difference (p=0.008). In cases where the distance between the AM and PL femoral tunnel guide wires was 12 mm, the bone bridge thickness was greater than 2 mm along the tunnel. Conclusion The technique for double bundle-anterior cruciate ligament (DB-ACL) reconstruction that we show here can avoid bone tunnel communication when AM and PL femoral guide wires are placed at least 10 mm apart, and 12 mm should be kept to preserve 2 mm bone bridge thickness. PMID:25323896

  13. Ozone-induced dissociation: elucidation of double bond position within mass-selected lipid ions.

    PubMed

    Thomas, Michael C; Mitchell, Todd W; Harman, David G; Deeley, Jane M; Nealon, Jessica R; Blanksby, Stephen J

    2008-01-01

    Ions formed from lipids during electrospray ionization of crude lipid extracts have been mass-selected within a quadrupole linear ion trap mass spectrometer and allowed to react with ozone vapor. Gas-phase ion-molecule reactions between unsaturated lipid ions and ozone are found to yield two primary product ions for each carbon-carbon double bond within the molecule. The mass-to-charge ratios of these chemically induced fragments are diagnostic of the position of unsaturation within the precursor ion. This novel analytical technique, dubbed ozone-induced dissociation (OzID), can be applied both in series and in parallel with conventional collision-induced dissociation (CID) to provide near-complete structural assignment of unknown lipids within complex mixtures without prior fractionation or derivatization. In this study, OzID is applied to a suite of complex lipid extracts from sources including human lens, bovine kidney, and commercial olive oil, thus demonstrating the technique to be applicable to a broad range of lipid classes including both neutral and acidic glycerophospholipids, sphingomyelins, and triacylglycerols. Gas-phase ozonolysis reactions are also observed with different types of precursor ions including [M+H]+, [M+Li]+, [M+Na]+, and [M-H]-: in each case yielding fragmentation data that allow double bond position to be unambiguously assigned. Within the human lens lipid extract, three sphingomyelin regioisomers, namely SM(d18:0/15Z-24:1), SM(d18:0/17Z-24:1), and SM(d18:0/19Z-24:1), and a novel phosphatidylethanolamine alkyl ether, GPEtn(11Z-18:1e/9Z-18:1), are identified using a combination of CID and OzID. These discoveries demonstrate that lipid identification based on CID alone belies the natural structural diversity in lipid biochemistry and illustrate the potential of OzID as a complementary approach within automated, high-throughput lipid analysis protocols.

  14. Ultraviolet exposure of thymocytes: selective inhibition of apoptosis.

    PubMed

    Ojeda, F; Guarda, M I; Lovengreen, C; Hidalgo, M A; Folch, H; Härtel, S; Maldonado, C

    2004-06-01

    To evaluate selective effects of ultraviolet (UV) irradiation on spontaneous and induced apoptosis in freshly extracted mice thymocytes. Cells were exposed to UV radiation with emission peaks of 365 nm (UVA) exposures of 1620-10200 J m(-2), of 312 nm (UVB) exposures of 34-1620 J m(-2) or of 254 nm (UVC) exposures of 1.5-1620 J m(-2), and incubated for 5.5 h with or without hydrocortisone, phorbol-12-myristate-13-acetate or anti-Fas antibody. Additionally, cells were irradiated with gamma-rays (5 Gy) before UVB exposure (408 J m(-2)) at different times. Apoptosis was quantified by DNA fragmentation. Up to an irradiation of 5000 J m(-2), UVA exposure did not show any effect on thymocyte apoptosis, while at 10200 J m(-2) irradiation, considerable DNA fragmentation was observed. In contrast, UVB and UVC irradiation clearly inhibited natural and cortisone-induced apoptosis. Moreover, UVB inhibited apoptosis triggered by phorbol-12-myristate-13-acetate and gamma-irradiation, but not by anti-Fas antibody. The response of mouse thymocytes in culture to UV irradiation strongly depends on the wavelength used. It is suggested that either a survival or an apoptotic pathway occurs depending on the physiological state of the cell, spectral composition of the UV light and cell type. The possible involvement of extracellular signal-regulated kinase and stress-activated protein kinase/c-Jun N-terminal kinase in the apoptotic pathway is discussed.

  15. The direct determination of double bond positions in lipid mixtures by liquid chromatography/in-line ozonolysis/mass spectrometry.

    PubMed

    Sun, Chenxing; Zhao, Yuan-Yuan; Curtis, Jonathan M

    2013-01-31

    The direct determination of double bond positions in unsaturated lipids using in-line ozonolysis-mass spectrometry (O(3)-MS) is described. In this experiment, ozone penetrates through the semi-permeable Teflon AF-2400 tubing containing a flow of a solution of fatty acid methyl esters (FAME). Unsaturated FAME are thus oxidized by the ozone and cleaved at the double bond positions. The ozonolysis products then flow directly into the atmospheric pressure photoionization (APPI) source of a mass spectrometer for analysis. Aldehyde products retaining the methyl ester group are indicative of the double bond positions in unsaturated FAME. For the first time, O(3)-MS is able to couple directly to high performance liquid chromatography (HPLC), making the double bond localization in lipid mixtures possible. The application of LC/O(3)-MS has been demonstrated for a fat sample from bovine adipose tissue. A total of 9 unsaturated FAME including 6 positional isomers were identified unambiguously, without comparison to standards. The in-line ozonolysis reaction apparatus is applicable to most mass spectrometers without instrumental modification; it is also directly compatible with various LC columns. The LC/O(3)-MS method described here is thus a practical, versatile and easy to use new approach to the direct determination of double bond positions in lipids, even in complex mixtures.

  16. S1P lyase in thymic perivascular spaces promotes egress of mature thymocytes via up-regulation of S1P receptor 1.

    PubMed

    Maeda, Yasuhiro; Yagi, Hideki; Takemoto, Kana; Utsumi, Hiroyuki; Fukunari, Atsushi; Sugahara, Kunio; Masuko, Takashi; Chiba, Kenji

    2014-05-01

    Sphingosine 1-phosphate (S1P) and S1P receptor 1 (S1P1) play an important role in the egress of mature CD4 or CD8 single-positive (SP) thymocytes from the thymus. Fingolimod hydrochloride (FTY720), an S1P1 functional antagonist, induced significant accumulation of CD62L(high)CD69(low) mature SP thymocytes in the thymic medulla. Immunohistochemical staining using anti-S1P1 antibody revealed that S1P1 is predominantly expressed on thymocytes in the thymic medulla and is strongly down-regulated even at 3h after FTY720 administration. 2-Acetyl-4-tetrahydroxybutylimidazole (THI), an S1P lyase inhibitor, also induced accumulation of mature SP thymocytes in the thymic medulla with an enlargement of the perivascular spaces (PVS). At 6h after THI administration, S1P1-expressing thymocytes reduced partially as if to form clusters and hardly existed in the proximity of CD31-expressing blood vessels in the thymic medulla, suggesting S1P lyase expression in the cells constructing thymic medullary PVS. To determine the cells expressing S1P lyase in the thymus, we newly established a mAb (YK19-2) specific for mouse S1P lyase. Immunohistochemical staining with YK19-2 revealed that S1P lyase is predominantly expressed in non-lymphoid thymic stromal cells in the thymic medulla. In the thymic medullary PVS, S1P lyase was expressed in ER-TR7-positive cells (reticular fibroblasts and pericytes) and CD31-positive vascular endothelial cells. Our findings suggest that S1P lyase expressed in the thymic medullary PVS keeps the tissue S1P concentration low around the vessels and promotes thymic egress via up-regulation of S1P1.

  17. Slant Wet Delays from GNSS observations - Precise Point Positioning vs. Double Difference Approach

    NASA Astrophysics Data System (ADS)

    Moeller, Gregor; Weber, Robert

    2015-04-01

    The tropospheric parameter SWD (Slant Wet Delay) is the path delay caused by the highly variable amount of humidity in the atmosphere at altitudes below 12 km. It can be derived from Numerical Weather Predication data or even more precisely from dual- or multi-frequency observations of a regional GNSS reference network. In order to find the most adequate processing strategy dual GNSS observations of a small network of reference stations were simulated and tropospheric parameters were estimated in Precise Point Positioning (PPP) and in Double Difference (DD) mode. In DD mode the integer character of the phase ambiguities remains which allows to fix them to their true values and to obtain the tropospheric zenith delay as well as north and east (N/E) gradients with highest precision over very short time periods. In PPP mode orbit and clock errors are not cancelled out which affects the quality of the tropospheric estimates. On the other hand it has the advantage that the GNSS observations are processed undifferenced. Latter is important because the Zero Difference Residuals (ZDR) contain the azimuthal-anisotropic part of the tropospheric delay which is not covered by the estimated parameters. From Double Difference Residuals (DDR) the ZDR can be recovered too but only conditionally since common tropospheric effects have been cancelled out in advance by differencing. In this presentation we show how good the anisotropic slant path delays can be obtained from GNSS observations processed using both concepts - the PPP and the DD approach. Therefore tropospheric zenith delays and N/E gradients were estimated and Pseudo-ZDR were reconstructed from DDR and afterwards compared with ZDR derived from the PPP solution. In addition it is shown how good both concepts are applicable for observations at very low elevation angles and under extreme weather conditions. The IGS final and ultra-rapid service products were taken into account to define the best strategy not only for post

  18. Characterization of circulating CD4(+) CD8(+) double positive and CD4(-) CD8(-) double negative T-lymphocyte in children with β-thalassemia major.

    PubMed

    Zahran, Asmaa M; Saad, Khaled; Elsayh, Khalid I; Alblihed, Mohamd A

    2017-03-01

    Infectious complications represent the second most common cause of mortality and a major cause of morbidity in β-thalassemia major (BTM), with a prevalence of 12-13%. The data on unconventional T-lymphocyte subsets in BTM children are limited. The aim of the present study was to investigate and evaluate phenotypic alterations in CD4(+)CD8(+) double positive (DP), CD4(-) CD8(-) double negative (DN), and natural killer T-lymphocytes (NKT) in BTM children in comparison to healthy controls. Our case control study included 80 children with BTM and 40 healthy children as controls. Assessment of unconventional T-lymphocyte populations was done using sensitive four-color flow cytometry (FACSCalibur). Our analysis of the data showed a significantly higher frequency CD4(+) CD8(+) (double-positive) T cells, CD4(-) CD8(-) (double negative) T cells, and natural killer T cells in the peripheral blood of both BTM groups (splenectomized and non-splenectomized) as compared to healthy controls, suggesting that these cells may play a role in the clinical course of BTM. The relationship of the unconventional T-lymphocytes to immune disorders in BTM children remains to be determined. Further longitudinal study with a larger sample size is warranted to elucidate the role these cells in BTM.

  19. Thymic medullary epithelium and thymocyte self tolerance require cooperation between CD28-CD80/86 and CD40-CD40L costimulatory pathways

    PubMed Central

    Williams, Joy A.; Zhang, Jingjing; Jeon, Hyein; Nitta, Takeshi; Ohigashi, Izumi; Klug, David; Kruhlak, Michael J.; Choudhury, Baishakhi; Sharrow, Susan O.; Granger, Larry; Adams, Anthony; Eckhaus, Michael A.; Jenkinson, S. Rhiannon; Richie, Ellen R.; Gress, Ronald E.; Takahama, Yousuke; Hodes, Richard J.

    2014-01-01

    A critical process during thymic development of the T cell repertoire is the induction of self-tolerance. Tolerance in developing T cells is highly dependent on medullary thymic epithelial cells (mTEC) and mTEC development in turn requires signals from mature single positive (SP) thymocytes, a bidirectional relationship termed thymus crosstalk. We show that CD28-CD80/86 and CD40-CD40L costimulatory interactions, which mediate negative selection and self-tolerance, upregulate expression of LTα, LTβ and RANK in the thymus and are necessary for medullary development. Combined absence of CD28-CD80/86 and CD40-CD40L results in profound deficiency in mTEC development comparable to that observed in the absence of SP thymocytes. This requirement for costimulatory signaling is maintained even in a TCR transgenic model of high affinity TCR-ligand interactions. CD4 thymocytes maturing in the altered thymic epithelial environment of CD40/CD80/86 KO mice are highly autoreactive in vitro and are lethal in congenic adoptive transfer in vivo, demonstrating a critical role for these costimulatory pathways in self-tolerance as well as thymic epithelial development. These findings demonstrate that cooperativity between CD28-CD80/86 and CD40-CD40L pathways is required for normal medullary epithelium and for maintenance of self-tolerance in thymocyte development. PMID:24337745

  20. Multiparameter double hole contrast detail phantom: Ability to detect image displacement due to off position anode stem

    NASA Astrophysics Data System (ADS)

    Pauzi, Nur Farahana; Majid, Zafri Azran Abdul; Sapuan, Abdul Halim; Azemin, Mohd Zulfaezal Che; Junet, Laila Kalidah

    2015-04-01

    Contrast Detail phantom is a quality control tool to analyze the performance of imaging devices. Currently, its function is solely to evaluate the contrast detail characteristic of imaging system. It consists of drilled hole which gives effect to the penetration of x-ray beam divergence to pass through the base of each hole. This effect will lead to false appearance of image from its original location but it does not being visualized in the radiograph. In this study, a new design of Contrast Detail phantom's hole which consists of double hole construction has been developed. It can detect the image displacement which is due to off position of anode stem from its original location. The double hole differs from previous milled hole, whereby it consists of combination of different hole diameters. Small hole diameter (3 mm) is positioned on top of larger hole diameter (10 mm). The thickness of double hole acrylic blocks is 13 mm. Result revealed` that, Multiparameter Double Hole Contrast Detail phantom can visualize the shifted flaw image quality produced by x-ray machine due to improper position of the anode stem which is attached to rotor and stator. The effective focal spot of x-ray beam also has been shifted from the center of collimator as a result of off-position anode stem. As a conclusion, the new design of double hole Contrast Detail phantom able to measure those parameters in a well manner.

  1. Multiparameter double hole contrast detail phantom: Ability to detect image displacement due to off position anode stem

    SciTech Connect

    Pauzi, Nur Farahana; Majid, Zafri Azran Abdul; Sapuan, Abdul Halim; Junet, Laila Kalidah; Azemin, Mohd Zulfaezal Che

    2015-04-24

    Contrast Detail phantom is a quality control tool to analyze the performance of imaging devices. Currently, its function is solely to evaluate the contrast detail characteristic of imaging system. It consists of drilled hole which gives effect to the penetration of x-ray beam divergence to pass through the base of each hole. This effect will lead to false appearance of image from its original location but it does not being visualized in the radiograph. In this study, a new design of Contrast Detail phantom’s hole which consists of double hole construction has been developed. It can detect the image displacement which is due to off position of anode stem from its original location. The double hole differs from previous milled hole, whereby it consists of combination of different hole diameters. Small hole diameter (3 mm) is positioned on top of larger hole diameter (10 mm). The thickness of double hole acrylic blocks is 13 mm. Result revealed that Multiparameter Double Hole Contrast Detail phantom can visualize the shifted flaw image quality produced by x-ray machine due to improper position of the anode stem which is attached to rotor and stator. The effective focal spot of x-ray beam also has been shifted from the center of collimator as a result of off-position anode stem. As a conclusion, the new design of double hole Contrast Detail phantom able to measure those parameters in a well manner.

  2. Hypergravity Provokes a Temporary Reduction in CD4+CD8+ Thymocyte Number and a Persistent Decrease in Medullary Thymic Epithelial Cell Frequency in Mice

    PubMed Central

    Miyauchi, Maki; Yoshinaga, Riko; Sasanuma, Hiroki; Kudo, Takashi; Shimbo, Miki; Shinohara, Masahiro; Obata, Koji; Inoue, Jun-ichiro; Shirakawa, Masaki; Shiba, Dai; Asahara, Hiroshi; Yoshida, Nobuaki; Takahashi, Satoru; Morita, Hironobu; Akiyama, Taishin

    2015-01-01

    Gravity change affects many immunological systems. We investigated the effects of hypergravity (2G) on murine thymic cells. Exposure of mice to 2G for three days reduced the frequency of CD4+CD8+ thymocytes (DP) and mature medullary thymic epithelial cells (mTECs), accompanied by an increment of keratin-5 and keratin-8 double-positive (K5+K8+) TECs that reportedly contain TEC progenitors. Whereas the reduction of DP was recovered by a 14-day exposure to 2G, the reduction of mature mTECs and the increment of K5+K8+ TEC persisted. Interestingly, a surgical lesion of the inner ear’s vestibular apparatus inhibited these hypergravity effects. Quantitative PCR analysis revealed that the gene expression of Aire and RANK that are critical for mTEC function and development were up-regulated by the 3-day exposure and subsequently down-regulated by the 14-day exposure to 2G. Unexpectedly, this dynamic change in mTEC gene expression was independent of the vestibular apparatus. Overall, data suggest that 2G causes a temporary reduction of DP and a persistent reduction of mature mTECs in a vestibular system-dependent manner, and also dysregulates mTEC gene expression without involving the vestibular system. These data might provide insight on the impact of gravity change on thymic functions during spaceflight and living. PMID:26513242

  3. Thermodynamic analysis of chain-melting transition temperatures for monounsaturated phospholipid membranes: dependence on cis-monoenoic double bond position.

    PubMed Central

    Marsh, D

    1999-01-01

    Unsaturated phospholipid is the membrane component that is essential to the dynamic environment needed for biomembrane function. The dependence of the chain-melting transition temperature, T(t), of phospholipid bilayer membranes on the position, n(u), of the cis double bond in the glycerophospholipid sn-2 chain can be described by an expression of the form T(t) = T(t)(c)(1 + h'(c)|n(u) - n(c)|)/(1 + s'(c)|n(u) - n(c)|), where n(c) is the chain position of the double bond corresponding to the minimum transition temperature, T(t)(c), for constant diacyl lipid chain lengths. This implies that the incremental transition enthalpy (and entropy) contributed by the sn-2 chain is greater for whichever of the chain segments, above or below the double-bond position, is the longer. The critical position, n(c), of the double bond is offset from the center of the sn-2 chain by an approximately constant amount, deltan(c) approximately 1. 5 C-atom units. The dependence of the parameters T(t)(c), h'(c), and s'(c) on sn-1 and sn-2 chain lengths can be interpreted consistently when allowance is made for the chain packing mismatch between the sn-1 and sn-2 chains. The length of the sn-2 chain is reduced by approximately 0.8 C-atom units by the cis double bond, in addition to a shortening by approximately 1.3 C-atom units by the bent configuration at the C-2 position. Based on this analysis, a general thermodynamic expression is proposed for the dependence of the chain-melting transition temperature on the position of the cis double bond and on the sn-1 and sn-2 chain lengths. The above treatment is restricted mostly to double-bond positions close to the center of the sn-2 chain. For double bonds positioned closer to the carboxyl or terminal methyl ends of the sn-2 chain, the effects on transition enthalpy can be considerably larger. They may be interpreted by the same formalism, but with different characteristic parameters, h'(c) and s'(c), such that the shorter of the chain segments

  4. Head Strap Double Fluid Level Device: An Innovative and User Friendly Design to Record Natural Head Position (NHP)

    PubMed Central

    Jose, Nidhin Philip; Shetty, Siddarth

    2015-01-01

    Head positions can be oriented in a standardized position when the patient stands upright and focusses his/her eyes into a point in infinity. This is the natural head position. This position offers the maximum reproducibility and correlates well with the clinical picture offered to the diagnostician. This article describes an innovative and user friendly method to record natural head position using the head strap double fluid level device, a design modified from the popular fluid level device by Showfety, Vig and Matteson. PMID:25738103

  5. Anchorage and lymphocyte function. Spreading-capacity distinguishes common thymocytes and peripheral T lymphocytes.

    PubMed Central

    Otteskog, P; Sundqvist, K G

    1983-01-01

    Contact of T-enriched human blood lymphocytes with an adhesive surface in the presence of Concanavalin A (Con A) almost immediately induced a sequence of motile changes in virtually all cells. The initial event in this spreading process was the formation of filopodia distinct from the microvilli of lymphocytes in suspension. The filopodia were accompanied by lamellipodia, ruffles and flattening of the nucleus. Contact with a nonadhesive substratum in the presence of Con A did not trigger this sequence of changes. Cytochalasin B and D or low temperature inhibited the contact-induced changes. With the exception of a small number of cells (5-15%), T-enriched lymphocytes that were allowed to settle in the absence of Con A showed a radius of action (area occupied by the cells/translational movement per hr) of 39 micrometers 2/ less than 1 micrometer. The small 'motile' population showed a radius of action of 74 micrometers 2/8 micrometers. The Con-A-mediated spreading-process yielded a radius of action of the lymphocytes of 117 micrometers 2/6 micrometers. This augmented radius of action markedly facilitated cell-cell interaction in a high frequency of the cells and appeared to be a prerequisite for such interactions at 'low' cell density. Thymocytes reactive with OKT 6 antibodies or belonging to the 'high-density' fraction of cells attached to a Con-A-coated surface to the same extent as peripheral OKT 3 positive lymphocytes, but did not exhibit the morphological changes characteristic of a spreading-process. In contrast, OKT 6 negative thymocytes or thymocytes with a relatively low density showed spreading indistinguishable from that of OKT 3 positive peripheral lymphocytes. These results characterize the spreading-process in human T lymphocytes and demonstrate its functional importance for interactions with the environment. Spreading-capacity appears to reflect the stage of maturation of T cells. Images Figure 1 Figure 2 Figure 3 Figure 4b Figure 4c Figure 7 PMID

  6. Dynamic Characteristics of Mechanical Ventilation System of Double Lungs with Bi-Level Positive Airway Pressure Model

    PubMed Central

    Shen, Dongkai; Zhang, Qian

    2016-01-01

    In recent studies on the dynamic characteristics of ventilation system, it was considered that human had only one lung, and the coupling effect of double lungs on the air flow can not be illustrated, which has been in regard to be vital to life support of patients. In this article, to illustrate coupling effect of double lungs on flow dynamics of mechanical ventilation system, a mathematical model of a mechanical ventilation system, which consists of double lungs and a bi-level positive airway pressure (BIPAP) controlled ventilator, was proposed. To verify the mathematical model, a prototype of BIPAP system with a double-lung simulators and a BIPAP ventilator was set up for experimental study. Lastly, the study on the influences of key parameters of BIPAP system on dynamic characteristics was carried out. The study can be referred to in the development of research on BIPAP ventilation treatment and real respiratory diagnostics. PMID:27660646

  7. Differences in the pattern of proliferative response with age in thymocytes undergoing spontaneous and induced proliferation.

    PubMed

    Hameed, M R; Coffman, F D; Cohen, S; Fresa, K L

    1989-10-15

    The proliferative capacity of thymocytes from C3H/HeJ mice decrease as the animals attain maturity. The proliferative response of thymocytes from 24- to 28-week-old mice to stimulation with concanavalin A (Con A) is only 20% of that observed at 4 weeks of age. The decreased proliferative capacity of thymocytes in response to Con A stimulation observed between 4 and 24 weeks of age closely correlates to the drop in thymic weight and cellularity observed during this period. In contrast, the spontaneous proliferative capacity of thymocytes, as well as proliferation of thymocytes in response to stimulation with phorbol myristate acetate (PMA) and ionomycin, drops only slightly during this period, as proliferation under these condition in thymocytes from 24- to 28-week-old mice is approximately 65-70% of that observed in 4-week-old animals. We have previously shown that cytoplasmic extracts from proliferating lymphoid cells contain a factor, termed the activator of DNA replication (ADR), which is capable of inducing DNA synthesis in isolated, quiescent nuclei. We show in this study that the decreased proliferative capacity of thymocytes during whole organism maturation and thymic involution is associated with decreased endogenous levels of ADR, while nuclear sensitivity of thymocyte to ADR was retained during these process. The diminution of ADR activity during thymic involution was quantitatively greater than the loss in proliferative capacity.

  8. Functional characteristics of a double positive feedback loop coupled with autorepression

    NASA Astrophysics Data System (ADS)

    Banerjee, Subhasis; Bose, Indrani

    2008-12-01

    We study the functional characteristics of a two-gene motif consisting of a double positive feedback loop and an autoregulatory negative feedback loop. The motif appears in the gene regulatory network controlling the functional activity of pancreatic β-cells. The model exhibits bistability and hysteresis in appropriate parameter regions. The two stable steady states correspond to low (OFF state) and high (ON state) protein levels, respectively. Using a deterministic approach, we show that the region of bistability increases in extent when the copy number of one of the genes is reduced from 2 to 1. The negative feedback loop has the effect of reducing the size of the bistable region. Loss of a gene copy, brought about by mutations, hampers the normal functioning of the β-cells giving rise to the genetic disorder, maturity-onset diabetes of the young (MODY). The diabetic phenotype makes its appearance when a sizable fraction of the β-cells is in the OFF state. Using stochastic simulation techniques we show that, on reduction of the gene copy number, there is a transition from the monostable ON to the ON state in the bistable region of the parameter space. Fluctuations in the protein levels, arising due to the stochastic nature of gene expression, can give rise to transitions between the ON and OFF states. We show that as the strength of autorepression increases, the ON → OFF state transitions become less probable whereas the reverse transitions are more probable. The implications of the results in the context of the occurrence of MODY are pointed out.

  9. The mechanism of epipodophyllotoxin-induced thymocyte apoptosis: possible role of a novel Ca(2+)-independent protein kinase.

    PubMed

    Ye, X; Georgoff, I; Fleisher, S; Coffman, F D; Cohen, S; Fresa, K L

    1993-10-15

    The epipodophyllotoxins, etoposide (VP-16) and teniposide (VM-26), inhibit topoisomerase II activity by stabilization of the cleavable complex between the enzyme and DNA and formation of protein-bound double-stranded DNA breaks. While it is thought that these agents are cytotoxic by preventing cells from completing the S phase or undergoing mitosis, recent evidence suggests that these agents are also potent inducers of programmed cell death or apoptosis in both normal and malignant cells. We have examined the intracellular pathway leading to epipodophyllotoxin-induced apoptosis in normal mouse thymocytes. Epipodophyllotoxin-induced apoptosis may proceed via a mechanism that is independent of inhibition of topoisomerase activity per se because novobiocin and coumermycin, which inhibit the ATPase subunit of topoisomerase II, were relatively inefficient inducers of apoptosis in these cells, under conditions where strong apoptosis by the epipodophyllotoxins and dexamethasone could be observed. In addition, camptothecin, which inhibits topoisomerase I by stabilization of the cleavable complex between that enzyme and DNA, was also a poor inducer of apoptosis in these cells. Our data suggest that epipodophyllotoxin-induced mouse thymocyte apoptosis, like that induced by dexamethasone, proceeds via a mechanism that involves protein kinase C (PKC) or a similar enzyme. Apoptosis induced by VM-26 or by dexamethasone was inhibited by 1-(5-isoquinolinylsulfonyl)-2- methylpiperazine dihydrochloride (H7), an inhibitor of both PKC and cAMP-dependent protein kinases, but was relatively unaffected by N-(2-guanidinoethyl)-5-isoquinolinesulfonamide (HA1004), a more specific inhibitor of cAMP-dependent protein kinases. A more specific inhibitor of PKC, sangivamycin, also inhibited both VM-26-induced and dexamethasone-induced apoptosis. Both VM-26- and dexamethasone-induced apoptosis were unaffected by EGTA, a calcium (Ca2+) chelator, under conditions that inhibited apoptosis induced by

  10. Activins and inhibins: Novel regulators of thymocyte development

    SciTech Connect

    Licona-Limon, Paula; Aleman-Muench, German; Macias-Silva, Marina; Garcia-Zepeda, Eduardo A.; Fortoul, Teresa I.; Soldevila, Gloria

    2009-04-03

    Activins and inhibins are members of the transforming growth factor-{beta} superfamily that act on different cell types and regulate a broad range of cellular processes including proliferation, differentiation, and apoptosis. Here, we provide the first evidence that activins and inhibins regulate specific checkpoints during thymocyte development. We demonstrate that both activin A and inhibin A promote the DN3-DN4 transition in vitro, although they differentially control the transition to the DP stage. Whereas activin A induces the accumulation of a CD8{sup +}CD24{sup hi}TCR{beta}{sup lo} intermediate subpopulation, inhibin A promotes the differentiation of DN4 to DP. In addition, both activin A and inhibin A appear to promote CD8{sup +}SP differentiation. Moreover, inhibin {alpha} null mice have delayed in vitro T cell development, showing both a decrease in the DN-DP transition and reduced thymocyte numbers, further supporting a role for inhibins in the control of developmental signals taking place during T cell differentiation in vivo.

  11. Stimulation of respiration in rat thymocytes induced by ionizing radiation.

    PubMed

    Gudz, T I; Pandelova, I G; Novgorodov, S A

    1994-04-01

    The effect of X irradiation on the respiration of rat thymocytes was studied. An increase in the rate of O2 uptake was observed 1 h after cells were irradiated with doses of 6-10 Gy. The radiation-induced increase in respiration could be blocked by oligomycin, an inhibitor of mitochondrial ATP synthase, suggesting control by increased cytoplasmic ATP turnover. The stimulation of respiration was not associated with changes in the activity of mitochondrial electron transfer enzymes or permeability of the inner membrane. Several inhibitors of processes which used ATP were screened for their effects on the basal respiration rate and on the radiation response. In irradiated thymocytes, an enhancement of inhibition of respiration by ouabain, La3+ and cycloheximide was observed. These results indicate that the radiation-induced stimulation of respiration is due to changes in ion homeostasis and protein synthesis. The effect of X irradiation was shown to be independent of the redox status of nonprotein thiols and was not associated with detectable changes in some products of lipid peroxidation. The radiation-induced decrease in activity of superoxide dismutase suggests free radical involvement in deleterious effects of radiation.

  12. Melatonin inhibits glucocorticoid receptor nuclear translocation in mouse thymocytes.

    PubMed

    Presman, Diego M; Hoijman, Esteban; Ceballos, Nora R; Galigniana, Mario D; Pecci, Adali

    2006-11-01

    The antiapoptotic effect of melatonin (MEL) has been described in several systems. In particular, MEL inhibits glucocorticoid-mediated apoptosis. Our group previously demonstrated that in the thymus, MEL inhibits the release of Cytochrome C from mitochondria and the dexamethasone-dependent increase of bax mRNA levels. In this study we analyzed the ability of MEL to regulate the activation of the glucocorticoid receptor (GR) in mouse thymocytes. We found that even though the methoxyindole does not affect the ligand binding capacity of the receptor, it impairs the steroid-dependent nuclear translocation of the GR and also prevents transformation by blocking the dissociation of the 90-kDa heat shock protein. Coincubation of the methoxyindole with dexamethasone did not affect the expression of a reporter gene in GR-transfected Cos-7 cells or HC11 and L929 mouse cell lines that express Mel-1a and retinoid-related orphan receptor-alpha (RORalpha) receptors. Therefore, the antagonistic effect of MEL seems to be specific for thymocytes, in a Mel 1a- and RORalpha-independent manner. In summary, the present results suggest a novel mechanism for the antagonistic action of MEL on GR-mediated effects, which involves the inhibition of 90-kDa heat shock protein dissociation and the cytoplasmic retention of the GR.

  13. Oxidative damage, bleomycin, and gamma radiation induce different types of DNA strand breaks in normal lymphocytes and thymocytes. A comet assay study.

    PubMed

    Benítez-Bribiesca, L; Sánchez-Suárez, P

    1999-01-01

    Most anticancer treatments such as chemo- and radiotherapy induce DNA damage and apoptosis in normal cells. The aim of this study was to assess the induction of single and double DNA strand breaks (ssb and dsb, respectively) and apoptosis in normal human lymphocytes and rat thymocytes subjected to the action of H2O2, bleomycin and ionizing radiation. Normal human peripheral thymocytes and young rat thymocytes were subjected to the following treatments: a) H2O2; b) bleomycin, and c) gamma-radiation, all with various doses. DNA strand breaks were studied with the alkaline and neutral comet assay for detection of ssb and dsb. Apoptosis was quantified morphologically and with DNA agarose gel electrophoresis. After H2O2 treatment, a dose-dependent increase of ssb was observed. Bleomycin treatment produced a moderate increase of ssb at lower concentrations and a striking increase of dsb at higher concentrations that coincided with the presence of apoptosis and DNA ladders. Gamma radiation initially induced the formation of ssb, and after three hours an increase of dsb in a dose-dependent manner. Apoptosis and DNA laddering appeared only 3 hours post-irradiation. The biomonitoring of DNA damage inflicted by antineoplastic agents can be easily performed with the comet assay and could be useful to monitor and modulate chemo- and radiotherapeutic regimes in cancer patients.

  14. EBV induces proliferation of immature human thymocytes in an IL-2-mediated response.

    PubMed

    Todd, S C; Tsoukas, C D

    1996-06-01

    The receptor for EBV, CD21 is expressed on a population of immature human thymocytes and facilitates infection of these cells by EBV. Thymocytes infected by EBV become responsive to exogenous rIL-2- or CD2-mediated stimulation in vitro. To address whether such costimulation may be provided by thymic presenting cells and to study the cellular effects of EBV infection, the present work utilizes thymocyte cultures containing autologous thymic presenting cells. In the presence of these presenting cells, EBV induces proliferation of thymocytes. EBV infection promotes the formation of adhesions between two populations of cells in an APC responder fashion, and separation of these two populations abrogates the proliferative response to EBV. The response is mediated by IL-2 because Ab blocking of the IL-2R inhibits proliferation as does cyclosporin A. EBV promotes an expansion in the number of CD4+8+ thymocytes, and the proliferating population is vulnerable to TCR/CD3-generated signals, indicating that the responding cells are phenotypically and functionally immature. Finally, addition of exogenous IL-2 to EBV-exposed thymocytes promotes a second wave of proliferation. Phenotypic characterization of the EBV-induced, IL-2-responding cells shows them to express reduced levels of CD1 and a transitional CD4(high)8(low) phenotype. These data characterize the cellular response to EBV infection in thymocytes and may offer insight into EBV-associated T lineage malignancies and autoimmune disorders.

  15. Structural and functional characteristics of the CD3 (T3) molecular complex on human thymocytes.

    PubMed

    Tsoukas, C D; Landgraf, B; Bentin, J; Lamberti, J F; Carson, D A; Vaughan, J H

    1987-06-01

    The CD3 (T3) molecular complex is noncovalently associated with the antigen receptor molecule on T cells. The mitogenic properties of anti-CD3 antibodies have suggested that this complex may be the transducer of the antigenic signal to the intracellular environment. In the present investigation, we studied some of the structural and functional characteristics of the CD3 complex on human thymocytes. In 11 specimens tested, we found that anti-CD3 antibodies react with 50 to 76% of the thymocytes. Two-color immunofluorescence analysis revealed that the majority (greater than 50%) of thymocytes express both CD3 and CD1 on their surfaces. The latter is a marker of immature thymocytes. However, a distinct subpopulation comprising 13 to 19% of the total cells displays only CD3, while an approximately equal percentage of cells expresses only CD1. The mitogenic potential of anti-CD3 antibodies on peripheral T cells is dependent on the presence of monocytes. Anti-CD3 antibodies by themselves cannot activate thymocytes, indicating that functionally active monocytes are absent from the thymocyte population. Even the addition of peripheral monocytes does not allow a response of thymocytes to anti-CD3 antibodies. However, when the anti-CD3 antibody 64.1 is added in the presence of exogenous rIL 2, a strong antibody and lymphokine dose-dependent response ensues. Only CD1- CD3+ thymocytes are stimulated by the addition of antibody and IL 2. The mere expression of CD3 on the CD1+ CD3+ subpopulation of thymocytes apparently is not sufficient to render the cells responsive to the signals of anti-CD3 and IL 2.

  16. Internalization of the TGF-β type I receptor into caveolin-1 and EEA1 double-positive early endosomes.

    PubMed

    He, Kangmin; Yan, Xiaohua; Li, Nan; Dang, Song; Xu, Li; Zhao, Bing; Li, Zijian; Lv, Zhizhen; Fang, Xiaohong; Zhang, Youyi; Chen, Ye-Guang

    2015-06-01

    Endocytosis and intracellular sorting of transforming growth factor-β (TGF-β) receptors play an important regulatory role in TGF-β signaling. Two major endocytic pathways, clathrin- and caveolae-mediated endocytosis, have been reported to independently mediate the internalization of TGF-β receptors. In this study, we demonstrate that the clathrin- and caveolae-mediated endocytic pathways can converge during TGF-β receptor endocytic trafficking. By tracking the intracellular dynamics of fluorescently-labeled TGF-β type I receptor (TβRI), we found that after mediating TβRI internalization, certain clathrin-coated vesicles and caveolar vesicles are fused underneath the plasma membrane, forming a novel type of caveolin-1 and clathrin double-positive vesicles. Under the regulation of Rab5, the fused vesicles are targeted to early endosomes and thus deliver the internalized TβRI to the caveolin-1 and EEA1 double-positive early endosomes (caveolin-1-positive early endosomes). We further showed that the caveolin-1-positive early endosomes are positive for Smad3/SARA, Rab11 and Smad7/Smurf2, and may act as a multifunctional device for TGF-β signaling and TGF-β receptor recycling and degradation. Therefore, these findings uncover a novel scenario of endocytosis, the direct fusion of clathrin-coated and caveolae vesicles during TGF-β receptor endocytic trafficking, which leads to the formation of the multifunctional sorting device, caveolin-1-positive early endosomes, for TGF-β receptors.

  17. Internalization of the TGF-β type I receptor into caveolin-1 and EEA1 double-positive early endosomes

    PubMed Central

    He, Kangmin; Yan, Xiaohua; Li, Nan; Dang, Song; Xu, Li; Zhao, Bing; Li, Zijian; Lv, Zhizhen; Fang, Xiaohong; Zhang, Youyi; Chen, Ye-Guang

    2015-01-01

    Endocytosis and intracellular sorting of transforming growth factor-β (TGF-β) receptors play an important regulatory role in TGF-β signaling. Two major endocytic pathways, clathrin- and caveolae-mediated endocytosis, have been reported to independently mediate the internalization of TGF-β receptors. In this study, we demonstrate that the clathrin- and caveolae-mediated endocytic pathways can converge during TGF-β receptor endocytic trafficking. By tracking the intracellular dynamics of fluorescently-labeled TGF-β type I receptor (TβRI), we found that after mediating TβRI internalization, certain clathrin-coated vesicles and caveolar vesicles are fused underneath the plasma membrane, forming a novel type of caveolin-1 and clathrin double-positive vesicles. Under the regulation of Rab5, the fused vesicles are targeted to early endosomes and thus deliver the internalized TβRI to the caveolin-1 and EEA1 double-positive early endosomes (caveolin-1-positive early endosomes). We further showed that the caveolin-1-positive early endosomes are positive for Smad3/SARA, Rab11 and Smad7/Smurf2, and may act as a multifunctional device for TGF-β signaling and TGF-β receptor recycling and degradation. Therefore, these findings uncover a novel scenario of endocytosis, the direct fusion of clathrin-coated and caveolae vesicles during TGF-β receptor endocytic trafficking, which leads to the formation of the multifunctional sorting device, caveolin-1-positive early endosomes, for TGF-β receptors. PMID:25998683

  18. A Human Immunodeficiency Virus Controller With a Large Population of CD4+CD8+ Double-Positive T Cells

    PubMed Central

    Durand, Christine M.; Buckheit, Robert W.; Salgado, Maria; Pohlmeyer, Christopher W.; Walker-Sperling, Victoria E.; Hegarty, Robert W.; Ambinder, Richard F.; Blankson, Joel N.

    2015-01-01

    Human immunodeficiency virus (HIV) controllers are patients who control viral replication without antiretroviral therapy. We present the case of an HIV controller who had CD4 and CD8 coexpressed on 40% of his T cells. Although a recent study found that double-positive T cells had superior antiviral capacity in HIV-1 controllers, in this case, the CD4+CD8+ T cells did not have strong antiviral activity. PMID:26380339

  19. A Human Immunodeficiency Virus Controller With a Large Population of CD4(+)CD8(+) Double-Positive T Cells.

    PubMed

    Durand, Christine M; Buckheit, Robert W; Salgado, Maria; Pohlmeyer, Christopher W; Walker-Sperling, Victoria E; Hegarty, Robert W; Ambinder, Richard F; Blankson, Joel N

    2015-04-01

    Human immunodeficiency virus (HIV) controllers are patients who control viral replication without antiretroviral therapy. We present the case of an HIV controller who had CD4 and CD8 coexpressed on 40% of his T cells. Although a recent study found that double-positive T cells had superior antiviral capacity in HIV-1 controllers, in this case, the CD4(+)CD8(+) T cells did not have strong antiviral activity.

  20. Double-Cross Instability: An Absolute Instability Caused by Counter-Propagating Positive- and Negative-Energy Waves

    SciTech Connect

    Brizard, A.J.; Morehead, J.J.; Kaufman, A.N.; Tracy, E.R.

    1996-08-01

    The resonant interaction of a negative-energy wave with a positive-energy wave gives rise to a linear instability. Whereas a single crossing of rays in a nonuniform medium leads to a {ital convectively} saturated instability, we show that a double crossing can yield an {ital absolute} instability, if the two rays are oppositely directed. We obtain expressions for the growth rate and the threshold, and present one application. {copyright} {ital 1996 The American Physical Society.}

  1. Multiyear measurements of Position Angle and Separation of selected binary stars from the Washington Double Star Catalog

    NASA Astrophysics Data System (ADS)

    Muller, Rafael J.; Cersosimo, Juan C.; Lopez, Andy J.; Vergara, Nelson; Torres, Brian; Mendoza, Lizyan; Ortiz, Deliris; Del Valle, Yashira; Espinosa, Gabriela; Reyes, Marjory

    2016-01-01

    We present here the multiyear data sets on separation and position angle of binary stars obtained at the NURO telescope, located east of Flagstaff Arizona at an elevation of 7200 feet. The data was analyzed at the Humacao University Observatory of the University of Puerto Rico and will be submitted for publication at the Journal of Double Star Observations. We describe the methodology for the analysis of the images we obtained.

  2. [Effect of irradiation on the degradation of rat thymocyte chromatin].

    PubMed

    Tsudzevich, B O; Parkhomets', Iu P; Andriĭchuk, T R; Iurkina, V V

    1998-01-01

    Genome instability of adaptive nature is formed under the experimental influence on a cell. Under critical conditions, strategy of organism is to damage the cells that cannot be restored and controlled by including the program of apoptosis. The ordered internucleosomal DNA degradation is considered to be one of the proof attributes of immunocompetent cell apoptosis. We investigated the effects of various doses of irradiation on the thymocytes chromatine fragmentation in 1,2,3 hours after a single X-ray exposure or after chronic influence in conditions of Chernobyl research base. By the means of electrophoresis in agarose and judging by polydeoxyribonucleotides accumulation we observed the "ladder pattern" of degradation in 3 hr after single 1 Gr irradiation (the smallest dose displaying the effect). We suppose that the influence of both chronic low-intensity irradiation taking place in Chernobyl and single X-ray exposure result in intensifying of DNA fragmentation in the cells of immunocompetent organs.

  3. Stimulatory effect on rat thymocytes proliferation and antimicrobial activity of two 6-(propan-2-yl)-4-methyl-morpholine-2,5-diones.

    PubMed

    Pavlovic, Voja; Djordjevic, Aleksandra; Cherneva, Emiliya; Yancheva, Denitsa; Smelcerovic, Andrija

    2012-03-01

    Recently we reported the identification and synthesis of cyclodidepsipeptides, 3,6-di(propan-2-yl)-4-methyl-morpholine-2,5-dione (PPM) and 3-(2-methylpropyl)-6-(propan-2-yl)-4-methyl-morpholine-2,5-dione (BPM), as potential precursors of enniatin B in Fusarium sporotrichioides. No data concerning biological activity of PPM and BPM have hitherto been published. The possible immunomodulatory effect and antimicrobial activity of PPM and BPM were investigated in this study, due to well known biological activities of enniatin B. The cytotoxicity effect of PPM and BPM on rat thymocytes demonstrated that increasing concentrations (0.1, 1, 10 μg/well) of PPM and BPM to cell culture, showed no significant effect on thymocytes toxicity. Simultaneously, incubation with studied cyclodidepsipeptides did not result with decreased mitochondrial membrane potential. Further, thymocytes exposure to increasing concentration of PPM and BPM was not able to induce significant reactive oxygen species (ROS) production in rat thymocytes. PPM and BPM administrations to cell culture in concentrations of 0.1 and 1 μg/well resulted with no significant increase of proliferative activity. However, significantly increased proliferative activity was detected with 10 μg of PPM (p<0.001) and BPM (p<0.05), as compared to their respective controls. The in vitro antimicrobial activity of PPM and BPM was tested against two Gram-positive and three Gram-negative bacteria. The results indicated that MIC values against tested strains ranged between 2.00 and 25.00 mg/ml. PPM showed much better activity against all tested bacteria in comparison with BPM. PPM was equally effective against both Gram-positive and Gram-negative bacteria, at the dose of 2.00 mg/ml.

  4. Different gamma delta T-cell receptors are expressed on thymocytes at different stages of development.

    PubMed Central

    Ito, K; Bonneville, M; Takagaki, Y; Nakanishi, N; Kanagawa, O; Krecko, E G; Tonegawa, S

    1989-01-01

    We have analyzed the structural diversity of the murine gamma delta T-cell receptor (TCR) heterodimer expressed on CD4- CD8- thymocyte populations and on TCR gamma delta-expressing hybridomas derived from thymocytes of fetal, newborn, and adult mice. We found that CD4- CD8- thymocytes derived from mice of different pre- and postnatal age preferentially express a gamma delta TCR encoded by different subsets of gamma and delta gene segments. This age-dependent differential expression of gamma delta TCR on thymocytes seems to be accomplished in part by a specific control of rearranged gamma genes operating at the level of transcription and/or RNA stability. We discuss the implications of these findings with respect to the recognition roles of the gamma delta TCR. Images PMID:2463632

  5. Abnormal in vitro thymocyte differentiation in a patient with severe combined immunodeficiency-Nezelof`s syndrome

    SciTech Connect

    Knutsen, A.P.; Wall, D.; Mueller, K.R.; Bouhasin, J.D.

    1996-05-01

    An in vitro coculture model system of CD34+ stem cells and allogenic cultured thymic epithelia fragments was used to evaluate thymocyte differentiation in a 9-month-old child of Amish descent with Nezelof syndrome. Though the patient`s stem cells differentiate to acquire normal expression of CD2 and CD7, later steps of maturation were abnormal. There was detectable but reduced expression of CD3 and CD4 phenotypes. CD44+ expression, however, was markedly reduced. CD44 is an adhesion molecule, interacting with the matrix ligands hyaluronan and fibronectin, and is expressed early in thymocyte differentiation and subsequently in mature T cells. It is hypothesized that abnormal expression of CD44 in a variant of severe combined immunodeficiency, Nezelof`s syndrome, interferes with normal thymocyte and thymic epithelial interaction, which leads to abnormal thymocyte differentiation. 35 refs., 2 figs., 3 tabs.

  6. CD81 expressed on human thymocytes mediates integrin activation and interleukin 2-dependent proliferation.

    PubMed

    Todd, S C; Lipps, S G; Crisa, L; Salomon, D R; Tsoukas, C D

    1996-11-01

    Lymphocyte recognition of antigen by the antigen-specific T cell receptor (TCR) and coreceptor complexes rapidly alters the cell's adhesive properties facilitating high avidity cell-ligand interactions necessary for lymphocyte development and function. Here, we report the expression of CD81 (target of antiproliferative antigen [TAPA]-1) on human thymocytes and the physical association of CD81 with CD4 and CD8 T cell coreceptors. Antibody ligation of CD81 on thymocytes promotes the rapid induction of integrin-mediated cell-cell adhesion via lymphocyte function-associated molecule-1 (LFA-1). Cross-linking CD81 is also shown to be costimulatory with signaling through the TCR/CD3 complex inducing interleukin 2-dependent thymocyte proliferation. These data suggest that a CD81-mediated pathway in thymocytes is involved in the regulation of both cell adhesion and activation.

  7. The production of alpha/beta and gamma/delta double negative (DN) T-cells and their role in the maintenance of pregnancy.

    PubMed

    Chapman, John C; Chapman, Fae M; Michael, Sandra D

    2015-07-12

    The ability of the thymus gland to convert bone marrow-derived progenitor cells into single positive (SP) T-cells is well known. In this review we present evidence that the thymus, in addition to producing SP T-cells, also has a pathway for the production of double negative (DN) T-cells. The existence of this pathway was noted during our examination of relevant literature to determine the cause of sex steroid-induced thymocyte loss. In conducting this search our objective was to answer the question of whether thymocyte loss is the end product of a typical interaction between the reproductive and immune systems, or evidence that the two systems are incompatible. We can now report that "thymocyte loss" is a normal process that occurs during the production of DN T-cells. The DN T-cell pathway is unique in that it is mediated by thymic mast cells, and becomes functional following puberty. Sex steroids initiate the development of the pathway by binding to an estrogen receptor alpha located in the outer membrane of the mast cells, causing their activation. This results in their uptake of extracellular calcium, and the production and subsequent release of histamine and serotonin. Lymphatic vessels, located in the subcapsular region of the thymus, respond to the two vasodilators by undergoing a substantial and preferential uptake of gamma/delta and alpha/beta DN T- cells. These T- cells exit the thymus via efferent lymphatic vessels and enter the lymphatic system.The DN pathway is responsible for the production of three subsets of gamma/delta DN T-cells and one subset of alpha/beta DN T-cells. In postpubertal animals approximately 35 % of total thymocytes exit the thymus as DN T-cells, regardless of sex. In pregnant females, their levels undergo a dramatic increase. Gamma/delta DN T-cells produce cytokines that are essential for the maintenance of pregnancy.

  8. Effect of syngeneic thymocytes on proliferation of the small intestinal epithelium in mice

    SciTech Connect

    Shmakov, A.N.; Aparovich, G.G.; Trufakin, V.A.

    1986-12-01

    This paper describes the study of the action of syngeneic thymocytes on proliferation of the epithelium of the mouse small intestine. The mice were injected with /sup 3/H-thymidine in the experiments. Under the experimental conditions presented here, syngeneic thymocytes can reduce the number of DNA-synthesizing cells in the intestinal epithelium, causing narrowing of the zone of proliferation and enlargement of the zone of differentiation of the enterocytes.

  9. "Two souls, two thoughts," two self-schemas: double consciousness can have positive academic consequences for African Americans.

    PubMed

    Brannon, Tiffany N; Markus, Hazel Rose; Taylor, Valerie Jones

    2015-04-01

    African Americans can experience a double consciousness-the two-ness of being an American and an African American. The present research hypothesized that: (a) double consciousness can function as 2 self-schemas-an independent self-schema tied to mainstream American culture and an interdependent self-schema tied to African American culture, and (b) U.S. educational settings can leverage an interdependent self-schema associated with African American culture through inclusive multicultural practices to facilitate positive academic consequences. First, a pilot experiment and Studies 1 and 2 provided evidence that double consciousness can be conceptualized as 2 self-schemas. That is, African Americans shifted their behavior (e.g., cooperation) in schema-relevant ways from more independent when primed with mainstream American culture to more interdependent when primed with African American culture. Then, Studies 3 and 4 demonstrated that incorporating African American culture within a university setting enhanced African Americans' persistence and performance on academic-relevant tasks. Finally, using the Gates Millennium Scholars dataset (Cohort 1), Study 5 conceptually replicated Studies 3 and 4 and provided support for one process that underlies the observed positive academic consequences. Specifically, Study 5 provided evidence that engagement with African American culture (e.g., involvement with cultural events/groups) on college campuses makes an interdependent self-schema more salient that increases African American students' sense of academic fit and identification, and, in turn, enhances academic performance (self-reported grades) and persistence (advanced degree enrollment in a long-term follow-up). The discussion examines double consciousness as a basic psychological phenomenon and suggests the intra- and intergroup benefits of inclusive multicultural settings.

  10. Functional characterization of human thymocyte subpopulations separated by density gradient centrifugation.

    PubMed Central

    Lederman, H M; Lee, J W; Cohen, A; Gelfand, E W

    1984-01-01

    A bovine serum albumin gradient was used to separate two populations of human thymocytes--a minority population (8%) of large thymocytes (LT) and a majority population (92%) of small thymocytes (ST). Fifty per cent of LT cells were in the S, G2 or M phases of the cell cycle compared to 5% of ST cells and 15% of unfractionated thymocytes. LT cells proliferated in response to T cell mitogens and included all of the T colony precursor cells (TCPC). In contrast, ST cells proliferated with mitogens only in the presence of added T cell growth factors and contained none of the thymocyte TCPC. ST cells neither helped nor suppressed the function of LT cells in any assay. This separation technique has provided a rapid method for isolating functionally distinct thymic lymphocyte subpopulations and permitted a further definition of the TCPC in the human thymus. Furthermore it should prove useful in studies of thymocytes at different stages of the cell cycle. PMID:6607793

  11. CD1+ human thymocytes proliferate in response to superantigen staphylococcal enterotoxin B1.

    PubMed

    Todd, S C; Baccala, R; Hedrick, J A; Theofilopoulos, A N; Tsoukas, C D

    1994-09-01

    Exposure of human thymocytes to superantigens results in the deletion of thymocytes expressing specific TCR-V beta genes. The factors that contribute to this deletion may relate to the inherent nature of the T cell at a given stage of development. In this paper, we demonstrate that CD1+ human cortical thymocytes are capable of proliferating in response to a bacterial superantigen (staphylococcal enterotoxin B (SEB)) in the presence of autologous CD2-/low thymic APCs. Phenotypic analysis of the responding populations revealed that the majority of the CD1+ cells were CD4+CD8low or CD8+CD4low cells. The response is triggered by low concentrations of SEB, requires the participation of the TCR and IL-2R molecules, and is inhibited by cyclosporin A. Thymocytes that express specific V beta genes are expanded, which results in an engagement profile that parallels that found in PBLs. Additionally, four V beta-chains that have not been reported previously are shown to engage SEB. Once stimulated, the thymocytes failed to respond to additional SEB; however, they could be induced to proliferative with IL-2, which suggests that these expanded populations had become anergic. These data represent the first demonstration of a human cortical thymocyte subpopulation that responds to superantigen by proliferation and subsequent anergy.

  12. Experimental glomerulonephritis is attenuated by CD8+ T cell chimerism and prevented by Mls-1-incompatible thymocytes.

    PubMed

    Sutmuller, M; Baelde, H J; Tysma, O M; de Heer, E; Bruijn, J A

    1998-07-01

    Chronic graft-versus-host disease (GvHD) in mice is a model resembling glomerulonephritis in human systemic lupus erythematosus. In the present study congenic mouse strains were used to investigate the pathogenetic role of (1) donor T cell subset chimerism and (2) donor thymocytes in this model. In GvHD employing minor lymphocyte-stimulating-1 (Mls-1)-compatible donors and recipients, full-blown immune complex glomerulonephritis was associated with a low-donor CD8(+) T cell chimerism. Injection of lymphocytes from Mls-1-negative donors (Mls-1(b)) into Mls-1-positive recipients (Mls-1(a)) induces a type of GvHD characterized by rapid self-limitation accompanied by the immediate inhibition of donor T cell chimerism and the absence of glomerulonephritis. However, omission of thymocytes from the donor inoculate does result in glomerular depositions containing immunoglobulins. These results suggest that donor CD8(+) T cell chimerism is associated with attenuation of immune complex glomerulonephritis, whereas Mls-1-incompatible donor T cell precursors prevent the disease. Copyright 1998 Academic Press.

  13. A voltage-gated sodium channel is essential for the positive selection of CD4(+) T cells.

    PubMed

    Lo, Wan-Lin; Donermeyer, David L; Allen, Paul M

    2012-09-01

    The sustained entry of Ca(2+) into CD4(+)CD8(+) double-positive thymocytes is required for positive selection. Here we identified a voltage-gated Na(+) channel (VGSC) that was essential for positive selection of CD4(+) T cells. Pharmacological inhibition of VGSC activity inhibited the sustained Ca(2+) influx induced by positively selecting ligands and the in vitro positive selection of CD4(+) but not CD8(+) T cells. In vivo short hairpin RNA (shRNA)-mediated knockdown of the gene encoding a regulatory β-subunit of a VGSC specifically inhibited the positive selection of CD4(+) T cells. Ectopic expression of VGSC in peripheral AND CD4(+) T cells bestowed the ability to respond to a positively selecting ligand, which directly demonstrated that VGSC expression was responsible for the enhanced sensitivity. Thus, active VGSCs in thymocytes provide a mechanism by which a weak positive selection signal can induce the sustained Ca(2+) signals required for CD4(+) T cell development.

  14. Vibratory response of a precision double-multi-layer monochromator positioning system using a generic modeling program with experimental verification.

    SciTech Connect

    Barraza, J.

    1998-07-29

    A generic vibratory response-modeling program has been developed as a tool for designing high-precision optical positioning systems. The systems are modeled as rigid-body structures connected by linear non-rigid elements such as complex actuators and bearings. The full dynamic properties of each non-rigid element are determined experimentally or theoretically, then integrated into the program as inertial and stiffness matrices. Thus, it is possible to have a suite of standardize structural elements for modeling many different positioning systems that use standardized components. This paper will present the application of this program to a double-multi-layer monochromator positioning system that utilizes standardized components. Calculated results are compared to experimental modal analysis results.

  15. Chicken thymocyte-specific antigens identified by monoclonal antibodies: characterization and distribution in normal tissues and in tumoral tissues from Marek's disease chicken.

    PubMed

    Mazzella, O; Cauchy, L; Coudert, F; Richard, J

    1986-01-01

    Monoclonal antibodies (MAbs) were obtained against purified thymocyte membrane extracts. Five MAbs TA3, TB1, TB6 (IgG1), TC4, and TA1 (IgG2a), were tested by immunofluorescence and by immunoperoxidase tests against normal cells from different organs, Marek's disease (MD) cell lines, and MD tumoral cells from chickens. Three of them, TA3, TB1, and TB6, reacted exclusively with lymphoid cells in both cortical and medullary areas of the thymus and with less than 8% bursa cells. They identified a protein of apparently 40 kD. The other two revealed antigenic determinants on most medullar thymocytes and some cortical thymocytes, and on some splenic and peripheral blood lymphocytes. They were positive with MD cell lines and cells deriving from MD tumors. TC4 and TA1 detected molecular masses of about 110 kD and 16 kD, respectively. No MAbs reacted with erythrocytes, bone marrow, liver, brain, and skin cells. Not all of the tested cells were stained after contact with an anti-chicken immunoglobulin serum. In this paper, we determine a specific antigen restricted to T cells from thymus and different markers belonging to the mature T cells. The latter are also present on MD cell lines and MD tumoral cells.

  16. A Role for the Tec Family Tyrosine Kinase Txk in T Cell Activation and Thymocyte Selection

    PubMed Central

    Sommers, Connie L.; Rabin, Ronald L.; Grinberg, Alexander; Tsay, Henry C.; Farber, Joshua; Love, Paul E.

    1999-01-01

    Recent data indicate that several members of the Tec family of protein tyrosine kinases function in antigen receptor signal transduction. Txk, a Tec family protein tyrosine kinase, is expressed in both immature and mature T cells and in mast cells. By overexpressing Txk in T cells throughout development, we found that Txk specifically augments the phospholipase C (PLC)-γ1–mediated calcium signal transduction pathway upon T cell antigen receptor (TCR) engagement. Although Txk is structurally different from inducible T cell kinase (Itk), another Tec family member expressed in T cells, expression of the Txk transgene could partially rescue defects in positive selection and signaling in itk−/− mice. Conversely, in the itk+/+ (wild-type) background, overexpression of Txk inhibited positive selection of TCR transgenic thymocytes, presumably due to induction of cell death. These results identify a role for Txk in TCR signal transduction, T cell development, and selection and suggest that the Tec family kinases Itk and Txk perform analogous functions. PMID:10562318

  17. Menatetrenone versus alfacalcidol in the treatment of Chinese postmenopausal women with osteoporosis: a multicenter, randomized, double-blinded, double-dummy, positive drug-controlled clinical trial

    PubMed Central

    Jiang, Yan; Zhang, Zhen-Lin; Zhang, Zhong-Lan; Zhu, Han-Min; Wu, Yi-Yong; Cheng, Qun; Wu, Feng-Li; Xing, Xiao-Ping; Liu, Jian-Li; Yu, Wei; Meng, Xun-Wu

    2014-01-01

    Objective To evaluate whether the efficacy and safety of menatetrenone for the treatment of osteoporosis is noninferior to alfacalcidol in Chinese postmenopausal women. Method This multicenter, randomized, double-blinded, double-dummy, noninferiority, positive drug-controlled clinical trial was conducted in five Chinese sites. Eligible Chinese women with postmenopausal osteoporosis (N=236) were randomized to Group M or Group A and received menatetrenone 45 mg/day or alfacalcidol 0.5 μg/day, respectively, for 1 year. Additionally, all patients received calcium 500 mg/day. Posttreatment bone mineral density (BMD), new fracture onsets, and serum osteocalcin (OC) and undercarboxylated OC (ucOC) levels were compared with the baseline value in patients of both groups. Results A total of 213 patients (90.3%) completed the study. After 1 year of treatment, BMD among patients in Group M significantly increased from baseline by 1.2% and 2.7% at the lumbar spine and trochanter, respectively (P<0.001); and the percentage increase of BMD in Group A was 2.2% and 1.8%, respectively (P<0.001). No difference was observed between groups. There were no changes in femoral neck BMD in both groups. Two patients (1.9%, 2/108) in Group M and four patients (3.8%, 4/105) in Group A had new fracture onsets (P>0.05). In Group M, OC and ucOC decreased from baseline by 38.7% and 82.3%, respectively (P<0.001). In Group A, OC and ucOC decreased by 25.8% and 34.8%, respectively (P<0.001). Decreases in serum OC and ucOC were more obvious in Group M than in Group A (P<0.001). The safety profile of menatetrenone was similar to alfacalcidol. Conclusion Menatetrenone is an effective and safe choice in the treatment of postmenopausal osteoporosis in Chinese women. PMID:24426779

  18. Embryonic age influences the capacity for cytokine induction in chicken thymocytes

    PubMed Central

    Peters, Michelle A; Browning, Glenn F; Washington, Elizabeth A; Crabb, Brendan S; Kaiser, Pete

    2003-01-01

    Thymocyte responses to functional activation are of relevance to the evaluation of the efficacy of in ovo immunotherapies and vaccines in chickens. In this study we have demonstrated differences in chicken thymocyte responses according to developmental age. RNA samples from stimulated and unstimulated chicken thymocytes were assayed for messenger RNA encoding the cytokines interleukin-1β (IL-1β), IL-2, interferon-α (IFN-α), IFN-β, IFN-γ and transforming growth factor-β4 (TGF-β4), and also components of the major histocompatibility complex (MHC), β2-microglobulin (β2M) and the MHC class I α-chain (MHC IA). At embryonic day 14 thymocytes were least responsive to functional activation and differences existed even between thymocyte populations at embryonic day 18 and day 1 post-hatch. The duration of proliferation in response to stimulation was found to increase with increasing embryonic age. Mitogen stimulation of embryonic day 18 and day 1 post-hatch thymocytes induced up-regulation of IFN-γ, IL-1β and TGF-β transcripts, and down-regulation of IFN-α, IFN-β and IL-2 transcripts, with a higher induction of IFN-γ, IL-1β and TGF-β transcripts in more immature T-cell-receptor-negative (TCR−) than TCR+ (TCR1+, TCR2+, or TCR3+) subsets. In contrast, in the mouse and human, both mature and immature thymocytes respond to mitogen stimulation with up-regulation of IL-2. Thymocytes from embryonic day 14 chicks responded to mitogen with a short burst of unsustained proliferation, and transcriptional down-regulation of the cytokines IL-2, IL-1β, IFN-α, IFN-β and IFN-γ. These results suggest that embryonic day 14 thymocytes are largely unresponsive to mitogen. Transcripts encoding TGF-β and type I interferons (IFN-α and IFN-β) were constitutively expressed at high levels in very early thymocytes at embryonic day 14. Thymocytes at embryonic days 14 and 18 and day 1 post-hatch responded to mitogen stimulation with up-regulation of MHC IA transcript. The

  19. Chromosome position determines the success of double-strand break repair.

    PubMed

    Lee, Cheng-Sheng; Wang, Ruoxi W; Chang, Hsiao-Han; Capurso, Daniel; Segal, Mark R; Haber, James E

    2016-01-12

    Repair of a chromosomal double-strand break (DSB) by gene conversion depends on the ability of the broken ends to encounter a donor sequence. To understand how chromosomal location of a target sequence affects DSB repair, we took advantage of genome-wide Hi-C analysis of yeast chromosomes to create a series of strains in which an induced site-specific DSB in budding yeast is repaired by a 2-kb donor sequence inserted at different locations. The efficiency of repair, measured by cell viability or competition between each donor and a reference site, showed a strong correlation (r = 0.85 and 0.79) with the contact frequencies of each donor with the DSB repair site. Repair efficiency depends on the distance between donor and recipient rather than any intrinsic limitation of a particular donor site. These results further demonstrate that the search for homology is the rate-limiting step in DSB repair and suggest that cells often fail to repair a DSB because they cannot locate a donor before other, apparently lethal, processes arise. The repair efficiency of a donor locus can be improved by four factors: slower 5' to 3' resection of the DSB ends, increased abundance of replication protein factor A (RPA), longer shared homology, or presence of a recombination enhancer element adjacent to a donor.

  20. Quantum dots-based double-color imaging of HER2 positive breast cancer invasion

    SciTech Connect

    Liu, Xiu-Li; Peng, Chun-Wei; Chen, Chuang; Yang, Xue-Qin; Hu, Ming-Bai; Xia, He-Shun; Liu, Shao-Ping; and others

    2011-06-10

    Highlights: {yields} HER2 level is closely related to the biologic behaviors of breast cancer cells. {yields} A new method to simultaneously image HER2 and type IV collagen was established. {yields} HER2 status and type IV collagen degradation predict breast cancer invasion. {yields} The complex interactions between tumor and its environment were revealed. -- Abstract: It has been well recognized that human epidermal growth factor receptor 2 (HER2) level in breast cancer (BC) is closely related to the malignant biologic behaviors of the tumor, including invasion and metastasis. Yet, there has been a lack of directly observable evidence to support such notion. Here we report a quantum dots (QDs)-based double-color imaging technique to simultaneously show the HER2 level on BC cells and the type IV collagen in the tumor matrix. In benign breast tumor, the type IV collagen was intact. With the increasing of HER2 expression level, there has been a progressive decrease in type IV collagen around the cancer nest. At HER2 (3+) expression level, there has virtually been a total destruction of type IV collagen. Moreover, HER2 (3+) BC cells also show direct invasion into the blood vessels. This novel imaging method provides direct observable evidence to support the theory that the HER2 expression level is directly related to BC invasion.

  1. The important role of positioning in nasogastric tube insertion in unconscious patients: A prospective, randomized, double-blind study.

    PubMed

    Zhao, Wangmiao; Ge, Chunyan; Zhang, Wanzeng; Sun, Zhaosheng; Li, Xiaowei

    2017-05-25

    To investigate whether positioning the body in a lateral decubitus position will facilitate nasogastric tube insertion in unconscious patients. Inserting a nasogastric tube into unconscious patients can be challenging because these patients cannot cooperate with the operator. The piriform sinus and arytenoid cartilage are the most commonly reported impaction sites. However, we found that the first impaction site was the backward displaced tongue when inserting a nasogastric tube in unconscious patients who often exhibited glossoptosis. Performing an intubation in the lateral decubitus position could make this procedure easy. This prospective, double-blind, parallel, randomized controlled trial was conducted in Hengshui City, Hebei Province, China. 110 cases of unconscious patients were enrolled. The patients were randomly assigned to a conventional group (group C) or a lateral decubitus position group (group L). In group C, the nasogastric tube was inserted while the patients were in a supine position using the conventional technique, and in group L, the tube was inserted in a lateral decubitus position or further tilting the body to a prone decubitus position 20-30 degrees. We discussed reasonable intubation methods in unconscious patients by comparing the success rate on first insertion, the overall success rate, the intubation time and the complication rates between the two groups. Group L had a higher success rate on first insertion and overall success rate than group C (P<0.05). The intubation time in group L was shorter than that in group C (P<0.001), and the complication rate in group L was lower than that in group C (P<0.05). The differences were statistically significant. The backward displaced tongue blocks the pharyngeal passage. Nasogastric tube insertions in the lateral decubitus position are recommended in unconscious patients because of the higher success rate, reduced intubation time and lower complication rate. This article is protected by

  2. Accuracy of relative positioning by interferometry with GPS Double-blind test results

    NASA Technical Reports Server (NTRS)

    Counselman, C. C., III; Gourevitch, S. A.; Herring, T. A.; King, B. W.; Shapiro, I. I.; Cappallo, R. J.; Rogers, A. E. E.; Whitney, A. R.; Greenspan, R. L.; Snyder, R. E.

    1983-01-01

    MITES (Miniature Interferometer Terminals for Earth Surveying) observations conducted on December 17 and 29, 1980, are analyzed. It is noted that the time span of the observations used on each day was 78 minutes, during which five satellites were always above 20 deg elevation. The observations are analyzed to determine the intersite position vectors by means of the algorithm described by Couselman and Gourevitch (1981). The average of the MITES results from the two days is presented. The rms differences between the two determinations of the components of the three vectors, which were about 65, 92, and 124 m long, were 8 mm for the north, 3 mm for the east, and 6 mm for the vertical. It is concluded that, at least for short distances, relative positioning by interferometry with GPS can be done reliably with subcentimeter accuracy.

  3. Nonylphenol induces thymocyte apoptosis through Fas/FasL pathway by mimicking estrogen in vivo.

    PubMed

    Yao, Genhong; Hou, Yayi

    2004-05-01

    Nonylphenol (NP) is the final biodegradation product of nonylphenol polyethoxylates, which are widely used surfactants in domestic and industrial products. Nonylphenol has been reported to have estrogenic activity and shown to have potential reproductive toxicity. However, its influence on immune system function remains unclear. In this study, we investigated the effects of nonylphenol on apoptosis and Fas/FasL gene expression in rat thymus. Nonylphenol were given orally by gavages at 125, 250, and 375mg/kg per day. Negative and positive controls were treated with the vehicle and E(2) 10ng/kg per day, respectively. Atrophy of thymus was determined by in situ morphological examination using hematoxylin and eosin staining. Apoptotic cells were identified by terminal deoxynucleotide transferase-mediated deoxy-UTP nick end labeling (TUNEL) assay. A semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) method was used to analyze Fas and FasL mRNA levels. The results showed that both nonylphenol and E(2) increased the rates of apoptotic death; reduced the expression of Fas; enhanced the expression of FasL. These findings demonstrated that nonylphenol with estrogen-like activity might affect the regulation of the immune function through thymocyte apoptosis. This apoptosis was mediated by altering the expression of Fas and FasL mRNA.

  4. Human natural killer cell committed thymocytes and their relation to the T cell lineage

    PubMed Central

    1993-01-01

    Recent studies have demonstrated that mature natural killer (NK) cells can be grown from human triple negative (TN; CD3-, CD4-, CD8-) thymocytes, suggesting that a common NK/T cell precursor exists within the thymus that can give rise to both NK cells and T cells under appropriate conditions. In the present study, we have investigated human fetal and postnatal thymus to determine whether NK cells and their precursors exist within this tissue and whether NK cells can be distinguished from T cell progenitors. Based on the surface expression of CD56 (an NK cell-associated antigen) and CD5 (a T cell-associated antigen), three phenotypically distinctive populations of TN thymocytes were identified. CD56+, CD5-; CD56-, CD5-, and CD56-, CD5+. The CD56+, CD5- population of TN thymocytes, although displaying a low cytolytic function against NK sensitive tumor cell targets, were similar in antigenic phenotype to fetal liver NK cells, gave rise to NK cell clones, and were unable to generate T cells in mouse fetal thymic organ cultures (mFTOC). This population of thymocytes represents a relatively mature population of lineage-committed NK cells. The CD56-, CD5- population of TN thymocytes were similar to thymic NK cells in antigenic phenotype and NK cell clonogenic potential. Clones derived from this population of TN thymocytes acquired CD56 surface expression and NK cell cytolytic function. CD56-, CD5- TN thymocytes thus contain a novel population of NK cell-committed precursors. The CD56-, CD5- population of TN thymocytes also contains a small percentage of CD34+ cells, which demonstrate no in vitro clonogenic potential, but possess T cell reconstituting capabilities in mFTOC. The majority of TN thymocytes do not express CD56, but coexpress CD34 and CD5. These CD56- , CD5+, CD34+ cells demonstrate no NK or T cell clonogenic potential, but are extremely efficient in repopulating mFTOC and differentiating into CD3+, CD4+, CD8+ T cells. The results of this investigation have

  5. High-intensity positive beams extracted from a compact double-chamber ion source

    SciTech Connect

    Huck, H.; Somacal, H.; Di Gregorio, D.E.; Fernandez Niello, J.O.; Igarzabal, M.; Di Paolo, H.; Reinoso, M.

    2005-06-15

    This work presents the design and development of a simple ion source, the associated ion extraction optics, and the beam transport of a low-energy and high-current proton accelerator. In its actual version, the ion source can deliver positive proton currents up to 100 mA. This rather high beam current is achieved by adding a small ionization chamber between the discharge chamber containing the filament and the extraction electrode of the ion source. Different parameters of the ion source and the injection beam line are evaluated by means of computer simulations to optimize the beam production and transmission.

  6. Undergraduate Observations of Separation and Position Angle of Double Stars ARY 6 AD and ARY 6 AE at Manzanita Observatory

    NASA Astrophysics Data System (ADS)

    Hoffert, Michael J.; Weise, Eric; Clow, Jenna; Hirzel, Jacquelyn; Leeder, Brett; Molyneux, Scott; Scutti, Nicholas; Spartalis, Sarah; Tokuhara, Corey

    2014-05-01

    Six beginning astronomy students, part of an undergraduate stellar astronomy course, one advanced undergraduate student assistant, and a professor measured the position angles and separations of Washington Double Stars (WDS) 05460 + 2119 (also known as ARY 6 AD and ARY 6 AE). The measurements were made at the Manzanita Observatory (116° 20'42" W, 32° 44' 5" N) of the Tierra Astronomical Institute on 10 Blackwood Rd. in Boulevard, California (www.youtube.com/watch?v=BHVdcMGBGDU), at an elevation of 4,500 ft. A Celestron 11" HD Edge telescope was used to measure the position angles and separations of ARY 6 AD and ARY 6 AE. The averages of our measurements are as follows: separation AD: trial 1 124.1 arcseconds and trial 2 124.5 arcseconds. The average of separation for AE: trial 1 73.3 arcseconds and trial 2 73.8 arcseconds. The averages of position angle for AD: trial 1 159.9 degrees and trial 2 161.3 degrees. The averages of position angle for AE: trial 1 232.6 degrees and trial 2 233.7 degrees.

  7. Interleukin 2 promotes growth and cytolytic activity in human T3+4-8- thymocytes.

    PubMed Central

    de la Hera, A; Toribio, M L; Marquez, C; Martinez, C

    1985-01-01

    Human thymocytes bearing T3 but neither T4 nor T8 antigens (T3+4-8- cells) were obtained after negative selection of thymocytes, either fresh or cultured in medium containing recombinant interleukin 2 (IL-2), by treatment with Na1/34, OKT4A and B9.4 monoclonal antibodies (which recognize T6, T4, and T8 antigens, respectively) and complement. Quantitative flow cytometry showed a 98% pure population of T3+4-8- lymphocytes, which included proliferating cells. The growth and maturation requirements of these thymocytes were characterized and related to the T3-receptor complex and IL-2 pathways, thought to be used by mature lymphocytes. The results show that addition of recombinant IL-2 promotes, in a dose-dependent way, proliferation and acquisition of effector functions by cultured T3+4-8- thymocytes, the growth being inhibitable by monoclonal antibody 33B73 (anti-Tac). Furthermore, cytolytic activity of T3+4-8- cells induced by recombinant IL-2 is specifically blocked by monoclonal antibody OKT3, showing that it operates via the T3-receptor complex and does not require either T4 or T8 molecules. The finding of in vitro responsiveness to recombinant IL-2 in T3+4-8- thymocytes suggests a role of IL-2 in the growth and maturation of cells committed to the T-cell lineage, during intrathymic differentiation, prior to expression of T4 and T8 molecules. PMID:3929254

  8. Curcumin attenuates Mancozeb-induced toxicity in rat thymocytes through mitochondrial survival pathway.

    PubMed

    Pavlovic, Voja; Cekic, Snezana; Ciric, Milan; Krtinic, Dane; Jovanovic, Jelena

    2016-02-01

    The widely used fungicide Mancozeb (Man) has been shown to cause genotoxic effects in rodents and toxicological manifestations in different cells, mainly by altering the antioxidant defense in cells. On the other hand, curcumin (Cur), a natural phenolic compound, is thought to possess anti-inflammatory and antioxidant properties. Here, we investigated the possible protective role of Cur on Man-induced toxicity in rat thymocytes and potential mechanism involved. Rat thymocytes were treated with Man(0.01 μg/ml) and/or increasing Cur(0.3, 1, 3 μM) concentrations and levels of cell viability, apoptosis, mitochondrial membrane potential (MMP),Bcl-2, Bax protein expression, caspase-3 and -9 activity and p38 MAPK signaling involvement were examined. Cells treated with Man displayed increased cell toxicity, hypodiploid cells, caspase-3 and -9 activity, Bax protein expression, followed with decreased MMP and Bcl-2 protein expression. Inhibition of p38 MAPK signaling pathway markedly reduced apoptosis rate and caspase-3 activity in thymocytes exposed to Man. Application of increasing Cur (1, 3 μM) concentrations resulted with significantly reduced cytotoxicity, apoptosis, caspase-3, -9 activity, Bax protein expression, together with increased MMP and Bcl-2 protein expression in rat thymocytes. These result suggest that certain Cur concentrations may mediate Man-induced rat thymocytes toxicity through mitochondrial survival pathway, which may be useful in preventing possible secondary immunological consequences induced by Man.

  9. Differentiation of human mature thymocytes: existence of a T3+4-8- intermediate stage.

    PubMed

    De la Hera, A; Toribio, M L; Marquez, C; Marcos, M A; Cabrero, E; Martinez-A, C

    1986-06-01

    A T3 complex-bearing subpopulation was characterized within an in vivo cycling T4-8- early thymocyte compartment which contains cells constitutively expressing interleukin 2 and transferrin receptors. We show differentiation in vitro of both mature subsets of thymocytes (T3+4+8- and T3+4-8+) from the above T4-8- compartment, their appearance being preceded by cells in a T3+4-8- intermediate stage. Furthermore, those mature thymocytes generated in vitro contain functionally competent cells which use T3, T4 and T8 structures for their cytolytic activity. The finding of T3+4-8- thymocytes in vivo, together with the observation that T3 antigen expression precedes that of T4 or T8 molecules in vitro, shows that T3 (and presumably Ti) is present early in ontogeny, and suggests that T3+4-8- cells constitute an "intermediate" stage relevant to the connection between early precursors and mature thymocytes during T lymphocyte ontogeny.

  10. Tnfaip8 is an essential gene for the regulation of glucocorticoid-mediated apoptosis of thymocytes.

    PubMed

    Woodward, M J; de Boer, J; Heidorn, S; Hubank, M; Kioussis, D; Williams, O; Brady, H J M

    2010-02-01

    Glucocorticoids have significant immunoregulatory actions on thymocytes and T cells and act by binding and activating cytosolic glucocorticoid receptors, which translocate to the nucleus and control gene expression through binding to specific response elements in target genes. Glucocorticoids promote cell death by activating an apoptotic program that requires transcriptional regulation. We set out to identify genes that are crucial to the process of glucocorticoid-mediated thymocyte apoptosis. Freshly isolated murine primary thymocytes were treated with dexamethasone, mRNA isolated and used to screen DNA microarrays. A set of candidate genes with upregulated expression was identified and selected members assayed in reconstituted fetal thymic organ culture (FTOC). Fetal liver-derived hematopoietic progenitor cells (HPCs) were infected with retroviruses expressing individual genes then used to repopulate depleted fetal thymic lobes. Reconstituted FTOCs expressing the gene Tnfaip8 were treated with dexamethasone and shown to be greatly sensitized to dexamethasone. Retrovirus-mediated RNA interference was applied to knock down Tnfaip8 expression in HPCs and these were used to reconstitute FTOCs. We observed that downregulating the expression of Tnfaip8 alone was sufficient to effectively protect thymocytes against glucocorticoid-induced apoptosis. We propose that Tnfaip8 is crucial in regulating glucocorticoid-mediated apoptosis of thymocytes.

  11. Osteosynthesis with Rush's double nail by the "Eiffel Tower" method in pseudarthrosis impacted in good position and retarded union.

    PubMed

    Zinghi, G F; Lanfranchi, R

    1980-04-01

    The "Eiffel Tower" method of nailing has not attracted the interest in Italy that it deserves, both because its utilisaton in fractures is difficult when closed reduction of the fragments is difficult, and because the attention of surgeons has been progressively directed towards osteosynthesis by open reduction. Our experience over many years, however, has convinced us that, in the diaphysis, only the intramedullary nail can provide the quick recovery that is not always forthcoming in the case of plating. We need only think of comminuted fractures, where the possible necrosis of one or more fragments demands a certain amount of prudence in allowing direct weight bearing. Therefore, in adopting the double Rush system, we extended its application to the intramedullary osteosynthesis of metaphyseal fractures. It was a short step from this to the surgical treatment of pseudarthrosis impacted in good position or retarded union, the results of which were encouraging, as demonstrated by the eighty-one cases reported in this paper.

  12. Shaped electric fields for fast optimal manipulation of electron spin and position in a double quantum dot

    NASA Astrophysics Data System (ADS)

    Budagosky, J. A.; Khomitsky, D. V.; Sherman, E. Ya.; Castro, Alberto

    2016-01-01

    We use quantum optimal control theory algorithms to design external electric fields that drive the coupled spin and orbital dynamics of an electron in a double quantum dot, subject to the spin-orbit coupling and Zeeman magnetic fields. We obtain time profiles of multifrequency electric field pulses which increase the rate of spin-flip transitions by several orders of magnitude in comparison with monochromatic fields, where the spin Rabi oscillations were predicted to be very slow. This precise (with fidelity higher than 1 ×10-4 ) and fast (at the time scale of the order of 0.1 ns, comparable with the Zeeman spin rotation and the interdot tunneling time) simultaneous control of the spin and position is achieved while keeping the electron in the four lowest tunneling- and Zeeman-split levels through the duration of the pulse. The proposed algorithms suggest effective applications in spintronics and quantum information devices.

  13. Hsp90 N- and C-terminal double inhibition synergistically suppresses Bcr-Abl-positive human leukemia cells.

    PubMed

    Chen, Chun; Zhuang, Yingting; Chen, Xianling; Chen, Xiaole; Li, Ding; Fan, Yingjuan; Xu, Jianhua; Chen, Yuanzhong; Wu, Lixian

    2017-02-07

    Heat shock protein 90 (Hsp90) contains amino (N)-terminal domain, carboxyl(C)-terminal domain, and middle domains, which activate Hsp90 chaperone function cooperatively in tumor cells. One terminal occupancy might influence another terminal binding with inhibitor. The Bcr-Abl kinase is one of the Hsp90 clients implicated in the pathogenesis of chronic myeloid leukemia (CML). Present studies demonstrate that double inhibition of the N- and C-terminal termini can disrupt Hsp90 chaperone function synergistically, but not antagonistically, in Bcr-Abl-positive human leukemia cells. Furthermore, both the N-terminal inhibitor 17-AAG and the C-terminal inhibitor cisplatin (CP) have the capacity to suppress progenitor cells; however, only CP is able to inhibit leukemia stem cells (LSCs) significantly, which implies that the combinational treatment is able to suppress human leukemia in different mature states.

  14. Schlafen, a new family of growth regulatory genes that affect thymocyte development.

    PubMed

    Schwarz, D A; Katayama, C D; Hedrick, S M

    1998-11-01

    The Schlafen (Slfn) family of genes are differentially regulated during thymocyte maturation and are preferentially expressed in the lymphoid tissues. Ectopic expression of the prototype member Slfn1 early in the T lineage profoundly alters cell growth and development. In these mice, the DP thymocytes fail to complete maturation, and, depending on the transgene dosage, the number of thymocytes is reduced to 1%-30% of normal. Furthermore, expression of the Schlafen family members in fibroblasts and thymoma cells either retards or ablates cell growth. The conceptual protein sequences deduced for each of the family members have no similarity to characterized proteins and must therefore participate in a heretofore unknown regulatory mechanism guiding both cell growth and T cell development.

  15. Involvement of histone phosphorylation in thymocyte apoptosis by protein phosphatase inhibitors.

    PubMed

    Lee, E; Nakatsuma, A; Hiraoka, R; Ishikawa, E; Enomoto, R; Yamauchi, A

    1999-07-01

    Incubation of rat thymocytes with the inhibitors of protein phosphatase such as calyculin A and okadaic acid resulted in an increase in DNA fragmentation. These effects were dependent on the concentration of the inhibitors and the incubation time. Analyses of the fragmented DNA revealed the production of approximately 50 kbp of DNA and a 180 bp DNA ladder. In addition, a laser scanning-microscopic analysis showed that these compounds caused nuclear condensation. Thus, these results demonstrated that protein phosphatase inhibitors induced thymocyte apoptosis. The inhibitors of protein phosphatase increased the phosphorylation of proteins of approximately 15 kDa. The phosphorylation of proteins preceded the DNA fragmentation induced by these inhibitors. Judging from acetic acid-urea-Triton X-100 gel electrophoresis, the phosphorylated proteins were histone H1 and H2A/H3. Therefore, these results suggest that phosphorylation of histones triggers the DNA fragmentation of thymocytes undergoing apoptosis.

  16. Domains of the TCR beta-chain required for early thymocyte development

    PubMed Central

    1996-01-01

    The T cell receptor beta (TCR beta) chain controls the developmental transition from CD4-CD8- to CD4+8+thymocytes. We show that the extracellular constant region and the transmembrane region, but not the variable domain or cytoplasmic tail of the TCR beta chain are required for this differentiation step. TCR beta mutant chains lacking the cytoplasmic tail can be found at the cell surface both in functional TCR/CD3 complexes and in a GPI-anchored monomeric form indicating that the cytoplasmic tail of the TCR beta chain functions as an ER retention signal. The concordance between cell surface expression of the mutant chains as TCR/CD3 complexes and their capacity to mediate thymocyte differentiation supports the CD3 mediated feedback model in which preTCR/CD3 complexes control the developmental transition from CD4-CD8- to CD4+CD8+thymocytes. PMID:8920871

  17. Recessive mutations in a common pathway block thymocyte apoptosis induced by multiple signals

    PubMed Central

    1994-01-01

    The glucocorticoid receptor (GR) is a ligand-regulated transcription factor that controls genes necessary to initiate glucocorticoid-induced thymocyte apoptosis. We have performed a genetic analysis of thymocyte cell death by isolating and characterizing a panel of GR+ dexamethasone- resistant mutants of the murine WEHI7.2 thymocyte cell line. These apoptosis-defective (Apt-) mutants were used to identify previously unknown early steps in the apoptotic pathway. The Apt- mutants contain nonglucocorticoid receptor, recessive mutations in genes that represent multiple complementation groups. These mutations block apoptosis induced by dexamethasone, gamma irradiation, and c-AMP treatment before the point where Bcl-2 exerts its protective effect. We propose that different signals share a common apoptotic pathway, and that the induction of apoptosis involves multiple precommitment steps that can be blocked by recessive mutations. PMID:7798323

  18. Celiac Disease-Related Inflammation Is Marked by Reduction of Nkp44/Nkp46-Double Positive Natural Killer Cells

    PubMed Central

    Marafini, Irene; Monteleone, Ivan; Di Fusco, Davide; Sedda, Silvia; Cupi, Maria Laura; Fina, Daniele; Paoluzi, Alessandro Omero; Pallone, Francesco; Monteleone, Giovanni

    2016-01-01

    Introduction and Aim Natural killer (NK) cells are a first line of defence against viruses and down-regulation of NK cell cytotoxic receptors represents one of the strategies by which viruses escape the host’s immune system. Since onset of celiac disease (CD), a gluten-driven enteropathy, has been associated with viral infections, we examined whether CD-associated inflammation is characterized by abnormal distribution of NK cell receptors involved in recognition of viral-infected cells. Materials and Methods Intraepithelial mononuclear cells, isolated from duodenal biopsies of active and inactive CD patients and healthy controls (CTR) and jejunal specimens of obese subjects undergoing gastro-intestinal bypass, were analysed for NK cell markers by flow-cytometry. Expression of granzyme B, interleukin (IL)-22 and tumor necrosis factor (TNF)-α was as assessed in freshly isolated and toll-like receptor (TLR) ligand-stimulated cells. Results The percentages of total NK cells and NKT cells did not significantly differ between CD patients and CTR. In active CD, the fractions of NKp30+ NK cells, NKG2D+ NK cells and NKG2D+ NKT cells were significantly increased as compared to inactive CD patients and CTR. In contrast, CD-associated inflammation was marked by diminished presence of NKG2A+ NK cells and NKG2A+ NKT cells. The fractions of NK cells and NKT cells expressing either NKp44 or NKp46 did not differ between CD and controls, but in CD less NK cells and NKT cells co-expressed these receptors. NKp44/NKp46-double positive cells produced granzyme B and IL-22 but not TNF-α and responded to TLR ligands with enhanced expression of granzyme B. Conclusions These data indicate that active phase of CD associates with reduced presence of NKp44/NKp46-double positive NK cells and NKT cells in the epithelial compartment. PMID:27171408

  19. High levels of functional endopeptidase 24.11 (CD10) activity on human thymocytes: preferential expression on immature subsets.

    PubMed Central

    Mari, B; Breittmayer, J P; Guerin, S; Belhacene, N; Peyron, J F; Deckert, M; Rossi, B; Auberger, P

    1994-01-01

    Although it is now well established that cells of the immune system express most of the exopeptidases described so far, little information is available concerning the identification and the characterization of the peptidases associated with the surface of human thymocytes. In the present study we have focused on CD10 expression on thymocytes using both FACS and enzymatic analysis. Unfractionated intact human thymocytes were shown to express significant levels of CD10-specific enzymatic activity, as assessed by the hydrolysis of the neutral endopeptidase (NEP) substrate Suc-Ala-Ala-Phe-pNA and of D-Ala2-Leu-enkephalin, a typical NEP substrate. CD10 activity was abolished by specific NEP inhibitors, including thiorphan, retrothiorphan and phosphoramidon. Moreover, high performance liquid chromatography (HPLC) analysis showed that intact thymocytes and purified NEP hydrolysed thymopentin, a thymic factor known to induce the maturation of prothymocytes into thymocytes. Finally, CD 10/NEP was preferentially associated with CD3- CD3low and immature CD4- CD8- thymocytes. The data demonstrate for the first time that human thymocytes express functional NEP and suggest a role for this enzyme in the maturation of human thymocytes. PMID:7959879

  20. Balancing Thymocyte Adhesion and Motility: A Functional Linkage Between β1 Lntegrins and The Motility Receptor RHAMM

    PubMed Central

    Gares, Sheryl L.

    2000-01-01

    Thymocyte differentiation involves several processes that occur in different anatomic sites within the thymus. Therefore, thymocytes must have the ability to respond to signals received from stromal cells and adopt either adhesive or motile behavior. We will discuss our data indicating human thymocytes use α4β1 integrin, α5β1 integrin and RHAMM to mediate these activities. Immature multinegative (MN; CD3–4–8–19-) thymocytes use α4β1 and α5β1 integrins to mediate weak and strong adhesion. This subset also uses α4β1 integrin to mediate motility. As thymocytes differentiate, they begin to express and use RHAMM to mediate motility in conjunction with α4β1 and α5β1 integrins. Motile thymocytes use β1 integrins to maintain weakly adhesive contacts with substrate to provide traction for locomoting cells, thus weak adhesion is a requirement of motile behavior. Hyaluronan (HA) is also required by thymocytes to mediate motility. HA binding to cell surface RHAMM redistributes intracellular RHAMM to the cell surface where it functions to mediate motility. We propose that the decision to maintain adhesive or motile behavior is based on the balance between low and high avidity binding conformations of β1 integrins on thymocytes and that RHAMM:HA interactions decrease high avidity binding conformations of integrins pushing the balance toward motile behavior. PMID:11097213

  1. Expression of CR2/EBV receptors on human thymocytes detected by monoclonal antibodies.

    PubMed

    Tsoukas, C D; Lambris, J D

    1988-08-01

    The biologic effects of the third component of complement, C3, are mediated via receptors which specifically bind the enzymatic degradation products resulting from the cleavage of C3. One of the products, C3d, has been associated with binding to the second complement receptor CR2 (CD21). This receptor, which is identical to the receptor for Epstein-Barr virus (EBV), has been primarily found on cells of the B lineage, but not on mature T cells or other cells of erythroid or myeloid lineages. In the present investigation, we report the presence of CR2 on human thymocytes. Indirect immunofluorescence analysis employing monoclonal anti-CR2 antibodies revealed a range of thymocyte reactivity from 15% to 63% in thirteen experiments using cells of different donors. Reactivity was always greater with the monoclonal anti-CR2 (CD21) antibody HB-5 than with two other antibodies which recognize distinct epitopes on the CR2 molecule. Two-color immunofluorescence analysis indicated that the brightest of the HB-5-stained thymocytes also reacted with the monoclonal anti-CD1 antibody T6 (immature thymocyte marker) while some of the duller HB-5-staining cells reacted with the monoclonal anti-CD3 antibody Leu-4 (mature thymocyte marker). Immunoprecipitation of CR2 on thymocytes with antibody HB-5 and polyacrylamide gel electrophoretic analysis revealed a protein of 145 kDa molecular mass which is consistent with the size of CR2 found on B lymphocytes. These findings raise several questions regarding the biologic role of CR2-EBV receptor on cells of the T lineage.

  2. Involvement of sodium in early phosphatidylserine exposure and phospholipid scrambling induced by P2X7 purinoceptor activation in thymocytes.

    PubMed

    Courageot, Marie-Pierre; Lépine, Sandrine; Hours, Michel; Giraud, Françoise; Sulpice, Jean-Claude

    2004-05-21

    Extracellular ATP (ATP(ec)), a possible effector in thymocyte selection, induces thymocyte death via purinoceptor activation. We show that ATP(ec) induced cell death by apoptosis, rather than lysis, and early phosphatidylserine (PS) exposure and phospholipid scrambling in a limited thymocyte population (35-40%). PS externalization resulted from the activation of the cationic channel P2X7 (formerly P2Z) receptor and was triggered in all thymocyte subsets although to different proportions in each one. Phospholipid movement was dependent on ATP(ec)-induced Ca(2+) and/or Na(+) influx. At physiological external Na(+) concentration, without external Ca(2+), PS was exposed in all ATP(ec)-responsive cells. In contrast, without external Na(+), physiological external Ca(2+) concentration promoted a submaximal response. Altogether these data show that Na(+) influx plays a major role in the rapid PS exposure induced by P2X7 receptor activation in thymocytes.

  3. SCL, LMO1 and Notch1 Reprogram Thymocytes into Self-Renewing Cells

    PubMed Central

    Rojas-Sutterlin, Shanti; Herblot, Sabine; Hébert, Josée; Sauvageau, Guy; Lemieux, Sébastien; Lécuyer, Eric; Veiga, Diogo F. T.; Hoang, Trang

    2014-01-01

    The molecular determinants that render specific populations of normal cells susceptible to oncogenic reprogramming into self-renewing cancer stem cells are poorly understood. Here, we exploit T-cell acute lymphoblastic leukemia (T-ALL) as a model to define the critical initiating events in this disease. First, thymocytes that are reprogrammed by the SCL and LMO1 oncogenic transcription factors into self-renewing pre-leukemic stem cells (pre-LSCs) remain non-malignant, as evidenced by their capacities to generate functional T cells. Second, we provide strong genetic evidence that SCL directly interacts with LMO1 to activate the transcription of a self-renewal program coordinated by LYL1. Moreover, LYL1 can substitute for SCL to reprogram thymocytes in concert with LMO1. In contrast, inhibition of E2A was not sufficient to substitute for SCL, indicating that thymocyte reprogramming requires transcription activation by SCL-LMO1. Third, only a specific subset of normal thymic cells, known as DN3 thymocytes, is susceptible to reprogramming. This is because physiological NOTCH1 signals are highest in DN3 cells compared to other thymocyte subsets. Consistent with this, overexpression of a ligand-independent hyperactive NOTCH1 allele in all immature thymocytes is sufficient to sensitize them to SCL-LMO1, thereby increasing the pool of self-renewing cells. Surprisingly, hyperactive NOTCH1 cannot reprogram thymocytes on its own, despite the fact that NOTCH1 is activated by gain of function mutations in more than 55% of T-ALL cases. Rather, elevating NOTCH1 triggers a parallel pathway involving Hes1 and Myc that dramatically enhances the activity of SCL-LMO1 We conclude that the acquisition of self-renewal and the genesis of pre-LSCs from thymocytes with a finite lifespan represent a critical first event in T-ALL. Finally, LYL1 and LMO1 or LMO2 are co-expressed in most human T-ALL samples, except the cortical T subtype. We therefore anticipate that the self-renewal network

  4. SCL, LMO1 and Notch1 reprogram thymocytes into self-renewing cells.

    PubMed

    Gerby, Bastien; Tremblay, Cedric S; Tremblay, Mathieu; Rojas-Sutterlin, Shanti; Herblot, Sabine; Hébert, Josée; Sauvageau, Guy; Lemieux, Sébastien; Lécuyer, Eric; Veiga, Diogo F T; Hoang, Trang

    2014-12-01

    The molecular determinants that render specific populations of normal cells susceptible to oncogenic reprogramming into self-renewing cancer stem cells are poorly understood. Here, we exploit T-cell acute lymphoblastic leukemia (T-ALL) as a model to define the critical initiating events in this disease. First, thymocytes that are reprogrammed by the SCL and LMO1 oncogenic transcription factors into self-renewing pre-leukemic stem cells (pre-LSCs) remain non-malignant, as evidenced by their capacities to generate functional T cells. Second, we provide strong genetic evidence that SCL directly interacts with LMO1 to activate the transcription of a self-renewal program coordinated by LYL1. Moreover, LYL1 can substitute for SCL to reprogram thymocytes in concert with LMO1. In contrast, inhibition of E2A was not sufficient to substitute for SCL, indicating that thymocyte reprogramming requires transcription activation by SCL-LMO1. Third, only a specific subset of normal thymic cells, known as DN3 thymocytes, is susceptible to reprogramming. This is because physiological NOTCH1 signals are highest in DN3 cells compared to other thymocyte subsets. Consistent with this, overexpression of a ligand-independent hyperactive NOTCH1 allele in all immature thymocytes is sufficient to sensitize them to SCL-LMO1, thereby increasing the pool of self-renewing cells. Surprisingly, hyperactive NOTCH1 cannot reprogram thymocytes on its own, despite the fact that NOTCH1 is activated by gain of function mutations in more than 55% of T-ALL cases. Rather, elevating NOTCH1 triggers a parallel pathway involving Hes1 and Myc that dramatically enhances the activity of SCL-LMO1 We conclude that the acquisition of self-renewal and the genesis of pre-LSCs from thymocytes with a finite lifespan represent a critical first event in T-ALL. Finally, LYL1 and LMO1 or LMO2 are co-expressed in most human T-ALL samples, except the cortical T subtype. We therefore anticipate that the self-renewal network

  5. Positioning.

    ERIC Educational Resources Information Center

    Conone, Ruth M.

    The key to positioning is the creation of a clear benefit image in the consumer's mind. One positioning strategy is creating in the prospect's mind a position that takes into consideration the company's or agency's strengths and weaknesses as well as those of its competitors. Another strategy is to gain entry into a position ladder owned by…

  6. Evolution of double positive autoregulatory feedback loops in CYCLOIDEA2 clade genes is associated with the origin of floral zygomorphy.

    PubMed

    Yang, Xia; Pang, Hong-Bo; Liu, Bo-Ling; Qiu, Zhi-Jing; Gao, Qiu; Wei, Lai; Dong, Yang; Wang, Yin-Zheng

    2012-05-01

    Members of the CYCLOIDEA2 (CYC2) clade of the TEOSINTE BRANCHED1, CYCLOIDEA, and PCF transcription factor genes are widely involved in controlling floral zygomorphy, a key innovation in angiosperm evolution, depending on their persistently asymmetric expression in the corresponding floral domains. However, it is unclear how this asymmetric expression is maintained throughout floral development. Selecting Primulina heterotricha as a model, we examined the expression and function of two CYC2 genes, CYC1C and CYC1D. We analyzed the role of their promoters in protein-DNA interactions and transcription activation using electrophoresis mobility shift assays, chromatin immunoprecipitation, and transient gene expression assays. We find that CYC1C and CYC1D positively autoregulate themselves and cross-regulate each other. Our results reveal a double positive autoregulatory feedback loop, evolved for a pair of CYC2 genes to maintain their expression in developing flowers. Further comparative genome analyses, together with the available expression and function data of CYC2 genes in the core eudicots, suggest that this mechanism might have led to the independent origins of floral zygomorphy, which are associated with plant-insect coevolution and the adaptive radiation of angiosperms.

  7. HER2-positive double primary tumor of gastric and breast cancer occur synchronously in a patient: A case report.

    PubMed

    Ouyang, Quchang; Tian, Can; Gao, Jianxiang; Huang, Jin; Fu, Hua; He, Jinsong; Yang, Jianbo

    2016-05-01

    The simultaneous occurrence of primary gastric cancer and breast cancer is rare, and the positive expression of human epidermal growth factor receptor (HER)2 in double primary carcinoma of gastric and breast cancer remains to be reported. The present study presented a 46-year-old woman complaining of irregular acid reflux and stomach discomfort. The stomach cancer was diagnosed by esophagogastroduodenoscopy examination of the pathological biopsies in 2010. The patient underwent a radical gastrectomy for gastric cancer, and postoperative pathological examination revealed moderately-poorly differentiated adenocarcinoma with HER2 positive expression. The tumor invaded into the entire thickness of the gastric wall and lymph nodes. The patient received five treatments of postoperative chemotherapy. In August 2011, the patient felt a lump in the right breast. Simple excision of the right breast mass was performed on September 2011, and postoperative pathological examination revealed the invasive ductal carcinoma of the right breast with HER2 amplification by fluorescent in situ hybridization assay. The patient was treated with postoperative chemotherapy and radiotherapy, and also Trastuzumab target therapy. The patient succumbed to aggressive disease progression in March 2012.

  8. Requirement of mitoses for the reversal of X-inactivation in cell hybrids between murine embryonal carcinoma cells and normal female thymocytes

    SciTech Connect

    Takagi, N. )

    1988-04-01

    By means of a 5-bromodeoxyuridine (BrdU) incorporation and acridine orange fluorescence staining method the authors studied reactivation of the inactivated X chromosome (X{sub i}) in newly formed cell hybrids between the near-diploid HPRT-deficient OTF9-63 murine embryonal carcinoma cell (ECC) with an XO sex chromosome constitution and the normal female mouse thymocyte. Synchronization of the late replicating S chromosome in such hybrid cells, indicative of reactivation, was found for the first time on Day 3, and the frequency of reactivation was attained 90% on Day 5. Inhibition of cell cycle progression either by methylglyoxal bis(guanylhydrazone) dihydrochloride, an inhibitor of polyamine metabolism, or by isoleucine-deficient medium after cell fusion delayed reactivation of the X{sub i}, which implied that the number of cell division cycles traversed by individual cells rather than the length of time after cell fusion is critical for the reactivation. Double-labeling experiments using ({sup 3}H)thymidine and BrdU indicated that hybrid cells had undergone three or four mitoses before reactivation of the X{sub i}. Most probably reactivation of the X{sub i} is consequent to reversion of the thymocyte genome to an undifferentiated state under the influence of OTF9 genome. DNA demethylation or dilution of X{sub i}-specific factors by mitoses may be involved in this process.

  9. Effectiveness of rivastigmine on positive, negative, and cognitive symptoms of schizophrenia: a double-blind clinical trial

    PubMed Central

    Shoja Shafti, Saeed; Azizi Khoei, Abbas

    2016-01-01

    Background: Several lines of evidence suggest that the cholinergic system may be disrupted in schizophrenia and so this may contribute to the cognitive impairments of schizophrenic patients. Because such deficits do not respond to neuroleptic treatment, different approaches have been done by acetylcholinesterase inhibitors (AChEIs). The objective of the present assessment was to evaluate the safety and clinical effects of rivastigmine, as an adjunctive drug, on the clinical symptoms of schizophrenia. Methods: A total of 46 patients with a diagnosis of schizophrenia entered into a 12-week, double-blind, clinical trial for random assignment to rivastigmine or placebo, as adjuvant to their current antipsychotic medication. Positive and Negative Symptom Scale (PANSS) and Mini Mental State Examination (MMSE) had been used as the primary outcome measures. Clinical Global Impressions- Improvement (CGI-I) Scale and Extrapyramidal Symptom Rating Scale (ESRS) had been used as the secondary measures. Treatment efficacy was evaluated by a Student’s t test and repeated-measures analysis of variance (ANOVA). Statistical significance was defined as a two-sided p value ⩽ 0.05. Cohen’s standard (d) and correlation measures of effect size (r) had been calculated for comparing baseline to endpoint changes. Results: According to the findings, except for significant enhancement of MMSE by rivastigmine (p < 0.001), no significant improvement in PANSS (negative symptoms), PANSS (positive symptoms), and PANSS (general psychopathology) was evident in the target group. Also, except for significant improvement of CGI-I by rivastigmine in intragroup analysis, no significant effectiveness was evident in between-group analysis or repeated-measures ANOVA. ESRS, also, did not show any significant alteration in either group. Effect size (ES) analysis showed a large improvement in MMSE by rivastigmine. Conclusions: According to the findings, while rivastigmine could not induce significant

  10. Early effects of carbon-ion irradiation on murine lymphocytes and thymocytes

    NASA Astrophysics Data System (ADS)

    Xie, Yi; Zhang, Hong; Dang, Bingrong; Bing, Tao; Hao, Jifang; Guo, Hongyun; Wang, Xiaohu

    To estimate the biological risks from space radiation encountered by cosmonauts in outer space, the effects from whole-body exposure to carbon ions or X-rays irradiations at 0, 0.39, 0.55 and 1 Gy at a dose rate of 0.2 Gy/min were investigated in BALB/c mice. The relative thymus and spleen weights were measured at 24 h after exposure, and the cell cycle distribution and percentage of apoptosis of thymocytes and spleen and peripheral blood lymphocytes were determined by flow cytometry. The data showed that exposure to carbon ions delayed cell progression of peripheral blood lymphocytes in S-phase, and delayed thymocytes and spleen lymphocytes in G 0/G 1-phase. Apoptosis of thymocytes and peripheral blood lymphocytes induced by carbon ions increased more rapidly with dose than was the case for X-rays. There were some differences between the relative weight loss of the thymus and the spleen with increasing dose of either carbon ions or X-rays. The results obtained provide evidence of dose- and organ-specific differences induced by carbon ions compared to X-rays, with increased apoptosis in peripheral blood lymphocytes and thymocytes, but not spleen lymphocytes. Our data may suggest that further work would be of interest to estimate risk of changes in immune function during particle radiation exposures in space travel.

  11. Transcriptome analysis of murine thymocytes reveals age-associated changes in thymic gene expression.

    PubMed

    Lustig, Ana; Carter, Arnell; Bertak, Dorothy; Enika, Divya; Vandanmagsar, Bolormaa; Wood, William; Becker, Kevin G; Weeraratna, Ashani T; Taub, Dennis D

    2009-01-01

    The decline in adaptive immunity, naïve T-cell output and a contraction in the peripheral T cell receptor (TCR) repertoire with age are largely attributable to thymic involution and the loss of critical cytokines and hormones within the thymic microenvironment. To assess the molecular changes associated with this loss of thymic function, we used cDNA microarray analyses to examine the transcriptomes of thymocytes from mice of various ages ranging from very young (1 month) to very old (24 months). Genes associated with various biological and molecular processes including oxidative phosphorylation, T- and B- cell receptor signaling and antigen presentation were observed to significantly change with thymocyte age. These include several immunoglobulin chains, chemokine and ribosomal proteins, annexin A2, vav 1 and several S100 signaling proteins. The increased expression of immunoglobulin genes in aged thymocytes could be attributed to the thymic B cells which were found to be actively producing IgG and IgM antibodies. Upon further examination, we found that purified thymic T cells derived from aged but not young thymi also exhibited IgM on their cell surface suggesting the possible presence of auto-antibodies on the surface thymocytes with advancing age. These studies provide valuable insight into the cellular and molecular mechanisms associated with thymic aging.

  12. Insulin effect on glucose transport in thymocytes and splenocytes from rats with metabolic syndrome

    PubMed Central

    2010-01-01

    Metabolic syndrome (MS) may comprise several clinical conditions such as obesity, diabetes and inflammatory disorders, which are characterized by metabolic imbalances. The study of glucose transport and regulation by insulin in lymphocytes is important, since the way they increase inflammation and susceptibility to infections are common in MS. We studied glucose internalization in isolated thymocytes and splenocytes, its regulation by insulin, and the role of three glucose transporters (Gluts) in control and in MS rats. Control glucose internalization and insulin responses were lower in splenocytes than in thymocytes. Control and insulin-induced glucose internalization in thymocytes declined with age, while transport by splenocyte continued to respond to insulin. Control thymocyte glucose internalization was blocked by antibodies against Glut 1 and 4, while the insulin response also was blocked by an anti-Glut 3 antibody. On four month old control and insulin-induced response, splenocyte transport was only blocked by Glut 1 and 4 antibodies. At six months splenocyte glucose internalization depended on Glut 1 and was less sensitive to the effects of an anti-Glut 4 antibody. In MS splenocytes the capacity of anti-Glut 1 antibodies to inhibit control and insulin-dependent glucose transport was less significant, and we found that in MS rats, glucose internalization was dependent on Glut 3 and Glut 4. In summary, the altered metabolic state present in MS rats shows signs of modulation of glucose internalization by the Glut1, Glut 3 and Glut 4 transporters, compared with its own age control. PMID:21044347

  13. Fish thymocyte viability, apoptosis and necrosis: In-vitro effects of organochlorine contaminants

    USGS Publications Warehouse

    Sweet, L.I.; Passino-Reader, D. R.; Meier, P.G.; Omann, G.M.

    1998-01-01

    The thymus is believed to be a central component of haematopoiesis and immune function in teleosts. Hence, chemically-elicited adverse effects to the thymus may result in immunomodulation and organ dysfunction. The objective of this research was to assess the levels of active (apoptotic) and passive (necrotic) cell death in untreated and organochlorine treated fish thymocytes. Lake trout (Salvelinus namaycush) thymocytes were challenged with Aroclor 1254 (concentration range 1.5-10.5??g ml-1) and alpha, beta, gamma, delta isomers of hexachlorocyclohexane (concentration range 10-100??M). The resulting maintenance or loss of viability was assessed by cytofluorometry (expression of phosphatidylserine and exclusion of propidium iodide) and confirmed with fluorescence microscopy. The results indicate that 20-60% of thymocytes in healthy fish undergo apoptosis, whereas thymocytes treated for 6-24 h with organochlorines exhibit increased levels of apoptotic cell death. This study demonstrates that given sufficient concentration, contact time and cellular receptors, organochlorines such as Aroclor 1254 and hexachlorocyclohexanes may induce direct or indirect toxicity, altered functionality, or cell death to an organ important for fish immunocompetence. ?? 1998 Academic Press Limited.

  14. Thymocytes may persist and differentiate without any input from bone marrow progenitors

    PubMed Central

    Peaudecerf, Laetitia; Lemos, Sara; Galgano, Alessia; Krenn, Gerald; Vasseur, Florence; Di Santo, James P.; Ezine, Sophie

    2012-01-01

    Thymus transplants can correct deficiencies of the thymus epithelium caused by the complete DiGeorge syndrome or FOXN1 mutations. However, thymus transplants were never used to correct T cell–intrinsic deficiencies because it is generally believed that thymocytes have short intrinsic lifespans. This notion is based on thymus transplantation experiments where it was shown that thymus-resident cells were rapidly replaced by progenitors originating in the bone marrow. In contrast, here we show that neonatal thymi transplanted into interleukin 7 receptor–deficient hosts harbor populations with extensive capacity to self-renew, and maintain continuous thymocyte generation and export. These thymus transplants reconstitute the full diversity of peripheral T cell repertoires one month after surgery, which is the earliest time point studied. Moreover, transplantation experiments performed across major histocompatibility barriers show that allogeneic transplanted thymi are not rejected, and allogeneic cells do not induce graft-versus-host disease; transplants induced partial or total protection to infection. These results challenge the current dogma that thymocytes cannot self-renew, and indicate a potential use of neonatal thymus transplants to correct T cell–intrinsic deficiencies. Finally, as found with mature T cells, they show that thymocyte survival is determined by the competition between incoming progenitors and resident cells. PMID:22778388

  15. Radical formation in the FMN-photosensitized reactions of unsaturated fatty acids bearing double bonds at different positions.

    PubMed

    Nishihama, Nao; Iwahashi, Hideo

    2016-08-15

    Although the reaction mechanisms through which flavin mononucleotide works as an endogenous photosensitizer have been investigated (Baier et al., 2006; Edwards and Silva, 2001; Pajares et al., 2001; Criado et al., 2003; Massad et al., 2008) [23-27], few studies have been performed for the reactions of flavin mononucleotide with unsaturated fatty acids. To examine the reactions of flavin mononucleotide with unsaturated fatty acids bearing a double bond at different positions, an electron spin resonance, a high performance liquid chromatography-electron spin resonance and a high performance liquid chromatography-electron spin resonance-mass spectrometry were employed. The control reaction mixtures contained 25μmolL(-1) of flavin mononucleotide, 1.0mmolL(-1) of FeSO4(NH4)2SO4, 10mmolL(-1) of cholic acid, 30mmolL(-1) of phosphate buffer (pH 7.4) and 0.1molL(-1) of α-(4-pyridyl-1-oxide)-N-tert-butylnitrone in deuterium oxide. In addition, it also contained 4.3mmolL(-1) of one of the following: (z)-11-octadecenoic acid, (z)-6-octadecenoic acid, (z)-9-octadecenoic acid or (z, z)-9, 12-octadecadienoic acid. The control reaction mixtures without FeSO4(NH4)2SO4 and α-(4-pyridyl-1-oxide)-N-tert-butylnitrone were exposed to the visible light at 436nm (7.8Jcm(-2)). After the irradiation, α-(4-pyridyl-1-oxide)-N-tert-butylnitrone was added. The reactions started from adding FeSO4(NH4)2SO4 and performed at 25°C for 1min. Electron spin resonance measurements of the control reaction mixtures showed prominent signals (α(N)=1.58mT and α(Hβ)=0.26mT). High performance liquid chromatography-electron spin resonance analyses of the control reaction mixtures showed prominent peaks at the retention times of 31.1min {(z)-6-octadecenoic acid}, 39.6min {(z)-9-octadecenoic acid}, 44.9min {(z)-11-octadecenoic acid} and 40.2min {(z, z)-9, 12-octadecadienoic acid}. High performance liquid chromatography-electron spin resonance-mass analyses of the control reaction mixtures showed that 4

  16. Standing striations as a series of double layers induced in a positive column by a transverse magnetic field

    SciTech Connect

    Toma, M.; Biborosch, L.; Curteanu, M.

    1995-12-31

    It was experimentally proved that the action of a static transverse magnetic field (TMF) on a cylindrical positive column (PC) can change its internal structure. As a result, a succession of luminous structures known as standing striations are observed. The excitation of striations in a PC (frequently in molecular gases) is usually explained taking into account the periodic changes in ionised rate. It is known that the ionization rate is a strong function of the electron temperature (more precisely, kinetic energy). Thus, the standing striations being the spatial periodic change in the ionization rate, are affected by electron velocity. It can easily observed that in a striated PC there is a periodic change in light intensity. This means that beside ionization processes there is also a periodic change in excitation processes. It was showed that standing striations are, in fact spatial sequences of space charge structures known as double layers (Dls). In the last time it was proved that there is a direct connection between the excitation processes and the ordered spatial arrangement of the electric charges inside the DLs. The aim of this paper is to emphasize that the appearance of standing striations can be adequately described by the model of DL generation in a collisional plasma, that takes into account the electron-neutral excitation processes.

  17. Population doublings and percentage of S100-positive cells as predictors of in vitro chondrogenicity of expanded human articular chondrocytes.

    PubMed

    Giovannini, Samoa; Diaz-Romero, Jose; Aigner, Thomas; Mainil-Varlet, Pierre; Nesic, Dobrila

    2010-02-01

    The aim of this study was to investigate the interconnection between the processes of proliferation, dedifferentiation, and intrinsic redifferentiation (chondrogenic) capacities of human articular chondrocyte (HAC), and to identify markers linking HAC dedifferentiation status with their chondrogenic potential. Cumulative population doublings (PD) of HAC expanded in monolayer culture were determined, and a threshold range of 3.57-4.19 PD was identified as indicative of HAC loss of intrinsic chondrogenic capacity in pellets incubated without added chondrogenic factors. While several specific gene and surface markers defined early HAC dedifferentiation process, no clear correlation with the loss of intrinsic chondrogenic potential could be established. CD90 expression during HAC monolayer culture revealed two subpopulations, with sorted CD90-negative cells showing lower proliferative capacity and higher chondrogenic potential compared to CD90-positive cells. Although these data further validated PD as critical for in vitro chondrogenesis, due to the early shift in expression, CD90 could not be considered for predicting chondrogenic potential of HAC expanded for several weeks. In contrast, an excellent mathematically modeled correlation was established between PD and the decline of HAC expressing the intracellular marker S100, providing a direct link between the number of cell divisions and dedifferentiation/loss of intrinsic chondrogenic capacity. Based on the dynamics of S100-positive HAC during expansion, we propose asymmetric cell division as a potential mechanism of HAC dedifferentiation, and S100 as a marker to assess chondrogenicity of HAC during expansion, of potential value for cell-based cartilage repair treatments. (c) 2009 Wiley-Liss, Inc.

  18. Efficacy of continuous positive airway pressure on middle ear atelectasis: A double-blind placebo-controlled clinical trial.

    PubMed

    Akbulut, Sevtap; Demir, Mehmet Gokhan; Salepci, Banu Musaffa; Gungor, Gulten Aktin; Demir, Necdet; Berk, Derya; Cakan, Dogan

    2016-07-01

    To investigate the effects of continuous positive airway pressure (CPAP) treatment on patients with middle ear atelectasis. Prospective, double-blind, randomized, placebo-controlled study. Fifty-four patients with middle ear atelectasis were randomized to receive CPAP treatment with a pressure level of either 14 cm H2 O (CPAP group) or 0 cm H2 O (placebo group) once per week for a period of 3 hours for 4 sessions. Outcome measures included otomicroscopic examination as well as tympanometric and audiometric evaluation. Patients were followed for 6 months. The CPAP group included 35 atelectatic ears, and the placebo group included 32 atelectatic ears. More ears recovered to normal tympanic membrane or regressed to grade 1 atelectasis in the CPAP group than in the placebo group during all follow-up visits (P < .05). There was a statistically significant increase in the middle ear pressure values of the patients in the CPAP group compared to the placebo group at week 5, month 3, and month 6 (P < .05). There was no significant difference in middle ear pressure values between follow-up visits in the CPAP group (P > .05). Significant improvement of pure-tone air-conduction threshold averages were found in the CPAP group compared to the placebo group at month 6 (P < .05). CPAP is a safe, well-tolerated way of applying positive pressure to the middle ear for patients with middle ear atelectasis. It contributes to significant improvement in middle ear pressure of these patients, also resulting in an improved degree of atelectasis. 1b. Laryngoscope, 126:1649-1655, 2016. © 2015 The American Laryngological, Rhinological and Otological Society, Inc.

  19. Antibody-directed double suicide gene therapy targeting of MUC1- positive leukemia cells in vitro and in vivo.

    PubMed

    Dong, Xiao-Ya; Wang, Wen-Qian; Zhao, Yu; Li, Xu-Dong; Fang, Zhi-Gang; Lin, Dong-Jun; Xiao, Ruo-Zhi; Huang, Ren-Wei; Pan, Guang-Jin; Liu, Jia-Jun

    2013-10-01

    Our aim was to specifically transfer the cytosine deaminase (CD) and thymidine kinase (TK) genes into mucin 1 (MUC1)-positive leukemia cells by anti-MUC1 antibody directed infection of replication-defective lentivirus and to evaluate the targeted cytotoxicity of double suicide genes to leukemia. The target gene vector (containing CD and TK) and envelope (containing GFP and anti-MUC1) and packaging plasmids were cotransfected into 293T cells to produce the recombinant lentivirus. Suicide genes in virus-infected leukemia cells (U937, Jurkat, and K562) were detected by western blot. The cytotoxicity and bystander effect in vitro and the therapeutic effect in vivo were detected after treatment with the prodrugs. The results revealed that combined treatment with prodrug 5-fluorocytosine (5-FC) and ganciclovir (GCV) inhibited leukemia cell growth and caused significant bystander effect than treatment with either prodrug alone. TK/GCV treatment alone induced degeneration and cell death while the effect of CD/5-FC alone mainly caused vacuolar degeneration and necrosis. The addictive effects of combinatorial use of GCV and 5-FC mainly induced swelling of the mitochondria followed by necrosis of the leukemia cells. In vivo experiments revealed that both single and combinatorial prodrug treatments could prolong the survival time of leukemic mice. In summary, anti-MUC1 antibody directed lentiviral vector successfully transduced dual suicide genes and exerted targeted cytotoxicity against MUC1 positive leukemia cells. This targeted lentiviral dual suicide gene delivering system provides a promising approach for clinical treatment of leukemia in future.

  20. Single venom-based immunotherapy effectively protects patients with double positive tests to honey bee and Vespula venom

    PubMed Central

    2013-01-01

    Background Referring to individuals with reactivity to honey bee and Vespula venom in diagnostic tests, the umbrella terms “double sensitization” or “double positivity” cover patients with true clinical double allergy and those allergic to a single venom with asymptomatic sensitization to the other. There is no international consensus on whether immunotherapy regimens should generally include both venoms in double sensitized patients. Objective We investigated the long-term outcome of single venom-based immunotherapy with regard to potential risk factors for treatment failure and specifically compared the risk of relapse in mono sensitized and double sensitized patients. Methods Re-sting data were obtained from 635 patients who had completed at least 3 years of immunotherapy between 1988 and 2008. The adequate venom for immunotherapy was selected using an algorithm based on clinical details and the results of diagnostic tests. Results Of 635 patients, 351 (55.3%) were double sensitized to both venoms. The overall re-exposure rate to Hymenoptera stings during and after immunotherapy was 62.4%; the relapse rate was 7.1% (6.0% in mono sensitized, 7.8% in double sensitized patients). Recurring anaphylaxis was statistically less severe than the index sting reaction (P = 0.004). Double sensitization was not significantly related to relapsing anaphylaxis (P = 0.56), but there was a tendency towards an increased risk of relapse in a subgroup of patients with equal reactivity to both venoms in diagnostic tests (P = 0.15). Conclusions Single venom-based immunotherapy over 3 to 5 years effectively and long-lastingly protects the vast majority of both mono sensitized and double sensitized Hymenoptera venom allergic patients. Double venom immunotherapy is indicated in clinically double allergic patients reporting systemic reactions to stings of both Hymenoptera and in those with equal reactivity to both venoms in diagnostic tests who have not reliably identified the

  1. CD40 expressed on thymic epithelial cells provides costimulation for proliferation but not for apoptosis of human thymocytes.

    PubMed

    Ruggiero, G; Martinez Cáceres, E; Voordouw, A; Noteboom, E; Graf, D; Kroczek, R A; Spits, H

    1996-05-15

    Human thymic epithelial cells express CD40, so we examined the possible role of CD40 in activation of thymocytes. We observed that both CD4+CD8- and CD4-CD8+ thymocytes proliferate after stimulation by anti-CD3 mAb in the presence of cultured thymic epithelial cells. Costimulation of CD4+ thymocytes by thymic epithelial cells is partly inhibited by an anti-CD40 mAb, but this mAb has no effect on costimulation of CD8+ thymocytes. The selective costimulatory ability of CD40 for CD4+ thymocytes was confirmed in experiments in which thymocytes were stimulated with anti-CD3 in the presence of murine P815 cells transfected with CD40 cDNA. The level of costimulation induced by P815-CD40 was comparable with that induced by P815 cells expressing CD80 (B7.1). Treatment of thymocytes with the Ca2+ ionophore ionomycin and the phorbol ester PMA or with anti-CD3 mAb resulted in up-regulation of the CD40 ligand, suggesting that this molecule is involved in CD40-mediated costimulation of human thymocytes. Costimulation of thymocytes by CD80 strongly increased anti-CD3-induced death of fetal thymocytes. In contrast, costimulation by CD40 did not increase anti-CD3-mediated apoptosis of these thymocytes. To confirm that CD40 does not affect anti-CD3-induced cell death, we established a variant of the Jurkat T leukemic cell line that constitutively expresses CD40L and analyzed the sensitivity of this cell line for activation-induced apoptosis. In contrast to CD80, CD40 failed to increase anti-CD3-mediated apoptosis in CD40L+ Jurkat cells, whereas both CD40 and CD80 strongly increased IL-2 production induced by anti-CD3. These findings suggest that costimulation by CD40 is involved in clonal expansion of CD4+ thymocytes but not in activation-induced cell death.

  2. [Effect of extremely low frequency electromagnetic radiation and ultra-violet radiation on aggregation of thymocytes and erythrocytes].

    PubMed

    Roshchupkin, D I; Kramarenko, G G; Anosov, A K

    1996-01-01

    Electromagnetic radiation of superhigh frequencies (46.12 and 46.19 GHz, 0.3-1 mV/cm2) at an incident dose of about 12 kJ/m2 enhances the ability of isolated rabbit thymocytes for aggregation interaction with homologous erythrocytes. In the case of 46.19 GHz frequency, the stimulatory effect disappears as radiation dose in increased. A radiation of 46.12 GHz stimulates thymocytes also at high radiation doses. Superhigh-frequency radiation enhances the sensitivity of thymocytes to the damaging effect of UV radiation.

  3. Canine peripheral blood CD4(+)CD8(+) double-positive Tcell subpopulations exhibit distinct Tcell phenotypes and effector functions.

    PubMed

    Rothe, Kathrin; Bismarck, Doris; Büttner, Mathias; Alber, Gottfried; von Buttlar, Heiner

    2017-03-01

    Canine peripheral blood CD4(+)CD8α(+) double-positive (dp) Tcells represent a small population of T lymphocytes that are not only derived from single-positive (sp) CD4(+) but also from CD8(+) Tcells. Dp Tcells can be subdivided in three different subpopulations according to their expression levels of CD4 and CD8α, i.e. the CD4(dim)CD8α(bright), CD4(bright)CD8α(bright), and the CD4(bright)CD8α(dim) subsets. Previously we could show that total canine peripheral CD4(+)CD8α(+) dp Tcells have features of activated Tcells. Here we demonstrate that the individual dp Tcell subsets distinctly differ in their activation status and phenotype. Thus, among the dp Tcell subsets the CD4(dim)CD8α(bright) subpopulation has the lowest and the CD4(bright)CD8α(bright) subset the highest frequency of CD25(+) cells pointing to the CD4(bright)CD8α(bright) subset with the highest activation status. Consistent with that, analysis of CD44 vs. CD62L expression demonstrates that the CD4(bright)CD8α(bright) subset contains the highest frequencies of effector-memory Tcells (TEM). Upon in vitro stimulation, the CD4(bright)CD8α(bright) and CD4(dim)CD8α(bright) dp Tcell subsets show the highest frequencies of IFN-γ producing cells. Expression of granzyme B was found exclusively in the CD4(dim)CD8α(bright) dp Tcell subset indicating cytotoxic potential of this dp Tcell subset. Furthermore, T-bet expression was restricted to the CD4(dim)CD8α(bright) subset, while Foxp3(+) regulatory Tcells were detected in the CD4(bright)CD8α(dim) dp Tcell subset. In conclusion, canine dp Tcells are phenotypically and functionally heterogeneous consistent with the diverse characteristics of their single-positive CD4(+) and CD8(+) Tcell progenitors. The data provide the basis for future in vivo studies to elucidate the role of individual dp Tcell subsets in host defense and/or immune pathological diseases of dogs.

  4. Clinical Significance of Tissue Factor and CD13 Double-Positive Microparticles in Sirs Patients with Trauma and Severe Sepsis.

    PubMed

    Matsumoto, Hisatake; Yamakawa, Kazuma; Ogura, Hiroshi; Koh, Taichin; Matsumoto, Naoya; Shimazu, Takeshi

    2017-04-01

    Activated immune cells such as monocytes are key factors in systemic inflammatory response syndrome (SIRS) following trauma and sepsis. Activated monocytes induce almost all tissue factor (TF) expression contributing to inflammation and coagulation. TF and CD13 double-positive microparticles (TF/CD13MPs) are predominantly released from these activated monocytes. This study aimed to evaluate TF/CD13MPs and assess their usefulness as a biomarker of pathogenesis in early SIRS following trauma and sepsis. This prospective study comprising 24 trauma patients, 25 severe sepsis patients, and 23 healthy controls was conducted from November 2012 to February 2015. Blood samples were collected from patients within 24 h after injury and diagnosis of severe sepsis and from healthy controls. Numbers of TF/CD13MPs were measured by flow cytometry immediately thereafter. Injury Severity Score (ISS) and Acute Physiology and Chronic Health Evaluation (APACHE) II and Sequential Organ Failure Assessment (SOFA) scores were calculated at patient enrollment. APACHE II and SOFA scores and International Society of Thrombosis and Haemostasis (ISTH) overt disseminated intravascular coagulation (DIC) diagnostic criteria algorithm were calculated at the time of enrollment of severe sepsis patients. Numbers of TF/CD13MPs were significantly increased in both trauma and severe sepsis patients versus controls and correlated significantly with ISS and APACHE II score in trauma patients and with APACHE II and ISTH DIC scores in severe sepsis patients. Increased numbers of TF/CD13MPs correlated significantly with severities in the acute phase in trauma and severe sepsis patients, suggesting that TF/CD13MPs are important in the pathogenesis of early SIRS following trauma and sepsis.

  5. Outcome of anti-thymocyte immunoglobulin plus cyclosporine A for severe aplastic anaemia with chronic hepatitis B virus infection.

    PubMed

    Chen, Miao; Zhuang, Junling; Zhou, Daobin; Xu, Ying; Zhao, Yongqiang; Wang, Shujie; Zhang, Wei; Duan, Minghui; Zhu, Tienan; Li, Jian; Cai, Huacong; Cao, Xinxin; Han, Bing

    2017-04-01

    The influence of chronic hepatitis B virus (HBV) infection on the efficacy of intensive immunosuppressive treatment (IST) of severe aplastic anaemia (SAA) patients remains unclear. Previous reports on this topic have been mostly case reports or have had a relatively short follow-up. Eight SAA patients carrying chronic HBV infection and 24 matched patients without HBV at a ratio of 1:3 were included in this retrospective analysis. The patients were treated with anti-thymocyte globulin (ATG) and cyclosporine A. Entecavir was or was not administered throughout the IST course to patients with positive or negative HBV-DNA results, respectively. No evident HBV reactivation developed. The overall response was 87.5% by 12 months, and the recurrence rate was 12.5%. There were no significant differences in overall response, overall survival and event-free survival between groups. Entecavir can effectively prevent reactivation of HBV in SAA patients with positive HBV-DNA who received intensive IST. Regular surveillance may be sufficient for HBV-DNA negative patients who should receive antiviral drugs immediately when their HBV-DNA status changes from negative to positive. The prognosis of SAA patients with chronic HBV infection after intensive IST treatment is not worse than those without HBV infection.

  6. Peroxynitrite-induced thymocyte apoptosis: the role of caspases and poly (ADP-ribose) synthetase (PARS) activation.

    PubMed Central

    Virág, L; Scott, G S; Cuzzocrea, S; Marmer, D; Salzman, A L; Szabó, C

    1998-01-01

    The mechanisms by which immature thymocyte apoptosis is induced during negative selection are poorly defined. Reports demonstrated that cross-linking of T-cell receptor leads to stromal cell activation, expression of inducible nitric oxide synthase (iNOS) and, subsequently, to thymocyte apoptosis. Therefore we examined, whether NO directly or indirectly, through peroxynitrite formation, causes thymocyte apoptosis. Immuno-histochemical detection of nitrotyrosine revealed in vivo peroxynitrite formation in the thymi of naive mice. Nitrotyrosine, the footprint of peroxynitrite, was predominantly found in the corticomedullary junction and the medulla of naive mice. In the thymi of mice deficient in the inducible isoform of nitric oxide synthase, considerably less nitrotyrosine was found. Exposure of thymocytes in vitro to low concentrations (10 microM) of peroxynitrite led to apoptosis, whereas higher concentrations (50 microM) resulted in intense cell death with the characteristics of necrosis. We also investigated the effect of poly (ADP-ribose) synthetase (PARS) inhibition on thymocyte apoptosis. Using the PARS inhibitor 3-aminobenzamide (3-AB), or thymocytes from PARS-deficient animals, we established that PARS determines the fate of thymocyte death. Suppression of cellular ATP levels, and the cellular necrosis in response to peroxynitrite were prevented by PARS inhibition. Therefore, in the absence of PARS, cells are diverted towards the pathway of apoptotic cell death. Similar results were obtained with H2O2 treatment, while apoptosis induced by non-oxidative stimuli such as dexamethasone or anti-FAS antibody was unaffected by PARS inhibition. In conclusion, we propose that peroxynitrite-induced apoptosis may play a role in the process of thymocyte negative selection. Furthermore, we propose that the physiological role of PARS cleavage by apopain during apoptosis may serve as an energy-conserving step, enabling the cell to complete the process of apoptosis

  7. Study of the role of cytosolic phospholipase A2 alpha in eicosanoid generation and thymocyte maturation in the thymus.

    PubMed

    Rousseau, Matthieu; Naika, Gajendra S; Perron, Jean; Jacques, Frederic; Gelb, Michael H; Boilard, Eric

    2015-01-01

    The thymus is a primary lymphoid organ, home of maturation and selection of thymocytes for generation of functional T-cells. Multiple factors are involved throughout the different stages of the maturation process to tightly regulate T-cell production. The metabolism of arachidonic acid by cyclooxygenases, lipoxygenases and specific isomerases generates eicosanoids, lipid mediators capable of triggering cellular responses. In this study, we determined the profile of expression of the eicosanoids present in the mouse thymus at different stages of thymocyte development. As the group IVA cytosolic phospholipase A2 (cPLA2α) catalyzes the hydrolysis of phospholipids, thereby generating arachidonic acid, we further verified its contribution by including cPLA2α deficient mice to our investigations. We found that a vast array of eicosanoids is expressed in the thymus, which expression is substantially modulated through thymocyte development. The cPLA2α was dispensable in the generation of most eicosanoids in the thymus and consistently, the ablation of the cPLA2α gene in mouse thymus and the culture of thymuses from human newborns in presence of the cPLA2α inhibitor pyrrophenone did not impact thymocyte maturation. This study provides information on the eicosanoid repertoire present during thymocyte development and suggests that thymocyte maturation can occur independently of cPLA2α.

  8. Study of the Role of Cytosolic Phospholipase A2 Alpha in Eicosanoid Generation and Thymocyte Maturation in the Thymus

    PubMed Central

    Rousseau, Matthieu; Naika, Gajendra S.; Perron, Jean; Jacques, Frederic; Gelb, Michael H.; Boilard, Eric

    2015-01-01

    The thymus is a primary lymphoid organ, home of maturation and selection of thymocytes for generation of functional T-cells. Multiple factors are involved throughout the different stages of the maturation process to tightly regulate T-cell production. The metabolism of arachidonic acid by cyclooxygenases, lipoxygenases and specific isomerases generates eicosanoids, lipid mediators capable of triggering cellular responses. In this study, we determined the profile of expression of the eicosanoids present in the mouse thymus at different stages of thymocyte development. As the group IVA cytosolic phospholipase A2 (cPLA2α) catalyzes the hydrolysis of phospholipids, thereby generating arachidonic acid, we further verified its contribution by including cPLA2α deficient mice to our investigations. We found that a vast array of eicosanoids is expressed in the thymus, which expression is substantially modulated through thymocyte development. The cPLA2α was dispensable in the generation of most eicosanoids in the thymus and consistently, the ablation of the cPLA2α gene in mouse thymus and the culture of thymuses from human newborns in presence of the cPLA2α inhibitor pyrrophenone did not impact thymocyte maturation. This study provides information on the eicosanoid repertoire present during thymocyte development and suggests that thymocyte maturation can occur independently of cPLA2α. PMID:25969996

  9. Recombinatorial biases and convergent recombination determine interindividual TCRβ sharing in murine thymocytes.

    PubMed

    Li, Hanjie; Ye, Congting; Ji, Guoli; Wu, Xiaohui; Xiang, Zhe; Li, Yuanyue; Cao, Yonghao; Liu, Xiaolong; Douek, Daniel C; Price, David A; Han, Jiahuai

    2012-09-01

    Overlap of TCR repertoires among individuals provides the molecular basis for public T cell responses. By deep-sequencing the TCRβ repertoires of CD4+CD8+ thymocytes from three individual mice, we observed that a substantial degree of TCRβ overlap, comprising ∼10-15% of all unique amino acid sequences and ∼5-10% of all unique nucleotide sequences across any two individuals, is already present at this early stage of T cell development. The majority of TCRβ sharing between individual thymocyte repertoires could be attributed to the process of convergent recombination, with additional contributions likely arising from recombinatorial biases; the role of selection during intrathymic development was negligible. These results indicate that the process of TCR gene recombination is the major determinant of clonotype sharing between individuals.

  10. N-(2-Mercaptoethyl)-1,3-Propanediamine (WR-1065) Protects Thymocytes from Programed Cell Death

    DTIC Science & Technology

    1992-03-15

    rldlation (1. 2) and gluIco- l). The pellet and supernatant DNA of unirradiated thy- corticoid 5reatmrnt 11,1 ol thymocytes. The pellet DNA mocytes...1065 showed no ionizing radiation. DNA damage occurs during irradiation cation-dependent DNA fragmentation (Table Ill), suggest- due to direct...Wolfe the induction of DNA damage In cultures Chinese hamster ovaryhelp In settin~g up the autoanalyzer. We thank W.Wle cella exposed to -y-radlatlon

  11. beta. -endorphin modulation of mitogen-stimulated calcium uptake by rat thymocytes

    SciTech Connect

    Hemmick, L.M.; Bidlack, J.M.

    1987-10-19

    Lymphocytes stimulated by mitogens or antigens exhibit an enhanced calcium uptake early in the proliferation or activation response. Modulation of this calcium uptake results in alterations of proliferation and immunocompetence. ..beta..-endorphin and other opioids affect several parameters of lymphocyte competence. Limited data are available concerning the mechanism(s) of these effects. This study examines whether a possible opioid mechanism is the modification of the early calcium influx into stimulated lymphocytes. The time course of both concanavalin A (Con A) and phytohemagglutinin (PHA)-stimulated /sup 45/Ca/sup 2 +/ uptake into thymocytes was characterized to determine the optimal time for testing the effects of opioids. BETA-Endorphin 1-31 significantly enhanced Con A-stimulated /sup 45/Ca/sup 2 +/ uptake into rat thymocytes. This peptide had no significant effect on PHA-simulated /sup 45/Ca/sup 2 +/ uptake or on basal thymocyte /sup 45/Ca/sup 2 +/ flux. The ..beta../sub h/-endorphin stimulatory effect was titratable in the range of 0.1 nM to 10 ..mu..M. Naloxone did not reverse the enhancement. Met-enkephalinamide and other opioid agonists did not duplicate the stimulatory effect. Thus, the ..beta../sub h/-endorphin 1-31 enhancement of Con A-stimulated /sup 45/Ca/sup 2 +/ uptake by rat thymocytes does not operate via classical opioid receptor mechanisms. ..beta../sub h/-endorphin 1-31 appears to be acting on a subset of T cells that are responsive to Con A but not to PHA. 30 references, 4 figures, 1 table.

  12. Macrophage activation syndrome: serological markers and treatment with anti-thymocyte globulin.

    PubMed

    Coca, Andreea; Bundy, Kemp W; Marston, Bethany; Huggins, Jennifer; Looney, R John

    2009-07-01

    Two patients presented at the University of Rochester Medical Center with a febrile illness, cytopenias, organ failure (liver failure or respiratory failure), and markedly elevated serum ferritin and sIL-2R. A diagnosis of probable macrophage activation syndrome was made. Both patients failed initial therapy with steroids and cyclosporine, either due to toxicity or lack of efficacy. Both patients responded dramatically to rabbit anti-thymocyte globulin (ATG).

  13. Expression of GTP-binding protein alpha subunits in human thymocytes.

    PubMed

    Kabouridis, P S; Waters, S T; Escobar, S; Stanners, J; Tsoukas, C D

    1995-03-09

    In this report, we investigate G protein alpha subunit diversity in human thymocytes, utilizing common properties shared by these genes and reverse transcription-polymerase chain reaction (RT-PCR). Sequence analysis of PCR amplified gene portions, indicate the presence of members from all four G-protein families that have been described thus far. The alpha subunit genes identified are: G alpha i1-3 and G alpha z but not G alpha o from the Gi family, G alpha s from the Gs family, G alpha 11, G alpha q, and G alpha 16 from the Gq family, and G alpha 12 and G alpha 13 from the G12 family. Also in this report we present the nucleotide and predicted amino acid sequences of the human G alpha 13 cloned from a thymocyte cDNA library. The sequence of the human G alpha 13 has not been previously reported. Comparison of this sequence with the reported murine G alpha 13 shows > 90% identity at the deduced amino acid sequence level. We conclude that thymocytes represent a useful experimental system for the study of G protein involvement in immune responses and lymphocyte development.

  14. Pre-treatment double- or triple-positive tumor markers are predictive of a poor outcome for patients undergoing radiofrequency ablation for hepatocellular carcinoma.

    PubMed

    Nitta, Hidetoshi; Nakagawa, Shigeki; Kaida, Takayoshi; Arima, Kota; Higashi, Takaaki; Taki, Katsunobu; Okabe, Hirohisa; Hayashi, Hiromitsu; Hashimoto, Daisuke; Chikamoto, Akira; Ishiko, Takatoshi; Beppu, Toru; Baba, Hideo

    2017-03-01

    We evaluated the therapeutic effect of radiofrequency ablation (RFA) on hepatocellular carcinoma (HCC) according to the number of positive tumor markers. The subjects of this study were 160 patients who underwent percutaneous and surgical RFA for HCC. Patients were divided into negative (n = 51), single- (n = 69), double- (n = 31), and triple-positive (n = 9) tumor marker groups according to the pre-treatment expression of these markers. We looked for any relationships among clinical parameters, outcomes, and tumor markers. The 3-year recurrence-free and overall survival rates of the negative, single-, double-, and triple-positive groups were 30, 19, 16, and 11 % (P = 0.02), and 94, 88, 67, and 37 % (P < 0.001), respectively. The 2-year local recurrence rates were 6.5, 0, 41.2, and 61.9 %, respectively (P < 0.001). Multivariate analysis revealed that a double- or triple-positive pre-treatment tumor marker profile was independently associated with local recurrence [hazard ratio (HR) 5.48, 95 % confidence interval (CI) 2.44-12.33, P < 0.001] and overall survival (HR 4.21, 95 % CI 1.89-9.37, P < 0.001). RFA may not be suitable for patients with HCC who have pre-treatment expression of ≥two of these tumor markers.

  15. Efficacy of rabbit anti-thymocyte globulin for steroid-resistant acute rejection after liver transplantation

    PubMed Central

    Lee, Jae Geun; Lee, Juhan; Lee, Jung Jun; Song, Seung Hwan; Ju, Man Ki; Choi, Gi Hong; Kim, Myoung Soo; Choi, Jin Sub; Kim, Soon Il; Joo, Dong Jin

    2016-01-01

    Abstract Acute cellular rejection after liver transplantation (LT) can be treated with steroid pulse therapy, but there is no ideal treatment for steroid-resistant acute rejection (SRAR). We aimed to determine the feasibility and potential complications of rabbit anti-thymocyte globulin (rATG) application to treat SRAR in liver transplant recipients. We retrospectively reviewed medical records of 429 recipients who underwent LT at Severance Hospital between January 2010 and March 2015. We compared clinical features and graft survival between patients with steroid-sensitive acute rejection (SSAR; n = 23) and SRAR (n = 11). We also analyzed complications and changes in laboratory findings after 2.5 mg/kg rATG treatment in patients with SRAR for 6 to 10 days. There were no significant differences in gender, age, model for end-stage liver disease score, Child–Turcotte–Pugh score, or original liver diseases between patients with SSAR and SRAR, although deceased donors were more frequently associated with the SRAR group (P = 0.004). All SRAR patients responded positively to rATG treatment; after treatment, the patients’ median AST levels decreased from 138 to 63 IU/L, and their median ALT levels dropped from 327 to 70 IU/L 1 day after rATG treatment (P = 0.022 and 0.017, respectively). Median aspartate aminotransferase (AST), alanine aminotransferase (ALT), and total bilirubin levels significantly decreased 1 month post-treatment (P = 0.038, 0.004, and 0.041, respectively). Median survival after LT was 23 months, and median survival after rATG was 22 months in patients with SRAR. Adverse effects included hepatitis C virus (HCV) reactivation, fungemia, and cytomegalovirus (CMV) infection. Nine SRAR patients survived with healthy liver function, 1 died from a traffic accident during follow-up, and 1 died from graft-versus-host disease and fungemia. Administration of rATG is an effective therapeutic option for SRAR with acceptable complications in liver

  16. Simultaneous spatial and angular positioning of plane specular samples by a novel double beam triangulation probe with full auto-compensation

    NASA Astrophysics Data System (ADS)

    Makai, Janos P.

    2016-02-01

    The positioning of a plane specular sample to be measured or processed is an important requirement in many fields of research and industry. Where a sample is to be processed either by electromagnetic waves or a particle beam of higher numerical aperture the irradiance or the particle number over unit area is position and angle dependent. Where optical properties of a sample are to be measured, such as in spectrophotometry, these parameters can depend on the angle of incidence and on the value of the irradiance, i.e. on the angular and spatial position of the sample. In some cases parameters of many samples have to be compared among each other or to those of a standard, this also requires the highly accurate positioning of each sample to the same position. This paper describes a method that is suitable for high accuracy alignment of specular plane samples both angularly and spatially. It applies a double beam triangulation probe, where the second beam serves not only as a reference beam to compensate for any changes of the transmitting media and that of the laser but also doubles the sensitivity of the probe. The method does not compete with interferometric methods, it is required only in special applications, but provides an absolute uncertainty for spatial positioning in the sub-micrometer range and an angular one in the 0.0003° range. Furthermore, the accuracy is tunable by the parameters of the setup.

  17. Aire knockdown in medullary thymic epithelial cells affects Aire protein, deregulates cell adhesion genes and decreases thymocyte interaction.

    PubMed

    Pezzi, Nicole; Assis, Amanda Freire; Cotrim-Sousa, Larissa Cotrim; Lopes, Gabriel Sarti; Mosella, Maritza Salas; Lima, Djalma Sousa; Bombonato-Prado, Karina F; Passos, Geraldo Aleixo

    2016-09-01

    We demonstrate that even a partial reduction of Aire mRNA levels by siRNA-induced Aire knockdown (Aire KD) has important consequences to medullary thymic epithelial cells (mTECs). Aire knockdown is sufficient to reduce Aire protein levels, impair its nuclear location, and cause an imbalance in large-scale gene expression, including genes that encode cell adhesion molecules. These genes drew our attention because adhesion molecules are implicated in the process of mTEC-thymocyte adhesion, which is critical for T cell development and the establishment of central self-tolerance. Accordingly, we consider the following: 1) mTECs contribute to the elimination of self-reactive thymocytes through adhesion; 2) Adhesion molecules play a crucial role during physical contact between these cells; and 3) Aire is an important transcriptional regulator in mTECs. However, its role in controlling mTEC-thymocyte adhesion remains unclear. Because Aire controls adhesion molecule genes, we hypothesized that the disruption of its expression could influence mTEC-thymocyte interaction. To test this hypothesis, we used a murine Aire(+) mTEC cell line as a model system to reproduce mTEC-thymocyte adhesion in vitro. Transcriptome analysis of the mTEC cell line revealed that Aire KD led to the down-modulation of more than 800 genes, including those encoding for proteins involved in cell adhesion, i.e., the extracellular matrix constituent Lama1, the CAM family adhesion molecules Vcam1 and Icam4, and those that encode peripheral tissue antigens. Thymocytes co-cultured with Aire KD mTECs had a significantly reduced capacity to adhere to these cells. This finding is the first direct evidence that Aire also plays a role in controlling mTEC-thymocyte adhesion.

  18. Insights into the mechanisms of thymus involution and regeneration by modeling the glucocorticoid-induced perturbation of thymocyte populations dynamics.

    PubMed

    Moleriu, Radu Dumitru; Zaharie, Daniela; Moatar-Moleriu, Lavinia Cristina; Gruia, Alexandra Teodora; Mic, Ani Aurora; Mic, Felix Aurel

    2014-05-07

    T-cells develop in the thymus and based on CD4 and CD8 expressions there are four main thymocyte populations in a normal mouse thymus. Currently, there are several mathematical models that describe the dynamics of thymocyte populations in a normal thymus, but only a few of them model the transient perturbation of their homeostasis. Our aim is to model the perturbation in the dynamics of each thymocyte population which is induced by the administration of a glucocorticoid, i.e. dexamethasone. The proposed approach relies on extending a four compartment thymus model based on differential equations by adding perturbation terms either globally (at the level of each equation) or locally (at the level of proliferation, death, and transfer rates). By fitting the perturbed model with experimental data on mice thymi collected before and after the administration of dexamethasone, it was possible to estimate the relevant parameters using a population-based stochastic search method. The fitted model is further used to conduct a quantitative analysis on the differentiated impact of dexamethasone on each T-cell population and on proliferation, death, and transfer processes. The obtained quantitative information on the perturbation could be used to explore and modify the flow of thymocytes between thymus compartments in order to elucidate the mechanisms of thymus involution and its subsequent regeneration. Since glucocorticoids are raised in many pathological situations, such a model could be useful in evaluating the impact of diseases on thymocyte dynamics in the thymus.

  19. Binding of anti-double stranded (ds) DNA-positive sera to denatured (d) DNA and synthetic poly[dA-dT] x poly[dA-dT] double stranded copolymer in an ELISA format.

    PubMed

    Batinić, D; Bozićević, M; Krstulović, A; Bosnić, D; Sentić, M; Markeljević, J; Malenica, B; Cikes, N; Marusić, M

    1996-04-01

    Using an ELISA assay anti-nuclear antibody-positive sera from 300 patients with various immune-related diseases and 64 anti-nuclear antibody-negative sera were analysed for binding to S1-nuclease-treated double stranded (ds) DNA. In addition, the pattern of reactivity of 50 selected anti-dsDNA-positive sera was established using denatured (d) DNA and poly[dA-dT] X poly[dA-dT] double-stranded alternating copolymer (dAT) as additional DNA antigens. None of the 64 anti-nuclear antibody-negative sera and 76 of the 300 anti-nuclear antibody-positive sera (25%) were anti-dsDNA-positive. Of the anti-nuclear antibody-positive and anti-dsDNA-positive sera, 48 (63%) were from systemic lupus erythematosus patients, and 7 (9%) from rheumatoid arthritis patients, whereas 21 patients (27.6%) suffered from various immune and non-immune related diseases. Anti-dsDNA-positive reactivity was highly correlated with dDNA and dAT reactivity (r = 0.906, p < 0.0001 and r = 0.93, p < 0.0001, respectively). Although the majority of the 50 selected (37 systemic lupus erythematosus and 13 non-systemic lupus erythematosus) anti-dsDNA-positive sera concomitantly bound to both additional antigens, 7 of these (14%) did not bind to dAT, and 2 (4%) did not bind to dDNA. Anti-dsDNA-positive sera (n = 37) showed a similar pattern, in which 8.1% and 2.7% of sera did not bind to dAT and to dDNA, respectively. In contrast, anti-dsDNA-negative sera from various immune-related diseases bound either ssDNA (12.5%) or dDNA and dAT (12.5%). These data suggest that dsDNA and dAT-based assays detect similar but not identical specificities in the sera of patients suffering from systemic lupus erythematosus and in a proportion of non-systemic lupus erythematosus patients.

  20. Mysm1 is required for interferon regulatory factor expression in maintaining HSC quiescence and thymocyte development

    PubMed Central

    Huang, X F; Nandakumar, V; Tumurkhuu, G; Wang, T; jiang, X; Hong, B; Jones, L; Won, H; Yoshii, H; Ozato, K; Masumi, A; Chen, S-Y

    2016-01-01

    Mysm1−/− mice have severely decreased cellularity in hematopoietic organs. We previously revealed that Mysm1 knockout impairs self-renewal and lineage reconstitution of HSCs by abolishing the recruitment of key transcriptional factors to the Gfi-1 locus, an intrinsic regulator of HSC function. The present study further defines a large LSKs in >8-week-old Mysm1−/− mice that exhibit increased proliferation and reduced cell lineage differentiation compared with those of WT LSKs. We found that IRF2 and IRF8, which are important for HSC homeostasis and commitment as transcription repressors, were expressed at lower levels in Mysm1−/− HSCs, and Mysm1 enhanced function of the IRF2 and IRF8 promoters, suggesting that Mysm1 governs the IRFs for HSC homeostasis. We further found that the lower expressions of IRF2 and IRF8 led to an enhanced transcription of p53 in Mysm1−/− HSCs, which was recently defined to have an important role in mediating Mysm1−/−-associated defects. The study also revealed that Mysm1−/− thymocytes exhibited lower IRF2 expression, but had higher Sca1 expression, which has a role in mediating thymocyte death. Furthermore, we found that the thymocytes from B16 melanoma-bearing mice, which display severe thymus atrophy at late tumor stages, exhibited reduced Mysm1 and IRF2 expression but enhanced Sca1 expression, suggesting that tumors may downregulate Mysm1 and IRF2 for thymic T-cell elimination. PMID:27277682

  1. DR3 Regulates Negative Selection during Thymocyte Development

    PubMed Central

    Wang, Eddie C. Y.; Thern, Anette; Denzel, Angela; Kitson, Jeremy; Farrow, Stuart N.; Owen, Michael J.

    2001-01-01

    DR3 (Ws1, Apo3, LARD, TRAMP, TNFSFR12) is a member of the death domain-containing tumor necrosis factor receptor (TNFR) superfamily, members of which mediate a variety of developmental events including the regulation of cell proliferation, differentiation, and apoptosis. We have investigated the in vivo role(s) of DR3 by generating mice congenitally deficient in the expression of the DR3 gene. We show that negative selection and anti-CD3-induced apoptosis are significantly impaired in DR3-null mice. In contrast, both superantigen-induced negative selection and positive selection are normal. The pre-T-cell receptor-mediated checkpoint, which is dependent on TNFR signaling, is also unaffected in DR3-deficient mice. These data reveal a nonredundant in vivo role for this TNF receptor family member in the removal of self-reactive T cells in the thymus. PMID:11313471

  2. [Change in glutathione content in rat thymocytes under apoptosis induced by H2O2 or irradiation].

    PubMed

    Koval', T V; Nazarova, O O; Matyshevs'ka, O P

    2008-01-01

    Glutathione (GSH) content as well as GSH-peroxidase and GSH-reductase activity in isolated rat thymocytes X-irradiated in a dose of 4.5 Gy or treated with 0.1 mM H2O2 were studied in a period preceding the appearance of apoptosis morphological symptoms. The early adaptive response of thymocytes to radiation - increase of both GSH content and glutathione peroxidase and glutathione reductase activity was revealed. On the contrary the rapid fall of GSH level in H2O2-treated thymocytes was observed simultaneousely with glutathione reductase inhibition and enhanced GSH consumption by glutathione peroxidase, this disbalance of GSH-dependent antioxidant system probably facilitates mitochondrial way of apoptosis.

  3. Lck regulates the tyrosine phosphorylation of the T cell receptor subunits and ZAP-70 in murine thymocytes

    PubMed Central

    1996-01-01

    The Src-family and Syk/ZAP-70 family of protein tyrosine kinases (PTK) are required for T cell receptor (TCR) functions. We provide evidence that the Src-family PTK Lck is responsible for regulating the constitutive tyrosine phosphorylation of the TCR zeta subunit in murine thymocytes. Moreover, ligation of the TCR expressed on thymocytes from Lck-deficient mice largely failed to induce the phosphorylation of TCR- zeta, CD3 epsilon, or ZAP-70. In contrast, we find that the TCR-zeta subunit is weakly constitutively tyrosine phosphorylated in peripheral T cells isolated from Lck-null mice. These data suggest that Lck has a functional role in regulation of TCR signal transduction in thymocytes. In peripheral T cells, other Src-family PTKs such as Fyn may partially compensate for the absence of Lck. PMID:8642247

  4. Single- and double-difference algorithms for position and time-delay calibration of transducer-elements in a sparse array.

    PubMed

    Li, Yue; Sharp, Ian; Hedley, Mark; Ho, Phil; Guo, Y Jay

    2007-06-01

    A method for the calibration of the position and time delay of transducer elements in a large, sparse array used for underwater, high-resolution, ultrasound imaging has been described in a previous work. This algorithm is based on the direct algorithm used in the global positioning system (GPS), but the wave propagation speed is treated as one of the to-be-calibrated parameters. In this article, the performance of two other commonly used GPS algorithms, namely the single-difference algorithm and the double-difference algorithm, is evaluated. The calibration of the propagation speed also is integrated into these two algorithms. Furthermore, a novel, least-squares method is proposed to calibrate the time delay associated with each transducer element for these two algorithms. The performances of these algorithms are theoretically analyzed and evaluated using numerical analysis and simulation study. The performance of the direct algorithm, the single-difference algorithm, and the double-difference algorithm is compared. It was found that the single-difference algorithm has the best performance among the three algorithms for the current application, and it is capable of calibrating the position and time delay of transducer elements to an accuracy of one-tenth of a wavelength.

  5. Collision-induced dissociation of fatty acid [M - 2H + Na]- ions: charge-directed fragmentation and assignment of double bond position.

    PubMed

    Thomas, Michael C; Altvater, Jens; Gallagher, Thomas J; Nette, Geoffrey W

    2014-11-01

    The collision-induced dissociation (CID) of cationic fatty acid-metal ion complexes has been extensively studied and, in general, provides rich structural information. In particular, charge-remote fragmentation processes are commonly observed allowing the assignment of double bond position. In a previous manuscript, we presented two methods to doubly deprotonate polyunsaturated fatty acids to form anionic fatty acid-sodium ion complexes, referred to as [M - 2H + Na] (-) ions. In the current manuscript, the CID behavior of these [M - 2H + Na] (-) ions is investigated for the first time. Significantly, we also present a deuterium-labeling experiment, which excludes the possibility that deprotonation occurs predominately at the α-carbon in the formation of fatty acid [M - H + NaF](-) ions. This supports our original proposal where deprotonation occurs at the bis-allylic positions of polyunsaturated fatty acids. CID spectra of polyunsaturated fatty acid [M - 2H + Na](-) ions display abundant product ions arising from acyl chain cleavages. Through the examination of fatty acid isomers, it is demonstrated that double bond position may be unequivocally determined for methylene-interrupted polyunsaturated fatty acids with three or more carbon-carbon double bonds. In addition, CID of [M - 2H + Na](-) ions was applied to 18:3 isomers of Nannochloropsis oculata and three isomers were tentatively identified: ∆(9,12,15)18:3, ∆(6,9,12)18:3, and ∆(5,8,11)18:3. We propose that structurally-informative product ions are formed via charge-driven fragmentation processes at the site of the resonance-stabilized carbanion as opposed to charge-remote fragmentation processes, which could be inferred if deprotonation occurred predominately at the α-carbon.

  6. Effects of selected polybrominated diphenyl ether flame retardants on lake trout (Salvelinus namaycush) thymocyte viability, apoptosis, and necrosis

    USGS Publications Warehouse

    Birchmeier, Kelly L.; Smith, Kimberly A.; Passino-Reader, Dora R.; Sweet, Leonard I.; Chernyak, Sergei M.; Adams, Jean V.; Omann, Geneva M.

    2005-01-01

    Polybrominated diphenyl ether (PBDE) flame-retardants have been identified as an emergent contaminants issue in many parts of the world. In vitro analyses were conducted to test the hypothesis that selected PBDEs congeners affect viability, apoptosis, and necrosis of thymocytes from laboratory-reared lake trout (Salvelinus namaycush). At current environmental levels (<1 mg/L), effects of the tested PBDEs on thymocytes were negligible. However, at 100 mg/L, major effects were seen for congener brominated diphenyl ether 47 (BDE-47) and minor effects were seen for congener BDE-99.

  7. [Combined simulated weightlessness and noise affect cell cycles and composition in rat thymocytes].

    PubMed

    Si, Shaoyan; Song, Shujun; Hua, Nan; Han, Haolun; Xu, Bingxin; Wang, Gang; Zhang, Chi; Wu, Wei

    2016-03-01

    To observe the combined effects of simulated weightlessness and noise on the cell cycles of thymocytes and cell compositions of thymus in rats and to explore the possible mechanism of immune function depression in space flight. SD rats were stimulated by simulated weightlessness and/or noise. On the 3rd, 7th and 14th day, the rats were weighed and then killed. The thymuses were taken, weighed and cell suspensions were made. Cell cycles and compositions in thymocytes were analyzed by flow cytometry. Compared with control group, the weights of rats were reduced in combined group and simulated weightlessness group, but the weights of rats increased or did not change in noise group. Rats in the three groups showed thymus atrophy. The ratio of cells increased in G0/G1 phase and decreased in S and G2/M phases. The ratios of CD4(-) CD8(-), CD4(+) CD8(-) and CD4(-) CD8(+) T lymphocytes increased and CD4(+) CD8(+) T lymphocytes decreased. However, these changes occurred at different time points in different groups. Simulated weightlessness and noise have significant effects on thymus, but the severity are different. The combined factors have superimposed effects. Maybe this is one of the reasons for depressed functions of T lymphocytes in space flight.

  8. Control of cell-cycle-associated tetrahydrobiopterin synthesis in rat thymocytes.

    PubMed

    Schott, K; Brand, K; Hatakeyama, K; Kagamiyama, H; Maier, J; Werner, T; Ziegler, I

    1992-05-01

    The cell-cycle progression of rat thymocytes from G0 through G1 to DNA synthesis is associated with a transient synthesis of H4biopterin, the concentration of which reaches a maximum at the time of S-phase entry and then decreases. This synthesis of H4biopterin is controlled by the specific activity of GTP cyclohydrolase I, which peaks in G1/S cells. In contrast, the catalytic activity of sepiapterin reductase remains constant throughout the cell-cycle. At G0 the steady state mRNA levels specific for GTP cyclohydrolase I and sepiapterin reductase, respectively, are below the limits of detection. Both accumulate as the thymocytes progress through the cell-cycle but lack cyclic down regulation. The data indicate that the variations in H4biopterin synthesis during the cell-cycle are caused by growth regulated increase in GTP cyclohydrolase I mRNA expression, with subsequent post-translational inactivation. This latter is likely due to the degree of enzyme phosphorylation.

  9. The effect of dichloroacetate on male rat thymus and on thymocyte cell cycle.

    PubMed

    Stanevičiūtė, Jurate; Urbonienė, Daiva; Valančiūtė, Angelija; Balnytė, Ingrida; Vitkauskienė, Astra; Grigalevičienė, Brigita; Stakišaitis, Donatas

    2016-12-01

    The study aim was to investigate the effect of dichloroacetate (DCA) on thymus and the thymocyte cycle in rats. Wistar male gonad-intact and castrated rats (4-5 weeks) were investigated in the following groups: (1) control; (2) treated with DCA; and (3) treated with the DCA and sodium valproate (NaVP) combination. Rats were treated for 4 weeks with DCA 200 mg/kg/day alone and 300 mg/kg/day of NaVP plus 200 mg/kg/day of DCA (every second week, beginning with NaVP). After the experiment, the thymus was weighted, and the thymus lobe was taken for thymocyte flow cytometry. In gonad-intact rats, the thymus weight of the control was higher than in rats treated with DCA (P <0.001) or with the NaVP-DCA combination (P <0.04); a comparison of thymus weight between DCA- and NaVP-DCA-treated groups revealed a higher thymus weight loss in the DCA-treated group (P <0.03). Flow cytometry shows that DCA treatment increased the percentage of cells in the G2-M phase (P <0.03) and reduced in G1-G0 (P <0.02). The DCA treatment effect was determined only in gonad-intact but not in castrated rats. The authors discuss the possible DCA and NaVP interaction mechanisms. © The Author(s) 2016.

  10. The Effect of Ascorbic Acid on Mancozeb-Induced Toxicity in Rat Thymocytes.

    PubMed

    Pavlovic, V; Cekic, S; Kamenov, B; Ciric, M; Krtinic, D

    2015-01-01

    Mancozeb, as a dithiocarbamate fungicide, has been found to exhibit toxicological manifestations in different cells, mainly by generation of free radicals which may alter antioxidant defence systems in cells. The effect of mancozeb on the cells of a primary lymphoid organ has not been studied. In the present study, the effects of mancozeb (0.2, 2 and 5 μg/ml) or mancozeb+ascorbic acid (100 μg/ml), or ascorbic acid alone or control medium alone on the levels of cell viability, apoptosis, intracellular reactive oxygen species production (ROS), mitochondrial membrane potential (MMP) and ATP levels in rat thymocytes were examined in vitro. Cells treated with mancozeb displayed a concentration-dependent increase of hypodiploid cells and ROS production followed by markedly decreased viability of the cells, MMP and ATP levels. Application of ascorbic acid significantly reduced cytotoxicity in cell cultures treated with 0.2 and 2 μg/ml of mancozeb, together with significantly decreased ROS levels and increased MMP and ATP levels. In cells treated with 5 μg/ml of mancozeb, ascorbic acid failed to reduce toxicity while simultaneously increasing the apoptosis rate of thymocytes. These results suggest that ROS plays a significant role in mancozeb-induced toxicity, through alteration of mitochondrial function. Ascorbic acid administration reduced the toxicity rate in cells treated with lower mancozeb concentrations, while it may have the ability to shift cells from necrosis to apoptosis in the presence of highest mancozeb concentrations.

  11. Pseudomonas aeruginosa and Its Bacterial Components Influence the Cytokine Response in Thymocytes and Splenocytes

    PubMed Central

    Zimmermann, Corinna; Mausberg, Anne K.; Dehmel, Thomas; Kieseier, Bernd C.; Hartung, Hans-Peter; Hofstetter, Harald H.

    2016-01-01

    Infections with Pseudomonas aeruginosa may cause many different diseases. The spectrum of such infections in general includes inflammation and bacterial sepsis. Hospital-acquired pneumonia, naturally resistant to a wide range of antibiotics, is associated with a particularly high mortality rate in mechanically ventilated patients. The pathogenesis of P. aeruginosa is complex and mediated by several virulence factors, as well as cell-associated factors. We have previously demonstrated that stimulation with different bacteria triggers the cytokine response of thymocytes. In this study, we investigated the effect of P. aeruginosa and its different components on the cytokine production of immature and mature immune cells. We found that the induced cytokine pattern in the thymus and the spleen after infections with P. aeruginosa is primarily mediated by lipopolysaccharide (LPS) of the outer cell membrane, but other components of the bacterium can influence the cytokine secretion as well. Stimulation with heat-killed P. aeruginosa and LPS does not influence the amount of cytokine-producing CD4+ T cells but instead suppresses the emergence of Th17 cells. However, stimulation with P. aeruginosa or its components triggers the interleukin-17 (IL-17) response both in thymocytes and in splenocytes. We conclude that infections with P. aeruginosa affect the cytokine secretion of immature and mature cells and that IL-17 and Th17 cells play only a minor role in the development of pathological systemic inflammatory disease conditions during P. aeruginosa infections. Therefore, other inflammatory immune responses must be responsible for septic reactions of the host. PMID:26902726

  12. Benidipine persistently inhibits delayed rectifier K(+)-channel currents in murine thymocytes.

    PubMed

    Kazama, Itsuro; Maruyama, Yoshio; Matsubara, Mitsunobu

    2013-02-01

    Lymphocytes predominantly express delayed rectifier K(+)-channels (Kv1.3) in their plasma membranes, and the channels play crucial roles in the lymphocyte activation and proliferation. Since 1,4-dihydropyridine (DHP) Ca(2+) channel blockers (CCBs), which are highly lipophilic, exert relatively stronger immunomodulatory effects than the other types of CCBs, they would affect the Kv1.3-channel currents in lymphocytes. In the present study, employing the standard patch-clamp whole-cell recording technique in murine thymocytes, we examined the effects of benidipine, one of the most lipophilic DHPs, on the channel currents and the membrane capacitance and compared them with those of nifedipine. Both drugs significantly suppressed the peak and the pulse-end currents of the channels with significant decreases in the membrane capacitance. However, the effects of benidipine were more marked than those of nifedipine and were irreversible after the drug withdrawal. This study demonstrated for the first time that DHP CCBs, such as nifedipine and benidipine, exert inhibitory effects on thymocyte Kv1.3-channel currents. The persistent effect of benidipine was thought to be associated with its sustained accumulation in the plasma membranes as detected by the long-lasting decrease in the membrane capacitance.

  13. The mitochondrial and death receptor pathways involved in the thymocytes apoptosis induced by aflatoxin B1

    PubMed Central

    Chi, Xiaofeng; Li, Xiaochong; Jiang, Min; Fang, Jing; Cui, Hengmin; Lai, Weimin; Zhou, Yi; Zhou, Shan

    2016-01-01

    Aflatoxin B1 (AFB1) is a potent immunosuppressive agent in endotherms, which can be related to the up-regulated apoptosis of immune organs. In this study, we investigated the roles of the mitochondrial, death receptor, and endoplasmic reticulum pathways in Aflatoxin B1 induced thymocytes apoptosis. Chickens were fed an aflatoxin B1 containing diet (0.6 mg/kg AFB1) for 3 weeks. Our results showed that (1) AFB1 diet induced the decrease of T-cell subsets, morphological changes, and excessive apoptosis of thymus. (2) The excessive apoptosis involved the mitochondrial pathway (up-regulation of Bax, Bak, cytC and down-regulation of Bcl-2 and Bcl-xL) and death receptor pathway (up-regulation of FasL, Fas and FADD). (3) Oxidative stress, an apoptosis inducer, was confirmed in the thymus. In conclusion, this is the first study to demonstrate that mitochondrial and death receptor pathways involved in AFB1 induced thymocytes apoptosis in broilers. PMID:26933817

  14. Vital roles of mTOR complex 2 in Notch-driven thymocyte differentiation and leukemia.

    PubMed

    Lee, Keunwook; Nam, Ki Taek; Cho, Sung Hoon; Gudapati, Prathyusha; Hwang, Yoonha; Park, Do-Sim; Potter, Ross; Chen, Jin; Volanakis, Emmanuel; Boothby, Mark

    2012-04-09

    Notch plays critical roles in both cell fate decisions and tumorigenesis. Notch receptor engagement initiates signaling cascades that include a phosphatidylinositol 3-kinase/target of rapamycin (TOR) pathway. Mammalian TOR (mTOR) participates in two distinct biochemical complexes, mTORC1 and mTORC2, and the relationship between mTORC2 and physiological outcomes dependent on Notch signaling is unknown. In this study, we report contributions of mTORC2 to thymic T-cell acute lymphoblastic leukemia (T-ALL) driven by Notch. Conditional deletion of Rictor, an essential component of mTORC2, impaired Notch-driven proliferation and differentiation of pre-T cells. Furthermore, NF-κB activity depended on the integrity of mTORC2 in thymocytes. Active Akt restored NF-κB activation, a normal rate of proliferation, and differentiation of Rictor-deficient pre-T cells. Strikingly, mTORC2 depletion lowered CCR7 expression in thymocytes and leukemic cells, accompanied by decreased tissue invasion and delayed mortality in T-ALL driven by Notch. Collectively, these findings reveal roles for mTORC2 in promoting thymic T cell development and T-ALL and indicate that mTORC2 is crucial for Notch signaling to regulate Akt and NF-κB.

  15. Susceptibility of neonatal T cells and adult thymocytes to peripheral tolerance to allogeneic stimuli

    PubMed Central

    do Canto, Fábio B; Lima, Celso; Teixeira, Ivan A; Bellio, Maria; Nóbrega, Alberto; Fucs, Rita

    2008-01-01

    We studied the tolerization of neonatal thymocytes (NT), neonatal splenocytes (NS) and adult thymocytes (AT), transferred to syngeneic nude (nu/nu) hosts previously injected with semi-allogeneic splenocytes, without any supportive immunosuppressive treatment. This protocol allows the study of peripheral tolerance in the absence of the thymus. BALB/c neonatal T cells and ATs were able to expand in syngeneic BALB/c nu/nu mice and functionally reconstituted an allogeneic response, rejecting (BALB/c × B6.Ba) F1 splenocytes transferred 3–4 weeks after injection of BALB/c cells. However, if (BALB/c × B6.Ba) F1 cells were injected into BALB/c nude hosts 30 days before transfer of NT, NS or AT cells, the F1 population was preserved and specific tolerance to B6 allografts was established. Furthermore, transfer to lymphopenic F1 nu/nu showed that tolerance could be established only for neonatal populations, showing that unique properties of neonatal T cells allow their tolerization in both lymphopenic and non-lymphopenic conditions, in the absence of suppressive immunotherapy. These results bring empirical support to the possibility of T-cell engraftment in immunodeficient patients showing partial identity with donor major histocompatibility complex (MHC) genes; the manipulation of immunological maturity of donor T cells may be the key for successful reconstitution of immunocompetence without induction of graft-versus-host disease. PMID:18462348

  16. Temporal increase in thymocyte negative selection parallels enhanced thymic SIRPα(+) DC function.

    PubMed

    Kroger, Charles J; Wang, Bo; Tisch, Roland

    2016-10-01

    Dysregulation of negative selection contributes to T-cell-mediated autoimmunity, such as type 1 diabetes. The events regulating thymic negative selection, however, are ill defined. Work by our group and others suggest that negative selection is inefficient early in ontogeny and increases with age. This study examines temporal changes in negative selection and the thymic DC compartment. Peptide-induced thymocyte deletion in vivo was reduced in newborn versus 4-week-old NOD mice, despite a similar sensitivity of the respective thymocytes to apoptosis induction. The temporal increase in negative selection corresponded with an elevated capacity of thymic antigen-presenting cells to stimulate T cells, along with altered subset composition and function of resident DC. The frequency of signal regulatory protein α(+) (SIRPα(+) ) and plasmacytoid DCs was increased concomitant with a decrease in CD8α(+) DC in 4-week-old NOD thymi. Importantly, 4-week-old versus newborn thymic SIRPα(+) DC exhibited increased antigen processing and presentation via the MHC class II but not class I pathway, coupled with an enhanced T-cell stimulatory capacity not seen in thymic plasmacytoid DC and CD8α(+) DC. These findings indicate that the efficiency of thymic DC-mediated negative selection is limited early after birth, and increases with age paralleling expansion of functionally superior thymic SIRPα(+) DC. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  17. Change in plasma membrane potential of rat thymocytes by tert-butylhydroquinone, a food additive: Possible risk on lymphocytes.

    PubMed

    Takeda, Maki; Oyama, Keisuke; Kamemura, Norio; Kanemaru, Kaori; Yuasa, Keizo; Yokoigawa, Kumio; Oyama, Yasuo

    2017-09-09

    Tertiary butylhydroquinone (TBHQ) is a food additive and has various beneficial actions under in vitro and in vivo experimental conditions. Therefore, it is necessary to collect additional data on the toxicity of TBHQ in order to avoid adverse effects during clinical applications. Changes in plasma membrane potential are associated with changes in physiological functions even in non-excitable cells such as lymphocytes. Thus, compounds that affect membrane potential may modify some lymphocytic functions. The effect of TBHQ on plasma membrane potential was examined in rat thymocytes using flow cytometric techniques. Treatment of rat thymocytes with TBHQ caused hyperpolarization and then depolarization. The TBHQ-induced hyperpolarization was due to the activation of Ca(2+)-dependent K(+) channels. TBHQ elevated intracellular Ca(2+) levels. The depolarization by TBHQ was caused by a nonspecific increase in membrane ionic permeability. Both the sustained depolarization and elevation of intracellular Ca(2+) level by TBHQ are thought to be adverse for thymocytes because such changes disturb membrane and intracellular signaling. The thymus is most active during neonatal and pre-adolescent periods. If TBHQ exerts adverse actions on thymocytes, it may result in an immunotoxic effect in neonates and adolescents. Copyright © 2017 Elsevier Ltd. All rights reserved.

  18. Thymic Nurse Cells Exhibit Epithelial Progenitor Phenotype and Create Unique Extra-Cytoplasmic Membrane Space for Thymocyte Selection

    PubMed Central

    Hendrix, Tonya M.; Chilukuri, Rajendra V.E.; Martinez, Marcia; Olushoga, Zachariah; Blake, Andrew; Brohi, Moazzam; Walker, Christopher; Samms, Michael; Guyden, Jerry C.

    2010-01-01

    Thymic nurse cells (TNCs) are epithelial cells in the thymic cortex that contain as many as fifty thymocytes within specialized cytoplasmic vacuoles. The function of this cell-in-cell interaction has created controversy since their discovery in 1980. Further, some skepticism exists about the idea that apoptotic thymocytes within the TNC complex result from negative selection, a process believed to occur exclusively within the medulla. In this report, we have microscopic evidence that defines a unique membranous environment wherein lipid raft aggregates around the αβTCR expressed on captured thymocytes and class II MHC molecules expressed on TNCs. Further, immunohistological examination of thymic sections show TNCs located within the cortico-medullary junction to express cytokeratins five and eight (K5 and K8), and the transcription factor Trp-63, the phenotype defined elsewhere as the thymic epithelial progenitor subset. Our results suggest that the microenvironment provided by TNCs plays an important role in thymocyte selection as well as the potential for TNCs to be involved in the maintenance of thymic epithelia. PMID:20035931

  19. Increased Fetal Thymocytes Apoptosis Contributes to Prenatal Nicotine Exposure-induced Th1/Th2 Imbalance in Male Offspring Mice

    PubMed Central

    Chen, Ting; Yan, You-e; Liu, Sha; Liu, Han-xiao; Yan, Hui-yi; Hou, Li-fang; Qu, Wen; Ping, Jie

    2016-01-01

    Nicotine, a definite risk factor during pregnancy, is an immunomodulator. This study was designed to investigate the effects of prenatal nicotine exposure (PNE) on the balance of Th1/Th2 in offspring, and further explore the developmental origin mechanisms from the perspective of fetal thymocytes apoptosis. Pregnant Balb/c mice were administered 1.5 mg/kg nicotine subcutaneously twice per day from gestational day (GD) 9 to GD18. Results showed that PNE could cause a Th2 shift in male offspring, manifested as increased ratio of IgG1/IgG2a, IL-4 production in serum, and IL-4/IFN-γ expression ratio in spleen. Increased apoptosis of total thymocytes and CD4SP and reduced cell proportion of CD4SP were found in PNE male offspring on postnatal day (PND) 14 and PND 49. In the fetuses, decreased body weight and organ index of fetal thymus, histological changes in fetal thymus, reduced CD4SP proportion and increased fetal thymocyte apoptosis were observed in nicotine group. The increased mRNA expression of genes involved in Fas-mediated apoptotic pathway and protein expression of Fas were also detected. In conclusion, PNE could cause a Th2 shift in male offspring mediated by reduced CD4+ T cells output, which may result from the increasing apoptosis of total thymocytes and CD4SP. PMID:27976742

  20. Lipids of heliobacteria are characterised by a high proportion of monoenoic fatty acids with variable double bond positions.

    PubMed

    Aase, B; Jantzen, E; Bryn, K; Ormerod, J

    1994-07-01

    The fatty acid composition and lipid pattern of six strains of heliobacteria have been analysed. The results were fairly uniform for all strains. Phosphatidyl ethanolamine and phosphatidyl glycerol were the dominating lipids found, with the former as the major one. No glycolipids were detected. The general fatty acid pattern was dominated by acids of chain length C16 to C18. An unusually large proportion of monoenoic acids was seen, with up to four positional isomers for each chain length. Methyl branched (iso) fatty acids were present, but not cyclopropyl or hydroxy fatty acids nor fatty alcohols.

  1. A positive but complex association between meiotic double-strand break hotspots and open chromatin in Saccharomyces cerevisiae.

    PubMed

    Berchowitz, Luke E; Hanlon, Sean E; Lieb, Jason D; Copenhaver, Gregory P

    2009-12-01

    During meiosis, chromatin undergoes extensive changes to facilitate recombination, homolog pairing, and chromosome segregation. To investigate the relationship between chromatin organization and meiotic processes, we used formaldehyde-assisted isolation of regulatory elements (FAIRE) to map open chromatin during the transition from mitosis to meiosis in the budding yeast Saccharomyces cerevisiae. We found that meiosis-induced opening of chromatin is associated with meiotic DSB hotpots. The positive association between open chromatin and DSB hotspots is most prominent 3 h into meiosis, when the early meiotic genes DMC1 and HOP1 exhibit maximum transcription and the early recombination genes SPO11 and RAD51 are strongly up-regulated. While the degree of chromatin openness is positively associated with the occurrence of recombination hotspots, many hotspots occur outside of open chromatin. Of particular interest, many DSB hotspots that fell outside of meiotic open chromatin nonetheless occurred in chromatin that had recently been open during mitotic growth. Finally, we find evidence for meiosis-specific opening of chromatin at the regions adjacent to boundaries of subtelomeric sequences, which exhibit specific crossover control patterns hypothesized to be regulated by chromatin.

  2. Estimates for the sensitivity and false-positive rates for second trimester serum screening for Down syndrome and trisomy 18 with adjustment for cross-identification and double-positive results.

    PubMed

    Benn, P A; Ying, J; Beazoglou, T; Egan, J F

    2001-01-01

    cross-identification and double-positives should be helpful for pre-test counseling and the evaluation of serum screening programs.

  3. Cutting Edge: Ikaros null thymocytes mature into the CD4 lineage with reduced TCR signal: A study using CD3{zeta} immunoreceptor tyrosine-based activation motif transgenic mice.

    PubMed

    Urban, Julie A; Brugmann, William; Winandy, Susan

    2009-04-01

    Positive selection is a critical T cell developmental checkpoint that is driven by TCR signals. Enhanced positive selection toward the CD4 lineage occurs in the absence of Ikaros. One explanation for this phenotype is that Ikaros establishes the TCR signaling threshold that must be overcome for positive selection to occur. In the current study, this possibility is explored through the use of CD3zeta ITAM transgenic mice that express a CD3 zeta-chain with zero, one, or three ITAMs and an MHC class II (DO11.10)- or MHC class I (H-Y)-restricted TCR transgene. Using this system, we demonstrate that in the absence of Ikaros, thymocytes are able to mature into the CD4 lineage with reduced TCR signaling potential compared with that required to drive the maturation of wild-type thymocytes. We also demonstrate that maturation into the CD8 lineage is enhanced under conditions of reduced TCR signaling potential in the absence of Ikaros.

  4. Treatment with anti-LFA-1 alpha monoclonal antibody selectively interferes with the maturation of CD4- 8+ thymocytes.

    PubMed

    Revilla, C; González, A L; Conde, C; López-Hoyos, M; Merino, J

    1997-04-01

    Maturation of T lymphocytes in the thymus is driven by signals provided by soluble factors and by the direct interaction between thymocytes and stromal cells. Although the interaction between T-cell receptor (TCR) and major histocompalibility complex (MHC) molecules on stromal cells is crucial for T-cell development, other accessory molecules seem to play a role in this process. In order to better understand the role of lymphocyte function-associated antigen-1 (LFA-1) and intercellular adhesion molecule-1 (ICAM-1) molecules in thymocyte maturation, mice were treated from birth with saturating doses of non-cytolytic-specific monoclonal antibodies. The effect of this treatment on thymocyte subpopulations and the expression of CD3 and TCR-alpha beta by these cells was investigated by flow cytometry. Our data demonstrated that the effective saturation of LFA-1 alpha chain in the thymus, but not ICAM-I or LFA-I beta chain, selectively interfered with the maturation of CD8+ T cells, as manifested by a marked reduction in the frequency of CD4-8+ thymocytes expressing high levels of CD3 and TCR-alpha beta. This selective reduction was also observed in peripheral blood mononuclear cells and spleen cells. The analysis of the frequencies of various V beta TCR showed that CD4-8+ thymocytes were globally affected by the treatment. These results underline the importance of the interaction between LFA-1 and its ligands in the maturation of CD8+ T cells and document the existence of different molecular requirements for the differentiation of CD4+ and CD8+ T cells.

  5. A multicenter randomized double-blind study: comparison of the Epley, Semont, and sham maneuvers for the treatment of posterior canal benign paroxysmal positional vertigo.

    PubMed

    Lee, Jong Dae; Shim, Dae Bo; Park, Hong Ju; Song, Chan Il; Kim, Min-Beom; Kim, Chang-Hee; Byun, Jae Yong; Hong, Sung Kwang; Kim, Tae Su; Park, Kye Hoon; Seo, Jae-Hyun; Shim, Byoung Soo; Lee, Joon Han; Lim, Hyun Woo; Jeon, Eun-Ju

    2014-01-01

    We evaluated the short-term efficacy of Epley, Semont, and sham maneuvers for resolving posterior canal benign paroxysmal positional vertigo (BPPV) in a prospective multicenter randomized double-blind controlled study. Subjects were randomly divided into three groups: Epley (36 patients), Semont (32 patients), and sham (Epley maneuver for the unaffected side, 31 patients). Out of 14 institutes which participated in this study, 5 institutes had previous experience of the Epley but not the Semont maneuver and the other 9 had previous experience of both maneuvers. Each maneuver was repeated twice if there was still positional vertigo or nystagmus on day 0, and the presence of nystagmus and vertigo on positional testing were evaluated immediately, 1 day, and 1 week after treatment. After the first maneuver, the Epley group showed a significantly higher resolution rate of positional nystagmus than the Semont or sham groups (63.9, 37.5, and 38.7%, respectively). After the second maneuver, the resolution rate (83.3%) of the Epley group was significantly higher than that (51.6%) of the sham group. At 1 day and 1 week after treatment, the resolution rate of the Epley group was significantly higher than those of the other groups. Similar results were seen for the resolution of positional vertigo. The Epley maneuver showed persistent resolution rates of positional vertigo and nystagmus without a fatigue phenomenon. The Epley maneuver was significantly more effective per maneuver than Semont or sham maneuvers for the short-term treatment of posterior canal BPPV. The Semont maneuver showed a higher success rate than the sham maneuver, but it was not significantly different.

  6. Comparison of a tube-holder (Rescuefix) versus tape-tying for minimizing double-lumen tube displacement during lateral positioning in thoracic surgery

    PubMed Central

    Byun, Sung Hye; Kang, Su Hwang; Kim, Jong Hae; Ryu, Taeha; Kim, Baek Jin; Jung, Jin Yong

    2016-01-01

    Abstract Background: Double-lumen endotracheal tubes (DLTs) are often displaced during change from the supine to the lateral decubitus position. The aim of this study was to determine whether Rescuefix, a recently developed tube-holder device, is more effective than the traditional tape-tying method for tube security during lateral positioning. Methods: Patients were randomly assigned to a Rescuefix (R) group (n = 22) or a tape (T) group (n = 22). After intubation with a left-sided DLT and adjustment of the appropriate DLT position using a fiberoptic bronchoscope, the DLT was fixed firmly at the side of the mouth by either Rescuefix or Durapore tape. “Tracheal depth” (from the tracheal carina to the elbow connector of the DLT) and “bronchial depth” (from the left bronchial carina to the elbow connector of the DLT) were measured in the supine position using the fiberoptic bronchoscope. After positional change, tracheal and bronchial depths were measured as described above. As the primary endpoint, displacement of the DLT during positional change was evaluated by obtaining the difference in depths measured when the patient was in the supine and lateral decubitus positions. In addition, after lateral positioning of the patient, any requirement for repositioning the DLT was recorded. Results: After lateral positioning, there were no significant differences in changes in tracheal and bronchial depths between the groups (tracheal depth 6.1 ± 4.4 mm [R group] and 9.1 ± 5.6 mm [T group], P = 0.058; bronchial depth 6.5 ± 4.4 mm [R group], and 8.5 ± 4.6 mm [T group], P = 0.132). Although the amount of change in tracheal and bronchial depths was not different between the groups, the need to reposition the DLT was significantly lower in the R group than in the T group (32% vs 68%, P = 0.016). Conclusion: This study demonstrated that use of Rescuefix did not reduce the amount of DLT displacement, but it did significantly

  7. Large Positive Thermal Expansion and Small Band Gap in Double-ReO3-Type Compound NaSbF6.

    PubMed

    Yang, C; Qu, B Y; Pan, S S; Zhang, L; Zhang, R R; Tong, P; Xiao, R C; Lin, J C; Guo, X G; Zhang, K; Tong, H Y; Lu, W J; Wu, Y; Lin, S; Song, W H; Sun, Y P

    2017-05-01

    Double-ReO3-type structure compound NaSbF6 undergoes a low-temperature rhombohedral to high-temperature cubic phase between 303 and 323 K, as revealed by temperature-dependent X-ray diffractions. Although many double-ReO3-type fluorides exhibit either low thermal expansion or negative thermal expansion (NTE), NaSbF6 exhibits positive thermal expansion (PTE) with a large volumetric coefficient of thermal expansion, αv = 62 ppm/K, in its cubic phase. Raman spectroscopy reveals that the low-frequency transverse vibration of fluorine atoms is stiffened in NaSbF6, compared with the typical NTE compound CaZrF6 with the same structure. The related weak contraction associated with the polyhedral rocking would be overcome by the notable elongation of the Na-F bond length on heating, thus leading to the large volumetric PTE. Unlike ScF3 and CaZrF6 which are insulators with a wide band gap, a relative small band gap of 3.76 eV was observed in NaSbF6. The small band gap can be attributed to the hybridization between the Sb 5s and F 2p orbitals.

  8. Changes in the cellular membrane surface coat of lymphocytes and thymocytes after incubation in vitro with cystein as revealed with electronmicroscopy.

    PubMed

    Borowicz, J; Olszewska, K; Roszkowski-Sliz, W; Ryzewski, J

    1977-01-01

    Changes in the cellular membrane surface coat of lymphocytes and thymocytes after incubation with cystein in vitro were revealed with electronmicroscope, while performing the reaction with Ruthenium Red and Concanavaline A. Lymphocytes and thymocytes not incubated with cystein to which reaction with Ruthenium red and Cocanavaline A was applied have shown a well developed and preserved surface coat of the cellular membrane. Contrary to this finding when lymphocytes and thymocytes were incubated with cystein and thereafter treated with Ruthenium Red and Concanavaline A no reaction product on the surface of the cellular membrane was observed. The experimental results could indicate on the influence of cystein on the glycoside bonds.

  9. Comparison between transthoracic lung ultrasound and a clinical method in confirming the position of double-lumen tube in thoracic anaesthesia. A pilot study.

    PubMed

    Álvarez-Díaz, N; Amador-García, I; Fuentes-Hernández, M; Dorta-Guerra, R

    2015-01-01

    To compare the ability of lung ultrasound and a clinical method in the confirmation of a selective bronchial intubation by left double-lumen tube in elective thoracic surgery. A prospective and blind, observational study was conducted in the setting of a university hospital operating room assigned for thoracic surgery. A single group of 105 consecutive patients from a total of 130, were included. After blind intubation, the position of the tube was confirmed by clinical and ultrasound assessment. Finally, the fiberoptic bronchoscopy confirmation as a reference standard was used to confirm the position of the tube. Under manual ventilation, by sequentially clamping the tracheal and bronchial limbs of the tube, clinical confirmation was made by auscultation, capnography, visualizing the chest wall expansion, and perceiving the lung compliance in the reservoir bag. Ultrasound confirmation was obtained by visualizing lung sliding, diaphragmatic movements, and the appearance of lung pulse sign. The sensitivity of the clinical method was 84.5%, with a specificity of 41.1%. The positive and negative likelihood ratio was 1.44 and 0.38, respectively. The sensitivity of the ultrasound method was 98.6%, specificity was 52.9%, with a positive likelihood ratio of 2.10 and a negative likelihood ratio of 0.03. Comparisons between the diagnostic performance of the 2 methods were calculated with McNemar's test. There was a significant difference in sensitivity between the ultrasound method and the clinical method (P=.002). Nevertheless, there was no statistically significant difference in specificity between both methods (P=.34). A p value<.01 was considered statistically significant. Lung ultrasound was superior to the clinical method in confirming the adequate position of the left double-lumen tube. On the other hand, in confirming the misplacement of the tube, differences between both methods could not be ensured. Copyright © 2014 Sociedad Española de Anestesiolog

  10. Double attention bias for positive and negative emotional faces in clinical depression: evidence from an eye-tracking study.

    PubMed

    Duque, Almudena; Vázquez, Carmelo

    2015-03-01

    According to cognitive models, attentional biases in depression play key roles in the onset and subsequent maintenance of the disorder. The present study examines the processing of emotional facial expressions (happy, angry, and sad) in depressed and non-depressed adults. Sixteen unmedicated patients with Major Depressive Disorder (MDD) and 34 never-depressed controls (ND) completed an eye-tracking task to assess different components of visual attention (orienting attention and maintenance of attention) in the processing of emotional faces. Compared to ND, participants with MDD showed a negative attentional bias in attentional maintenance indices (i.e. first fixation duration and total fixation time) for sad faces. This attentional bias was positively associated with the severity of depressive symptoms. Furthermore, the MDD group spent a marginally less amount of time viewing happy faces compared with the ND group. No differences were found between the groups with respect to angry faces and orienting attention indices. The current study is limited by its cross-sectional design. These results support the notion that attentional biases in depression are specific to depression-related information and that they operate in later stages in the deployment of attention. Copyright © 2014 Elsevier Ltd. All rights reserved.

  11. Detecting contaminant-induced apoptosis and necrosis in lake trout thymocytes via flow cytometry.

    USGS Publications Warehouse

    Sweet, Leonard I.; Passino-Reader, Dora R.; Meier, Peter G.; Omann, Geneva M.; Stolen, J.S.; Fletcher, T.C.; Rowley, A.F.; Zelikoff, J.T.; Kaattari, S.L.; Smith, S.A.

    1997-01-01

    This chapter details the cytofluorometric techniques employed to assess levels of active (apoptosis) and passive (necrotic) cell death in untreated and contaminant-treated fish thymocytes. The thymus is believed to be a central component of hematopoiesis and immune function in teleosts (Abelli et al., 1996). Hence, chemically-elicited adverse effects to the thymus may result in immunomodulation and organ dysfunction. However, it is not well documented that environmental contaminants induce apoptosis, or programmed cell death. There is some evidence suggesting that low level exposure to waterborne contaminants can specifically induce cell death in the olfactory epithelium of rainbow trout (Julliard et al., 1996). Presently, only limited information is available in the literature regarding apoptotic death in piscine immune cells (Alford et al., 1994; Greenlee et al., 1991).

  12. Anti-thymocyte globulin induced non-cardiogenic pulmonary edema during renal transplantation.

    PubMed

    Parikh, Beena K; Bhosale, Guruprasad P; Shah, Veena R

    2011-10-01

    Non-cardiogenic pulmonary edema (NCPE) is a clinical syndrome characterized by simultaneous presence of severe hypoxemia, bilateral alveolar infiltrates on chest radiograph, without evidence of left atrial hypertension/congestive heart failure/fluid overload. The diagnosis of drugrelated NCPE relies upon documented exclusion of other causes of NCPE like gastric aspiration, sepsis, trauma, negative pressure pulmonary edema. We describe a 28year-old, 50 kg male with ASA risk III posted for laparoscopic renal transplantation, who developed NCPE after 4 hours of administration of rabbit anti-human thymocyte immunoglobulin (ATG). He was successfully treated with mechanical ventilatory support and adjuvant therapy. This report emphasizes that this fatal complication may occur with use of ATG.

  13. Carnosine prevents necrotic and apoptotic death of rat thymocytes via ouabain sensitive Na/K-ATPase

    PubMed Central

    Smolyaninova, Larisa V.; Dergalev, Alexander A.; Kulebyakin, Konstantin Y.; Carpenter, David O.; Boldyrev, Alexander A.

    2012-01-01

    It is known that ouabain, a selective inhibitor of Na/K-ATPase, can cause not only activation of signal cascades, which regulate the cell viability, but also can cause free radical accumulation, which can evoke the oxidative stress. We have shown that nanomolar concentrations of ouabain result in the temporary increase in the level of intracellular free radicals but the millimolar concentration of ouabain induces a stable intracellular accumulation of free radicals in rat thymocytes. The increasing level of free radicals resulting from both low and high concentrations of ouabain can be attenuated by the antioxidant, carnosine. Moreover the long-term incubation with ouabain leads to the cell death by necrosis and apoptosis. Ouabain-mediated apoptosis and necrosis were also abolished by carnosine. PMID:22763713

  14. Fulvestrant 500 mg vs 250 mg in postmenopausal women with estrogen receptor-positive advanced breast cancer: a randomized, double-blind registrational trial in China.

    PubMed

    Zhang, Qingyuan; Shao, Zhimin; Shen, Kunwei; Li, Li; Feng, Jifeng; Tong, Zhongsheng; Gu, Kangsheng; Wang, Xiaojia; Xu, Binghe; Sun, Guofang; Chen, Huifang; Rukazenkov, Yuri; Jiang, Zefei

    2016-08-30

    The international CONFIRM study showed that fulvestrant 500 mg improved progression-free survival (PFS) vs fulvestrant 250 mg in postmenopausal women with estrogen receptor (ER)-positive locally advanced/metastatic breast cancer (LA/MBC). In this randomized, double-blind study, postmenopausal Chinese women with ER-positive LA/MBC and progression after endocrine therapy received fulvestrant 500 mg (days 0, 14, 28, and every 28 days thereafter) or fulvestrant 250 mg (every 28 days). Consistency with the international study was assumed if the hazard ratio (HR) for comparison of PFS (primary endpoint) was < 1 (stratified log-rank test). The study was not powered to assess between-group differences.In total, 221 patients were randomized (fulvestrant 500 mg: n = 111; fulvestrant 250 mg: n = 110). Baseline characteristics were balanced. Median PFS was 8.0 months with fulvestrant 500 mg vs 4.0 months with 250 mg (HR = 0.75; 95% confidence interval [CI] 0.54-1.03; P = 0.078). PFS (HR; 95% CI) favored fulvestrant 500 mg in post-antiestrogen (0.86; 0.54-1.37) and post-aromatase inhibitor (0.65; 0.42-1.03) settings. No new safety considerations were observed. These results are consistent with the international CONFIRM study, supporting the superior clinical benefit of fulvestrant 500 mg in women with ER-positive LA/MBC experiencing progression following prior endocrine therapy.

  15. Domain-Specific and Stage-Intrinsic Changes in Tcrb Conformation During Thymocyte Development

    PubMed Central

    Majumder, Kinjal; Rupp, Levi J.; Yang-Iott, Katherine S.; Koues, Olivia I.; Kyle, Katherine E.; Bassing, Craig H.; Oltz, Eugene M.

    2015-01-01

    Considerable crosstalk exists between mechanisms controlling genome architecture and gene expression. Antigen receptor loci are excellent models for these processes because they are regulated at both conformational and transcriptional levels to facilitate their assembly by V(D)J recombination. Upon commitment to the DN stage of T cell development, Tcrb adopts a compact conformation that promotes long-range recombination between Vβ gene segments (Trbvs) and their DβJβ targets. Formation of a functional VβDβJβ join signals for robust proliferation of DN thymocytes and their differentiation into DP cells, where Trbv recombination is squelched (allelic exclusion). DP differentiation also is accompanied by decontraction of Tcrb, which has been thought to separate the entire Trbv cluster from DβJβ segments (spatial segregation-based model for allelic exclusion). However, DP cells also repress transcription of unrearranged Trbvs, which may contribute to allelic exclusion. We performed a more detailed study of developmental changes in Tcrb topology and found that only the most distal portion of the Trbv cluster separates from DβJβ segments in DP thymocytes, leaving most Trbvs spatially available for rearrangement. Preferential dissociation of distal Trbvs is independent of robust proliferation or changes in transcription, chromatin, or architectural factors, which are coordinately regulated across the entire Trbv cluster. Segregation of distal Trbvs also occurs on alleles harboring a functional VβDβJβ join, suggesting this process is independent of rearrangement status and is DP-intrinsic. Our finding that most Trbvs remain associated with DβJβ targets in DP cells revises allelic exclusion models from their current conformation-dominant to a transcription-dominant formulation. PMID:26101321

  16. Some characteristics of membrane Cd²+ transport in rat thymocytes: an analysis using Fluo-3.

    PubMed

    Kawanai, Takuya; Fujinaga, Masahiro; Koizumi, Kazuki; Kurotani, Isao; Hashimoto, Erika; Satoh, Masaya; Imai, Shoji; Miyoshi, Norikazu; Oyama, Yasuo

    2011-10-01

    Although cadmium-induced apoptosis of lymphocytes is one of common features in the immunotoxicity of cadmium, the membrane pathway for intracellular cadmium accumulation is not fully elucidated. To characterize membrane Cd(2+) transport of rat thymocytes, the change in intracellular Cd(2+) concentration under various conditions was examined by the use of Fluo-3, a fluorescent probe for monitoring the change in intracellular concentration of divalent metal cations. The membrane Cd(2+) transport was estimated by the augmentation of Fluo-3 fluorescence induced by bath application of CdCl(2). Lowering temperature strongly suppressed the augmentation of Fluo-3 fluorescence by CdCl(2), suggesting that the metabolic process can be involved in membrane Cd(2+) transport. External acidification (decreasing pH) and membrane depolarization by adding KCl attenuated the augmentation, indicating the requirement of electrochemical driving force for membrane Cd(2+) transport into the cells. Bath application of CaCl(2) and ZnCl(2) equally decreased the augmentation, suggesting their competition with Cd(2+) at the membrane transport. The augmentation by CdCl(2) was lesser in the cells treated with N-ethylmaleimide inducing chemical depletion of cellular thiols. The result suggests the contribution of sulfhydryl groups to membrane Cd(2+) transport. Taken together, it is suggested that the cells possess a temperature-sensitive membrane Cd(2+) pathway, driven by electrochemical gradient of Cd(2+) and transmembrane potential, with competitive binding site. Based on the characteristics described above, it is unlikely that the membrane Cd(2+) transport in rat thymocytes is attributed to a single transport system although it has characteristics that are similar to those of divalent cation transporter 1.

  17. Effects of glutamate positive modulators on cognitive deficits in schizophrenia: a systematic review and meta-analysis of double-blind randomized controlled trials

    PubMed Central

    Iwata, Y; Nakajima, S; Suzuki, T; Keefe, RSE; Plitman, E; Chung, JK; Caravaggio, F; Mimura, M; Graff-Guerrero, A; Uchida, H

    2017-01-01

    Hypofunction of N-methyl-D-aspartate (NMDA) receptors has been proposed to have an important role in the cognitive impairments observed in schizophrenia. Although glutamate modulators may be effective in reversing such difficult-to-treat conditions, the results of individual studies thus far have been inconsistent. We conducted a systematic review and meta-analysis to examine whether glutamate positive modulators have beneficial effects on cognitive functions in patients with schizophrenia. A literature search was conducted to identify double-blind randomized placebo-controlled trials in schizophrenia or related disorders, using Embase, Medline, and PsycINFO (last search: February 2015). The effects of glutamate positive modulators on cognitive deficits were evaluated for overall cognitive function and eight cognitive domains by calculating standardized mean differences (SMDs) between active drugs and placebo added to antipsychotics. Seventeen studies (N = 1391) were included. Glutamate positive modulators were not superior to placebo in terms of overall cognitive function (SMD = 0.08, 95% confidence interval = − 0.06 to 0.23) (11 studies, n = 858) nor each of eight cognitive domains (SMDs = − 0.03 to 0.11) (n = 367–940) in this population. Subgroup analyses by diagnosis (schizophrenia only studies), concomitant antipsychotics, or pathway of drugs to enhance the glutamatergic neurotransmission (glycine allosteric site of NMDA receptors or α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors) suggested no procognitive effect of glutamate positive modulators. Further, no effect was found in individual compounds on cognition. In conclusion, glutamate positive modulators may not be effective in reversing overall cognitive impairments in patients with schizophrenia as adjunctive therapies. PMID:26077694

  18. Ikaros is required to survive positive selection and to maintain clonal diversity during T-cell development in the thymus

    PubMed Central

    Tinsley, Kevin W.; Hong, Changwan; Luckey, Megan A.; Park, Joo-Young; Kim, Grace Y.; Yoon, Hee-won; Keller, Hilary R.; Sacks, Andrew J.; Feigenbaum, Lionel

    2013-01-01

    The zinc-finger protein Ikaros is a key player in T-cell development and a potent tumor suppressor in thymocytes. To understand the molecular basis of its function, we disabled Ikaros activity in vivo using a dominant negative Ikaros transgene (DN-IkTg). In DN-IkTg mice, T-cell development was severely suppressed, and positively selected thymocytes clonally expanded, resulting in a small thymus with a heavily skewed T-cell receptor (TCR) repertoire. Notably, DN-IkTg induced vigorous proliferation concomitant to downregulation of antiapoptotic factor expression such as Bcl2. Ikaros activity was required during positive selection, and specifically at the CD4+CD8lo intermediate stage of thymocyte differentiation, where it prevented persistent TCR signals from inducing aberrant proliferation and expansion. In particular, DN-IkTg induced the accumulation of CD4 single-positive (SP) thymocytes with a developmentally transitional phenotype, and it imposed a developmental arrest accompanied by massive apoptosis. Thus, we identified an in vivo requirement for Ikaros function, which is to suppress the proliferative potential of persistent TCR signals and to promote the survival and differentiation of positively selected thymocytes. PMID:23908463

  19. Post-transplant cyclophosphamide versus anti-thymocyte globulin as graft- versus-host disease prophylaxis in haploidentical transplant

    PubMed Central

    Ruggeri, Annalisa; Sun, Yuqian; Labopin, Myriam; Bacigalupo, Andrea; Lorentino, Francesca; Arcese, William; Santarone, Stella; Gülbas, Zafer; Blaise, Didier; Messina, Giuseppe; Ghavamzadeh, Ardeshi; Malard, Florent; Bruno, Benedetto; Diez-Martin, Jose Luis; Koc, Yener; Ciceri, Fabio; Mohty, Mohamad; Nagler, Arnon

    2017-01-01

    Severe graft-versus-host disease is a major barrier for non-T-cell-depleted haploidentical stem cell transplantation. There is no consensus on the optimal graft-versus-host disease prophylaxis. This study compared the two most commonly used graft-versus-host disease prophylaxis regimens (post-transplant cyclophosphamide-based vs. the anti-thymocyte globulin-based) in adults with acute myeloid leukemia reported to the European Society for Blood and Bone Marrow Transplantation. A total of 308 patients were analyzed; 193 received post-transplant cyclophosphamide-based regimen and 115 anti-thymocyte globulin-based regimen as anti-graft-versus-host disease prophylaxis. The post-transplant cyclophosphamide-based regimen was more likely to be associated to bone marrow as graft source (60% vs. 40%; P=0.01). Patients in the post-transplant cyclophosphamide-based regimen group had significantly less grade 3–4 acute graft-versus-host disease than those in the anti-thymocyte globulin-based group (5% vs. 12%, respectively; P=0.01), comparable to chronic graft-versus-host disease. Multivariate analysis showed that non-relapse mortality was lower in the post-transplant cyclophosphamide-based regimen group [22% vs. 30%, Hazard ratio (HR) 1.77(95%CI: 1.09–2.86); P=0.02] with no difference in relapse incidence. Patients receiving post-transplant cyclophosphamide-based regimen had better graft-versus-host disease-free, relapse-free survival [HR 1.45 (95%CI: 1.04–2.02); P=0.03] and leukemia-free survival [HR 1.48 (95%CI: 1.03–2.12); P=0.03] than those in the anti-thymocyte globulin-based group. In the multivariate analysis, there was also a trend for a higher overall survival [HR 1.43 (95%CI: 0.98–2.09); P=0.06] for post-transplant cyclophosphamide-based regimen versus the anti-thymocyte globulin-based group. Notably, center experience was also associated with non-relapse mortality and graft-versus-host disease-free, relapse-free survival. Haplo-SCT using a post

  20. In vivo stimulation of sugar uptake in rat thymocytes. An extranuclear action of 3,5,3'-triiodothyronine.

    PubMed Central

    Segal, J; Ingbar, S H

    1985-01-01

    In previous studies we have demonstrated that 3,5,3'-triiodothyronine (T3) in vitro produces a prompt increase in the uptake of the sugar analogue 2-deoxyglucose (2-DG) by freshly isolated rat thymocytes. This effect is prompt, being evident at 20 min after addition of T3, is independent of new protein synthesis, and can be elicited by physiologic concentrations of the hormone. In the present studies, we have sought to determine whether physiologic doses of T3 are capable of inducing an increase in 2-DG uptake in the thymocytes of the living animal. Therefore, 26-28-d-old female rats were injected with increasing doses of i.v. T3, followed 60 min later by 3H-labeled 2-DG. 30 min later, animals were killed, thymocytes were isolated, and their 3H content determined. Uptake of [3H]2-DG was increased by T3 in a dose-dependent manner. The lowest effective dose was 10 ng/100 g of body weight (30% above control) and the maximally effective dose 1 microgram/100 g of body weight (116% above control). The effect of T3 was independent of new protein synthesis in that it was not blocked by a dose of cycloheximide that inhibited the incorporation of [3H]leucine into thymocyte protein by 92-95%. Comparable studies with various thyronine analogues revealed the following rank order of potency: L-T3 greater than L-3,5,3'5'-tetraiodothyronine (L-T4) greater than D-T3 greater than or equal to D-T4 greater than L-3,3'5'-triiodothyronine greater than 3'-isopropyl-3,5-L-diiodothyronine (T2) = 3,5-L-T2. DL-thyronine was without effect. These studies indicate that T3 in physiologic doses acts in vivo to increase the uptake of sugar by rat thymocytes by a mechanism that is extranuclear in origin, in that it is independent of new protein synthesis. The findings support the conclusion that the previously demonstrated effects of T3 on thymocyte sugar uptake in vitro, which seem clearly to be mediated at the level of the plasma membrane, have physiologic relevance. PMID:4056041

  1. Thymic Medullary Epithelial Cell Differentiation, Thymocyte Emigration, and the Control of Autoimmunity Require Lympho–Epithelial Cross Talk via LTβR

    PubMed Central

    Boehm, Thomas; Scheu, Stefanie; Pfeffer, Klaus; Bleul, Conrad C.

    2003-01-01

    Thymocytes depend on the interaction with thymic epithelial cells for the generation of a diverse, nonautoreactive T cell repertoire. In turn, thymic epithelial cells acquire their three-dimensional cellular organization via instructive signals from developing thymocytes. The nature of these signals has been elusive so far. We show that thymocytes and medullary epithelial cells (MECs) communicate via the lymphotoxin β receptor (LTβR) signaling axis. Normal differentiation of thymic MECs requires LTβR ligand on thymocytes and LTβR together with nuclear factor–κB-inducing kinase (Nik) in thymic epithelial cells. Impaired lympho–epithelial cross talk in the absence of the LTβR causes aberrant differentiation and reduced numbers of thymic MECs, leads to the retention of mature T lymphocytes, and is associated with autoimmune phenomena, suggesting an unexpected role for LTβR signaling in central tolerance induction. PMID:12953095

  2. Class II major histocompatibility complex molecules regulate the development of the T4+T8- inducer phenotype of cultured human thymocytes.

    PubMed Central

    Blue, M L; Daley, J F; Levine, H; Schlossman, S F

    1985-01-01

    We demonstrate that a variety of Ia+ cells has the ability to promote the development of human T4+T8- thymocytes in vitro. Prolonged thymocyte culture in the absence of Ia+ accessory cells results in a predominantly T8+T4- cell population. The generation of T4+ cells in the presence of irradiated Ia+ cells could be suppressed up to 70% by a monoclonal antibody directed against a nonpolymorphic epitope on HLA-DR. Using two-color fluorescence sorting techniques, we were able to identify the activated T4+T8+ thymocyte as the cell that interacts with Ia and gives rise to the T4+T8- cell subset. These results directly and specifically implicate class II major histocompatibility complex molecules in the differentiative pathway of the human thymocyte. Images PMID:2933749

  3. A two-amino-acid substitution in the transcription factor RORγt disrupts its function in TH17 differentiation but not in thymocyte development.

    PubMed

    He, Zhiheng; Ma, Jian; Wang, Ruiqing; Zhang, Jing; Huang, Zhaofeng; Wang, Fei; Sen, Subha; Rothenberg, Ellen V; Sun, Zuoming

    2017-10-01

    The transcription factor RORγt regulates differentiation of the TH17 subset of helper T cells, thymic T cell development and lymph-node genesis. Although elimination of RORγt prevents TH17 cell-mediated experimental autoimmune encephalomyelitis (EAE), it also disrupts thymocyte development, which could lead to lethal thymic lymphoma. Here we identified a two-amino-acid substitution in RORγt (RORγt(M)) that 'preferentially' disrupted TH17 differentiation but not thymocyte development. Mice expressing RORγt(M) were resistant to EAE associated with defective TH17 differentiation but maintained normal thymocyte development and normal lymph-node genesis, except for Peyer's patches. RORγt(M) showed less ubiquitination at Lys69 that was selectively required for TH17 differentiation but not T cell development. This study will inform the development of treatments that selectively target TH17 cell-mediated autoimmunity but do not affect thymocyte development or induce lymphoma.

  4. Double negative feedback loop between reprogramming factor LIN28 and microRNA let-7 regulates aldehyde dehydrogenase 1-positive cancer stem cells

    PubMed Central

    Yang, Xiaojun; Lin, Xiaojuan; Zhong, Xiaomin; Kaur, Sippy; Li, Ning; Liang, Shun; Lassus, Heini; Wang, Liping; Katsaros, Dionyssios; Montone, Kathleen; Zhao, Xia; Zhang, Youcheng; Bützow, Ralf; Coukos, George; Zhang, Lin

    2010-01-01

    A relatively rare aldehyde dehydrogenase 1 (ALDH1) positive “stem cell-like” subpopulation of tumor cells has the unique ability to initiate and perpetuate tumor growth; moreover it is highly resistant to chemotherapy and significantly associated with poor clinical outcomes. The development of more effective therapies for cancer requires targeting of this cell population. Using cDNA microarray analysis, we identified that the expression of the C. elegans lin-28 homolog (LIN28) was positively correlated with the percentage of ALDH1+ tumor cells; this was further validated in an independent set of tissue arrays (n=197). Both lose-of-function and gain-of-function studies demonstrated that LIN28 plays a critical role in the maintenance of ALDH1+ tumor cells. In addition, we found that there is a double negative feedback loop between LIN28 and let-7 in tumor cells, and that let-7 negatively regulates ALDH1+ tumor cells. Finally, we report that a LIN28/let-7 loop modulates self renewal and differentiation of mammary gland epithelial progenitor cells. Our data provide evidence that cancer stem cells may arise through a “reprogramming-like” mechanism. A rebalancing of the LIN28/let-7 regulatory loop could be a novel therapeutic strategy to target ALDH1+ cancer stem cells. PMID:21045151

  5. Evolution of Double Positive Autoregulatory Feedback Loops in CYCLOIDEA2 Clade Genes Is Associated with the Origin of Floral Zygomorphy[W

    PubMed Central

    Yang, Xia; Pang, Hong-Bo; Liu, Bo-Ling; Qiu, Zhi-Jing; Gao, Qiu; Wei, Lai; Dong, Yang; Wang, Yin-Zheng

    2012-01-01

    Members of the CYCLOIDEA2 (CYC2) clade of the TEOSINTE BRANCHED1, CYCLOIDEA, and PCF transcription factor genes are widely involved in controlling floral zygomorphy, a key innovation in angiosperm evolution, depending on their persistently asymmetric expression in the corresponding floral domains. However, it is unclear how this asymmetric expression is maintained throughout floral development. Selecting Primulina heterotricha as a model, we examined the expression and function of two CYC2 genes, CYC1C and CYC1D. We analyzed the role of their promoters in protein–DNA interactions and transcription activation using electrophoresis mobility shift assays, chromatin immunoprecipitation, and transient gene expression assays. We find that CYC1C and CYC1D positively autoregulate themselves and cross-regulate each other. Our results reveal a double positive autoregulatory feedback loop, evolved for a pair of CYC2 genes to maintain their expression in developing flowers. Further comparative genome analyses, together with the available expression and function data of CYC2 genes in the core eudicots, suggest that this mechanism might have led to the independent origins of floral zygomorphy, which are associated with plant–insect coevolution and the adaptive radiation of angiosperms. PMID:22649271

  6. Undergraduate Observations of Separation and Position Angle of Double Stars WDS J05460+2119AB (ARY 6AD and ARY 6 AE) at Manzanita Observatory (Abstract)

    NASA Astrophysics Data System (ADS)

    HOffert, M. J.; Weise, E.; Clow, J.; Hirzel, J.; Leeder, B.; Molyneux, S.; Scutti, N.; Spartalis, S.; Takuhara, C.

    2014-12-01

    (Abstract only) Six beginning astronomy students, part of an undergraduate stellar astronomy course, one advanced undergraduate student assistant, and a professor measured the position angles and separations of Washington Double Stars (WDS) J05460+2119 (= WDS J05460+2119AB; also known as ARY 6 AD and ARY 6 AE). The measurements were made at the Manzanita Observatory (116º 20' 42" W, 32º 44' 5" N) of the Tierra Astronomical Institute on 10 Blackwood Road in Boulevard, California (www.youtube.com/watch?v=BHVdeMGBGDU), at an elevation of 4,500 ft. A Celestron 11-inch HD Edge telescope was used to measure the position angles and separations of ARY 6 AD and ARY 6 AE. The averages of our measurements are as follows: separation AD: trial 1 124.1 arcseconds and trial 2 124.5 arcseconds; separation AE: trial 1 73.3 arcseconds and trial 2 73.8 arcseconds. The averages of positon angle for AD: trial 1 159.9 degrees and trial 2 161.3 degrees, for AE: trial 1 232.6 degrees and trial 2 233.7 degrees.

  7. Diphosphates at the 5' end of the positive strand of yeast L-A double-stranded RNA virus as a molecular self-identity tag.

    PubMed

    Fujimura, Tsutomu; Esteban, Rosa

    2016-10-01

    The 5'end of RNA conveys important information on self-identity. In mammalian cells, double-stranded RNA (dsRNA) with 5'di- or triphosphates generated during virus infection is recognized as foreign and elicits the host innate immune response. Here, we analyze the 5' ends of the dsRNA genome of the yeast L-A virus. The positive strand has largely diphosphates with a minor amount of triphosphates, while the negative strand has only diphosphates. Although the virus can produce capped transcripts by cap snatching, neither strand carried a cap structure, suggesting that only non-capped transcripts serve as genomic RNA for encapsidation. We also found that the 5' diphosphates of the positive but not the negative strand within the dsRNA genome are crucial for transcription in vitro. Furthermore, the presence of a cap structure in the dsRNA abrogated its template activity. Given that the 5' diphosphates of the transcripts are also essential for cap acquisition and that host cytosolic RNAs (mRNA, rRNA, and tRNA) are uniformly devoid of 5' pp-structures, the L-A virus takes advantage of its 5' terminal diphosphates, using them as a self-identity tag to propagate in the host cytoplasm.

  8. Differentiation of CD3-4-8- human fetal thymocytes in vivo: characterization of a CD3-4+8- intermediate

    PubMed Central

    1993-01-01

    Human thymocyte differentiation was examined by injecting fetal thymic progenitor populations into human thymic xenografts in SCID-hu mice. Thymic progenitors were fluorescently labeled with the lipophilic dye PKH2. The phenotypes of their progeny could be identified by flow cytometric analysis of cells with a very high fluorescent PKH2 signal. Intrathymic injection of purified triple negative (TN) CD3-4-8- thymocytes resulted in the sequential appearance of CD3-4+8-, CD3-4+8+, and CD3+4+8+ cells, with the subsequent appearance of small numbers of phenotypically mature CD3+4+8- and CD3+4-8+ cells over a 4-d period. Sorted CD3-4+8- thymocytes injected intrathymically rapidly differentiated to CD4+8+ cells. CD4+8+ fetal thymocytes in cell cycle differentiated into phenotypically mature CD3+4+8- and CD3+4-8+ populations, whereas nondividing CD4+8+ cells failed to differentiate after intrathymic transfer. The number of cell divisions that occurred between the injection of TN thymocytes and their progeny at different time points was estimated based on the decrease in the intensity of the PKH2 label. The average length of the cell cycle for the TN population was calculated to be 24 h. The SCID-hu model thus provides a useful tool for studying the kinetics of cell division and differentiation of human thymocytes in vivo. PMID:8315382

  9. HMG-CoA reductase inhibitors pravastatin, lovastatin and simvastatin suppress delayed rectifier K(+)-channel currents in murine thymocytes.

    PubMed

    Kazama, Itsuro; Baba, Asuka; Maruyama, Yoshio

    2014-08-01

    Since lymphocytes predominantly express delayed rectifier K(+)-channels (Kv1.3) that trigger lymphocyte activation, statins, which exert immunosuppressive effects, would affect the channel currents. Employing the patch-clamp technique in murine thymocytes, we examined the effects of statins on Kv1.3-channel currents and the membrane capacitance (Cm). Pravastatin significantly suppressed the pulse-end currents of the channels. Lovastatin and simvastatin also suppressed the peak currents, significantly decreasing the Cm. This study demonstrated for the first time that statins inhibit thymocyte Kv1.3-channels. The slow inactivation patterns induced by lovastatin and simvastatin may be associated with their accumulation in the plasma membranes. Copyright © 2014 Institute of Pharmacology, Polish Academy of Sciences. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.

  10. Transgenic expression of microRNA-181d augments the stress-sensitivity of CD4(+)CD8(+) thymocytes.

    PubMed

    Belkaya, Serkan; van Oers, Nicolai S C

    2014-01-01

    Physiological stress resulting from infections, trauma, surgery, alcoholism, malnutrition, and/or pregnancy results in a substantial depletion of immature CD4(+)CD8(+) thymocytes. We previously identified 18 distinct stress-responsive microRNAs (miRs) in the thymus upon systemic stress induced by lipopolysaccharide (LPS) or the synthetic glucocorticoid, dexamethasone (Dex). MiRs are short, non-coding RNAs that play critical roles in the immune system by targeting diverse mRNAs, suggesting that their modulation in the thymus in response to stress could impact thymopoiesis. MiR-181d is one such stress-responsive miR, exhibiting a 15-fold down-regulation in expression. We utilized both transgenic and gene-targeting approaches to study the impact of miR-181d on thymopoiesis under normal and stress conditions. The over-expression of miR-181d in developing thymocytes reduced the total number of immature CD4(+)CD8(+) thymocytes. LPS or Dex injections caused a 4-fold greater loss of these cells when compared with the wild type controls. A knockout mouse was developed to selectively eliminate miR-181d, leaving the closely spaced and contiguous family member miR-181c intact. The targeted elimination of just miR-181d resulted in a thymus stress-responsiveness similar to wild-type mice. These experiments suggest that one or more of three other miR-181 family members have overlapping or compensatory functions. Gene expression comparisons of thymocytes from the wild type versus transgenic mice indicated that miR-181d targets a number of stress, metabolic, and signaling pathways. These findings demonstrate that selected miRs enhance stress-mediated thymic involution in vivo.

  11. Formation of large molecular weight fragments of DNA is a key committed step of apoptosis in thymocytes.

    PubMed

    Cohen, G M; Sun, X M; Fearnhead, H; MacFarlane, M; Brown, D G; Snowden, R T; Dinsdale, D

    1994-07-15

    Apoptosis is a process in which cells die in a controlled manner and apparently participate in their own demise. It is best characterized morphologically by condensation of chromatin and biochemically by cleavage of chromatin at internucleosomal regions to yield a classical DNA ladder pattern. Apoptosis was induced in thymocytes by exposure to either the glucocorticoid, dexamethasone, or DNA topoisomerase II inhibitor, etoposide. We describe the formation of large m.w. fragments of DNA, 30 to 50 kilobase pairs in length, in a population of these thymocytes at an early stage of apoptosis before internucleosomal cleavage of DNA. These fragments are absent in normal thymocytes and their formation is dependent on protein synthesis. Their appearance is coincident with the commitment of these cells to apoptosis. The formation of these large fragments is associated with the condensation of chromatin, abutting the nuclear membrane, recognized as one of the earliest ultrastructural signs of apoptosis. Subsequent cleavage of these large fragments gives rise to oligonucleosomal fragments and is independent of protein synthesis. We propose that the formation of large fragments of DNA represents a key committed step in apoptosis, and that it is from these fragments that the archetypal DNA ladders associated with apoptosis are derived.

  12. Melatonin protects rat thymus against oxidative stress caused by exposure to microwaves and modulates proliferation/apoptosis of thymocytes.

    PubMed

    Sokolovic, Dusan; Djordjevic, Branka; Kocic, Gordana; Veljkovic, Andrej; Marinkovic, Milena; Basic, Jelena; Jevtovic-Stoimenov, Tatjana; Stanojkovic, Zoran; Sokolovic, Danka M; Pavlovic, Voja; Djindjic, Boris; Krstic, Dejan

    2013-03-01

    The aim of the study was to evaluate the effect of melatonin on oxidative stress, DNA fragmentation, apoptsis and proliferation in thymus tissue of rats exposed to microwaves. Wistar rats were divided in four groups: I - treated with saline; II - treated with melatonin; III - microwaves exposed; IV - microwaves exposed and melatonin treated. Melatonin (2 mg/kg i.p.) was administered daily. Animals were sacrificed after 20, 40 and 60 days. A significant increase in malondialdehyde and carbonyl group content, as well as decrease in catalase and increase in xanthine oxidase activity were registered under microwave exposure. Melatonin prevented the increase in malondialdehyde and carbonyl group content, and reversed the effect on catalase and xanthine oxidase activity. Both, alkaline and acid DNase activity were increased due to microwave exposure. Furthermore, microwaves caused increase in apoptosis rate (detected using Annexin V-FITC/PI kit) and reduced proliferative capacity of thymocytes (induced by ConA). However, melatonin caused decrease in alkaline and acid DNase activity, decrease in apoptotic rate and increase in proliferation rate of thymocytes. Melatonin exerts protective effects on rat thymocytes by modulating processes of apoptosis and proliferation, and causes decrease in DNA fragmentation and oxidative stress intensity under exposure to microwaves.

  13. Betaglycan (TβRIII) Is Expressed in the Thymus and Regulates T Cell Development by Protecting Thymocytes from Apoptosis

    PubMed Central

    Aleman-Muench, German R.; Mendoza, Valentin; Stenvers, Kaye; Garcia-Zepeda, Eduardo A.; Lopez-Casillas, Fernando; Raman, Chander; Soldevila, Gloria

    2012-01-01

    TGF-β type III receptor (TβRIII) is a coreceptor for TGFβ family members required for high-affinity binding of these ligands to their receptors, potentiating their cellular functions. TGF-β [1]–[3], bone morphogenetic proteins (BMP2/4) and inhibins regulate different checkpoints during T cell differentiation. Although TβRIII is expressed on hematopoietic cells, the role of this receptor in the immune system remains elusive. Here, we provide the first evidence that TβRIII is developmentally expressed during T cell ontogeny, and plays a crucial role in thymocyte differentiation. Blocking of endogenous TβRIII in fetal thymic organ cultures led to a delay in DN-DP transition. In addition, in vitro development of TβRIII−/− thymic lobes also showed a significant reduction in absolute thymocyte numbers, which correlated with increased thymocyte apoptosis, resembling the phenotype reported in Inhibin α −/− thymic lobes. These data suggest that Inhibins and TβRIII may function as a molecular pair regulating T cell development. PMID:22952931

  14. [Changes in functional activity of the synthetic apparatus of rat thymocytes under acute and chronic gamma-irradiation].

    PubMed

    Sergievich, L A; Karnaukhova, N A

    2002-01-01

    The changes in functional activity of rat thymocyte synthetic apparatus (synthetic activity) under acute (7.5 Gy) and continuous (dose rates 14.4 and 0.43 cGy/day) gamma-irradiation were studied by the fluorescent microspectral analysis. It has been shown that after the acute irradiation the changes in synthetic activity occurred in three main stages. The stages reflect the depression and activation of synthetic processes that is due to interphase and reproductive cell death and urgent recovery of thymus cellularity and secondary repopulating. Under continuous irradiation with a dose rate 14.4 cGy/day in long-term period both the decrease of thymocyte synthetic activity (in most animals) and activation (in the animals with pronounced symptoms of radiation damage) were observed. This reflects the depression processes in immune system and augmentation of immunoreactivity due to mass antigen influence of transformed cells and infectious agents on thymocytes. Under low dose ionizing irradiation (dose rate 0.43 cGy/day) the undulating changes in synthetic processes in thymus cells were observed. This depends on the recurrence of depression and recovery processes in the blood-forming tissue.

  15. UEA-I-binding to thymic medullary epithelial cells selectively reduces numbers of cortical TCRalphabeta+ thymocytes in FTOCs.

    PubMed

    Graziano, M; St-Pierre, Y; Potworowski, E F

    2001-07-02

    Thymic medullary epithelial cells (TMECs) constitute a major stromal cell type, the function of which is incompletely understood. Some TMECs express L-fucose-glycosylated proteins on their plasma membrane; these have been shown to specifically bind the lectin UEA-I. We exploited this observation to investigate the consequences of in situ blockage of TMECs in FTOCs by UEA-I. In UEA-I-treated FTOCs, we noted a decreased cellularity among TCRalphabeta+ but not TCRgammadelta+ cells. In fact, CD3- and CD3lo cortical cells were markedly depleted, while CD3hi cells were unaffected. Since the affected cell subsets are in a different compartment from that where UEA-I binding occurs, it is likely that the effect is mediated through a soluble factor. Two possible mechanisms are proposed: a reduced activation of either TMECs or of medullary thymocytes which normally bind to them, results in lowered production of soluble factors responsible for cortical thymocyte proliferation. Alternately, the binding of UEA-I to TMECs could activate the latter to produce signals inhibitory to cortical thymocytes.

  16. Complementation in trans of altered thymocyte development in knock-in mice expressing mutant forms of SLP76

    PubMed Central

    Jordan, Martha S.; Smith, Jennifer E.; Burns, Jeremy C.; Austin, Jessica-Elise T.; Nichols, Kim E.; Aschenbrenner, Anna C.; Koretzky, Gary A.

    2008-01-01

    Summary The adaptor protein SLP76 directs signaling downstream of the TCR and is essential for thymocyte development. SLP76 contains three tyrosines in its N-terminus that are critical for its function. To define the role of these residues in thymocyte development, we generated two lines of KI mice, one expressing a mutation in tyrosine 145 (Y145F) and a second harboring two point mutations at tyrosines 112 and 128 (Y112/128F). We show here that while thymocyte development requires both Y145 and Y112/128-generated signals, selection is more dependent upon Y145. While several proximal TCR signaling events were defective in both KI mice, phosphorylation of Vav1 and activation of Itk-dependent pathways were differentially affected by mutations at Y112/128 or Y145, respectively. Analysis of mice expressing one Y145F and one Y112/128F allele revealed that these mutants could complement one another in trans, demonstrating cooperativity between two or more SLP76 molecules. PMID:18342008

  17. A positive correlation between mercury and oxidative stress-related gene expression (GPX3 and GSTM3) is measured in female Double-crested Cormorant blood.

    PubMed

    Gibson, Laura A; Lavoie, Raphael A; Bissegger, Sonja; Campbell, Linda M; Langlois, Valerie S

    2014-08-01

    Mercury (Hg) is a widespread contaminant that has been shown to induce a wide range of adverse health effects in birds including reproductive, physiological and neurological impairments. Here we explored the relationship between blood total Hg concentrations ([THg]) and oxidative stress gene induction in the aquatic piscivorous Double-crested Cormorants (Phalacrocorax auritus) using a non-lethal technique, i.e., blood gene expression analysis. P. auritus blood was sampled at five sites across the Great Lakes basin, Ontario, Canada and was analyzed for [THg]. To assess cellular stress, the expression of glutathione peroxidases 1 and 3 (GPX1, GPX3), superoxide dismutase 1 (SOD1), heat-shock protein 70 kd-8 (HSP70-8) and glutathione S-transferase µ3 (GSTM3) were measured in whole blood samples using real-time RT-PCR. Results showed a significantly positive correlation between female blood [THg] and both GPX3 and GSTM3 expression. Different levels of oxidative stress experienced by males and females during the breeding season may be influencing the differential oxidative stress responses to blood [THg] observed in this study. Overall, these results suggest that Hg may lead to oxidative stress as some of the cellular stress-related genes were altered in the blood of female P. auritus and that blood gene expression analysis is a successful approach to assess bird health condition.

  18. [A multi-center, randomized, double-blind, positive controlled clinical trial of salbutamol sulfate MDI without CFC in the treatment of asthma].

    PubMed

    2014-03-01

    To evaluate the efficacy and safety of domestically produced Salbutamol Sulfate metered dose inhaler (MDI) without chloro-fluro-carbon (CFC) in the treatment of asthmatic patients. A muticenter, randomized, double-blind, positive controlled clinical trial was conducted in asthmatic patients. The participants were randomized divided into trial group treated with domestically produced Salbutamol Sulfate Aerosol MDI (200 microg single-dose) and control group treated with imported Salbutamol Sulfate Aerosol MDI (200 microg single-dose). The lung function was measured by spirometry and a scoring questionaire before medications and 15, 30, 120 and 240 min post medications. A total of 238 patients were enrolled in the trial. Compared with baselines, the forced expiratory volume in 1 s (FEV1) of the participants in both groups increased significantly 15, 30, 120 and 240 min after medications (P < 0.05). There was no significant difference in the changes of FEV1 between the two groups (P > 0.05). Similarly, the forced vital capacity (FVC) and asthma symptomatic scores of the participants in both groups improved significantly after administration of medications (P < 0.05). There was no significant difference in the ratio of adverse reactions between the two groups (P > 0.05). In both groups, drugs were well tolerated. The domestically produced Salbutamol Sulfate Aerosol MDI without CFC is effective and safe for treating asthma.

  19. The position of cytochrome b(559) relative to Q(A) in photosystem II studied by electron-electron double resonance (ELDOR).

    PubMed

    Kuroiwa, S; Tonaka, M; Kawamori, A; Akabori, K

    2000-11-20

    The electron-electron double resonance (ELDOR) method was applied to measure the dipole interaction between cytochrome (Cyt) b(+)(559) and the primary acceptor quinone (Q(-)(A)), observed at g=2.0045 with the peak to peak width of about 9 G, in Photosystem II (PS II) in which the non-heme Fe(2+) was substituted by Zn(2+). The paramagnetic centers of Cyt b(+)(559)Y(D)Q(-)(A) were trapped by illumination at 273 K for 8 min, followed by dark adaptation for 3 min and freezing into 77 K. The distance between the pair Cyt b(+)(559)-Q(-)(A) was estimated from the dipole interaction constant fitted to the observed ELDOR time profile to be 40+/-1 A. In the membrane oriented PS II particles the angle between the vector from Q(A) to Cyt b(559) and the membrane normal was determined to be 80+/-5 degrees. The position of Cyt b(559) relative to Q(A) suggests that the heme plane is located on the stromal side of the thylakoid membrane. ELDOR was not observed for Cyt b(+)(559) Y(D) spin pair, suggesting the distance between them is more than 50 A.

  20. Double-blind randomized trial on short-term efficacy of the Semont maneuver for the treatment of posterior canal benign paroxysmal positional vertigo.

    PubMed

    Mandalà, Marco; Santoro, Giovanni Paolo; Asprella Libonati, Giacinto; Casani, Augusto Pietro; Faralli, Mario; Giannoni, Beatrice; Gufoni, Mauro; Marcelli, Vincenzo; Marchetti, Pierpaolo; Pepponi, Emanuela; Vannucchi, Paolo; Nuti, Daniele

    2012-05-01

    The need for Class I and II studies on the efficacy of Semont's liberatory maneuver (SLM) in the treatment of posterior canal benign paroxysmal positional vertigo (PC-BPPV) motivated the present double-blind randomized trial on the short-term efficacy of SLM. A total of 342 patients with unilateral PC-BPPV were recruited for a multicenter study. Patients were randomly assigned to treatment by SLM (n = 174) or sham treatment (n = 168). Subjects were followed up twice (1 and 24 h) with the Dix-Hallpike maneuver by blinded examiners. At the 1 and 24 h follow-up, 79.3 and 86.8%, respectively, of patients undergoing SLM had recovered from vertigo, compared to none of the patients undergoing the sham maneuver (p < 0.0001). Patients who manifested liberatory nystagmus at the end of SLM showed a significantly higher percentage of recovery (87.1 vs. 55.7%; p < 0.0001). To the best of our knowledge, this is the first Class I study on the efficacy of SLM. SLM proved highly effective with respect to the sham maneuver (p < 0.0001). Liberatory nystagmus was demonstrated to be a useful prognostic factor for the efficacy of treatment. The present Class I study of efficacy of SLM changes the level of recommendation of the maneuver for treating PC-BPPV from level C to level B.

  1. Absence of QTc Prolongation with Domperidone: A Randomized, Double-Blind, Placebo- and Positive-Controlled Thorough QT/QTc Study in Healthy Volunteers

    PubMed Central

    Biewenga, Jeike; Keung, Chi; Solanki, Bhavna; Natarajan, Jaya; Leitz, Gerhard; Deleu, Sofie; Soons, Paul

    2015-01-01

    Domperidone effects on QTc duration were assessed in a single-center, double-blind, four-way crossover study of 44 healthy participants randomized to one of four treatment sequences consisting of four treatment periods separated by 4–9 days washout. On Day 1 of each 4-day period, participants began oral domperidone 10 or 20 mg q.i.d., matching placebo q.i.d., or single-dose moxifloxacin 400 mg (positive control)/placebo q.i.d. In each period, triplicate 12-lead electrocardiograms were recorded at baseline (30, 20, and 10 minutes predose), 8 timepoints after dosing on Days 1 and 4, and predose on Day 4. In mixed effects models, the largest difference for domperidone in least squares means for change from baseline QTcP versus placebo was 3.4 milliseconds (20 mg q.i.d., Day 4), 90% CI: 1.0–5.9, and <10 milliseconds at all timepoints for both domperidone dosages. Moxifloxacin response confirmed assay sensitivity. Participants achieved expected domperidone plasma exposures. No significant exposure-response relationship was found for QTc increase per ng/mL domperidone (90% CI of the slope estimate included zero at mean Cmax on Day 1 or Day 4). In summary, domperidone at doses up to 80 mg/day did not cause clinically relevant QTc interval prolongation. PMID:26097791

  2. Positive regulation of meiotic DNA double-strand break formation by activation of the DNA damage checkpoint kinase Mec1(ATR).

    PubMed

    Gray, Stephen; Allison, Rachal M; Garcia, Valerie; Goldman, Alastair S H; Neale, Matthew J

    2013-07-31

    During meiosis, formation and repair of programmed DNA double-strand breaks (DSBs) create genetic exchange between homologous chromosomes-a process that is critical for reductional meiotic chromosome segregation and the production of genetically diverse sexually reproducing populations. Meiotic DSB formation is a complex process, requiring numerous proteins, of which Spo11 is the evolutionarily conserved catalytic subunit. Precisely how Spo11 and its accessory proteins function or are regulated is unclear. Here, we use Saccharomyces cerevisiae to reveal that meiotic DSB formation is modulated by the Mec1(ATR) branch of the DNA damage signalling cascade, promoting DSB formation when Spo11-mediated catalysis is compromised. Activation of the positive feedback pathway correlates with the formation of single-stranded DNA (ssDNA) recombination intermediates and activation of the downstream kinase, Mek1. We show that the requirement for checkpoint activation can be rescued by prolonging meiotic prophase by deleting the NDT80 transcription factor, and that even transient prophase arrest caused by Ndt80 depletion is sufficient to restore meiotic spore viability in checkpoint mutants. Our observations are unexpected given recent reports that the complementary kinase pathway Tel1(ATM) acts to inhibit DSB formation. We propose that such antagonistic regulation of DSB formation by Mec1 and Tel1 creates a regulatory mechanism, where the absolute frequency of DSBs is maintained at a level optimal for genetic exchange and efficient chromosome segregation.

  3. Screening the allergenic repertoires of wheat and maize with sera from double-blind, placebo-controlled food challenge positive patients.

    PubMed

    Weichel, M; Vergoossen, N J; Bonomi, S; Scibilia, J; Ortolani, C; Ballmer-Weber, B K; Pastorello, E A; Crameri, R

    2006-01-01

    Food allergy to wheat and maize is an increasing factor of deterioration of life quality, especially childhood and can, in rare cases, even induce anaphylaxis. Although omega-5 gliadin from wheat and maize lipid transfer protein have been characterized as major cereal allergens on the molecular level, the list of food allergens is far to be complete. To identify the IgE-binding repertoires of wheat and maize we screened respective cDNA libraries displayed on phage surface with sera from patients with a confirmed food allergy. The study included six patients with a positive double-blind, placebo-controlled food challenge (DBPCFC) to wheat, nine patients with a positive DBPCFC to maize, and six patients with anaphylactic reactions after ingestion of wheat. The enriched sequences encoding IgE-binding proteins showed heterogeneous repertoires for both, wheat and maize. The selected wheat repertoire yielded 12, the maize repertoire 11 open reading frames. Among these we identified allergens belonging to already characterized allergens families, such as gliadin, profilin and beta-expansin. Besides, we found novel proteins with high cross-reactive potential, such as thioredoxins, as well as sequences that had so far not been related to cereal allergy at all. The IgE-binding capacity of some selected proteins was evaluated in vitro and cross-reactivity was demonstrated by competition ELISA. With regard to the heterogeneity of the characterized sequences as well as to the biochemical nature of the new allergens detected we conclude that wheat and maize-related food allergy is more complex than so far anticipated.

  4. CD40L confers helper functions to human intra-melanoma class-I-restricted CD4+CD8+ double positive T cells

    PubMed Central

    Parrot, Tiphaine; Oger, Romain; Benlalam, Houssem; Raingeard de la Blétière, Diane; Jouand, Nicolas; Coutolleau, Anne; Preisser, Laurence; Khammari, Amir; Dréno, Brigitte; Guardiola, Philippe; Delneste, Yves; Labarrière, Nathalie; Gervois, Nadine

    2016-01-01

    ABSTRACT Although CD4+CD8+ double positive (DP) T cells represent a small fraction of peripheral T lymphocytes in healthy human donors, their frequency is often increased under pathological conditions (in blood and targeted tissues). In solid cancers such as melanoma, we previously demonstrated an enrichment of tumor reactive CD4lowCD8highαβ DP T cells among tumor-infiltrating lymphocytes of unknown function. Similarly to their single positive (SP) CD8+ counterparts, intra-melanoma DP T cells recognized melanoma cell lines in an HLA-class-I restricted context. However, they presented a poor cytotoxic activity but a strong production of diverse Th1 and Th2 cytokines. The aim of this study was to clearly define the role of intra-melanoma CD4lowCD8highαβ DP T cells in the antitumor immune response. Based on a comparative transcriptome analysis between intra-melanoma SP CD4+, SP CD8+ and DP autologous melanoma-infiltrating T-cell compartments, we evidenced an overexpression of the CD40L co-stimulatory molecule on activated DP T cells. We showed that, like SP CD4+ T cells, and through CD40L involvement, DP T cells are able to induce both proliferation and differentiation of B lymphocytes and maturation of functional DCs able to efficiently prime cytotoxic melanoma-specific CD8 T-cell responses. Taken together, these results highlight the helper potential of atypical DP T cells and their role in potentiating antitumor response. PMID:28123891

  5. Anaplastic large cell lymphoma arises in thymocytes and requires transient TCR expression for thymic egress

    PubMed Central

    Malcolm, Tim I. M.; Villarese, Patrick; Fairbairn, Camilla J.; Lamant, Laurence; Trinquand, Amélie; Hook, C. Elizabeth; Burke, G. A. Amos; Brugières, Laurence; Hughes, Katherine; Payet, Dominique; Merkel, Olaf; Schiefer, Ana-Iris; Ashankyty, Ibraheem; Mian, Shahid; Wasik, Mariusz; Turner, Martin; Kenner, Lukas; Asnafi, Vahid; Macintyre, Elizabeth; Turner, Suzanne D.

    2016-01-01

    Anaplastic large cell lymphoma (ALCL) is a peripheral T-cell lymphoma presenting mostly in children and young adults. The natural progression of this disease is largely unknown as is the identity of its true cell of origin. Here we present a model of peripheral ALCL pathogenesis where the malignancy is initiated in early thymocytes, before T-cell receptor (TCR) β-rearrangement, which is bypassed in CD4/NPM–ALK transgenic mice following Notch1 expression. However, we find that a TCR is required for thymic egress and development of peripheral murine tumours, yet this TCR must be downregulated for T-cell lymphomagenesis. In keeping with this, clonal TCR rearrangements in human ALCL are predominantly in-frame, but often aberrant, with clonal TCRα but no comparable clonal TCRβ rearrangement, yielding events that would not normally be permissive for survival during thymic development. Children affected by ALCL may thus harbour thymic lymphoma-initiating cells capable of seeding relapse after chemotherapy. PMID:26753883

  6. A physical map linking the five CD1 human thymocyte differentiation antigen genes.

    PubMed Central

    Yu, C Y; Milstein, C

    1989-01-01

    Human CD1 is a family of thymocyte differentiation antigens which consist of heavy chains with mol. wts between 43 and 49 kd binding to beta 2 microglobulin. They are distant relatives of the major histocompatibility complex (MHC) class I and II products. Five human CD1 genes have been described. Three (CD1A, -B and -C) code for the serologically defined CD1a, -b and -c antigens. The protein products of the other two genes, CD1D and CD1E, remain unknown. All CD1 genes are located on chromosome 1 and hence are independent of the MHC locus. In this paper, the tight linkage of the CD1 genes has been established by pulse field gel electrophoresis, cosmid cloning and walking techniques. The 190 kb of DNA linking all five CD1 genes has been spanned by 14 overlapping cosmids. The order of the genes in the CD1 complex is CD1D-CD1A-CD1C-CD1B-CD1E, and, with the exception of CD1B, they are arranged in the same transcriptional orientation. The genes are evenly spaced in the complex except for the distance between CD1D and CD1A, which is two to three times greater than the average. Images PMID:2583117

  7. Inhibiting the repair of DNA damage induced by gamma irradiation in rat thymocytes

    SciTech Connect

    Smit, J.A.; Stark, J.H.

    1994-01-01

    This study assessed the ability of 11 established and potential radiosensitizing agents to retard the repair of radiation-induced DNA damage with a view to enhancing the immunosuppressive effects of in vivo lymphoid irradiation. The capability of irradiated rat thymocytes to repair DNA damage was assessed by an adaptation of the fluorimetric unwinding method. Three compounds, 3-aminobenzamide (3-AB), novobiocin and flavone-8-acetic acid (FAA), inhibited repair significantly. We also report the effect of low-dose irradiation combined with repair inhibitors on the relationship between DNA strand breaks, fragmentation, cell viability and use of nicotinamide adenine dinucleotide (NAD). DNA fragmentation was increased by 1 mM/l FAA, 1 mM/l novobiocin and 50 {mu}M/l RS-61443 within 3 h of incubation. The latter two compounds also proved cytotoxic. All three drugs augmented the effect of ionizing radiation on the use of NAD. Of the agents investigated, FAA showed the most promise for augmenting the immunosuppressive action of irradiation at nontoxic, pharmacokinetically achievable concentrations. 33 refs., 1 fig., 2 tabs.

  8. Pilot study of anti-thymocyte globulin plus mycophenolate mofetil in recent-onset diffuse scleroderma.

    PubMed

    Stratton, R J; Wilson, H; Black, C M

    2001-01-01

    To assess the safety and efficacy of anti-thymocyte globulin (ATG) followed by mycophenolate mofetil (MMF) in the treatment of diffuse scleroderma. A pilot study of 13 patients with recent-onset diffuse scleroderma was carried out. Patients received ATG for 5 days, followed by MMF for 12 months. We recorded adverse events, scleroderma skin score, hand contractures, EuroQol score, scleroderma functional assessment, pulmonary function studies, echocardiogram and plasma creatinine concentration. Mean skin score decreased during the study from 28 at baseline to 17 after 12 months of MMF (P<0.01). Hand contractures worsened during the study. Mean measurements of systemic disease remained stable. One patient died after a scleroderma renal crisis. Five patients developed serum sickness after ATG treatment, but this was controlled by corticosteroid therapy. MMF therapy was well tolerated. ATG and MMF appear safe in scleroderma. The improvement in skin score and the apparent stability of systemic disease during the study period suggest that controlled studies of these agents are justified.

  9. Effect of concanavalin A on expression of cell surface sialyltransferase activity of mouse thymocytes.

    PubMed Central

    Painter, R G; White, A

    1976-01-01

    Incubation of mouse thymocytes with mitogenic concentrations of concanavalin A causes a 2-fold increase in cell-surface-associated (but not total cell) sialyltransferase activity (ectosialyltransferase, CMP-N-acetylneuraminate:D-galactosyl-glycoprotein N-acetylneuraminyltransferase, EC 2.4.99.1) as judged by incorporation of [14C]sialic acid into endogenous cell acceptors and into added desialylated fetuin acceptor. The concanavalin-A-induced enhancement of enzymic activity is essentially complete within 1 hr after addition of mitogen and remains at elevated levels for 12 hr, declining rapidly thereafter. Intact cells labeled previously with [14C]sialic acid and then incubated briefly with hydrolytic enzymes, including neuraminidase and insoluble trypsin, released 43-66% of total cell-associated radioactivity without appreciably changing cell viability. Alterations in sialyltransferase activity due to concanavalin A treatment could not be explained by a mitogen-mediated (a) uptake of radioactive precursors, (b) cell death, (c) increased product catabolism, or (d) activation of sialyltransferase by mitogen binding to the enzyme. Furthermore, the process does not require active protein synthesis. The results are consistent with a rapid concanavalin-A-induced exposure of potential enzymic activity that was previously inaccessible to substrate. PMID:1083027

  10. Ozone-induced dissociation on a traveling wave high-resolution mass spectrometer for determination of double-bond position in lipids.

    PubMed

    Vu, Ngoc; Brown, Jeffery; Giles, Kevin; Zhang, Qibin

    2017-09-15

    The position of C=C within fatty acyl chains affects the biological function of lipids. Ozone-induced dissociation mass spectrometry (OzID-MS) has great potential in determination of lipid double-bond position, but has generally been implemented on low-resolution ion trap mass spectrometers. In addition, most of the OzID-MS experiments carried out so far were focused on the sodiated adducts of lipids; fragmentation of the most commonly observed protonated ions generated in LC/MS-based lipidomics workflow has been less explored. Ozone generated in line from an ozone generator was connected to the trap and transfer gas supply line of a Synapt G2 high-resolution mass spectrometer. Protonated ions of different phosphatidylcholines (PC) were generated by electrospray ionization through direct infusion. Different parameters, including traveling wave height and velocity, trap entrance and DC potential, were adjusted to maximize the OzID efficiency. sn-positional isomers and cis/trans isomers of lipids were compared for their reactivity with ozone. Traveling wave height and velocity were tuned to prolong the encounter time between lipid ions and ozone, and resulted in improved OzID efficiency, as did increasing trapping region DC and entrance potential. Under optimized settings, at least 1000 times enhancement in OzID efficiency was achieved compared to that under default settings for monounsaturated PC standards. Monounsaturated C=C in the sn-2 PC isomer reacted faster with ozone than the sn-1 isomer. Similarly, the C=C in trans PC reacted faster than in cis PC. This is the first implementation of OzID in the trap and transfer region of a traveling wave enabled high-resolution mass spectrometer. The OzID reaction efficiency is significantly improved by slowing down ions in the trap region for their prolonged interaction with ozone. This will facilitate application of high-resolution OzID-MS in lipidomics. Copyright © 2017 John Wiley & Sons, Ltd.

  11. Extraordinary positional cervical spinal cord compression in extension position as a rare cause of postoperative progressive myelopathy after cervical posterior laminoplasty detected using the extension/flexion positional CT myelography: one case after laminectomy following failure of a single-door laminoplasty/one case after double-door laminoplasty without interlaminar spacers.

    PubMed

    Fujiwara, Yasushi; Manabe, Hideki; Harada, Takahiro; Izumi, Bunichiro; Adachi, Nobuo

    2017-05-01

    Posterior cervical laminectomies and laminoplasties are common treatments for cervical spondylotic myelopathy. However, recent studies demonstrated that positional spinal cord compression occurred after cervical laminectomies and caused postoperative progressive myelopathy. Although there were no such reports after laminoplasties, we report two cases in which symptomatic extraordinary positional spinal cord compression occurred after laminoplasties in this paper. This study included two patients who showed progressive myelopathy: one case after a laminectomy following failure of a single-door laminoplasty and one case after a double-door laminoplasty without interlaminar spacers. The MRIs showed mild cord compression in the neutral position in both cases. However, the patients could not extend their necks, because it triggered severe neck pain and numbness. Therefore, the positional CT myelography (CTM) was taken in the flexion and extension positions, and it showed severe spinal cord compression only in the extension position. Posterior instrumented fusions were performed for both patients, which improved their symptoms. This paper demonstrates that postoperative positional spinal cord compression during neck extension caused a progressive myelopathy even after laminoplasty. When myelopathy symptoms worsen after laminoplasties, we recommend positional CTM/MRI evaluation, even though there is no apparent cord compression in the neutral MRI.

  12. Double positivity for HPV DNA/p16 in tonsillar and base of tongue cancer improves prognostication: Insights from a large population-based study.

    PubMed

    Garnaes, Emilie; Frederiksen, Kirsten; Kiss, Katalin; Andersen, Luise; Therkildsen, Marianne H; Franzmann, Maria B; Specht, Lena; Andersen, Elo; Norrild, Bodil; Kjaer, Susanne K; von Buchwald, Christian

    2016-12-01

    The aim was to explore the overall survival (OS) for palatine tonsillar squamous cell carcinoma (TSCC), subdivided, according to certainty of tonsillar tumour origin, into specified tonsillar squamous cell carcinomas (STSCCs) and nonspecified tonsillar squamous cell carcinomas (NSTSCCs), and base of tongue squamous cell carcinoma (BSCC) when stratifying for HPV DNA status, p16 expression and combined HPV/p16 status. We included all patients (n = 797) diagnosed with TSCCs and BSCCs in Eastern Denmark as registered in the Danish Head and Neck Cancer Group (DAHANCA) database and the Danish Pathology Databank, 2000-2010. Patients were treated according to national guidelines (radiotherapy +/- concomitant cisplatin). All specimens were analysed using HPV DNA PCR and p16 immunohistochemistry. Clinical information was retrieved from the DAHANCA database and the Danish National Patient Registry. Information on vital status was obtained from the Danish Civil Registration System. We observed improved OS for HPV+/p16+ BSCCs compared to HPV-/p16- (hazard ratio for death [HR], 0.15; 95% CI, 0.09-0.24). Among STSCCs, HPV+/p16+ showed the lowest HR (0.19, 95% CI, 0.13-0.29); whereas, HPV-/p16+ showed an intermediate HR (0.39; 95% CI, 0.22-0.70). For NSTSCCs, HPV+/p16+ and HPV-/p16+ showed similar OS (HRs, 0.39; 95% CI, 0.26-0.59; and 0.48; 95% CI, 0.24-0.95, respectively). Combined HPV+/p16+ was a significantly better prognostic marker in BSCCs and STSCCs than HPV DNA and p16, alone (all p-values < 0.05). Whereas, combined testing in NSTSCC was not better than p16 (p = 0.53), alone. In conclusion, double positivity for HPV/p16 in conjunction with the certainty of tumour site improved prognosis. © 2016 UICC.

  13. Five-degree, 10-degree, and 20-degree reverse Trendelenburg position during functional endoscopic sinus surgery: a double-blind randomized controlled trial.

    PubMed

    Gan, Eng Cern; Habib, Al-Rahim R; Rajwani, Alykhan; Javer, Amin R

    2014-01-01

    Using the reverse Trendelenburg position (RTP) during functional endoscopic sinus surgery (FESS) is a safe, simple, and cost-free method that has been found to reduce intraoperative blood loss. However, the critical angle of RTP that produces the least amount of bleeding without compromising surgical technique and safety remains unanswered. The objective of this study was to assess the effects of 5-degree, 10-degree, and 20-degree RTP (5-RTP, 10-RTP, and 20-RTP, respectively) on intraoperative bleeding during FESS. This double-blind randomized controlled trial involved 75 patients with chronic rhinosinusitis (CRS) with and without nasal polyposis undergoing FESS. Twenty-five patients were enrolled into each group: 5-RTP, 10-RTP, and 20-RTP. Boezaart endoscopic field-of-view score (BS), total blood loss (TBL), mean arterial blood pressure (MABP), operating time, and blood loss per minute were recorded. An intention-to-treat analysis was used, with a Bonferroni adjustment for multiple comparisons. Intervention groups were comparable in age, sex, nasal polyposis, and disease severity. Mean values of BS and TBL were as follows: 5-RTP (2.0, 231 mL), 10-RTP (1.8, 230 mL), and 20-RTP (1.4, 135 mL). The differences in means were significant for BS (p < 0.01) and TBL (p = 0.03). There was no significant difference in MABP (p = 0.85), operating time (p = 0.10), or blood loss per minute (p = 0.11) between the 3 groups. Pairwise comparison between 5-RTP vs 20-RTP found significant difference in BS (p < 0.01) but not TBL (p = 0.04). Significance was not found in similar comparisons of 10-RTP vs 20-RTP and 5-RTP vs 10-RTP (p > 0.03). FESS in 20-RTP produced the best BS and lowest blood loss without compromising surgical technique. © 2013 ARS-AAOA, LLC.

  14. Secondary prevention of cardiogenic arterial thromboembolism in the cat: The double-blind, randomized, positive-controlled feline arterial thromboembolism; clopidogrel vs. aspirin trial (FAT CAT).

    PubMed

    Hogan, Daniel F; Fox, Philip R; Jacob, Kristin; Keene, Bruce; Laste, Nancy J; Rosenthal, Steven; Sederquist, Kimberly; Weng, Hsin-Yi

    2015-12-01

    To determine if clopidogrel administration is associated with a reduced likelihood of recurrent cardiogenic arterial thromboembolism (CATE) in cats compared to aspirin administration. Secondary aims were to determine if clopidogrel administration had an effect on the composite endpoint of recurrent CATE and cardiac death and to identify adverse effects of chronic clopidogrel or aspirin therapy. Seventy-five cats that survived a CATE event. Multicenter, double-blind, randomized, positive-controlled study. Cats were assigned to clopidogrel (18.75 mg/cat PO q 24 h) or aspirin (81 mg/cat PO q 72 h). Kaplan-Meier survival curves were created for each endpoint and the log rank test performed to compare treatment groups with respect to time to event and the likelihood of the event occurring. The mean age of all cats was 8.0 ± 3.5 yr and 57/75 (76%) were male (p < 0.001); 62/75 (83%) were mixed breed with the remainder including Persian, Abyssinian, American Shorthair, Bengal, Birman, Himalayan, Maine Coon, Ragdoll, Snowshoe, and Sphynx breeds. Only 15% (11/75) of cats had a history of heart disease recorded prior to the CATE event. Clopidogrel administration was associated with significantly reduced likelihood of recurrent CATE compared to aspirin (p = 0.024) and had a longer median time to recurrence [443 (95% CI 185-990) days vs. 192 (95% CI 62-364) days, respectively]. Clopidogrel was also associated with a significantly reduced likelihood of the composite endpoint of recurrent CATE or cardiac death (p = 0.033) with a longer median time to event [346 (95% CI 146-495) days vs. 128 (95% CI 58-243) days]. Clopidogrel administration significantly reduces the likelihood of recurrent CATE compared with aspirin in cats; both drugs were well tolerated. Copyright © 2015 Elsevier B.V. All rights reserved.

  15. Coreceptor signal strength regulates positive selection but does not determine CD4/CD8 lineage choice in a physiologic in vivo model.

    PubMed

    Erman, Batu; Alag, Amala S; Dahle, Oyvind; van Laethem, François; Sarafova, Sophia D; Guinter, Terry I; Sharrow, Susan O; Grinberg, Alexander; Love, Paul E; Singer, Alfred

    2006-11-15

    TCR signals drive thymocyte development, but it remains controversial what impact, if any, the intensity of those signals have on T cell differentiation in the thymus. In this study, we assess the impact of CD8 coreceptor signal strength on positive selection and CD4/CD8 lineage choice using novel gene knockin mice in which the endogenous CD8alpha gene has been re-engineered to encode the stronger signaling cytoplasmic tail of CD4, with the re-engineered CD8alpha gene referred to as CD8.4. We found that stronger signaling CD8.4 coreceptors specifically improved the efficiency of CD8-dependent positive selection and quantitatively increased the number of MHC class I (MHC-I)-specific thymocytes signaled to differentiate into CD8+ T cells, even for thymocytes expressing a single, transgenic TCR. Importantly, however, stronger signaling CD8.4 coreceptors did not alter the CD8 lineage choice of any MHC-I-specific thymocytes, even MHC-I-specific thymocytes expressing the high-affinity F5 transgenic TCR. This study documents in a physiologic in vivo model that coreceptor signal strength alters TCR-signaling thresholds for positive selection and so is a major determinant of the CD4:CD8 ratio, but it does not influence CD4/CD8 lineage choice.

  16. Immunomodulatory role of Emblica officinalis in arsenic induced oxidative damage and apoptosis in thymocytes of mice

    PubMed Central

    2013-01-01

    Background Arsenic is widely distributed in the environment and has been found to be associated with the various health related problems including skin lesions, cancer, cardiovascular and immunological disorders. The fruit extract of Emblica officinalis (amla) has been shown to have anti-oxidative and immunomodulatory properties. In view of increasing health risk of arsenic, the present study has been carried out to investigate the protective effect of amla against arsenic induced oxidative stress and apoptosis in thymocytes of mice. Methods Mice were exposed to arsenic (sodium arsenite 3 mg/kg body weight p.o.) or amla (500 mg/kg body weight p.o.) or simultaneously with arsenic and amla for 28 days. The antioxidant enzyme assays were carried out using spectrophotometer and generation of ROS, apoptotic parameters, change in cell cycle were carried out using flow cytometer following the standard protocols. Results Arsenic exposure to mice caused a significant increase in the lipid peroxidation, ROS production and decreased cell viability, levels of reduced glutathione, the activity of superoxide dismutase, catalase, cytochrome c oxidase and mitochondrial membrane potential in the thymus as compared to controls. Increased activity of caspase-3 linked with apoptosis assessed by the cell cycle analysis and annexin V/PI binding was also observed in mice exposed to arsenic as compared to controls. Co-treatment with arsenic and amla decreased the levels of lipid peroxidation, ROS production, activity of caspase-3, apoptosis and increased cell viability, levels of antioxidant enzymes, cytochrome c oxidase and mitochondrial membrane potential as compared to mice treated with arsenic alone. Conclusions The results of the present study exhibits that arsenic induced oxidative stress and apoptosis significantly protected by co-treatment with amla that could be due to its strong antioxidant potential. PMID:23889914

  17. TNF regulates thymocyte production by apoptosis and proliferation of the triple negative (CD3-CD4-CD8-) subset.

    PubMed

    Baseta, J G; Stutman, O

    2000-11-15

    TNF is a proinflammatory cytokine with opposing death/no-death effects in vivo and in vitro. Our studies showed that TNF regulates mouse thymocyte production, inducing both apoptosis and proliferation of the most immature CD3(-)CD4(-)CD8(-) triple negative (TN) subset within a broad range of dosages (10(1)-10(5) pg/ml) in the presence of IL-7. TNF apoptosis affected only the TN3 (CD44(-)CD25(+)) and TN4 (CD44(-)CD25(-)) subsets that expressed both TNFR-p55 and -p75. Although each TNFR alone could mediate TNF apoptosis, maximal apoptosis was seen in C57BL/6J wild type, which expressed both TNFRs. TNF also induced proliferation of TN3 cells at higher doses (10(4)-10(5) pg/ml) mediated only by TNFR-p75. Both anti-TNFR-p55 and -TNFR-p75 mAb inhibited apoptosis but only anti-p75 inhibited proliferation. TNF also regulated TN proliferation to IL-7 because TNFR knockout (KO), TNF KO, and TNF/lymphotoxin alpha and beta triple KO mice showed 2- to 3-fold increased responses not seen in C57BL/6J wild type. In vivo, TNFR KO mice showed thymic hypertrophy with a 60% increase in total thymocytes, with no effect on the CD4/CD8 subsets. We conclude that TNF maintains homeostatic control of total thymocyte production by negative selection of TN3 and TN4 prothymocytes and down-regulation of their proliferation to endogenous IL-7.

  18. V(D)J recombination generates a high frequency of nonstandard TCR D[delta]-associated rearrangements in thymocytes

    SciTech Connect

    Carroll, A.M.; Slack, J.K.; Mu, Xiaochun )

    1993-03-15

    The standard products of V(D)J recombination are coding junctions, which encode Ag receptor polypeptide, and their commonly excised reciprocal products, signal junctions. Additional nonstandard products also have been detected, mostly in artificial recombination substrate studies. The occurrence of nonstandard products, including pseudonormal, hybrid, and open/shut junctions, indicates significant indeterminacy of the V(D)J recombinase. However, the incidence of nonstandard products of endogenous Ag receptor genes in vivo has not been specifically addressed. The data presented here show that for the TCR-[delta] locus, D element-associated recombination in mouse thymocytes results in a high incidence of nonstandard recombination products. D[delta]1-D[delta]2 rearrangements, both chromosome retained and excised episomal products, were studied by polymerase chain reaction amplification, cloning, and sequence analysis. The proximity of D[delta]1 and D[delta]2 elements, and the fact that both are flanked by 5[prime] and 3[prime] recombination signal sequences with 12-bp and 23-bp spacers, respectively, results in frequent pseudonormal joining. The resulting products are signal junctions retained on the chromosome. Excised episomal products include coding junctions, hybrid junctions formed in apparent violation of the 12/23 spacer rule, and standard signal junctions; some signal junctions show evidence of imprecise cleavage. Evidence for open/shut and/or oligonucleotide capture events was also seen. Similar rearrangements were detectable in thymocytes of mutant scid mice. These findings indicate a high degree of indeterminancy of V(D)J recombinase-mediated D[delta]1-D[delta]2 rearrangement in both wild-type and scid thymocytes. This indeterminacy affects the productive potential of TCR-[delta] loci. 45 refs., 4 figs.

  19. Cell proliferation and thymocyte subset reconstitution in sublethally irradiated mice: Compared kinetics of endogenous and intrathymically transferred progenitors

    SciTech Connect

    Penit, C.; Ezine, S.

    1989-07-01

    After sublethal (6 Gy) whole-body irradiation, the C57BL/Ba (Thy-1.1) murine thymus regenerated in two waves, on days 3-10 and 25-32, separated by a severe relapse. The second phase of depletion-reconstitution reproduced the first one, in a less synchronous manner. The depletion affected all cell subsets, but CD4+ CD8- cells decreased later than immature cells. Cell proliferation, measured by BrdUrd incorporation, started on day 3 after irradiation and concerned CD4- CD8-, CD4- CD8+, and CD4+ CD8+ cells, sequentially. CD4+ CD8- cells never represented a significant percentage of cycling cells. When irradiation was immediately followed by an intrathymic injection of 10(5) C57BL/Ka (Thy-1.2) bone marrow cells, the relapse in thymus reconstitution was no longer observed. Detected with anti-Thy-1.2 antibodies, donor cells started cycling on day 14 and showed only one wave of proliferation. In these chimeras, recipient thymocytes behave exactly like thymocytes of solely irradiated mice. Intrathymically transferred CD4- CD8- thymocytes 10(5) showed the same proliferation kinetics as endogenous cells, with a peak in number on day 10 but completely disappeared from the thymus on days 14-21. These data reflect maturational differences between intrathymic and bone marrow precursor cells and suggest different radiosensitivities not linked to proliferative status. The resting state of the thymus immigrants was shown by the absence of Thy-1 acquisition by bone marrow cells continuously labeled for 10 days with BrdUrd in vivo before intrathymic transfer. When such labeled bone marrow cells were injected in the thymus, only the minor BrdUrd- subset gave rise to Thy-1+ cells.

  20. Epitope-specific immunotherapy targeting CD4-positive T cells in coeliac disease: two randomised, double-blind, placebo-controlled phase 1 studies.

    PubMed

    Goel, Gautam; King, Tim; Daveson, A James; Andrews, Jane M; Krishnarajah, Janakan; Krause, Richard; Brown, Gregor J E; Fogel, Ronald; Barish, Charles F; Epstein, Roger; Kinney, Timothy P; Miner, Philip B; Tye-Din, Jason A; Girardin, Adam; Taavela, Juha; Popp, Alina; Sidney, John; Mäki, Markku; Goldstein, Kaela E; Griffin, Patrick H; Wang, Suyue; Dzuris, John L; Williams, Leslie J; Sette, Alessandro; Xavier, Ramnik J; Sollid, Ludvig M; Jabri, Bana; Anderson, Robert P

    2017-07-01

    A gluten-free diet is the only means to manage coeliac disease, a permanent immune intolerance to gluten. We developed a therapeutic vaccine, Nexvax2, designed to treat coeliac disease. Nexvax2 is an adjuvant-free mix of three peptides that include immunodominant epitopes for gluten-specific CD4-positive T cells. The vaccine is intended to engage and render gluten-specific CD4-positive T cells unresponsive to further antigenic stimulation. We assessed the safety and pharmacodynamics of the vaccine in patients with coeliac disease on a gluten-free diet. We did two randomised, double-blind, placebo-controlled, phase 1 studies at 12 community sites in Australia, New Zealand, and the USA, in HLA-DQ2·5-positive patients aged 18-70 years who had coeliac disease and were on a gluten-free diet. In the screening period for ascending dose cohorts, participants were randomly assigned (1:1) by central randomisation with a simple block method to a double-blind crossover, placebo-controlled oral gluten challenge. Participants with a negative interferon γ release assay to Nexvax2 peptides after the screening oral gluten challenge were discontinued before dosing. For the biopsy cohorts, the screening period included an endoscopy, and participants with duodenal histology who had a Marsh score of greater than 1 were discontinued before dosing. Participants were subsequently randomly assigned to either Nexvax2 or placebo in ascending dose cohorts (2:1) and in biopsy cohorts (1:1) by central randomisation with a simple block method. In the three-dose study, participants received either Nexvax2 60 μg, 90 μg, or 150 μg weekly, or placebo over 15 days; in a fourth biopsy cohort, patients received either Nexvax2 at the maximum tolerated dose (MTD) or placebo. In the 16-dose study, participants received Nexvax2 150 μg or 300 μg or placebo twice weekly over 53 days; in a third biopsy cohort, patients also received either Nexvax2 at the MTD or placebo. In the 4-week post

  1. Tthyd, a new thymocyte alloantigen linked to Igh-1. Implications for a switch mechanism for T cell antigen receptors.

    PubMed

    Owen, F L; Spurll, G M; Panageas, E

    1982-01-01

    Tthyd is an alloantigen coded for by a gene(s) near the immunoglobulin locus on chromosome 12 in the mouse. This T cell-specific antigen may be the third member of a family of antigen receptors on T cells encoded by a cluster of genes in the IgT-C region. This antigen is preferentially expressed on thymocytes in contrast to Tindd or Tsud that are expressed on peripheral T cells. The hypothesis that T cell receptors undergo a switch in surface isotype upon maturation is discussed.

  2. Tthyd, a new thymocyte alloantigen linked to Igh-1. Implications for a switch mechanism for T cell antigen receptors

    PubMed Central

    1982-01-01

    Tthyd is an alloantigen coded for by a gene(s) near the immunoglobulin locus on chromosome 12 in the mouse. This T cell-specific antigen may be the third member of a family of antigen receptors on T cells encoded by a cluster of genes in the IgT-C region. This antigen is preferentially expressed on thymocytes in contrast to Tindd or Tsud that are expressed on peripheral T cells. The hypothesis that T cell receptors undergo a switch in surface isotype upon maturation is discussed. PMID:6976417

  3. Identification of Stage-Specific Gene Modulation during Early Thymocyte Development by Whole-Genome Profiling Analysis after Aryl Hydrocarbon Receptor Activation

    PubMed Central

    Mills, Jeffrey H.; Lai, Zhi-Wei; Singh, Kameshwar P.; Middleton, Frank A.; Gasiewicz, Thomas A.; Silverstone, Allen E.

    2010-01-01

    The aryl hydrocarbon receptor (AHR) is a basic helix-loop-helix transcription factor, implicated as an important modulator of the immune system and of early thymocyte development. We have shown previously that AHR activation by the environmental contaminant and potent AHR agonist 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) leads to a significant decline in the percentage of S-phase cells in the CD3−CD4−CD8− triple-negative stage (TN) 3 and TN4 T-cell committed thymocytes 9 to 12 h after exposure. In the more immature TN1- or TN2-stage cells, no effect on cell cycle was observed. To identify early molecular targets, which could provide insight into how the AHR acts as a modulator of thymocyte development and cell cycle regulation, we performed gene-profiling experiments using RNA isolated from four intrathymic progenitor populations in which the AHR was activated for 6 or 12 h. This microarray analysis of AHR activation identified 108 distinct gene probes that were significantly modulated in the TN1–4 thymocyte progenitor stages. Although most of the genes identified have specific AHR recognition sequences, only seven genes were altered exclusively in the two T-cell committed stages of early thymocyte development (TN3 and TN4) in which the decline of S-phase cells is seen. Moreover, all seven of these genes were reduced in expression, and five of the seven are associated with cell cycle regulatory processes. These seven genes are novel targets for modulation by the TCDD-activated AHR and may be involved in the observed cell-cycle arrest and suppression of early thymocyte development. PMID:20159946

  4. Identification of stage-specific gene modulation during early thymocyte development by whole-genome profiling analysis after aryl hydrocarbon receptor activation.

    PubMed

    Laiosa, Michael D; Mills, Jeffrey H; Lai, Zhi-Wei; Singh, Kameshwar P; Middleton, Frank A; Gasiewicz, Thomas A; Silverstone, Allen E

    2010-05-01

    The aryl hydrocarbon receptor (AHR) is a basic helix-loop-helix transcription factor, implicated as an important modulator of the immune system and of early thymocyte development. We have shown previously that AHR activation by the environmental contaminant and potent AHR agonist 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) leads to a significant decline in the percentage of S-phase cells in the CD3(-)CD4(-)CD8(-) triple-negative stage (TN) 3 and TN4 T-cell committed thymocytes 9 to 12 h after exposure. In the more immature TN1- or TN2-stage cells, no effect on cell cycle was observed. To identify early molecular targets, which could provide insight into how the AHR acts as a modulator of thymocyte development and cell cycle regulation, we performed gene-profiling experiments using RNA isolated from four intrathymic progenitor populations in which the AHR was activated for 6 or 12 h. This microarray analysis of AHR activation identified 108 distinct gene probes that were significantly modulated in the TN1-4 thymocyte progenitor stages. Although most of the genes identified have specific AHR recognition sequences, only seven genes were altered exclusively in the two T-cell committed stages of early thymocyte development (TN3 and TN4) in which the decline of S-phase cells is seen. Moreover, all seven of these genes were reduced in expression, and five of the seven are associated with cell cycle regulatory processes. These seven genes are novel targets for modulation by the TCDD-activated AHR and may be involved in the observed cell-cycle arrest and suppression of early thymocyte development.

  5. Adenosine produced from adenine nucleotides through an interaction between apoptotic cells and engulfing macrophages contributes to the appearance of transglutaminase 2 in dying thymocytes.

    PubMed

    Sándor, Katalin; Pallai, Anna; Duró, Edina; Legendre, Pascal; Couillin, Isabelle; Sághy, Tibor; Szondy, Zsuzsa

    2017-03-01

    Transglutaminase 2 (TG2) has been known for a long time to be associated with the in vivo apoptosis program of various cell types, including T cells. Though the expression of the enzyme is strongly induced in mouse thymocytes following apoptosis induction in vivo, no significant induction of TG2 can be detected, when thymocytes are induced to die by the same stimuli in vitro indicating that signals arriving from the tissue environment are required for the proper in vivo induction of the enzyme. Previous studies from our laboratory have demonstrated that two of these signals, transforming growth factor-β (TGF-β) and retinoids, are produced by macrophages engulfing apoptotic cells. However, in addition to TGF-β and retinoids, engulfing macrophages produce adenosine as well. Here, we show that in vitro adenosine, adenosine, and retinoic acid or adenosine, TGF-β and retinoic acids together can significantly enhance the TG2 mRNA expression in dying thymocytes. The effect of adenosine is mediated via adenosine A2A receptors (A2ARs) and the A2AR-triggered adenylate cyclase signaling pathway. In accordance, loss of A2ARs in A2AR null mice significantly attenuates the in vivo induction of TG2 following apoptosis induction in the thymus indicating that adenosine indeed contributes in vivo to the apoptosis-related appearance of the enzyme. We also demonstrate that adenosine is produced extracellularly during engulfment of apoptotic thymocytes, partly from adenine nucleotides released via thymocyte pannexin-1 channels. Our data reveal a novel crosstalk between macrophages and apoptotic cells, in which apoptotic cell uptake-related adenosine production contributes to the appearance of TG2 in the dying thymocytes.

  6. Double triplex real-time PCR assay for simultaneous detection of Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus hominis, and Staphylococcus haemolyticus and determination of their methicillin resistance directly from positive blood culture bottles.

    PubMed

    Kilic, Abdullah; Basustaoglu, A Celal

    2011-12-01

    We developed and validated here a double triplex real-time PCR assay to simultaneously detect and identify Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus hominis, Staphylococcus haemolyticus and their methicillin resistance in a single reaction directly from Gram-positive cocci-in-clusters (GPCs)-positive blood culture bottles. From August 15, 2009 through February 15, 2010, 238 GPC-positive samples were collected and identified by conventional methods as 11 methicillin-resistant S. aureus (MRSA), 28 methicillin-susceptible S. aureus (MSSA), 176 MR coagulase-negative staphylococci (MRCoNS), 21 MSCoNS and two Enterococcus faecalis. The double triplex real-time PCR assay was targeted and detected tuf, nuc and mecA genes in the first tube and atlE, gap and mvaA genes in the second tube which could be run simultaneously. The detection limit of the assay was found at 10(3) CFU/ml for the atleE gene, 10(4) CFU/ml for the mva gene and 10(5) CFU/ml for gap, nuc, mecA and tuf genes based on seeding experiments. All Staphylococcus species except two S. epidermidis were correctly identified by the assay. The double triplex real-time PCR assay quickly and accurately detects S. aureus, S. epidermidis, S. hominis and S. haemolyticus and their methicillin resistance in a single reaction directly from positive blood culture bottles within 83 min.

  7. Surface Oxidation under Ambient Air—Not Only a Fast and Economical Method to Identify Double Bond Positions in Unsaturated Lipids But Also a Reminder of Proper Lipid Processing

    PubMed Central

    2015-01-01

    A simple, fast approach elucidated carbon–carbon double bond positions in unsaturated lipids. Lipids were deposited onto various surfaces and the products from their oxidation in ambient air were observed by electrospray ionization (ESI) mass spectrometry (MS). The most common oxidative products, aldehydes, were detected as transformations at the cleaved double bond positions. Ozonides and carboxylic acids were generated in certain lipids. Investigations of the conditions controlling the appearance of these products indicated that the surface oxidation depends on light and ambient air. Since the lipid oxidation was slower in a high concentration of ozone, singlet oxygen appeared to be a parallel oxidant for unsaturated lipids. The 3-hydroxyl group in the sphingoid base of sulfatides offered some protection from oxidation for the Δ4,5-double bond, slowing its oxidation rate relative to that of the isolated double bond in the N-linked fatty acyl chain. Direct sampling by thin-layer chromatography (TLC)-ESI-MS provides a powerful approach to elucidate detailed structural information on biological samples. Co-localization of the starting lipids and their oxidation products after TLC separation allowed assignment of the native unsaturation sites. Phosphatidylserine and N,N-dimethyl phosphatidylethanolamine isomers in a bovine brain total lipid extract were distinguished on the basis of their oxidation products. Meanwhile, the findings reported herein reveal a potential pitfall in the assignment of structures to lipids extracted from TLC plates because of artifactual oxidation after the plate development. PMID:24832382

  8. Surface oxidation under ambient air--not only a fast and economical method to identify double bond positions in unsaturated lipids but also a reminder of proper lipid processing.

    PubMed

    Zhou, Ying; Park, Hyejung; Kim, Philseok; Jiang, Yan; Costello, Catherine E

    2014-06-17

    A simple, fast approach elucidated carbon-carbon double bond positions in unsaturated lipids. Lipids were deposited onto various surfaces and the products from their oxidation in ambient air were observed by electrospray ionization (ESI) mass spectrometry (MS). The most common oxidative products, aldehydes, were detected as transformations at the cleaved double bond positions. Ozonides and carboxylic acids were generated in certain lipids. Investigations of the conditions controlling the appearance of these products indicated that the surface oxidation depends on light and ambient air. Since the lipid oxidation was slower in a high concentration of ozone, singlet oxygen appeared to be a parallel oxidant for unsaturated lipids. The 3-hydroxyl group in the sphingoid base of sulfatides offered some protection from oxidation for the Δ4,5-double bond, slowing its oxidation rate relative to that of the isolated double bond in the N-linked fatty acyl chain. Direct sampling by thin-layer chromatography (TLC)-ESI-MS provides a powerful approach to elucidate detailed structural information on biological samples. Co-localization of the starting lipids and their oxidation products after TLC separation allowed assignment of the native unsaturation sites. Phosphatidylserine and N,N-dimethyl phosphatidylethanolamine isomers in a bovine brain total lipid extract were distinguished on the basis of their oxidation products. Meanwhile, the findings reported herein reveal a potential pitfall in the assignment of structures to lipids extracted from TLC plates because of artifactual oxidation after the plate development.

  9. A carbohydrate fraction, AIP1, from Artemisia iwayomogi down-regulates Fas gene expression and suppresses apoptotic death of the thymocytes induced by 2,3,7,8-tectrachlorodibenzo-p-dioxin.

    PubMed

    Ji, Hee Jung; Yeo, Hee Kyoung; Lee, Nam Hee; Hwang, Jung Suk; Koo, Kyung Ah; Cheong, Seon Woo; Park, Joo Hung; Oh, Gap Soo; Yoon, Chun Sik; Youn, Hyun Joo

    2005-02-01

    Apoptotic death of mouse thymocytes in vitro, as induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), involves the up-regulation of Fas gene expression, while a carbohydrate fraction, AIP1, from Artemisia iwayomogi suppresses the death of thymocytes in culture along with the down-regulation of Fas gene expression. We have now investigated whether the AIP1 fraction modulates TCDD-induced thymocyte death. When treated with TCDD and AIP1 fraction together, the thymocytes do not show apoptosis induced by the TCDD treatment. The AIP1 supplementation to the TCDD treatment also down-regulates the TCDD-induced Fas gene up-regulation. These findings indicate that the AIP1 fraction suppresses TCDD-induced thymocyte apoptosis through the modulation of Fas gene expression.

  10. Thymic Nurse Cells Participate in Heterotypic Internalization and Repertoire Selection of Immature Thymocytes; Their Removal from the Thymus of Autoimmune Animals May be Important to Disease Etiology

    PubMed Central

    Guyden, J.C.; Martinez, M.; Chilukuri, R.V.E.; Reid, V.; Kelly, F.; Samms, M.-O.D.

    2016-01-01

    Thymic nurse cells (TNCs) are specialized epithelial cells that reside in the thymic cortex. The initial report of their discovery in 1980 showed TNCs to contain up to 200 thymocytes within specialized vacuoles in their cytoplasm. Much has been reported since that time to determine the function of this heterotypic internalization event that exists between TNCs and developing thymocytes. In this review, we discuss the literature reported that describes the internalization event and the role TNCs play during T cell development in the thymus as well as why these multicellular complexes may be important in inhibiting the development of autoimmune diseases.

  11. Regulation of thymocyte development through CD3. I. Timepoint of ligation of CD3 epsilon determines clonal deletion or induction of developmental program

    PubMed Central

    1993-01-01

    Several recent observations suggest that successful rearrangement of the T cell receptor (TCR) beta locus induces several important events in thymocyte maturation. Allelic exclusion is achieved by interruption of further rearrangement of the beta locus, and CD4-8- interleukin (IL)- 2R+ cells enter the CD4+8+IL-2R- stage. The actual molecular events regulating this important control point are unknown, but may be related to the expression of the TCR-beta locus in immature CD4-8- thymocytes. It is not clear whether maturation is induced by intracellular appearance of TCR-beta chain or by signal transduction through an immature TCR complex on the thymocyte membrane, possibly involving TCR- beta chain homodimers and CD3. Here we show that early addition of anti- CD3 mAb to fetal thymic organ cultures induces all known events associated with the acquisition of the CD4+8+ stage. Expression of CD4 and CD8 is accelerated, IL-2R alpha is downregulated, and the cells fail to produce TCR-beta, possibly based on premature cessation of beta gene rearrangement. Upon stimulation with anti-CD3 antibodies, we see calcium mobilization in 15% of all CD4-8- thymocytes with no detectable surface TCR expression. These results suggest that functional CD3 is expressed on immature thymocytes at very low concentrations before the appearance of a complete TCR-beta chain. Ligation of CD3 at this stage may mimic the maturation signal normally generated by the immature TCR- beta homodimer-CD3 complex. The results are consistent with the notion that acquisition of the CD4+8+ stage involves signal transduction through an immature TCR complex. Later in thymocyte development, ligation of CD3 results in deletion of CD4+8+ cells. Thus, signal transduction through CD3 may result in entirely different cellular responses, depending on the stage of thymocyte differentiation. These results suggest an involvement of CD3 as a link in signal transduction for at least two different decision points in the

  12. Yttrium decreases the intracellular Zn2+ concentration in rat thymocytes by attenuating a temperature-sensitive Zn2+ influx.

    PubMed

    Takahashi, Yusuke; Kanemaru, Kaori; Imai, Shoji; Miyoshi, Norikazu; Kawanai, Takuya; Oyama, Yasuo

    2012-09-01

    Yttrium is used in the production of various electronic devices because the alloy it contains enhances or modifies the properties of other elements. In order to study the cytotoxic action of yttrium, the effect of yttrium chloride (YCl(3)) on the intracellular Zn(2+) level was examined in rat thymocytes using a flow cytometer with FluoZin-3-AM and propidium iodide. The application of YCl(3) significantly decreased the intensity of the FluoZin-3 fluorescence, suggesting a decrease in the intracellular Zn(2+) level or quenching of the FluoZin-3 fluorescence by Y(3+). However, since Y(3+) did not attenuate the FluoZin-3 fluorescence under cell-free conditions, the latter suggestion was ruled out. Rat thymocytes possess a temperature-sensitive membrane pathway that carries Zn(2+) into the cells. The application of YCl(3) attenuated the FluoZin-3 fluorescence augmented by externally applied ZnCl(2) in a concentration-dependent manner. This suggested that Y(3+) inhibited the Zn(2+) influx, resulting in the decrease in the intracellular Zn(2+) level. Yttrium may induce dyshomeostasis of intracellular Zn(2+), leading to some cytotoxic actions.

  13. Cremophor EL, a non-ionic surfactant, promotes Ca(2+)-dependent process of cell death in rat thymocytes.

    PubMed

    Yamaguchi, Jun-Ya; Nishimura, Yumiko; Kanada, Aimi; Kobayashi, Masako; Mishima, Kyoko; Tatsuishi, Tomoko; Iwase, Kyoko; Oyama, Yasuo

    2005-08-01

    Cremophor EL, a surfactant for pharmaceutical products, augments the cytotoxicity of hydrogen peroxide in rat thymocytes [Iwase, K., Oyama, Y., Tatsuishi, T., Yamaguchi1, J., Nishimura1, Y., Kanada, A., Kobayashi, M., Maemura, Y., Ishida, S., Okano, Y., 2004. Cremophor EL augments the cytotoxicity of hydrogen peroxide in lymphocytes dissociated from rat thymus glands. Toxicol. Lett. 154, 143-148]. The effect of cremophor EL on Ca(2+)-dependent process of cell death has been examined using a flow cytometer since hydrogen peroxide increases intracellular Ca2+ concentration. Cremophor EL at clinically-relevant concentrations greatly increased the population of dead cells in rat thymocytes simultaneously treated with A23187, a calcium ionophore increasing intracellular Ca2+ concentration. Removal of Ca2+ from external solution diminished the cremophor EL-induced increase in the dead cell population. Result suggests that Ca(2+)-dependent process is involved in the cremophor EL-induced decrease in the cell viability in the simultaneous presence of A23187. The population of cells with hypodiploidal DNA was not increased by the application of cremophor EL and A23187 although the cell viability was greatly decreased, indicating that the type of cell death is necrosis. It is suggested that cremophor EL at clinically-relevant concentrations augments the Ca(2+)-dependent process of necrosis.

  14. Estimation of increased concentration of intracellular Cd(2+) by fluo-3 in rat thymocytes exposed to CdCl(2).

    PubMed

    Oyama, Yasuo; Arata, Tomohiro; Chikahisa, Lumi; Soeda, Fumio; Takahama, Kazuo

    2002-03-01

    Cadmium, an environmental pollutant, has been reported to induce apoptosis in murine lymphocytes. To reveal the mechanism of cadmium-induced apoptosis, one of important questions is whether cadmium increases intracellular concentration of Ca(2+) ([Ca(2+)](i)), Cd(2+) ([Cd(2+)](i)) or both. It is difficult to detect the increase in [Ca(2+)](i) using Ca(2+)-chelator-based fluorescent Ca(2+) indicators in the presence of Cd(2+) because of their sensitivity to Cd(2+). Therefore, the study on membrane response such as Ca(2+)-dependent hyperpolarization gives a clue to reveal whether the [Ca(2+)](i) or [Cd(2+)](i) is increased. Cadmium at concentrations of 3 μM or more dose-dependently augmented fluo-3 fluorescence in rat thymocytes, presumably suggesting an increased [Ca(2+)](i). However, the membranes were not hyperpolarized although the cells possess Ca(2+)-dependent K(+) channels. One may argue that cadmium inhibits Ca(2+)-dependent K(+) channels so that cadmium fails to hyperpolarize the membranes. It is unlikely because the [Ca(2+)](i) increased by A23187, a calcium ionophore, elicited the hyperpolarization in the presence of Cd(2+). Furthermore, the profile of cytotoxicity induced by cadmium, examined by ethidium bromide and annexin V-FITC, was different from that induced by A23187. Taken together, it is concluded that the application of cadmium increases the [Cd(2+)](i) rather than the [Ca(2+)](i) in rat thymocytes, resulting in the induction of cytotoxicity.

  15. Ceramide synthase 2 facilitates S1P-dependent egress of thymocytes into the circulation in mice.

    PubMed

    Rieck, Michael; Kremser, Christiane; Jobin, Katarzyna; Mettke, Elisabeth; Kurts, Christian; Gräler, Markus; Willecke, Klaus; Kolanus, Waldemar

    2017-02-15

    Well-defined gradients of the lipid mediator sphingosine-1-phosphate (S1P) direct chemotactic egress of mature thymocytes from the thymus into the circulation. Although it is known that these gradients result from low S1P levels in the thymic parenchyma and high S1P concentrations at the exit sites and in the plasma, the biochemical mechanisms that regulate these differential S1P levels remain unclear. Several studies demonstrated that ceramide synthase 2 (Cers2) regulates the levels of the S1P precursor sphingosine. We, therefore, investigated whether Cers2 is involved in the regulation of S1P gradients and S1P-dependent egress into the circulation. By analyzing Cers2-deficient mice, we demonstrate that Cers2 limits the levels of S1P in thymus and blood to maintain functional S1P gradients that mediate thymocyte emigration into the circulation. This function is specific for Cers2, as we also show that Cers4 is not involved in the regulation of thymic egress. Our study identified Cers2 as an important regulator of S1P-dependent thymic egress, and thus contributes to the understanding of how S1P gradients are maintained in vivo.

  16. The organotin-induced thymus atrophy, characterized by depletion of CD4+ CD8+ thymocytes, is preceded by a reduction of the immature CD4- CD8+ TcR alpha beta-/low CD2high thymoblast subset.

    PubMed Central

    Pieters, R H; Bol, M; Lam, B W; Seinen, W; Penninks, A H

    1992-01-01

    Thymic changes in the rat induced by the thymus atrophy-inducing organotin compound di-n-butyltin dichloride (DBTC) were examined using FACS analyses. The number of CD4+CD8+ thymocytes was reduced by DBTC treatment from Day 2 onwards and reached minimum level on Days 4 and 5 after dosing. On these days the CD4-CD8- and both the CD4-CD8+ and CD4+CD8- subsets were not affected. On Day 2 we observed a reduced proportion of transferrin receptor (CD71)-positive CD4-OX44- cells, representing the cycling immature CD4-CD8+ cells, and of CD71+OX44- cells, representing the cycling CD4+CD8+ cells, but not of CD71+CD4-CD8- cells. When compared to controls, the FSChigh cell population of DBTC-treated rats contained less CD4-OX44- and OX44- cells, which were further characterized as CD2high and T-cell receptor (TcR)alpha beta- low. Moreover, fewer TcR alpha beta high cells were detected in the OX44- thymoblast subset of DBTC-treated rats. The number of CD4-CD8- thymoblasts appeared marginally decreased while the numbers of CD4+OX44+ cells, representing mature CD4+ cells, were not affected. These data indicate that DBTC causes a preferential initial depletion of immature CD4-CD8+CD2high TcR alpha beta-low thymoblasts. This initial event may result in a decreased formation of CD4+CD8+ thymoblasts and of small CD4+CD8+ thymocytes. These characteristics of the initially depleted subset indicate a specific anti-proliferative effect of DBTC and may give clues for the mechanism involved in the induction of thymus atrophy. PMID:1353062

  17. A role for T3+4-6-8- transitional thymocytes in the differentiation of mature and functional T cells from human prothymocytes.

    PubMed Central

    Toribio, M L; Martinez, C; Marcos, M A; Marquez, C; Cabrero, E; de la Hera, A

    1986-01-01

    In vivo, immunocompetent T lymphocytes are only detected late in ontogeny, among mature thymocytes expressing either T4 (L3T4 in mouse) or T8 (Lyt-2) surface glycoproteins. We have previously shown, however, that there are functional precursors among T3+4-6-8- human thymocytes in vivo. Here we report on the in vitro differentiation of prothymocytes into T3+4-6-8- and mature T cells. T11+3-4-6-8- prothymocytes (0.5% of total thymocytes, greater than 98% pure) were obtained after treatment of thymocytes with OKT3 (T3), OKT4A (T4), Na1/34 (T6), and B9.4 (T8) monoclonal antibodies plus complement. During culture, the prothymocyte precursors acquire first T3 and then either T4 or T8, but not T6. The largest subpopulation in the thymus, T4+6+8+ cells, are not detected among the in vitro T-cell precursors. During culture, the precursors acquire cytolytic activity as soon as they express either the T3+4-6-8- or the mature (T3+4+8- or T3+4-8+) phenotypes. We suggest that T3+4-6-8- cells are a productive, transitional stage in T-lymphocyte development. PMID:3092222

  18. A population of early fetal thymocytes expressing Fc gamma RII/III contains precursors of T lymphocytes and natural killer cells.

    PubMed

    Rodewald, H R; Moingeon, P; Lucich, J L; Dosiou, C; Lopez, P; Reinherz, E L

    1992-04-03

    We have identified a dominant fetal thymocyte population at day 14.5 of gestation in the mouse that lacks CD4 and CD8 but expresses Fc gamma RII/III several days prior to acquisition of the T cell receptor (TCR) in vivo. If maintained in a thymic microenvironment, this population of CD4-CD8-TCR-Fc gamma RII/III+ thymocytes differentiates first into CD4+CD8+TCRlowFc gamma RII/III- thymocytes and subsequently CD4+CD8-TCRhighFc gamma RII/III- and CD4-CD8+TCRhighFc gamma RII/III- mature Ti alpha-beta lineage T cells. However, if removed from the thymus, the CD4-CD8-TCR-Fc gamma RII/III+ thymocyte population selectively generates functional natural killer (NK) cells in vivo as well as in vitro. These findings show that a cellular pool of Fc gamma RII/III+ precursors gives rise to T and NK lineages in a microenvironment-dependent manner. Moreover, they suggest a hitherto unrecognized role for Fc receptors on primitive T cells.

  19. QUANTITATION OF ABERRANT INTERLOCUS T-CELL RECEPTOR REARRANGEMENTS IN MOUSE THYMOCYTES AND THE EFFECT OF THE HERBICIDE 2,4- DICHLOROPHENOXYACETIC ACID

    EPA Science Inventory

    Quantitation of aberrant interlocus T-cell receptor rearrangements in mouse thymocytes and the effect of the herbicide 2,4- Dichlorophenoxyacetic acid

    Small studies in human populations have suggested a correlation between the frequency of errors in antigen receptor gene a...

  20. QUANTITATION OF ABERRANT INTERLOCUS T-CELL RECEPTOR REARRANGEMENTS IN MOUSE THYMOCYTES AND THE EFFECT OF THE HERBICIDE 2,4- DICHLOROPHENOXYACETIC ACID

    EPA Science Inventory

    Quantitation of aberrant interlocus T-cell receptor rearrangements in mouse thymocytes and the effect of the herbicide 2,4- Dichlorophenoxyacetic acid

    Small studies in human populations have suggested a correlation between the frequency of errors in antigen receptor gene a...

  1. Detection of a novel population of fetal thymocytes characterized by preferential emigration and a TCRgammadelta+ T cell fate after dioxin exposure.

    PubMed

    Majora, Marc; Frericks, Markus; Temchura, Vladimir; Reichmann, Gaby; Esser, Charlotte

    2005-11-01

    T cell maturation into TCRalphabeta(+) or TCRgammadelta(+) cells from common immature CD4(-)CD8(-)(DN) precursors occurs in the thymus, and is controlled through ordered regulation of genes. The aryl hydrocarbon receptor (AHR), a latent cytoplasmic transcription factor, affects thymocyte maturation and differentiation at several stages, also including DN cells. We analyzed in murine fetal thymus organ cultures (FTOC) the outcome of AHR-signaling and found a higher frequency of DN TCRgammadelta(+) cells in the presence of the AHR-activating ligand TCDD. We detected a novel population of CD25(int/lo)CD44(hi) cells associated with preferential emigration and a TCRgammadelta(+) T cell fate of thymocytes. Sorted DN TCRgammadelta(+) emigrants could proliferate if IL-2 was available. Moreover, they suppressed the proliferation of co-cultivated, activated CD4(+) T cells. Gene expression profiles of purified DN emigrants from TCDD*FTOC revealed 295 modulated genes, 10% of which are genes of the immune system. For instance, RAG-1, TdT, and Gfi-1 were downregulated, yet genes indicative of mature thymocytes were upregulated. In conclusion, we have detected changes in the differentiation programme of fetal DN thymocytes after ligand-activation of the AHR. In particular, we observed a higher frequency of DN TCRgammadelta(+) cells with high emigration potential, and possible regulatory functions.

  2. Mtf-1 lymphoma-susceptibility locus affects retention of large thymocytes with high ROS levels in mice after {gamma}-irradiation

    SciTech Connect

    Maruyama, Masaki; Yamamoto, Takashi; Kohara, Yuki; Katsuragi, Yoshinori; Mishima, Yukio; Aoyagi, Yutaka; Kominami, Ryo; E-mail: rykomina@med.niigata-u.ac.jp

    2007-03-02

    Mouse strains exhibit different susceptibilities to {gamma}-ray-induced thymic lymphomas. Our previous study identified Mtf-1 (metal responsive transcription factor-1) as a candidate susceptibility gene, which is involved in the radiation-induced signaling pathway that regulates the cellular reactive oxygen species (ROS). To reveal the mechanism for the increased susceptibility conferred by Mtf-1 locus, we examined early effects of {gamma}-ray on ROS levels in vivo and its difference between Mtf-1 susceptible and resistant congenic mice. Here, we show the detection of clonally growing thymocytes at 4 weeks after irradiation, indicating the start of clonal expansion at a very early stage. We also show that large thymocytes with higher ROS levels and a proliferation capacity were more numerous in the Mtf-1 susceptible mice than the resistant mice when examined at 7 days after irradiation, although such tendency was not found in mice lacking one allele of Bcl11b tumor suppressor gene. This high retention of the large thymocytes, at a high risk for ROS-induced mutation, is a compensatory proliferation and regeneration response to depletion of the thymocytes after irradiation and the response is likely to augment the development of prelymphoma cells leading to thymic lymphomas.

  3. TNF-α Is Involved in the Abnormal Thymocyte Migration during Experimental Trypanosoma cruzi Infection and Favors the Export of Immature Cells

    PubMed Central

    Pérez, Ana Rosa; Berbert, Luiz Ricardo; Lepletier, Ailin; Revelli, Silvia; Bottasso, Oscar; Silva-Barbosa, Suse Dayse; Savino, Wilson

    2012-01-01

    Previous studies revealed a significant production of inflammatory cytokines together with severe thymic atrophy and thymocyte migratory disturbances during experimental Chagas disease. Migratory activity of thymocytes and mature T cells seem to be finely tuned by cytokines, chemokines and extracellular matrix (ECM) components. Systemic TNF-α is enhanced during infection and appears to be crucial in the response against the parasite. However, it also seems to be involved in disease pathology, since it is implicated in the arrival of T cells to effector sites, including the myocardium. Herein, we analyzed the role of TNF-α in the migratory activity of thymocytes in Trypanosoma cruzi (T. cruzi) acutely-infected mice. We found increased expression and deposition of TNF-α in the thymus of infected animals compared to controls, accompanied by increased co-localization of fibronectin, a cell migration-related ECM molecule, whose contents in the thymus of infected mice is also augmented. In-vivo studies showed an enhanced export of thymocytes in T. cruzi-infected mice, as ascertained by intrathymic injection of FITC alone or in combination with TNF-α. The increase of immature CD4+CD8+ T cells in secondary lymphoid organs was even more clear-cut when TNF-α was co-injected with FITC. Ex-vivo transmigration assays also revealed higher number of migrating cells when TNF-α was added onto fibronectin lattices, with higher input of all thymocyte subsets, including immature CD4+CD8+. Infected animals also exhibit enhanced levels of expression of both mRNA TNF-α receptors in the CD4+CD8+ subpopulation. Our findings suggest that in T. cruzi acute infection, when TNF-α is complexed with fibronectin, it favours the altered migration of thymocytes, promoting the release of mature and immature T cells to different compartments of the immune system. Conceptually, this work reinforces the notion that thymocyte migration is a multivectorial biological event in health and disease

  4. Isolation of human CD4/CD8 double-positive, graft-versus-host disease-protective, minor histocompatibility antigen-specific regulatory T cells and of a novel HLA-DR7-restricted HY-specific CD4 clone.

    PubMed

    Eljaafari, Assia; Yuruker, Ozel; Ferrand, Christophe; Farre, Annie; Addey, Caroline; Tartelin, Marie-Laure; Thomas, Xavier; Tiberghien, Pierre; Simpson, Elizabeth; Rigal, Dominique; Scott, Diane

    2013-01-01

    Minor histocompatibility (H) Ags are classically described as self-peptides derived from intracellular proteins that are expressed at the cell surface by MHC class I and class II molecules and that induce T cell alloresponses. We have isolated three different T cell populations from a skin biopsy of a patient suffering from acute graft-versus-host disease following sex-mismatched HLA-identical bone marrow transplantation. The first population was: 1) CD4(+)/CD8(+) double-positive; 2) specific for an HLA class I-restricted autosomal Ag; 3) expressed a Tr1 profile with high levels of IL-10, but low IL-2 and IFN-γ; and 4) exerted regulatory function in the presence of recipient APCs. The second was CD8 positive, specific for an HLA class I-restricted autosomally encoded minor H Ag, but was only weakly cytotoxic. The third was CD4 single positive, specific for an HLA-DR7-restricted HY epitope and exerted both proliferative and cytotoxic functions. Identification of the peptide recognized by these latter cells revealed a new human HY epitope, TGKIINFIKFDTGNL, encoded by RPS4Y and restricted by HLA-DR7. In this paper, we show human CD4/CD8 double-positive, acute graft-versus-host disease-protective, minor H Ag-specific regulatory T cells and identify a novel HLA-DR7/ HY T cell epitope, encoded by RPS4Y, a potential new therapeutic target.

  5. Towards positional cloning in Brassica napus: generation and analysis of doubled haploid B. rapa possessing the B. napus pol CMS and Rfp nuclear restorer gene.

    PubMed

    Formanová, Natasa; Li, Xiu-Qing; Ferrie, Alison M R; Depauw, Mary; Keller, Wilf A; Landry, Benoit; Brown, Gregory G

    2006-05-01

    The Polima (pol) system of cytoplasmic male sterility (CMS) and its fertility restorer gene Rfp are used in hybrid rapeseed production in Brassica napus. To facilitate map-based cloning of the Rfp gene, we have successfully transferred the pol cytoplasm and Rfp from the amphidiploid B. napus to the diploid species B. rapa and generated a doubled haploid pol cytoplasm B. rapa population that segregates for the Rfp gene. This was achieved through interspecific crosses, in vitro rescue of hybrid embryos, backcrosses, and microspore culture. Male fertility conditioned by Rfp was shown to co-segregate in this population with Rfp-specific mitochondrial transcript modifications and with DNA markers previously shown to be linked to Rfp in B. napus. The selfed-progeny of one doubled haploid plant were confirmed to be characteristic B. rapa diploids by cytogenetic analysis. Clones recovered from a genomic library derived from this plant line using the RFLP probe cRF1 fell into several distinct physical contigs, one of which contained Rfp-linked polymorphic restriction fragments detected by this probe. This indicates that chromosomal DNA segments anchored in the Rfp region can be recovered from this library and that the library may therefore prove to be a useful resource for the eventual isolation of the Rfp gene.

  6. Ouabain-induced alterations in ABCB1 of mesenteric lymph nodes and thymocytes of rats and mice

    PubMed Central

    Lima, Daniel Boff; Valente, Raphael Carmo; Capella, Marcia Alves Marques

    2016-01-01

    Ouabain is a glycoside with immunomodulating properties, and recent studies have suggested its use in adjuvant therapy for cancer treatment. Ouabain is known to modulate the immune system in vitro, and previous studies have revealed that ouabain can modulate the expression and activity of ABCB1, a protein associated with multidrug resistance present in immune system. Therefore, the present study investigated alterations in the expression and activity of ABCB1 in the thymi, peripheral blood monocytes and lymph nodes of Wistar rats and Swiss mice treated acutely or chronically with ouabain. A decrease of almost 45% in the monocyte count and an increase of 55% in the basophil count were observed. A significant decrease (75% reduction) in the amount of cells with ABCB1 activity was found in the thymocytes of ouabain-treated rats and mice. The possible implications of these results for cancer treatment are discussed. PMID:28105236

  7. Carnosine prevents necrotic and apoptotic death of rat thymocytes via ouabain-sensitive Na/K-ATPase.

    PubMed

    Smolyaninova, Larisa V; Dergalev, Alexander A; Kulebyakin, Konstantin Y; Carpenter, David O; Boldyrev, Alexander A

    2013-01-01

    It is known that ouabain, a selective inhibitor of Na/K-ATPase, not only can cause the activation of signal cascades, which regulate the cell viability, but also can cause the accumulation of free radicals, which can evoke the oxidative stress. We have shown that the nanomolar concentrations of ouabain result in the temporary increase in the level of intracellular free radicals, but the millimolar concentration of ouabain induces a stable intracellular accumulation of free radicals in rat thymocytes. The increasing level of free radicals resulting from both low and high concentrations of ouabain can be attenuated by the antioxidant, carnosine. Moreover, the long-term incubation with ouabain leads to the cell death by necrosis and apoptosis. Ouabain-mediated apoptosis and necrosis were also abolished by carnosine.

  8. Ouabain-induced alterations in ABCB1 of mesenteric lymph nodes and thymocytes of rats and mice.

    PubMed

    Lima, Daniel Boff; Valente, Raphael Carmo; Capella, Marcia Alves Marques

    2016-12-01

    Ouabain is a glycoside with immunomodulating properties, and recent studies have suggested its use in adjuvant therapy for cancer treatment. Ouabain is known to modulate the immune system in vitro, and previous studies have revealed that ouabain can modulate the expression and activity of ABCB1, a protein associated with multidrug resistance present in immune system. Therefore, the present study investigated alterations in the expression and activity of ABCB1 in the thymi, peripheral blood monocytes and lymph nodes of Wistar rats and Swiss mice treated acutely or chronically with ouabain. A decrease of almost 45% in the monocyte count and an increase of 55% in the basophil count were observed. A significant decrease (75% reduction) in the amount of cells with ABCB1 activity was found in the thymocytes of ouabain-treated rats and mice. The possible implications of these results for cancer treatment are discussed.

  9. Asparagine-linked sugar chains of glycoproteins in calf thymocyte plasma membrane. Isolation and fractionation of oligosaccharides liberated by hydrazinolysis.

    PubMed

    Yoshima, H; Takasaki, S; Kobata, A

    1980-07-01

    The plasma membrane glycoproteins of calf thymocytes were converted to glycopeptides by exhaustive pronase digestion. Glycopeptides with asparagine-linked sugar chains were separated from those with mucine-type sugar chains by Bio-Gel P-10 column chromatography. The asparagine-linked sugar chains were released as oligosaccharides from the peptide moiety by hydrazinolysis and labeled by reduction with NaB[3H]4. The radioactive oligosaccharides were fractionated into fifteen acidic components and ten neutral components by combination of paper electrophoresis and Bio-Gel P-4 column chromatography. The acidic nature of all fifteen acidic components can be ascribed to their N-acetylneuraminic acid residues. The Bio-Gel P-4 column chromatographic patterns of the neutral oligosaccharide fraction and of the neutral fraction obtained on sialidase treatment of the pooled acidic oligosaccharide fraction were totally different, indicating that the acidic oligosaccharides are not simple sialyl derivatives of the neutral oligosaccharides.

  10. Determining the DUF55-domain structure of human thymocyte nuclear protein 1 from crystals partially twinned by tetartohedry

    SciTech Connect

    Yu, Feng; Song, Aixin; Xu, Chunyan; Sun, Lihua; Li, Jian; Tang, Lin; Yu, Minmin; Yeates, Todd O.; Hu, Hongyu; He, Jianhua

    2009-06-06

    Human thymocyte nuclear protein 1 (hTHYN1) contains a unique DUF55 domain of 167 residues (55-221), but its cellular function is unclear. Crystals of DUF55 belong to the trigonal space group P3{sub 1}, but twinning causes the data to approach an apparent 622 symmetry. Two datasets to 2.3 {angstrom} resolution were collected. Statistical analysis confirmed that both datasets were partially twinned by tetartohedry. Tetartohedral twin fractions were estimated. After the structure was determined, only one twofold axis of rotational pseudosymmetry was found in the crystal structure. Using the DALI program, a YTH domain, which is a potential RNA binding domain from human YTH domain-containing protein 2, was identified to have the most similar three-dimensional fold to DUF55. It is implied that DUF55 might be a potential RNA-related domain.

  11. The receptor tyrosine kinase MerTK activates phospholipase C γ2 during recognition of apoptotic thymocytes by murine macrophages

    PubMed Central

    Todt, Jill C.; Hu, Bin; Curtis, Jeffrey L.

    2008-01-01

    Apoptotic leukocytes must be cleared efficiently by macrophages (Mø). Apoptotic cell phagocytosis by Mø requires the receptor tyrosine kinase (RTK) MerTK (also known as c-Mer and Tyro12), the phosphatidylserine receptor (PS-R), and the classical protein kinase C (PKC) isoform βII, which translocates to Mø membrane and cytoskeletal fractions in a PS-R-dependent fashion. How these molecules cooperate to induce phagocytosis is unknown. Because the phosphatidylinositol-specific phospholipase (PI-PLC) PLC γ2 is downstream of RTKs in some cell types and can activate classical PKCs, we hypothesized that MerTK signals via PLC γ2. To test this hypothesis, we examined the interaction of MerTK and PLC γ2 in resident murine PMø and in the murine Mø cell line J774A.1 (J774) following exposure to apoptotic thymocytes. We found that, as with PMø, J774 phagocytosis of apoptotic thymocytes was inhibited by antibody against MerTK. Western blotting and immunoprecipitation showed that exposure to apoptotic cells produced three time-dependent changes in PMø and J774: (1) tyrosine phosphorylation of MerTK; (2) association of PLC γ2 with MerTK; and (3) tyrosine phosphorylation of PLC γ2. Phosphorylation of PLC γ2 and its association with MerTK was also induced by cross-linking MerTK using antibody. A PI-PLC appears to be required for phagocytosis of apoptotic cells because the PI-PLC inhibitor Et-18-OCH3 and the PLC inhibitor U73122, but not the inactive control U73343, blocked phagocytosis without impairing adhesion. On apoptotic cell adhesion to Mø, MerTK signals at least in part via PLC γ2. PMID:14704368

  12. Differential expression of cyclin-dependent kinase 6 in cortical thymocytes and T-cell lymphoblastic lymphoma/leukemia.

    PubMed Central

    Chilosi, M.; Doglioni, C.; Yan, Z.; Lestani, M.; Menestrina, F.; Sorio, C.; Benedetti, A.; Vinante, F.; Pizzolo, G.; Inghirami, G.

    1998-01-01

    Cyclin-dependent kinase-6 (CDK6) is the earliest inducible member of the CDK family in human T lymphocytes, involved in growth factor stimulation and cell cycle progression. CDK6 is one of the targets of p16 and p15, CDK inhibitors encoded by MTS1 and MTS2, two tumor suppressor genes that are frequently deleted in T-cell leukemia. In this study we have investigated CDK6 expression in normal and neoplastic lymphoid tissues using immunohistochemistry and flow cytometry. In normal (six samples) and hyperplastic (four samples) thymuses, strong CDK6 expression was observed in a discrete proportion of cortical thymocytes (10 to 15%), mainly located in the peripheral (subcapsular) zone of the cortex. All tested cases of T-cell lymphoblastic lymphoma/leukemia (T-LBL/ALL) showed strong CDK6 expression in the majority (up to 100%) of neoplastic lymphoid cells. Western blot analysis confirmed the expected CDK6 protein size (40 kd). According to Southern blot analysis, CDK6 overexpression in neoplastic T lymphoblasts was not due to gene amplification. In all other lymphomas investigated (28 peripheral T-cell non-Hodgkin's lympohomas (T-NHLs), 7 CD30+ anaplastic NHLs, 22 high-grade B-NHLs, 15 low-grade B-NHLs, 25 B-cell precursor ALLs), CDK6 was not expressed or expressed at low levels, with the only exception of three nasal angiocentric T-NHLs, all exhibiting CDK6 immunoreactivity comparable to that observed in T-LBL/ALL. These data provide evidence that CDK6 is abnormally expressed in T-LBL/ALL and may be involved in the pathogenesis of this malignancy. In addition, the quantitative difference of CDK6 expression between neoplastic and non-neoplastic cortical thymocytes can be potentially useful in the differential diagnosis of thymic neoplasms on histological and cytological specimens. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 7 Figure 8 PMID:9422538

  13. Clinical association of mixed connective tissue disease and granulomatosis with polyangiitis: a case report and systematic screening of anti-U1RNP and anti-PR3 auto-antibody double positivity in ten European hospitals.

    PubMed

    Tubery, Amandine; Fortenfant, Françoise; Combe, Bernard; Abreu, Isabelle; Bossuyt, Xavier; Chretien, Pascale; Desplat-Jégo, Sophie; Fabien, Nicole; Hue, Sophie; Johanet, Catherine; Lakomy, Daniela; Vincent, Thierry; Daïen, Claire I

    2016-12-01

    We report here the case of a 50-years-old man treated for mixed connective tissue disease (MCTD) positive for anti-U1 ribonucleoprotein (U1RNP) antibodies who secondarily developed a granulomatosis with polyangiitis (GPA) associated with anti-proteinase 3 anti-neutrophil cytoplasmic antibodies (PR3-ANCA). We then evaluated the frequency of the association between anti-U1RNP and anti-PR3-ANCA antibodies by a systematic retrospective study in ten European hospitals. Overall, out of 11,921 samples analyzed for both auto-antibodies, 18 cases of anti-U1RNP and anti-PR3-ANCA double positivity were found and only one patient presented with both MCTD and GPA symptoms. Our retrospective analysis indicates that anti-U1RNP and anti-PR3-ANCA antibodies double positivity is infrequent and very rarely associated with both MTCD and GPA. Our observation describes for the first time the coexistence of MTCD and severe GPA in a Caucasian patient. Association of anti-U1RNP and ANCA antibodies was rarely reported in the literature. Eleven cases of MCTD and ANCA vasculitis have been reported to date, with only two cases with anti-PR3-ANCA association, and only one vasculitis. The seven other cases reported in the literature presented with an association of MCTD and microscopic polyangiitis which appears to be a more frequent presentation than MTCD associated with GPA.

  14. Would doubling the human lifespan be a net positive or negative for us, either as individuals or as a society? Point-counterpoint.

    PubMed

    Stock, Gregory B; Callahan, Daniel

    2005-12-01

    There is a significant possibility that over the next few decades science will make discoveries of a kind that might allow the doubling of the average human life span, from roughly 76 years now to 150. This development would, for many, represent the realization of a dream: that of enabling people to live much longer lives than at present, holding back death, which has often been seen as an ancient, unbeatable enemy. It would also raise a large number of unprecedented individual and social problems: Would we really want to live to 150? Is such a goal ethical? What would this putative longevity do to our present social structures and arrangements? Would we get a better society or a worse one?

  15. Effects of Systemic Administration of Dexmedetomidine on Intraocular Pressure and Ocular Perfusion Pressure during Laparoscopic Surgery in a Steep Trendelenburg Position: Prospective, Randomized, Double-Blinded Study.

    PubMed

    Joo, Jin; Koh, Hyunjung; Lee, Kusang; Lee, Jaemin

    2016-06-01

    Increased intraocular pressure (IOP) during surgery is a risk factor for postoperative ophthalmological complications. We assessed the efficacy of systemically infused dexmedetomidine in preventing the increase in IOP caused by a steep Trendelenburg position, and evaluated the influence of underlying hypertension on IOP during surgery. Sixty patients undergoing laparoscopic surgery in a steep Trendelenburg position were included. Patients in the dexmedetomidine group received a 1.0 µg/kg IV loading dose of dexmedetomidine before anesthesia, followed by an infusion of 0.5 µg/kg/hr throughout the operation. Patients in the saline group were infused with the same volume of normal saline. IOP and ocular perfusion pressure (OPP) were measured 16 times pre- and intraoperatively. In the saline group, IOP increased in the steep Trendelenburg position, and was 11.3 mmHg higher at the end of the time at the position compared with the baseline value (before anesthetic induction). This increase in IOP was attenuated in the dexmedetomidine group, for which IOP was only 4.2 mmHg higher (P < 0.001 vs. the saline group). The steep Trendelenburg position was associated with a decrease in OPP; the degree of decrease was comparable for both groups. In intragroup comparisons between patients with underlying hypertension and normotensive patients, the values of IOP at every time point were comparable. Dexmedetomidine infusion attenuated the increase in IOP during laparoscopic surgery in a steep Trendelenburg position, without further decreasing the OPP. Systemic hypertension did not seem to be associated with any additional increase in IOP during surgery. (Registration at the Clinical Research Information Service of Korea National Institute of Health ID: KCT0001482).

  16. Effects of Systemic Administration of Dexmedetomidine on Intraocular Pressure and Ocular Perfusion Pressure during Laparoscopic Surgery in a Steep Trendelenburg Position: Prospective, Randomized, Double-Blinded Study

    PubMed Central

    2016-01-01

    Increased intraocular pressure (IOP) during surgery is a risk factor for postoperative ophthalmological complications. We assessed the efficacy of systemically infused dexmedetomidine in preventing the increase in IOP caused by a steep Trendelenburg position, and evaluated the influence of underlying hypertension on IOP during surgery. Sixty patients undergoing laparoscopic surgery in a steep Trendelenburg position were included. Patients in the dexmedetomidine group received a 1.0 µg/kg IV loading dose of dexmedetomidine before anesthesia, followed by an infusion of 0.5 µg/kg/hr throughout the operation. Patients in the saline group were infused with the same volume of normal saline. IOP and ocular perfusion pressure (OPP) were measured 16 times pre- and intraoperatively. In the saline group, IOP increased in the steep Trendelenburg position, and was 11.3 mmHg higher at the end of the time at the position compared with the baseline value (before anesthetic induction). This increase in IOP was attenuated in the dexmedetomidine group, for which IOP was only 4.2 mmHg higher (P < 0.001 vs. the saline group). The steep Trendelenburg position was associated with a decrease in OPP; the degree of decrease was comparable for both groups. In intragroup comparisons between patients with underlying hypertension and normotensive patients, the values of IOP at every time point were comparable. Dexmedetomidine infusion attenuated the increase in IOP during laparoscopic surgery in a steep Trendelenburg position, without further decreasing the OPP. Systemic hypertension did not seem to be associated with any additional increase in IOP during surgery. (Registration at the Clinical Research Information Service of Korea National Institute of Health ID: KCT0001482) PMID:27247511

  17. D-cycloserine increases positive symptoms in chronic schizophrenic patients when administered in addition to antipsychotics: a double-blind, parallel, placebo-controlled study.

    PubMed

    van Berckel, B N; Evenblij, C N; van Loon, B J; Maas, M F; van der Geld, M A; Wynne, H J; van Ree, J M; Kahn, R S

    1999-08-01

    A hypofunction of the glutamatergic system and NMDA receptors in schizophrenia has been hypothesized. Therefore, stimulation of these receptors could be of benefit to patients with schizophrenia. D-cycloserine has been used for this purpose. This study reports the effects of 100 mg D-cycloserine, when added to typical antipsychotics in chronic schizophrenic patients exhibiting prominent negative symptoms, using a placebo-controlled, double-blind, parallel, design. D-cycloserine slightly worsened psychotic symptoms and general psychopathology as compared to placebo. D-cycloserine failed to change negative symptoms and had no effect on extrapyramidal symptoms. The exacerbation of schizophrenic symptoms may be explained by the antagonistic effects of this dose of D-cycloserine at the glycine recognition site of the NMDA receptor due to competition with the endogenous agonist glycine. Another explanation for the increase in psychopathology may be an interaction with the effects of antipsychotics on NMDA mediated neurotransmission. Thus, D-cycloserine in this study did not ameliorate schizophrenic symptoms. However, the fact that they actually worsened suggests that NMDA systems may be involved in the pathogenesis of schizophrenia. Further placebo-controlled studies with lower dosages of D-cycloserine, preferably in drug-free patients, are necessary to evaluate if D-cycloserine is of use for the treatment of patients with schizophrenia.

  18. Short-term Curcuminoid Supplementation for Chronic Pulmonary Complications due to Sulfur Mustard Intoxication: Positive Results of a Randomized Double-blind Placebo-controlled Trial.

    PubMed

    Panahi, Y; Ghanei, M; Bashiri, S; Hajihashemi, A; Sahebkar, A

    2015-11-01

    Pulmonary problems are among the most frequent chronic complications of sulfur mustard (SM) intoxication and are often accompanied by deregulated production of pro-inflammatory cytokines. Curcuminoids, comprising curcumin, demethoxycurcumin and bisdemethoxycurcumin, are phytochemicals with remarkable anti-inflammatory properties that are derived from dried rhizomes of the plant Curcuma longa L. (turmeric). The present pilot study aimed to investigate the clinical effects of supplementation with curcuminoids on markers of pulmonary function and systemic inflammation in SM-intoxicated subjects. In a randomized double-blind placebo-controlled trial, 89 male subjects who were suffering from chronic SM-induced pulmonary complications were recruited and assigned to either curcuminoids (500 mg TID per oral; n=45) or placebo (n=44) for a period of 4 weeks. Efficacy measures were changes in the spirometric parameters (FVC, FEV1, FEV1/FVC) and serum levels of inflammatory mediators including interleukins 6 (IL-6) and 8 (IL-8), tumor necrosis factor-α (TNFα), transforming growth factor-β (TGFβ), high-sensitivity C-reactive protein (hs-CRP), calcitonin gene related peptide (CGRP), substance P and monocyte chemotactic protein-1 (MCP-1). 78 subjects completed the trial. Although FEV1 and FVC remained comparable between the groups, there was a greater effect of curcuminoids vs. placebo in improving FEV1/FVC (p=0.002). Curcuminoids were also significantly more efficacious compared to placebo in modulating all assessed inflammatory mediators: IL-6 (p<0.001), IL-8 (p=0.035), TNFα (p<0.001), TGFβ (p<0.001), substance P (p=0.016), hs-CRP (p<0.001), CGRP (p<0.001) and MCP-1 (p<0.001). Curcuminoids were safe and well-tolerated throughout the trial. Short-term adjunctive therapy with curcuminoids can suppress systemic inflammation in patients suffering from SM-induced chronic pulmonary complications. © Georg Thieme Verlag KG Stuttgart · New York.

  19. The NIFTY study: a multicentre randomised double-blind placebo-controlled trial of nifedipine maintenance tocolysis in fetal fibronectin-positive women in threatened preterm labour.

    PubMed

    Parry, Emma; Roos, Carolien; Stone, Peter; Hayward, Lynsey; Mol, Ben Willem; McCowan, Lesley

    2014-06-01

    In an unselected group of women with signs of preterm labour, maintenance tocolysis is not effective in the prevention of preterm birth and does not improve neonatal outcome. Among women with signs of preterm labour, those who are fetal fibronectin positive have an increased risk of preterm birth. We investigated whether maintenance tocolysis with nifedipine would delay delivery and improve neonatal outcome in women with threatened preterm labour and a positive fetal fibronectin status. Women with a singleton pregnancy in threatened preterm labour (24(+0) to 33(+6)  weeks) with a positive fetal fibronectin test were randomised to nifedipine or placebo. Study medication was continued until 36 completed weeks' gestation. The primary endpoint was prolongation of pregnancy of seven days. Secondary endpoints were gestational age at delivery and length of NICU admission. Of the 60 participants, 29 received nifedipine and 31 placebo. Prolongation of pregnancy by >7 days occurred in 22/29 (76%) in the nifedipine group and 25/31 (81%) in the placebo group (relative risks, RR 0.94 [0.72-1.2]). Gestational age at delivery was 36.1 ± 5.1 weeks for nifedipine and 36.8 ± 3.6 weeks for placebo (P = 0.027). Length of NICU admission [median (interquartile ranges, IQR)] was 27 (24-41) days and 16 (8-37) days in nifedipine and placebo groups, respectively (P = 0.17). In women with threatened preterm labour who are fetal fibronectin positive, maintenance tocolysis with nifedipine does not seem to prolong pregnancy, nor reduce length of NICU admission. © 2014 The Royal Australian and New Zealand College of Obstetricians and Gynaecologists.

  20. In vitro toxicity and interactions of environmental contaminants (Arochlor 1254 and mercury) and immunomodulatory agents (lipopolysaccharide and cortisol) on thymocytes from lake trout (Salvelinus namaycush)

    USGS Publications Warehouse

    Miller, Gregory G.; Sweet, Leonard I.; Adams, Jean V.; Omann, Geneva M.; Passino-Reader, Dora R.; Meier, Peter G.

    2002-01-01

    The immunotoxicity of chemical combinations commonly encountered by the lake trout (Salvelinus namaycush) immune system was the focus of this study. It was hypothesised that combinations of an environmental contaminant (mercuric chloride or Aroclor 1254) and an immunomodulatory agent (bacterial endotoxin or cortisol) might interact to produce a greater toxicity than that of the environmental contaminant alone at concentrations typically encountered in piscine blood and other tissues. Thus lake trout thymocytes were isolated and treated with mercuric chloride or Aroclor 1254 in the presence and absence of cortisol or lipopolysaccharide. Incubations were performed for 6 or 20 h at 4° C or 10° C. Lipopolysaccharide did not affect the toxicity of either contaminant. In contrast, cortisol enhanced the toxicity of both environmental contaminants. Hence, stressors that lead to increased cortisol production, but not lipopolysaccharide directly, may increase the toxicity of mercury and Aroclor 1254 to lake trout thymocytes.

  1. Comparative effects of technical toxaphene, 2,5-dichloro-3-biphenylol and octabromodiphenylether on cell viability, [Ca2+]i levels and membrane fluidity in mouse thymocytes.

    PubMed

    Sandal, Suleyman; Yilmaz, Bayram; Chen, Chang-Hwei; Carpenter, David O

    2004-08-01

    Flow cytometric studies of mouse thymocytes show that technical toxaphene (10-20 ppm) and 2,5-dichloro-3-biphenylol (PCB 9-OH) (5-10 ppm) kill cells and cause an increase in intracellular calcium concentration, [Ca2+]i, whereas commercial octabromodiphenylether (OBDE) has no effect. The cell death is not a result of the rise of [Ca2+]i, since the divalent cation ionophore, ionomycin, causes a large elevation in [Ca2+]i without cell death. We have studied effects of these compounds on membrane fluorescence polarization, a measure of membrane fluidity, using 1,6-diphenyl-1,3,5-hexatriene (DPH). We find that toxaphene causes a decrease in membrane fluidity in the concentration range associated with cell death, whereas PCB 9-OH causes an increase in fluidity and OBDE has no effect. These observations suggest that alterations of membrane fluidity of thymocytes, whether it be an increase or decrease, can cause cytotoxicity.

  2. In vitro toxicity and interactions of environmental contaminants (Arochlor 1254 and mercury) and immunomodulatory agents (lipopolysaccharide and cortisol) on thymocytes from lake trout (Salvelinus namaycush).

    PubMed

    Miller, Gregory G; Sweet, Leonard I; Adams, Jean V; Omann, Geneva M; Passino-Reader, Dora R; Meier, Peter G

    2002-07-01

    The immunotoxicity of chemical combinations commonly encountered by the lake trout (Salvelinus namaycush) immune system was the focus of this study. It was hypothesised that combinations of an environmental contaminant (mercuric chloride or Aroclor 1254) and an immunomodulatory agent (bacterial endotoxin or cortisol) might interact to produce a greater toxicity than that of the environmental contaminant alone at concentrations typically encountered in piscine blood and other tissues. Thus lake trout thymocytes were isolated and treated with mercuric chloride or Aroclor 1254 in the presence and absence of cortisol or lipopolysaccharide. Incubations were performed for 6 or 20 h at 4 degrees C or 10 degrees C. Lipopolysaccharide did not affect the toxicity of either contaminant. In contrast, cortisol enhanced the toxicity of both environmental contaminants. Hence, stressors that lead to increased cortisol production, but not lipopolysaccharide directly, may increase the toxicity of mercury and Aroclor 1254 to lake trout thymocytes.

  3. In vitro co-culture systems for studying molecular basis of cellular interaction between Aire-expressing medullary thymic epithelial cells and fresh thymocytes

    PubMed Central

    Yamaguchi, Yoshitaka; Kudoh, Jun; Yoshida, Tetsuhiko; Shimizu, Nobuyoshi

    2014-01-01

    ABSTRACT We previously established three mouse cell lines (Aire+TEC1, Aire+TEC2 and Aire+DC) from the medullary thymic epithelial cells (mTECs) and dendritic cells (mDCs). These cells constitutively expressed “autoimmune regulator (Aire) gene” and they exhibited various features of self antigen-presenting cells (self-APCs) present in the thymic medullary region. Here, we confirmed our previous observation that Aire+ thymic epithelial cells adhere to fresh thymocytes and kill them by inducing apoptosis, thus potentially reproducing in vitro some aspects of the negative selection of T cells in vivo. In this system, a single Aire+ cell appeared able to kill ∼30 thymocytes within 24 hrs. Moreover, we observed that ectopic expression of peripheral tissue-specific antigens (TSAs), and expression of several surface markers involved in mTEC development, increased as Aire+ cell density increases toward confluency. Thus, these Aire+ cells appear to behave like differentiating mTECs as if they pass through the developmental stages from intermediate state toward mature state. Surprisingly, an in vitro co-culture system consisting of Aire+ cells and fractionated sub-populations of fresh thymocytes implied the possible existence of two distinct subtypes of thymocytes (named as CD4+ killer and CD4− rescuer) that may determine the fate (dead or alive) of the differentiating Aire+mTECs. Thus, our in vitro co-culture system appears to mimic a part of “in vivo thymic crosstalk”. PMID:25326516

  4. In vitro co-culture systems for studying molecular basis of cellular interaction between Aire-expressing medullary thymic epithelial cells and fresh thymocytes.

    PubMed

    Yamaguchi, Yoshitaka; Kudoh, Jun; Yoshida, Tetsuhiko; Shimizu, Nobuyoshi

    2014-10-17

    We previously established three mouse cell lines (Aire(+)TEC1, Aire(+)TEC2 and Aire(+)DC) from the medullary thymic epithelial cells (mTECs) and dendritic cells (mDCs). These cells constitutively expressed "autoimmune regulator (Aire) gene" and they exhibited various features of self antigen-presenting cells (self-APCs) present in the thymic medullary region. Here, we confirmed our previous observation that Aire(+) thymic epithelial cells adhere to fresh thymocytes and kill them by inducing apoptosis, thus potentially reproducing in vitro some aspects of the negative selection of T cells in vivo. In this system, a single Aire(+) cell appeared able to kill ∼30 thymocytes within 24 hrs. Moreover, we observed that ectopic expression of peripheral tissue-specific antigens (TSAs), and expression of several surface markers involved in mTEC development, increased as Aire(+) cell density increases toward confluency. Thus, these Aire(+) cells appear to behave like differentiating mTECs as if they pass through the developmental stages from intermediate state toward mature state. Surprisingly, an in vitro co-culture system consisting of Aire(+) cells and fractionated sub-populations of fresh thymocytes implied the possible existence of two distinct subtypes of thymocytes (named as CD4(+) killer and CD4(-) rescuer) that may determine the fate (dead or alive) of the differentiating Aire(+)mTECs. Thus, our in vitro co-culture system appears to mimic a part of "in vivo thymic crosstalk". © 2014. Published by The Company of Biologists Ltd.

  5. Immature thymocytes become sensitive to calcium-mediated apoptosis with the onset of CD8, CD4, and the T cell receptor expression: a role for bcl-2?

    PubMed

    Andjelić, S; Jain, N; Nikolić-Zugić, J

    1993-11-01

    During intrathymic negative selection by clonal deletion, crosslinking of the T cell receptor (TCR) induces cell death by delivering an apoptotic signal(s) to the nucleus along a calcium-dependent pathway. We investigated the reactivity of early precursor-containing thymocytes to Ca(2+)-induced signals, and discovered a breakpoint in their sensitivity to calcium-mediated cell death (CMCD). CD25+CD8-4- TCR- (triple negative [TN]) thymocytes stimulated with a calcium ionophore maintain their viability and precursor activity. By contrast, their immediate progeny, CD25-CD8lo4loTCR alpha beta lo (triple low [TL]) cells react to calcium elevation by abrogation of precursor activity and apoptotic cell death. This developmental difference is specific for CMCD, since both CD25+TN and CD25-TL cells are susceptible to steroid-induced apoptosis. The presence of bcl-2 mRNA correlates directly to the resistance to CMCD-CD25+ TN cells express it and CD25-TL cells do not. These experiments show that thymocytes become sensitive to Ca(2+)-induced apoptosis as soon as they begin to express molecules that mediate thymic selection, and suggest that a concomitant downregulation of bcl-2 may mediate this phenomenon.

  6. beta-sitosterol decreases irradiation-induced thymocyte early damage by regulation of the intracellular redox balance and maintenance of mitochondrial membrane stability.

    PubMed

    Li, Chun Rong; Zhou, Zhe; Lin, Ru Xin; Zhu, Dan; Sun, Yu Ning; Tian, Lin Lin; Li, Lu; Gao, Yue; Wang, Sheng Qi

    2007-10-15

    Both radiation injury and oxidation toxicity occur when cells are exposed to ion irradiation (IR), ultimately leading to apoptosis. This study was designed to determine the effect of beta-sitosterol (BSS) on early cellular damage in irradiated thymocytes and a possible mechanism of effect on irradiation-mediated activation of the apoptotic pathways. Thymocytes were irradiated (6 Gy) with or without BSS. Cell apoptosis and apoptosis-related proteins were evaluated. BSS decreased irradiation-induced cell death and nuclear DNA strand breaks while attenuating intracellular reactive oxygen species (ROS) and increasing the activities of antioxidant enzymes, including superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx). BSS decreased the release of cytochrome c from mitochondria to the cytosol and the mitochondrio-nuclear translocation of apoptosis-inducing factor (AIF). Furthermore, BSS partially inhibited the radiation-induced increase of cleaved caspase 3 and cleaved PARP, and attenuated the activation of JNK and AP-1. In addition, evidence suggests that ROS generated by irradiation are involved in this course of cell damage. The results indicate that BSS confers a radioprotective effect on thymocytes by regulation of the intracellular redox balance which is carried out via the scavenging of ROS and maintenance of mitochondrial membrane stability.

  7. Immature thymocytes become sensitive to calcium-mediated apoptosis with the onset of CD8, CD4, and the T cell receptor expression: a role for bcl-2?

    PubMed Central

    1993-01-01

    During intrathymic negative selection by clonal deletion, crosslinking of the T cell receptor (TCR) induces cell death by delivering an apoptotic signal(s) to the nucleus along a calcium-dependent pathway. We investigated the reactivity of early precursor-containing thymocytes to Ca(2+)-induced signals, and discovered a breakpoint in their sensitivity to calcium-mediated cell death (CMCD). CD25+CD8-4- TCR- (triple negative [TN]) thymocytes stimulated with a calcium ionophore maintain their viability and precursor activity. By contrast, their immediate progeny, CD25-CD8lo4loTCR alpha beta lo (triple low [TL]) cells react to calcium elevation by abrogation of precursor activity and apoptotic cell death. This developmental difference is specific for CMCD, since both CD25+TN and CD25-TL cells are susceptible to steroid- induced apoptosis. The presence of bcl-2 mRNA correlates directly to the resistance to CMCD-CD25+ TN cells express it and CD25-TL cells do not. These experiments show that thymocytes become sensitive to Ca(2+)- induced apoptosis as soon as they begin to express molecules that mediate thymic selection, and suggest that a concomitant downregulation of bcl-2 may mediate this phenomenon. PMID:8228820

  8. Drug induced hypersensitivity syndrome by triple therapy of peginterferon alpha2b, ribavirin and telaprevir in patient with double positive for HBV and HCV.

    PubMed

    Takagi, Hitoshi; Hoshino, Takashi; Naganuma, Atsushi; Koitabashi, Eri; Uehara, Sanae; Sakamoto, Naomi; Kudo, Tomohiro; Ryusaki, Keiichirou; Kakizaki, Satoru; Okamoto, Hiroaki

    2013-10-01

    Sixty year-old male positive for both HCV-RNA and HBsAg was treated by triple therapy of peginterferon alpha2b, ribavirin and telaprevir. Eight weeks after the beginning of the therapy, the patient developed drug induced hypersensitivity syndrome (DIHS) with general erythema multiforme and 64 times anti-HHV6 antibody elevation. Sixty milligram of prednisolone was administered with gradual dose reduction and the skin lesion was improved. HBV-DNA and transaminase elevated one week after the steroid induction and entecavir improved them. DIHS itself and the aggravation of hepatitis B by corticosteroid should be kept in mind in cases with dual infection of HBV and HCV treated by antivirals including telaprevir.

  9. A Randomized, Double-blind, Non-inferiority Trial of Magnesium Sulphate versus Dexamethasone for Prevention of Postoperative Sore Throat after Lumbar Spinal Surgery in the Prone Position

    PubMed Central

    Park, Jin Ha; Shim, Jae-Kwang; Song, Jong-Wook; Jang, Jaewon; Kim, Ji Hoon; Kwak, Young-Lan

    2015-01-01

    Background: Postoperative sore throat (POST) is a frequent complication of tracheal intubation, particularly after surgery in the prone position. We designed this study to validate the non-inferiority of magnesium sulphate against dexamethasone for prevention of POST after lumbar spinal surgery. Methods: One hundred and forty-six patients were randomly allocated to receive either magnesium or dexamethasone. Before anesthetic induction, the magnesium group (n = 73) received magnesium sulphate 30 mg/kg followed by 10 mg/kg/h by continuous infusion until the end of surgery. The dexamethasone group (n = 73) received dexamethasone 8 mg. The primary endpoint was the overall incidence of POST, which was assessed serially over 48 hr postoperatively. The predefined margin of non-inferiority for magnesium against dexamethasone was 15%. Results: Overall incidences of POST at rest (50.7% versus 49.3% in the magnesium and dexamethasone group, respectively, p = 0.869) and swallowing (65.8% versus 61.6% in the magnesium and dexamethasone group, respectively, p = 0.606) were not different between the groups. The upper limit of the 90% confidence interval, which must be lower than the predefined margin of non-inferiority to prove the non-inferiority of magnesium sulphate against dexamethasone, for at rest and swallowing were 14.97% (p = 0.0496) and 17.19% (p = 0.0854), respectively. The incidences and severities of POST and hoarseness were also not different between the groups throughout the study period. Conclusions: Prophylactic magnesium sulphate appears to be non-inferior to dexamethasone for the prevention of POST at rest in patients undergoing lumbar spinal surgery in the prone position. PMID:26516308

  10. Phase III double-blind, placebo-controlled, prospective randomized trial of adjuvant tamoxifen vs. tamoxifen and fenretinide in postmenopausal women with positive receptors (EB193): an intergroup trial coordinated by the Eastern Cooperative Oncology Group.

    PubMed

    Rao, Ruta D; Cobleigh, Melody A; Gray, Robert; Graham, Mark L; Norton, Larry; Martino, Silvana; Budd, George Thomas; Ingle, James N; Wood, William C

    2011-12-01

    Fenretinide and tamoxifen have additive antitumor effects preclinically. We performed a randomized, placebo-controlled, double-blind adjuvant trial in breast cancer patients treated for 5 years with tamoxifen, with or without fenretinide. Between October 1995 and October 1999, 426 postmenopausal women with hormone receptor-positive breast cancer were randomized. Patients were monitored for efficacy and toxicity. Four hundred and nineteen patients were evaluable. The study was terminated early due to slow accrual. There were no significant differences between treatment groups in DFS, TTR or survival. More patients stopped treatment early on the fenretinide arm than on placebo (P = 0.02). Grade 3/4 toxicities, including visual problems and musculoskeletal complaints were more common in patients receiving fenretinide (P = 0.007). A Night Blindness Questionnaire was used to monitor nyctalopia, which was slightly, but not significantly, more common on fenretinide. In this underpowered study, no significant difference was observed in efficacy between treatment groups. This trial provides important toxicity information about fenretinide, a retinoid that has been used in the prevention setting, because it is the only placebo-controlled, double-blind randomized study ever performed.

  11. Phase III Double-Blind, Placebo-Controlled, Prospective Randomized Trial of Adjuvant Tamoxifen vs. Tamoxifen and Fenretinide in Postmenopausal Women with Positive Receptors (EB193): An Intergroup Trial Coordinated by the Eastern Cooperative Oncology Group

    PubMed Central

    Rao, Ruta D.; Cobleigh, Melody A.; Gray, Robert; Graham, Mark L.; Norton, Larry; Martino, Silvana; Budd, George Thomas; Ingle, James N.; Wood, William C.

    2016-01-01

    Purpose Fenretinide and tamoxifen have additive antitumor effects preclinically. We performed a randomized, placebo-controlled, double-blind adjuvant trial in breast cancer patients treated for five years with tamoxifen, with or without fenretinide. Patients and methods Between October 1995 and October 1999, 426 postmenopausal women with hormone receptor-positive breast cancer were randomized. Patients were monitored for efficacy and toxicity. Results Four hundred nineteen patients were evaluable. The study was terminated early due to slow accrual. There were no significant differences between treatment groups in DFS, TTR or survival. More patients stopped treatment early on the fenretinide arm than on placebo (p=.02). Grade 3/4 toxicities, including visual problems and musculoskeletal complaints were more common in patients receiving fenretinide (p=.007). A Night Blindness Questionnaire was used to monitor nyctalopia, which was slightly, but not significantly, more common on fenretinide. Conclusions In this underpowered study, no significant difference was observed in efficacy between treatment groups. This trial provides important toxicity information about fenretinide, a retinoid which has been used in the prevention setting, because it is the only placebo-controlled, double-blind randomized study ever performed. PMID:20878269

  12. Pharmacokinetic and pharmacodynamic characterisation of JNJ-40411813, a positive allosteric modulator of mGluR2, in two randomised, double-blind phase-I studies.

    PubMed

    Salih, Hiba; Anghelescu, Ion; Kezic, Iva; Sinha, Vikash; Hoeben, Eef; Van Nueten, Luc; De Smedt, Heidi; De Boer, Peter

    2015-04-01

    Metabotropic glutamate receptor-2 positive allosteric modulator, JNJ-40411813 (ADX71149), was characterised for clinical effects in healthy volunteers in two phase-1 studies. In study 1, healthy men received 50-, 100-, 150- or 225 mg and women received 100 mg JNJ-40411813 (n=6, each cohort) or placebo (n=2, each cohort) twice daily for seven days; smoking men (n=30) received placebo twice daily on days 1-7, 100 mg JNJ-40411813 (n=20) or placebo (n=10) on days 8-14. In study 2, healthy men received intravenous 0.005 mg/kg S(+) ketamine over 60 min at 3 (n=24; cohort 1), 12 h (n=8; cohort 3), and 24 h (n=8; cohort 2) after a single oral dose of 500 mg JNJ-40411813 or placebo. The pharmacokinetics and effects of JNJ-40411813 on cognition and subjective awareness were evaluated. Plasma JNJ-40411813 exposure was dose-dependent, t max ranged from 3-4 h and t 1/2 19.4-34.2 h across the dose levels. JNJ-40411813 significantly (p=0.02) reduced continuity of attention score (150 mg dose) and ameliorated smoking withdrawal-induced changes in power of attention and quality of episodic memory versus placebo. A modest reduction in alertness was observed at 150-225 mg doses, JNJ-40411813 (500 mg) reduced S(+) ketamine-induced negative symptoms by approximately 43% and 30% in cohorts 1 and 3, respectively. JNJ-40411813 was generally well-tolerated.

  13. Nanomolar concentrations of zinc pyrithione increase cell susceptibility to oxidative stress induced by hydrogen peroxide in rat thymocytes.

    PubMed

    Oyama, Tomohiro M; Saito, Minoru; Yonezawa, Takayasu; Okano, Yoshiro; Oyama, Yasuo

    2012-06-01

    Zinc pyrithione is used as an antifouling agent. However, the environmental impacts of zinc pyrithione have recently been of concern. Zinc induces diverse actions during oxidative stress; therefore, we examined the effect of zinc pyrithione on rat thymocytes suffering from oxidative stress using appropriate fluorescent probes. The cytotoxicity of zinc pyrithione was not observed when the cells were incubated with 3 μM zinc pyrithione for 3 h. However, zinc pyrithione at nanomolar concentrations (10 nM or more) significantly increased the lethality of cells suffering from oxidative stress induced by 3 mM H(2)O(2). The application of zinc pyrithione alone at nanomolar concentrations increased intracellular Zn(2+) level and the cellular content of superoxide anions, and decreased the cellular content of nonprotein thiols. The simultaneous application of nanomolar zinc pyrithione and micromolar H(2)O(2) synergistically increased the intracellular Zn(2+) level. Therefore, zinc pyrithione at nanomolar concentrations may exert severe cytotoxic action on cells simultaneously exposed to chemicals that induce oxidative stress. If so, zinc pyrithione leaked from antifouling materials into surrounding environments would be a risk factor for aquatic ecosystems. Alternatively, zinc pyrithione under conditions of oxidative stress may become more potent antifouling ingredient.

  14. Exuberant cortical thymocyte proliferation mimicking T-lymphoblastic lymphoma within recurrent large inguinal lymph node masses of localized Castleman disease.

    PubMed

    Kansal, Rina; Nathwani, Bharat N; Yiakoumis, Xanthi; Moschogiannis, Maria; Sachanas, Sotirios; Stefanaki, Kalliopi; Pangalis, Gerassimos A

    2015-07-01

    We report a 13-year-old adolescent girl, the youngest thus far, with "an indolent T-lymphoblastic" proliferation (~10%) that uniquely presented within recurrent, large inguinal lymph node masses in a predominating (90%) background of Castleman disease. These nodal masses were resected thrice; the patient is well 5 years after diagnosis without further treatment. Histologically, the features of Castleman disease, hyaline vascular type, were present. Importantly, the interfollicular T-lymphoblastic component occurred as multiple clusters and islands of variable shapes and sizes composed of small "lymphoblasts" indistinguishable from normal cortical thymocytes but without thymic epithelial cells. Immunohistochemically, these lymphoblasts were consistent with the intermediate stage of T-cell differentiation (TdT(+)CD34(-)CD99(+)CD1a(+)CD2(+)CD3(+)CD4(+)CD8(+)CD5(+)CD7(+)CD10(+) [subset]), with 80% Ki-67. Molecularly, the T cells were nonclonal. Our case provides evidence for the benign nature of this highly unusual and poorly understood entity; because the current terminology can be readily misinterpreted as an indolent lymphoblastic lymphoma, we suggest a new term accurately reflecting this entity. Copyright © 2015 Elsevier Inc. All rights reserved.

  15. Effects of ultraviolet radiation on the production of epidermal derived thymocyte-activating factor/interleukin-1

    SciTech Connect

    Gahring, L.C.

    1986-01-01

    The effect of both a single exposure and multiple exposures to ultraviolet radiation (UVR) on the production and/or release of epidermal cell derived thymocyte activating factor/interleukin-1 (ETAF/IL-1) were investigated. A single exposure to UVR enhances the release of ETAF/IL-1 both in vitro and in vivo. This conclusion was based on the observation that: (1) in vitro UV-irradiation of the keratinocyte cell line elevated the release of ETAF/IL-1 on a per cell basis; (2) exposure of animals to UVR stimulated a number of in vivo biologic responses which are known to be mediated by ETAF/IL-1; and (3) ETAF/IL-1 could be detected in the serum of UVR exposed but not normal animals. Exposure of mice to UV-irradiation on a daily basis induced a desensitized state in which animals were found to be refractory to further stimulation with this inflammatory agent. A similar desensitization or tolerance was also shown to be induced by multiple intraperitoneal injections of bacterial lipopolysaccharide (LPS). Desensitization, induced by either LPS or UVR, was found to be regulated at the site of interaction between the cells capable of ETAF/IL-1 production and the exogenous inflammatory stimulus. The results indicate that the regulatory mechanisms for ETAF/IL-1 production which are employed by the keratinocyte are distinct from those regulating ETAF/Il-1 production by the macrophage.

  16. Concurrent V(D)J recombination and DNA end instability increase interchromosomal trans-rearrangements in ATM-deficient thymocytes.

    PubMed

    Bowen, Steven; Wangsa, Darawalee; Ried, Thomas; Livak, Ferenc; Hodes, Richard J

    2013-04-01

    During the CD4(-)CD8(-) (DN) stage of T-cell development, RAG-dependent DNA breaks and V(D)J recombination occur at three T-cell receptor (TCR) loci: TCRβ, TCRγ and TCRδ. During this stage, abnormal trans-rearrangements also take place between TCR loci, occurring at increased frequency in absence of the DNA damage response mediator ataxia telangiectasia mutated (ATM). Here, we use this model of physiologic trans-rearrangement to study factors that predispose to rearrangement and the role of ATM in preventing chromosomal translocations. The frequency of DN thymocytes with DNA damage foci at multiple TCR loci simultaneously is increased 2- to 3-fold in the absence of ATM. However, trans-rearrangement is increased 10 000- to 100 000-fold, indicating that ATM function extends beyond timely resolution of DNA breaks. RAG-mediated synaptic complex formation occurs between recombination signal sequences with unequal 12 and 23 base spacer sequences (12/23 rule). TCR trans-rearrangements violate this rule, as we observed similar frequencies of 12/23 and aberrant 12/12 or 23/23 recombination products. This suggests that trans-rearrangements are not the result of trans-synaptic complex formation, but they are instead because of unstable cis synaptic complexes that form simultaneously at distinct TCR loci. Thus, ATM suppresses trans-rearrangement primarily through stabilization of DNA breaks at TCR loci.

  17. A dynamic niche provides Kit ligand in a stage-specific manner to the earliest thymocyte progenitors

    PubMed Central

    Buono, Mario; Facchini, Raffaella; Matsuoka, Sahoko; Thongjuea, Supat; Waithe, Dominique; Luis, Tiago C.; Giustacchini, Alice; Besmer, Peter; Mead, Adam J.; Jacobsen, Sten Eirik W.; Nerlov, Claus

    2016-01-01

    Thymic T-cell development is initiated from bone marrow-derived multi-potent thymus seeding progenitors (TSPs). During the early stages of thymocyte differentiation progenitors become T-cell restricted. However, the cellular environments supporting these critical initial stages of T-cell development within the thymic cortex are not known. We here use the dependence of early, c-Kit–expressing thymic progenitors on Kit ligand (KitL) to show that CD4–CD8–c-Kit+CD25– DN1-stage progenitors associate with, and depend on the membrane-bound form of KitL (mKitL) provided by, a cortex-specific KitL-expressing vascular endothelial cell (VEC) population. In contrast, the subsequent CD4–CD8–c-Kit+CD25+ DN2 stage progenitors associate selectively with cortical thymic epithelial cells (cTECs) and depend on cTEC-presented mKitL. These results show that the dynamic process of early thymic progenitor differentiation is paralleled by migration-dependent changes to the supporting niche, and identify VECs as a thymic niche cell, with mKitL as a critical ligand. PMID:26780297

  18. Proteomic profiling of rapid non-genomic and concomitant genomic effects of acute restraint stress on rat thymocytes.

    PubMed

    Billing, Anja M; Revets, Dominique; Hoffmann, Céline; Turner, Jonathan D; Vernocchi, Sara; Muller, Claude P

    2012-04-03

    In order to investigate rapid non-genomic effects of acute stress, rats were restrained for 15 min which was sufficient to activate the hypothalamus-pituitary-adrenal (HPA) axis but too short to induce massive genomic effects of cortisol. Subcellular fractions of thymocytes (cytosol, nucleus, membrane) were investigated using quantitative 2D DIGE with MALDI-TOF/TOF mass spectrometry. In total, 108 proteins with differential subcellular localizations were identified. The specificity of the changes induced by psychological stress was reflected by the prominent modulation of proteins involved in the HPA and sympathoadrenal medullar (SAM) axis such as HMGB1 and NHERF1. Intracellular trafficking was characterized by a dominant protein exodus from the cytosol. Real translocation was observed for 9 proteins with 6 that shuttled from the cytosol to the nucleus (HYOU1, HNRPF, HNRPC, STRAP, PSA1, PPA1) and 3 from the nucleus to the cytosol (HMGB1, NHERF1, PSMA1). Proteins showing subcellular reshuffling were largely involved in transcription and translation processes (39 of 108) with a significant enrichment of RNA splicing factors. Bioinformatics analysis revealed significant enrichment for protein kinase A and 14-3-3 signaling, probably reflecting real non-genomic effects. This is the first study investigating rapid effects of stress-induced HPA activation in vivo at the proteome level. Copyright © 2012 Elsevier B.V. All rights reserved.

  19. Immunosuppressive therapy with horse anti-thymocyte globulin and cyclosporine as treatment for fulminant aplastic anemia in children.

    PubMed

    Yagasaki, Hiroshi; Shichino, Hiroyuki; Ohara, Akira; Kobayashi, Ryoji; Yabe, Hiromasa; Ohga, Shouichi; Hamamoto, Kazuko; Ohtsuka, Yoshitoshi; Shimada, Hiroyuki; Inoue, Masami; Muramatsu, Hideki; Takahashi, Yoshiyuki; Kojima, Seiji

    2014-05-01

    Patients with severe aplastic anemia (SAA) and an absolute neutrophil count (ANC) of 0 typically have fatal outcomes. We defined fulminant AA (FAA) as ANC = 0 for at least 2 weeks prior to and after immunosuppressive therapy (IST). We analyzed the outcomes of 35 children with FAA among 288 children who enrolled in a prospective study for AA (AA-97 study). AA was classified as FAA (n = 35), very SAA (vSAA; n = 129), or SAA (n = 124). All of the children received the IST with horse anti-thymocyte globulin (ATG) and cyclosporine (CsA). A significantly lower response rate at 6 months was seen in children with FAA when compared to those with vSAA or SAA (40.0, 63.6, and 63.7 %, respectively; p = 0.027). Of 20 nonresponder patients in the FAA group, 11 were rescued by alternative donor transplantation, and 5 patients showed a late response after 6 months. Consequently, no significant difference was noted in overall survival when comparing the FAA, vSAA, and SAA groups (88.5, 95.8, and 96.8 %). These findings indicate that IST with ATG and CsA is justified as a first-line treatment for children with FAA who lack a human leukocyte antigen-matched sibling donor.

  20. Double helicenes

    NASA Astrophysics Data System (ADS)

    Bachrach, Steven M.

    2016-12-01

    The even double helicenes with 4-12 phenyl groups in each helix were examined at B3LYP-D3/6-311G(d). The double helicenes with 4-10 phenyl rings are less than twice as strained as their component helicenes; the strain results from twisting about the shared naphthyl moiety, with accompanying loss of aromaticity. These compounds should be reasonable synthetic targets, and computed NMR shifts are provided to aid in their characterization.

  1. Double Negative Materials (DNM), Phenomena and Applications

    DTIC Science & Technology

    2009-07-01

    Nanoparticles Formed by Pairs Of Concentric Double-Negative (DNG), Single-Negative ( SNG ) and/or Double-Positive (DPS) Metamaterial Layers.” J. Appl...material RRL Rapid Research Letters SHG second-harmonic generation SNG single-negative SSR split-ring resonator A-1 Appendix A. October 2008...Pairs of Concentric Double-Negative (DNG), Single-Negative ( SNG ), and/or Double-Positive (DPS) Metamaterial Layers.” J. Appl. Phys. 97, no. 9 (May

  2. Maize food allergy: lipid-transfer proteins, endochitinases, and alpha-zein precursor are relevant maize allergens in double-blind placebo-controlled maize-challenge-positive patients.

    PubMed

    Pastorello, Elide A; Farioli, Laura; Pravettoni, Valerio; Scibilia, Joseph; Conti, Amedeo; Fortunato, Donatella; Borgonovo, Linda; Bonomi, Simona; Primavesi, Laura; Ballmer-Weber, Barbara

    2009-09-01

    Italian patients with maize anaphylaxis have been shown to have IgE toward two major maize allergens: an alpha-amylase inhibitor and a 9-kDa LTP. A complete study on maize food allergens in patients with positive maize double-blind, placebo-controlled food challenge (DBPCFC) is lacking. The objective was to utilize the three maize protein fractions to identify and characterize the most relevant IgE-binding proteins recognized by the sera of Italian and Swiss patients with either a positive maize-DBPCFC or a history of maize-induced anaphylaxis. Osborne's protein fractions of maize were extracted to obtain water-soluble, total zein, and total protein fractions. Protein IgE-binding capacity was investigated by SDS-PAGE immunoblotting using the sera from DBPCFC-positive patients and from patients with maize-induced anaphylaxis. Purified maize LTP was used to inhibit the IgE immunoblotting of the three protein fractions. IgE immunoblotting demonstrated that the 9-kDa LTP was recognized by all the Italian patients and by none of the Swiss patients. Other allergens were: 14-kDa alpha-amylase inhibitor, 30-kDa endochitinases A and -B, 19 kDa zein-beta precursor, and 26 kDa zein-alpha precursor; a newly described allergen, the globulin-2 precursor, identified in the total protein fraction. It is noteworthy that maize LTP and endochitinase were cross-reactive with grape LTP and one grape endochitinase. LTP was found to be the only major allergen in Italian patients with either positive maize challenge or a history of maize-induced anaphylaxis. We have identified other maize allergens in subjects with maize food allergy, as grape cross-reactive endochitinase, however, the clinical significance of these proteins needs to be investigated in larger groups of patients with allergy to these food items.

  3. L-theanine relieves positive, activation, and anxiety symptoms in patients with schizophrenia and schizoaffective disorder: an 8-week, randomized, double-blind, placebo-controlled, 2-center study.

    PubMed

    Ritsner, Michael S; Miodownik, Chanoch; Ratner, Yael; Shleifer, Tatyana; Mar, Maria; Pintov, Leonid; Lerner, Vladimir

    2011-01-01

    L-theanine is a unique amino acid present almost exclusively in the tea plant. It possesses neuroprotective, mood-enhancing, and relaxation properties. This is a first study designed to evaluate the efficacy and tolerability of L-theanine augmentation of antipsychotic treatment of patients with chronic schizophrenia and schizoaffective disorder. 60 patients with DSM-IV schizophrenia or schizoaffective disorder participated in an 8-week, double-blind, randomized, placebo-controlled study. 400 mg/d of L-theanine was added to ongoing antipsychotic treatment from February 2006 until October 2008. The outcome measures were the Positive and Negative Syndrome Scale (PANSS), the Hamilton Anxiety Rating Scale (HARS), the Cambridge Neuropsychological Test Automated Battery (CANTAB) for neurocognitive functioning, and additional measures of general functioning, side effects, and quality of life. 40 patients completed the study protocol. Compared with placebo, L-theanine augmentation was associated with reduction of anxiety (P = .015; measured by the HARS scale) and positive (P = .009) and general psychopathology (P < .001) scores (measured by the PANSS 3-dimensional model). According to the 5-dimension model of psychopathology, L-theanine produced significant reductions on PANSS positive (P = .004) and activation factor (P = .006) scores compared to placebo. The effect sizes (Cohen d) for these differences ranged from modest to moderate (0.09-0.39). PANSS negative and CANTAB task scores, general functioning, side effect, and quality of life measures were not affected by L-theanine augmentation. L-theanine was found to be a safe and well-tolerated medication. L-theanine augmentation of antipsychotic therapy can ameliorate positive, activation, and anxiety symptoms in schizophrenia and schizoaffective disorder patients. Further long-term studies of L-theanine are needed to substantiate the clinically significant benefits of L-theanine augmentation. © Copyright 2011 Physicians

  4. Diethylstilbestrol alters positive and negative selection of T cells in the thymus and modulates T-cell repertoire in the periphery

    SciTech Connect

    Brown, Nicole; Nagarkatti, Mitzi; Nagarkatti, Prakash S. . E-mail: pnagark@hsc.vcu.edu

    2006-04-15

    Prenatal exposure to diethylstilbestrol (DES) is known to cause altered immune functions and increased susceptibility to autoimmune disease in humans. In the current study, we investigated the effects of DES on T-cell differentiation in the thymus using the HY-TCR transgenic (Tg) mouse model in which the female mice exhibit positive selection of T cells bearing the Tg TCR, while the male mice show negative selection of such T cells. In female HY-TCR-Tg mice, exposure to DES showed more pronounced decrease in thymic cellularity when compared to male mice. Additionally, female mice also showed a significant decrease in the proportion of double-positive (DP) T cells in the thymus and HY-TCR-specific CD8{sup +} T cells in the periphery. Male mice exhibiting negative selection also showed decreased thymic cellularity following DES exposure. Moreover, the male mice showed increased proportion of double-negative (DN) T cells in the thymus and decreased proportion of CD8{sup +} T cells. The density of expression of HY-TCR on CD8{sup +} cells was increased following DES exposure in both females and males. Finally, the proliferative response of thymocytes to mitogens and peripheral lymph node T cells to male H-Y antigen was significantly altered in female and male mice following DES treatment. Taken together, these data suggest that DES alters T-cell differentiation in the thymus by interfering with positive and negative selection processes, which in turn modulates the T-cell repertoire in the periphery.

  5. Crossreactive αβ T cell receptors are the predominant targets of thymocyte negative selection

    PubMed Central

    McDonald, Benjamin D.; Bunker, Jeffrey J.; Erickson, Steven A.; Oh-Hora, Masatsugu; Bendelac, Albert

    2015-01-01

    SUMMARY The precise impact of thymic positive and negative selection on the T cell receptor (TCR) repertoire remains controversial. Here, we used unbiased, high-throughput cloning and retroviral expression of individual preselection TCRs to provide a direct assessment of these processes at the clonal level in vivo. We found that 15% of random TCRs induced signaling and directed positive (7.5%) or negative (7.5%) selection, depending on strength of signal, whereas the remaining 85% failed to induce signaling or selection. Most negatively selected TCRs exhibited promiscuous crossreactivity toward multiple other major histocompatibility complex (MHC) haplotypes. In contrast, TCRs that were positively selected or non-selected were minimally crossreactive. Negative selection of crossreactive TCRs led to clonal deletion but also recycling into intestinal CD4−CD8β− intraepithelial lymphocytes (iIELs). Thus, broadly crossreactive TCRs arise at low frequency in the pre-selection repertoire but constitute the primary drivers of thymic negative selection and iIEL lineage differentiation. PMID:26522985

  6. Identification of a Late Stage of Small Noncycling pTα−  Pre-T Cells as Immediate Precursors of T Cell Receptor α/β+  Thymocytes

    PubMed Central

    Trigueros, César; Ramiro, Almudena R.; Carrasco, Yolanda R.; de Yebenes, Virginia G.; Albar, Juan P.; Toribio, María L.

    1998-01-01

    During thymocyte development, progression from T cell receptor (TCR)β to TCRα rearrangement is mediated by a CD3-associated pre-TCR composed of the TCRβ chain paired with pre-TCRα (pTα). A major issue is how surface expression of the pre-TCR is regulated during normal thymocyte development to control transition through this checkpoint. Here, we show that developmental expression of pTα is time- and stage-specific, and is confined in vivo to a limited subset of large cycling human pre-T cells that coexpress low density CD3. This restricted expression pattern allowed the identification of a novel subset of small CD3− thymocytes lacking surface pTα, but expressing cytoplasmic TCRβ, that represent late noncycling pre-T cells in which recombination activating gene reexpression and downregulation of T early α transcription are coincident events associated with cell cycle arrest, and immediately preceding TCRα gene expression. Importantly, thymocytes at this late pre-T cell stage are shown to be functional intermediates between large pTα+ pre-T cells and TCRα/β+ thymocytes. The results support a developmental model in which pre-TCR–expressing pre-T cells are brought into cycle, rapidly downregulate surface pre-TCR, and finally become small resting pre-T cells, before the onset of TCRα gene expression. PMID:9782117

  7. Apple Valley Double Star Workshop

    NASA Astrophysics Data System (ADS)

    Brewer, Mark

    2015-05-01

    The High Desert Astronomical Society hosts an annual double star workshop, where participants measure the position angles and separations of double stars. Following the New Generation Science Standards (NGSS), adopted by the California State Board of Education, participants are assigned to teams where they learn the process of telescope set-up and operation, the gathering of data, and the reduction of the data. Team results are compared to the latest epoch listed in the Washington Double Star Catalog (WDS) and papers are written for publication in the Journal of Double Star Observations (JDSO). Each team presents a PowerPoint presentation to their peers about actual hands-on astronomical research.

  8. Psychomotor and subjective effects of bilastine, hydroxyzine, and cetirizine, in combination with alcohol: a randomized, double-blind, crossover, and positive-controlled and placebo-controlled Phase I clinical trials.

    PubMed

    García-Gea, Consuelo; Martínez, Joan; Ballester, Maria Rosa; Gich, Ignasi; Valiente, Román; Antonijoan, Rosa Maria

    2014-03-01

    The aim of this study was to compare the effects of concomitant administration of alcohol and bilastine versus alcohol alone on the central nervous system. Twenty-four healthy young volunteers of both sexes participated in a randomized, double-blind, double-dummy, crossover, and positive-controlled and placebo-controlled clinical trials. At 1-week intervals, subjects received six different treatments: (i) placebo; (ii) alcohol 0.8 g/kg alone (ALC); (iii) ALC in combination with: bilastine 20 mg (B20 + A); (iv) bilastine 80 mg (B80 + A); (v) cetirizine 10 mg (CET + A); and (vi) hydroxyzine 25 mg (HYD + A). Psychomotor performance tests (fine motor, finger tapping, nystagmus, critical flicker-fusion frequency, temporal estimation, 'd2' cancellation, and simple reaction time) and subjective self-reports (drunkenness, drowsiness, mental slowness, clumsiness, anger, attentiveness, competence, happiness, hostility, interest, and extroversion) were carried out at baseline and multiple points thereafter. All active treatments induced a significant psychomotor impairment. The greatest and most lasting impairment was observed with HYD + A followed by B80 + A and CET + A. In contrast, objective measures showed less impairment with B20 + A and ALC, both with a similar magnitude. Self-reports showed a subjective perception of performance impairment in all active treatments. Concomitant administration of bilastine (at therapeutic dose) and alcohol does not produce greater central nervous system depressant effects than ACL alone. Copyright © 2014 John Wiley & Sons, Ltd.

  9. Induction by anti-thymocyte globulins in kidney transplantation: a review of the literature and current usage.

    PubMed

    Malvezzi, Paolo; Jouve, Thomas; Rostaing, Lionel

    2015-10-01

    Preventing acute rejection (AR) after kidney transplantation is of utmost importance because an AR can have a negative impact on long-term allograft survival. Directory of Open Access Journals (DOAJ), Google Scholar, PubMed, EBSCO, and Web of Science have been searched. At the moment this can be done by using rabbit anti-thymocyte globulins (rATGs) as an induction therapy. However, because rATGs are associated with some deleterious side-effects, such as the opportunistic infections cytomegalovirus (CMV) and de novo post-transplant cancer, it is very important they are used optimally, i.e., at minimal doses that avoid many side-effects but still retain optimal treatment efficacy. Recent data show that the risk of CMV infection can be minimized using tacrolimus plus everolimus, and not tacrolimus plus mycophenolic acid, as the maintenance immunosuppression. The use of rATG is particularly valuable in; (a) sensitized patients; (b) in recipients from an expanded-criteria donor, thus enabling the introduction of calcineurin inhibitors at reduced doses; and (c) for patients where steroid avoidance is contemplated. However, we also need to consider that rATG may increase the risk of de novo cancer, even though recent data indicate this is unlikely and that any risk can be reduced by using mammalian target of rapamycin (mTOR) inhibitors instead of mycophenolic acid combined with low-dose calcineurin inhibitors. Even though rATGs do not improve long-term kidney-allograft survival, they may help reduce calcineurin-inhibitor dosage during the early post-transplant period and minimize the risk of AR.

  10. Increase in intracellular Zn2+ concentration by thimerosal in rat thymocytes: intracellular Zn2+ release induced by oxidative stress.

    PubMed

    Hashimoto, Erika; Oyama, Toshihisa B; Oyama, Keisuke; Nishimura, Yumiko; Oyama, Tomohiro M; Ueha-Ishibashi, Toshiko; Okano, Yoshiro; Oyama, Yasuo

    2009-09-01

    Thimerosal (TMR), an ethylmercury-containing preservative in pharmaceutical products, was recently reported to increase intracellular Zn(2+) concentration. Therefore, some health concerns about the toxicity of TMR remain because of physiological and pathological roles of Zn(2+). To reveal the property of TMR-induced increase in intracellular Zn(2+) concentration, the effect of TMR on FluoZin-3 fluorescence, an indicator of intracellular Zn(2+), of rat thymocytes was examined. TMR at concentrations ranging from 0.3 microM to 10 microM increased the intensity of FluoZin-3 fluorescence in a concentration-dependent manner under external Ca(2+)- and Zn(2+)-free condition. The threshold concentration was 0.3-1 microM. The increase in the intensity was significant when TMR concentration was 1 microM or more. N,N,N',N'-Tetrakis(2-pyridylmethyl)ethylenediamine (TPEN), a chelator for intracellular Zn(2+), completely attenuated the TMR-induced augmentation of FluoZin-3 fluorescence. Hydrogen peroxide (H(2)O(2)) and N-ethylmaleimide, reducing cellular thiol content, significantly increased FluoZin-3 fluorescence intensity and decreased 5-chloromethylfluorescein (5-CMF) fluorescence intensity, an indicator for cellular thiol. The correlation coefficient between TMR-induced augmentation of FluoZin-3 fluorescence and attenuation of 5-CMF fluorescence was -0.882. TMR also attenuated the 5-CMF fluorescence in the presence of TPEN. Simultaneous application of H(2)O(2) and TMR synergistically augmented the FluoZin-3 fluorescence. It is suggested that TMR increases intracellular Zn(2+) concentration via decreasing cellular thiol content.

  11. Triclosan, an antibacterial agent, increases intracellular Zn(2+) concentration in rat thymocytes: its relation to oxidative stress.

    PubMed

    Tamura, Ikumi; Kanbara, Yasuhiro; Saito, Minoru; Horimoto, Kanna; Satoh, Masaya; Yamamoto, Hiroshi; Oyama, Yasuo

    2012-01-01

    Triclosan is used as an antibacterial agent in household items and personal care products. Since this compound is found in maternal milk of humans and bodies of wild animals, there is growing concern among some consumer groups and scientific community that triclosan is adverse for humans and wild animals. In order to estimate adverse actions of triclosan, the effects of triclosan on intracellular Zn(2+) concentration and cellular thiol content were studied in rat thymocytes by the use of flow cytometer with appropriate fluorescent probes. Triclosan at 1-3 μM (sublethal concentrations) increased the intensity of FluoZin-3 fluorescence (intracellular Zn(2+) concentration) and decreased the intensity of 5-chloromethylfluorescein (5-CMF) fluorescence (cellular thiol content). Negative correlation (r=-0.985) between triclosan-induced changes in FluoZin-3 and 5-CMF fluorescences was found. Removal of external Zn(2+) did not significantly affect the triclosan-induced augmentation of FluoZin-3 fluorescence, suggesting an intracellular Zn(2+) release by triclosan. These actions of triclosan were similar to those of H(2)O(2) and triclosan significantly potentiated the cytotoxicity of H(2)O(2). Therefore, the results may suggest that triclosan at sublethal concentrations induces oxidative stress that decreases cellular thiol content, resulting in an increase in intracellular Zn(2+) concentration by Zn(2+) release from intracellular store(s). Since recent studies show many physiological roles of intracellular Zn(2+) in cellular functions, the triclosan-induced disturbance of cellular Zn(2+) homeostasis may induce adverse actions on the cells. Copyright © 2011 Elsevier Ltd. All rights reserved.

  12. Innate-like CD4 T cells selected by thymocytes suppress adaptive immune responses against bacterial infections

    PubMed Central

    Qiao, Yu; Gray, Brian M.; Sofi, Mohammed H.; Bauler, Laura D.; Eaton, Kathryn A.; O'Riordan, Mary X. D.; Chang, Cheong-Hee

    2012-01-01

    We have reported a new innate-like CD4 T cell population that expresses cell surface makers of effector/memory cells and produce Th1 and Th2 cytokines immediately upon activation. Unlike conventional CD4 T cells that are selected by thymic epithelial cells, these CD4 T cells, named T-CD4 T cells, are selected by MHC class II expressing thymocytes. Previously, we showed that the presence of T-CD4 T cells protected mice from airway inflammation suggesting an immune regulatory role of T-CD4 T cells. To further understand the function of T-CD4 T cells, we investigated immune responses mediated by T-CD4 T cells during bacterial infection because the generation of antigen specific CD4 T cells contributes to clearance of infection and for the development of immune memory. The current study shows a suppressive effect of T-CD4 T cells on both CD8 and CD4 T cell-mediated immune responses during Listeria and Helicobacter infections. In the mouse model of Listeria monocytogenes infection, T-CD4 T cells resulted in decreasedfrequency of Listeria-specific CD8 T cells and the killing activity of them. Furthermore, mice with T-CD4 T cells developed poor immune memory, demonstrated by reduced expansion of antigen-specific T cells and high bacterial burden upon re-infection. Similarly, the presence of T-CD4 T cells suppressed the generation of antigen-specific CD4 T cells in Helicobacter pylori infected mice. Thus, our studies reveal a novel function of T-CD4 T cells in suppressing anti-bacterial immunity. PMID:23264931

  13. [Double responses].

    PubMed

    Motté, G; Dinanian, S; Sebag, C; Drieu, L; Slama, M

    1995-12-01

    Double response is a rare electrocardiographic phenomenon requiring two atrioventricular conduction pathways with very different electrophysiological properties. Double ventricular responses are the usual manifestation: an atrial depolarisation (spontaneous or provoked, anticipated or not) is followed by a first ventricular response dependent on an accessory pathway or a rapid nodal pathway and then a second response resulting from sufficiently delayed transmission through a nodal pathway for the ventricles to have recovered their excitability when the second wave of activation reaches them. A simple curiosity when isolated and occurring under unusual conditions, particularly during electrophysiological investigation of the Wolff-Parkinson-White syndrome, the double response may initiate symptomatic non-reentrant junctional tachycardia when associated with nodal duality and repeating from atria in sinus rhythm. The functional incapacity and resistance to antiarrhythmic therapy may require referral for ablation of the slow pathway.

  14. Effect of suppressive acyclovir administered to HSV-2 positive mothers from week 28 to 36 weeks of pregnancy on adverse obstetric outcomes: a double-blind randomised placebo-controlled trial.

    PubMed

    Nakubulwa, Sarah; Kaye, Dan K; Bwanga, Freddie; Tumwesigye, Nazarius Mbona; Nakku-Joloba, Edith; Mirembe, Florence

    2017-03-03

    Acyclovir (ACV) given to HSV-2 positive women after 36 weeks reduces adverse outcomes but its benefit at lower gestation was undocumented. We determined the effect of oral acyclovir administered from 28 to 36 weeks on premature rupture of membranes (PROM) primarily and preterm delivery risk. This was a randomized, double-blind placebo-controlled trial among 200 HSV-2 positive pregnant women at 28 weeks of gestation at Mulago Hospital, Uganda. Participants were assigned randomly (1:1) to take either acyclovir 400 mg orally twice daily (intervention) or placebo (control) from 28 to 36 weeks. Both arms received acyclovir after 36 weeks until delivery. Development of Pre-PROM by 36 weeks and preterm delivery were outcomes. One hundred women were randomised to acyclovir and 100 to placebo arms between January 2014 and February 2015. There was tendency towards reduction of incidence of PROM at 36 weeks but this was not statistically significant (4.0% versus 10.0%; RR 0.35; 95% 0.11-1.10) in the acyclovir and placebo arms respectively. However, there was a significant reduction in the incidence of preterm delivery (11.1% versus 23.5%; RR 0.41; 95% 0.20-0.85) in the acyclovir and placebo arms respectively. Oral acyclovir given to HSV-2 positive pregnant women from 28 to 36 weeks reduced incidence of preterm delivery but did not significantly reduce incidence of pre-PROM. www.pactr.org, PACTR201311000558197 .

  15. Single-cell analysis of thymocyte differentiation: identification of transcription factor interactions and a major stochastic component in αβ-lineage commitment.

    PubMed

    Boudil, Amine; Skhiri, Lamia; Candéias, Serge; Pasqualetto, Valérie; Legrand, Agnès; Bedora-Faure, Marie; Gautreau-Rolland, Laetitia; Rocha, Benedita; Ezine, Sophie

    2013-01-01

    T cell commitment and αβ/γδ lineage specification in the thymus involves interactions between many different genes. Characterization of these interactions thus requires a multiparameter analysis of individual thymocytes. We developed two efficient single-cell methods: (i) the quantitative evaluation of the co-expression levels of nine different genes, with a plating efficiency of 99-100% and a detection limit of 2 mRNA molecules/cell; and (ii) single-cell differentiation cultures, in the presence of OP9 cells transfected with the thymus Notch1 ligand DeltaL4. We show that during T cell commitment, Gata3 has a fundamental, dose-dependent role in maintaining Notch1 expression, with thymocytes becoming T-cell-committed when they co-express Notch1, Gata3 and Bc11b. Of the transcription factor expression patterns studied here, only that of Bcl11b was suggestive of a role in Pu1 down-regulation. Individual thymocytes became αβ/γδ lineage-committed at very different stages (from the TN2a stage onwards). However, 20% of TN3 cells are not αβ/γδ-lineage committed and TN4 cells comprise two main subpopulations with different degrees of maturity. The existence of a correlation between differentiation potential and expression of the pre-TCR showed that 83% of αβ-committed cells do not express the pre-TCR and revealed a major stochastic component in αβ-lineage specification.

  16. Single-Cell Analysis of Thymocyte Differentiation: Identification of Transcription Factor Interactions and a Major Stochastic Component in αβ-Lineage Commitment

    PubMed Central

    Boudil, Amine; Skhiri, Lamia; Candéias, Serge; Pasqualetto, Valérie; Legrand, Agnès; Bedora-Faure, Marie; Gautreau-Rolland, Laetitia; Rocha, Benedita; Ezine, Sophie

    2013-01-01

    T cell commitment and αβ/γδ lineage specification in the thymus involves interactions between many different genes. Characterization of these interactions thus requires a multiparameter analysis of individual thymocytes. We developed two efficient single-cell methods: (i) the quantitative evaluation of the co-expression levels of nine different genes, with a plating efficiency of 99–100% and a detection limit of 2 mRNA molecules/cell; and (ii) single-cell differentiation cultures, in the presence of OP9 cells transfected with the thymus Notch1 ligand DeltaL4. We show that during T cell commitment, Gata3 has a fundamental, dose-dependent role in maintaining Notch1 expression, with thymocytes becoming T-cell-committed when they co-express Notch1, Gata3 and Bc11b. Of the transcription factor expression patterns studied here, only that of Bcl11b was suggestive of a role in Pu1 down-regulation. Individual thymocytes became αβ/γδ lineage-committed at very different stages (from the TN2a stage onwards). However, 20% of TN3 cells are not αβ/γδ-lineage committed and TN4 cells comprise two main subpopulations with different degrees of maturity. The existence of a correlation between differentiation potential and expression of the pre-TCR showed that 83% of αβ-committed cells do not express the pre-TCR and revealed a major stochastic component in αβ-lineage specification. PMID:24098325

  17. Ephrin-B-dependent thymic epithelial cell-thymocyte interactions are necessary for correct T cell differentiation and thymus histology organization: relevance for thymic cortex development.

    PubMed

    Cejalvo, Teresa; Munoz, Juan J; Tobajas, Esther; Fanlo, Lucía; Alfaro, David; García-Ceca, Javier; Zapata, Agustín

    2013-03-15

    Previous analysis on the thymus of erythropoietin-producing hepatocyte kinases (Eph) B knockout mice and chimeras revealed that Eph-Eph receptor-interacting proteins (ephrins) are expressed both on T cells and thymic epithelial cells (TECs) and play a role in defining the thymus microenvironments. In the current study, we have used the Cre-LoxP system to selectively delete ephrin-B1 and/or ephrin-B2 in either thymocytes (EfnB1(thy/thy), EfnB2(thy/thy), and EfnB1(thy/thy)EfnB2(thy/thy) mice) or TECs (EfnB1(tec/tec), EfnB2(tec/tec), and EfnB1(tec/tec)EfnB2(tec/tec) mice) and determine the relevance of these Eph ligands in T cell differentiation and thymus histology. Our results indicate that ephrin-B1 and ephrin-B2 expressed on thymocytes play an autonomous role in T cell development and, expressed on TECs, their nonautonomous roles are partially overlapping. The effects of the lack of ephrin-B1 and/or ephrin-B2 on either thymocytes or TECs are more severe and specific on thymic epithelium, contribute to the cell intermingling necessary for thymus organization, and affect cortical TEC subpopulation phenotype and location. Moreover, ephrin-B1 and ephrin-B2 seem to be involved in the temporal appearance of distinct cortical TECs subsets defined by different Ly51 levels of expression on the ontogeny.

  18. Passive smoking exposure from partners as a risk factor for ER+/PR+ double positive breast cancer in never-smoking Chinese urban women: a hospital-based matched case control study.

    PubMed

    Tong, Jian-hua; Li, Zhi; Shi, Jing; Li, He-ming; Wang, Yan; Fu, Ling-yu; Liu, Yun-peng

    2014-01-01

    The relationship between passive smoking exposure (PSE) and breast cancer risk is of major interest. To evaluate the relationship between PSE from partners and breast cancer risk stratified by hormone-receptor (HR) status in Chinese urban women population. Hospital-based matched case control study. Chinese urban breast cancer patients without current or previous active smoking history in China Medical University 1st Hospital, Liaoning Province, China between Jan 2009 and Nov 2009. Each breast cancer patient was matched 1∶1 with healthy controls by gender and age (±2 years) from the same hospital. The authors used unconditional logistic regression analyses to estimate odds ratio for women with PSE from partners and breast cancer risk. 312 pairs were included in the study. Women who endured PSE had significantly increased risk of breast cancer (adjusted OR: 1.46; 95% CI: 1.05-2.03; P = 0.027), comparing with unexposed women. Women who exposed to >5 cigarettes/day also had significant increased risk (adjusted OR: 1.99; 95% CI: 1.28-3.10; P = 0.002), as were women exposed to passive smoke for 16-25 years (adjusted OR: 1.87 95% CI: 1.22-2.86; P = 0.004), and those exposed to > 4 pack-years (adjusted OR: 1.71 95% CI: 1.17-2.50; P = 0.004). Similar trends were significant for estrogen receptor (ER)/progesterone receptor (PR) double positive subgroup(adjusted OR: 1.71; 2.20; 1.99; 1.92, respectively), but not for ER+/PR-, ER-/PR+, or ER-/PR- subgroups. limitations of the hospital-based retrospective study, lack of information on entire lifetime PSE and low statistical power. Our findings provide further evidence that PSE from partners contributes to increased risk of breast cancer, especially for ER/PR double positive breast cancer, in Chinese urban women.

  19. Anti-alpha 4 integrin antibody induces apoptosis in murine thymocytes and staphylococcal enterotoxin B-activated lymph node T cells.

    PubMed Central

    Tchilian, E Z; Owen, J J; Jenkinson, E J

    1997-01-01

    We have shown that an antibody (9C10) to the alpha 4 integrin induces apoptosis in murine immature CD4+ CD8+ thymocytes and in activated (but not resting) mature lymph node T cells. In both cases, apoptosis is blocked by the highly selective protein kinase C (PKC) inhibitor Ro31.8425, suggesting that 9C10 induces signalling through the alpha 4 integrin resulting in PKC activation leading to apoptosis. Overall, our results indicate the potential role of the alpha 4 integrin-mediated interactions in apoptosis induction during T-cell development and following mature T-cell activation. PMID:9486103

  20. Double Crater

    NASA Image and Video Library

    2012-03-23

    A double crater, called a crater doublet, is seen in the bottom right part of this image from NASA Dawn spacecraft of asteroid Vesta. This crater doublet was likely formed by the simultaneous impact of two fragments of a split projectile.

  1. Distinct phases in the positive selection of CD8+ T cells distinguished by intrathymic migration and T-cell receptor signaling patterns

    PubMed Central

    Ross, Jenny O.; Melichar, Heather J.; Au-Yeung, Byron B.; Herzmark, Paul; Weiss, Arthur; Robey, Ellen A.

    2014-01-01

    Positive selection of CD8 T cells in the thymus is thought to be a multistep process lasting 3–4 d; however, the discrete steps involved are poorly understood. Here, we examine phenotypic changes, calcium signaling, and intrathymic migration in a synchronized cohort of MHC class I-specific thymocytes undergoing positive selection in situ. Transient elevations in intracellular calcium concentration ([Ca2+]i) and migratory pauses occurred throughout the first 24 h of positive selection, becoming progressively briefer and accompanied by a gradual shift in basal [Ca2+]i over time. Changes in chemokine-receptor expression and relocalization from the cortex to medulla occurred between 12 and 24 h after the initial encounter with positive-selecting ligands, a time frame at which the majority of thymocytes retain CD4 and CD8 expression and still require T-cell receptor (TCR) signaling to efficiently complete positive selection. Our results identify distinct phases in the positive selection of MHC class I-specific thymocytes that are distinguished by their TCR-signaling pattern and intrathymic location and provide a framework for understanding the multistep process of positive selection in the thymus. PMID:24927565

  2. Comparative efficacy of a recombinant feline interferon omega in refractory cases of calicivirus-positive cats with caudal stomatitis: a randomised, multi-centre, controlled, double-blind study in 39 cats.

    PubMed

    Hennet, Philippe R; Camy, Guy A L; McGahie, David M; Albouy, Maxime V

    2011-08-01

    Chronic caudal stomatitis with alveolar/buccal mucositis in calicivirus-positive cats is the most severe presentation of feline chronic gingivostomatitis. Refractory cases are helped by antibiotic and anti-inflammatory treatments often including glucocorticoids. In order to evaluate the comparative efficacy of oromucosal administration of recombinant feline interferon omega (rFeIFN-ω) versus oral administration of glucocorticoids, a randomised, multi-centre, controlled, double-blind study was performed in 39 cats. The progression of behavioural, clinical and lesional scores was assessed over 90 days. Daily oromucosal treatment with 0.1 MU of rFeIFN-ω was associated with a significant improvement of clinical lesions (caudal stomatitis and alveolar/buccal mucositis) and a decrease of pain scores from D0 to D90. Although no such statistical improvement was noticed in the prednisolone group, there was, however, no significant difference between the two groups for most of the parameters, except pain at D60 and D90.

  3. Treatment of older patients with HER2-positive metastatic breast cancer with pertuzumab, trastuzumab, and docetaxel: subgroup analyses from a randomized, double-blind, placebo-controlled phase III trial (CLEOPATRA).

    PubMed

    Miles, David; Baselga, José; Amadori, Dino; Sunpaweravong, Patrapim; Semiglazov, Vladimir; Knott, Adam; Clark, Emma; Ross, Graham; Swain, Sandra M

    2013-11-01

    Although the incidence of cancer increases with age, older patients are under-represented in cancer treatment trials, resulting in limited data availability in this patient population. Here we present results from pre-defined subgroup analyses conducted by age group (<65 vs ≥ 65 years) from a randomized, double-blind, placebo-controlled phase III trial in patients with HER2-positive metastatic breast cancer. Patients who had not received previous chemotherapy or biological therapy for HER2-positive locally recurrent, unresectable or metastatic breast cancer were randomly assigned to treatment with placebo, trastuzumab, and docetaxel or with pertuzumab, trastuzumab, and docetaxel. Primary endpoint was independently assessed progression-free survival. We performed pre-specified subgroup analyses of progression-free survival according to age. The study is registered with ClinicalTrials.gov, NCT00567190. 808 patients were enrolled. Of those, 127 patients were 65 years of age or older (placebo arm: 67, pertuzumab arm: 60). Patients in both age groups experienced progression-free survival benefit with treatment in the pertuzumab arm (<65 years: HR: 0.65; 95 % CI 0.53-0.80; ≥65 years: HR: 0.52; 95 % CI 0.31-0.86). Diarrhoea, fatigue, asthenia, decreased appetite, vomiting, and dysgeusia were reported more frequently in patients 65 years of age or older compared with younger patients. Neutropenia and febrile neutropenia were reported less frequently in the older age group. The efficacy and safety data reported in CLEOPATRA suggest that the combined use of pertuzumab, trastuzumab, and docetaxel should not be limited by patient age.

  4. Double Layers in Astrophysics

    NASA Technical Reports Server (NTRS)

    Williams, Alton C. (Editor); Moorehead, Tauna W. (Editor)

    1987-01-01

    Topics addressed include: laboratory double layers; ion-acoustic double layers; pumping potential wells; ion phase-space vortices; weak double layers; electric fields and double layers in plasmas; auroral double layers; double layer formation in a plasma; beamed emission from gamma-ray burst source; double layers and extragalactic jets; and electric potential between plasma sheet clouds.

  5. Double screening

    SciTech Connect

    Gratia, Pierre; Hu, Wayne; Joyce, Austin; Ribeiro, Raquel H.

    2016-06-15

    Attempts to modify gravity in the infrared typically require a screening mechanism to ensure consistency with local tests of gravity. These screening mechanisms fit into three broad classes; we investigate theories which are capable of exhibiting more than one type of screening. Specifically, we focus on a simple model which exhibits both Vainshtein and kinetic screening. We point out that due to the two characteristic length scales in the problem, the type of screening that dominates depends on the mass of the sourcing object, allowing for different phenomenology at different scales. We consider embedding this double screening phenomenology in a broader cosmological scenario and show that the simplest examples that exhibit double screening are radiatively stable.

  6. Mitogen-stimulated glucose transport in thymocytes. Possible role of Ca++ and antagonism by adenosine 3':5'-monophosphate

    PubMed Central

    1977-01-01

    The plant lectin, concanavalin A (Con-A), and the ionophore, A-23187 (specific for divalent cations), stimulated glucose transport in rat thymocytes. Con-A stimulation developed more slowly and was somewhat less extensive than that of stimulation developed more slowly and was somewhat less extensive than that of A-23187. Both responses showed saturation dose dependencies. The two responses were poorly additive, suggesting that A-23187 may saturate regulatory processes shared by the two stimulatory mechanisms. Doses of methylisobutylxanthine (MIX) and prostaglandin E2 which raised adenosine 3':5'-monophosphate (cAMP) levels in these cells also antagonized the Con-A stimulation of glucose transport but did not inhibit basal glucose transport or the A-23187 stimulation. Dibutyryl-cAMP and 8-bromo-cAMP also natagonized Con-A stimulation without inhibiting basal glucose transport. MIX antagonized high Con-A doses about as strongly as it did low Con-A doses, suggesting that MIX did not compete in the Con-A binding step or other process saturable by Con-A. [3H-A1Con-A binding was not affected by MIX. The stimulatory effects of Con-A and A-23187 were reduced by reduction of Ca++ in the medium. Both Con-A and A-23187 enhanced 45Ca++ influx and cellular Ca++ content. The A-23187 dose, which was saturating for glucose transport stimulation, enhanced Ca++ influx and cellular Ca++ content more than did the Con-A dose which was saturating for glucose transport stimulation. The dose fo MIX which specifically antagonized Con-A stimulation of glucose transport proved also to reduce Ca++ influx and cellular Ca++ in the presence of Con-A but not in the presence of A-23187. Thus, glucose transport correlates rather well with cellular Ca++. These results are compatible with the view that Ca++ in a cellular compartment can promote glucose transport, the Con-A's enhancement of Ca++ entry contributes to its stimulation of glucose transport, and the MIX antagonized Con-A action at least

  7. [Responses of thymocytes and splenocytes to low-intensity extremely high-frequency electromagnetic radiation in normal mice and in mice with systemic inflammation].

    PubMed

    Gapeev, A B; Sirota, N P; Kudriavtsev, A A; Chemeris, N K

    2010-01-01

    Changes in T cell subsets and expression of cytokine genes in thymocytes and splenocytes after exposure of BAL/c mice to low-intensity extremely high-frequency electromagnetic radiation (42.2 GHz, 0.1 mW/cm2, exposure duration 20 min) under normal conditions and in systemic inflammation were studied using flow cytometry and the methods of reverse transcription and real-time polymerase chain reaction. It was found that the number of CD4+ and CD8+ T cells statistically significantly increased in the thymus and considerably decreased in the spleen of exposed animals. Apparently, the exposure of animals leads to an intensification of the host defense, by activating the T-cellular immunity. As for effector functions, the increased expression of IL-1beta and IFNgamma genes in thymocytes and essentially enhanced expression of IL-1beta, IL-10, and TNFalpha genes in splenocytes were observed in mice exposed against the background of a progressive inflammatory process. The experimental data obtained specify that the directed (anti-inflammatory) response of an organism to a specific combination of effective exposure parameters of electromagnetic radiation can be realized by the activation of particular immunocompetent cells and changes in the cytokine profile.

  8. Non-canonical antagonism of PI3K by the kinase Itpkb delays thymocyte β-selection and renders it Notch-dependent

    PubMed Central

    Westernberg, Luise; Conche, Claire; Huang, Yina Hsing; Rigaud, Stephanie; Deng, Yisong; Siegemund, Sabine; Mukherjee, Sayak; Nosaka, Lyn'Al; Das, Jayajit; Sauer, Karsten

    2016-01-01

    β-selection is the most pivotal event determining αβ T cell fate. Here, surface-expression of a pre-T cell receptor (pre-TCR) induces thymocyte metabolic activation, proliferation, survival and differentiation. Besides the pre-TCR, β-selection also requires co-stimulatory signals from Notch receptors - key cell fate determinants in eukaryotes. Here, we show that this Notch-dependence is established through antagonistic signaling by the pre-TCR/Notch effector, phosphoinositide 3-kinase (PI3K), and by inositol-trisphosphate 3-kinase B (Itpkb). Canonically, PI3K is counteracted by the lipid-phosphatases Pten and Inpp5d/SHIP-1. In contrast, Itpkb dampens pre-TCR induced PI3K/Akt signaling by producing IP4, a soluble antagonist of the Akt-activating PI3K-product PIP3. Itpkb-/- thymocytes are pre-TCR hyperresponsive, hyperactivate Akt, downstream mTOR and metabolism, undergo an accelerated β-selection and can develop to CD4+CD8+ cells without Notch. This is reversed by inhibition of Akt, mTOR or glucose metabolism. Thus, non-canonical PI3K-antagonism by Itpkb restricts pre-TCR induced metabolic activation to enforce coincidence-detection of pre-TCR expression and Notch-engagement. DOI: http://dx.doi.org/10.7554/eLife.10786.001 PMID:26880557

  9. An endogenous calcium-dependent, caspase-independent intranuclear degradation pathway in thymocyte nuclei: Antagonism by physiological concentrations of K{sup +} ions

    SciTech Connect

    Ajiro, Kozo; Bortner, Carl D.; Westmoreland, Jim; Cidlowski, John A.

    2008-04-01

    Calcium ions have been implicated in apoptosis for many years, however the precise role of this ion in the cell death process remains incomplete. We have extensively examined the role of Ca{sup 2+} on nuclear degradation in vitro using highly purified nuclei isolated from non-apoptotic rat thymocytes. We show that these nuclei are devoid of CAD (caspase-activated DNase), and DNA degradation occurs independent of caspase activity. Serine proteases rather than caspase-3 appear necessary for this Ca{sup 2+}-dependent DNA degradation in nuclei. We analyzed nuclei treated with various concentrations of Ca{sup 2+} in the presence of both a physiological (140 mM) and apoptotic (40 mM) concentration of KCl. Our results show that a 5-fold increase in Ca{sup 2+} is required to induce DNA degradation at the physiological KCl concentration compared to the lower, apoptotic concentration of the cation. Ca{sup 2+}-induced internucleosomal DNA degradation was also accompanied by the release of histones, however the apoptotic-specific phosphorylation of histone H2B does not occur in these isolated nuclei. Interestingly, physiological concentrations of K{sup +} inhibit both Ca{sup 2+}-dependent DNA degradation and histone release suggesting that a reduction of intracellular K{sup +} is necessary for this apoptosis-associated nuclear degradation in cells. Together, these data define an inherent caspase-independent catabolic pathway in thymocyte nuclei that is sensitive to physiological concentrations of intracellular cations.

  10. Effect of Dexmedetomidine on Heart Rate-Corrected QT and Tpeak-Tend Intervals During Robot-Assisted Laparoscopic Prostatectomy With Steep Trendelenburg Position: A Prospective, Randomized, Double-Blinded, Controlled Study.

    PubMed

    Kim, Na Young; Han, Dong Woo; Koh, Jae Chul; Rha, Koon Ho; Hong, Jung Hwa; Park, Jong Min; Kim, So Yeon

    2016-05-01

    Intraperitoneal insufflation of carbon dioxide may affect the sympathetic activity that leads to changes in ventricular repolarization. This in turn can result in changes of heart rate-corrected QT (QTc) interval and Tpeak-Tend (Tp-e) interval. Dexmedetomidine is a highly selective α2-receptor agonist and has potential antiarrhythmic properties. This prospective, randomized, double-blinded, controlled study evaluated the effects of dexmedetomidine administration on QTc and Tp-e intervals during robot-assisted laparoscopic prostatectomy with steep Trendelenburg position.Fifty patients scheduled for robot-assisted laparoscopic prostatectomy randomly received either a continuous infusion of dexmedetomidine at a rate of 0.3 μg/kg/hour, from anesthetic induction until the end of the Trendelenburg position (dexmedetomidine group; n = 25), or the same volume of normal saline (control group; n = 25). Anesthesia was maintained with sevoflurane and remifentanil. The primary and secondary goals were to evaluate the effect of dexmedetomidine on the QTc and Tp-e interval changes. Mean arterial pressure, heart rate, end-tidal CO2, and end-tidal sevoflurane concentrations were assessed as well.Forty-seven patients (94%) completed the study. Dexmedetomidine significantly attenuated QTc interval prolongation and reduced the Tp-e interval, even though the baseline values of the QTc and Tp-e intervals were similar between the 2 groups (PGroup × Time = 0.001 and 0.014, respectively). Twenty-two patients (96%) in the control group and 13 (54%) in the dexmedetomidine group had QTc interval prolongation of >20 ms from the baseline value during surgery (P = 0.001). The maximum QTc interval prolongation from the baseline value during surgery was 46 ± 21 ms in the control group and 24 ± 21 ms in the dexmedetomidine group (mean ± SD, P = 0.001). Mean arterial pressure and heart rate were comparable between the groups.Continuous infusion of

  11. The efficacy and safety of sodium hyaluronate injection (Adant®) in treating degenerative osteoarthritis: a multi-center, randomized, double-blind, positive-drug parallel-controlled and non-inferiority clinical study.

    PubMed

    Xin, Yang; Jianhao, Lin; Tiansheng, Sun; Yongqiang, Hao; Weimin, Fan; Ming, Chen; Tiezheng, Sun; Jianhua, Yao; Liang, Xuan; Xiaoyuan, Gu; Yongping, Cao

    2016-03-01

    To compare the efficacy and safety of two different sodium hyaluronate drugs in treating degenerative osteoarthritis (OA) of the knee. This randomized, multi-center, double-blind, positive-drug, parallel-controlled study included 229 patients aged ≥ 45 years who were clinically diagnosed with degenerative OA of the knee. The patients were randomly assigned to receive for 5 consecutive weeks a once-weekly intra-articular injection of the investigational drug Adant®, which is manufactured by fermentation, or the control drug Artz®, which is manufactured by extraction of cockscomb. The follow-up examinations were conducted 1, 2, 3, 4 and 6 weeks after the first injection. The primary efficacy parameter was the decrease in the visual analog scale (VAS) scores of pain on movement caused by load-bearing, and the secondary efficacy parameter was the decrease in the Lequesne index. The intra-articular injections of Adant® and Artz® produced a significant reduction in the VAS scores for pain on movement (50.4 and 50.3 mm, respectively) and in the Lequesne index. There were no significant differences in efficacy and safety between the two drugs and non-inferiority in VAS score decreases was confirmed. The results of this study show that both Adant® and Artz® are effective for the treatment of OA and that there were no statistical differences between them in the VAS scores of pain on movement, Lequesne index or safety during the observation period with short-time follow up. © 2015 Asia Pacific League of Associations for Rheumatology and Wiley Publishing Asia Pty Ltd.

  12. Age-dependent variation in the proportion and number of intestinal lymphocyte subsets, especially natural killer T cells, double-positive CD4+ CD8+ cells and B220+ T cells, in mice

    PubMed Central

    Ishimoto, Yuiko; Tomiyama-Miyaji, Chikako; Watanabe, Hisami; Yokoyama, Hisashi; Ebe, Kazuto; Tsubata, Shunsuke; Aoyagi, Yutaka; Abo, TORU

    2004-01-01

    The age-dependent variation in the proportion and number of lymphocyte subsets was examined at various extrathymic sites, including the liver, small intestine, colon and appendix in mice. In comparison with young mice (4 weeks of age), the number of total lymphocytes yielded by all tested organs was greater in adult (9 weeks) and old (40 weeks) mice. The major lymphocyte subset that expanded with age was interleukin-2 receptor (IL-2R) β+ CD3int cells (50% of them expressed NK1.1) in the liver, whereas it was CD3+ IL-2Rβ− NK1.1− cells at all intraepithelial sites in the intestine. Although NK1.1+ CD3+ cells were present at intraepithelial sites in the intestine, the proportion of this subset was rather low. The ratio of CD4 to CD8 tended to decrease among natural killer T (NKT) cells and T cells at all intraepithelial sites in the intestine with age. A unique population of double-positive CD4+ CD8+ cells in the small intestine increased in old mice. B220+ T cells were found mainly in the appendix and colon, and the proportion of these T cells decreased in old mice. Conventional NKT cells were very few in Jα281−/− and CD1d−/− mice in the liver, while NKT cells which existed in the appendix remained unchanged even in these mice. This was because unconventional CD8+ NKT cells were present in the intestine. The present results suggest that despite the fact that both the liver and intraepithelial sites in the intestine carry many extrathymic T cells, the distribution of lymphocyte subsets and their age-associated variation are site-specific. PMID:15500624

  13. Recovery from chemically induced thymus atrophy starts with CD4- CD8- CD2high TcR alpha beta-/low thymocytes and results in an increased formation of CD4- CD8- TcR alpha beta high thymocytes.

    PubMed Central

    Pieters, R H; Bol, M; Lam, B W; Seinen, W; Bloksma, N; Penninks, A H

    1993-01-01

    Regeneration of the thymus was studied in rats that were treated with a single oral dose of the organotin compound di-n-butyltin dichloride (DBTC). After an initial maximum depletion of cortical BrdU+ thymocytes on day 2 after treatment, repopulation appeared to start on day 3 as indicated by an increased number of BrdU+ cells in the subcapsular region. On day 5, when thymocyte depletion was most pronounced, a relative increase of BrdU+ cells was observed all over the cortex. In comparison with controls, the thymoblast population on day 5 appeared to harbour increased numbers of CD4- CD8- and immature CD4- CD8+ CD53- thymoblasts, while the number of CD4+ CD8+ blasts had decreased. In comparison with day 3, however, the number of CD4+ CD8+ blasts had increased again. Results together have been interpreted as indicative for thymus regeneration starting from CD4- CD8- blasts which differentiate to immature CD4- CD8+ and then to CD4+ CD8+ blasts. Further characterization revealed that the majority of the CD4- CD8- and CD4- CD8+ CD53- blasts expressed high levels of CD2 and no or low levels of T-cell receptor (TcR) alpha beta. The high expression of CD2 on repopulating thymoblasts may be an additional indication of their activated state and for a role of interaction with the ligand LFA-3 on thymic epithelial cells during this phase of thymocyte differentiation. The number of CD4- CD8- TcR alpha beta high cells was increased on day 5 after dosing. The origin of this population and the possible implication of its development during thymus regeneration after chemically induced thymus atrophy are discussed. Images Figure 1 Figure 4 PMID:8098700

  14. Neratinib after trastuzumab-based adjuvant therapy in patients with HER2-positive breast cancer (ExteNET): a multicentre, randomised, double-blind, placebo-controlled, phase 3 trial.

    PubMed

    Chan, Arlene; Delaloge, Suzette; Holmes, Frankie A; Moy, Beverly; Iwata, Hiroji; Harvey, Vernon J; Robert, Nicholas J; Silovski, Tajana; Gokmen, Erhan; von Minckwitz, Gunter; Ejlertsen, Bent; Chia, Stephen K L; Mansi, Janine; Barrios, Carlos H; Gnant, Michael; Buyse, Marc; Gore, Ira; Smith, John; Harker, Graydon; Masuda, Norikazu; Petrakova, Katarina; Zotano, Angel Guerrero; Iannotti, Nicholas; Rodriguez, Gladys; Tassone, Pierfrancesco; Wong, Alvin; Bryce, Richard; Ye, Yining; Yao, Bin; Martin, Miguel

    2016-03-01

    Neratinib, an irreversible tyrosine-kinase inhibitor of HER1, HER2, and HER4, has clinical activity in patients with HER2-positive metastatic breast cancer. We aimed to investigate the efficacy and safety of 12 months of neratinib after trastuzumab-based adjuvant therapy in patients with early-stage HER2-positive breast cancer. We did this multicentre, randomised, double-blind, placebo-controlled, phase 3 trial at 495 centres in Europe, Asia, Australia, New Zealand, and North and South America. Eligible women (aged ≥18 years, or ≥20 years in Japan) had stage 1-3 HER2-positive breast cancer and had completed neoadjuvant and adjuvant trastuzumab therapy up to 2 years before randomisation. Inclusion criteria were amended on Feb 25, 2010, to include patients with stage 2-3 HER2-positive breast cancer who had completed trastuzumab therapy up to 1 year previously. Patients were randomly assigned (1:1) to receive oral neratinib 240 mg per day or matching placebo. The randomisation sequence was generated with permuted blocks stratified by hormone receptor status (hormone receptor-positive [oestrogen or progesterone receptor-positive or both] vs hormone receptor-negative [oestrogen and progesterone receptor-negative]), nodal status (0, 1-3, or ≥4), and trastuzumab adjuvant regimen (sequentially vs concurrently with chemotherapy), then implemented centrally via an interactive voice and web-response system. Patients, investigators, and trial sponsors were masked to treatment allocation. The primary outcome was invasive disease-free survival, as defined in the original protocol, at 2 years after randomisation. Analysis was by intention to treat. This trial is registered with ClinicalTrials.gov, number NCT00878709. Between July 9, 2009, and Oct 24, 2011, we randomly assigned 2840 women to receive neratinib (n=1420) or placebo (n=1420). Median follow-up time was 24 months (IQR 20-25) in the neratinib group and 24 months (22-25) in the placebo group. At 2 year follow-up, 70

  15. Sensitive and high resolution in situ hybridization to human chromosomes using biotin labelled probes: assignment of the human thymocyte CD1 antigen genes to chromosome 1.

    PubMed Central

    Albertson, D G; Fishpool, R; Sherrington, P; Nacheva, E; Milstein, C

    1988-01-01

    A method for in situ hybridization originally developed for mapping genes in the nematode, Caenorhabditis elegans has been adapted for high resolution cytological mapping of genes in the human. The probe DNAs are labelled by incorporation of biotin dUTP and the site of hybridization detected by immunofluorescence. For the accurate assignment of the hybridization signal to chromosome bands, visualized by staining with Hoechst 33258, a heterologous ribosomal DNA probe is also included in the hybridization reaction. These rDNA signals are used as fiducial markers when aligning the two fluorescent images. We demonstrate the method by assignment of the human thymocyte CD1 antigen genes to human chromosome 1q22-23. Images PMID:3053166

  16. Triclocarban-induced change in intracellular Ca²⁺ level in rat thymocytes: cytometric analysis with Fluo-3 under Zn²⁺-free conditions.

    PubMed

    Miura, Yukari; Chen, Xiaohui; Yamada, Saki; Sugihara, Aya; Enkhjargal, Molomjamts; Sun, Yuanzhi; Kuroda, Keiko; Satoh, Masaya; Oyama, Yasuo

    2014-03-01

    Triclocarban (TCC) is an antimicrobial used in personal hygiene products. Recent health concerns arose after TCC was detected in the blood of human subjects who showered with soap containing TCC. In this study, the effect of TCC on intracellular Ca(2+) concentration in rat thymocytes was examined using Fluo-3, an indicator of intracellular Ca(2+). TCC at concentrations ranging from 0.1 μM to 3 μM increased intracellular Ca(2+) concentration biphasically: first by releasing Ca(2+) from intracellular Ca(2+) stores and then inducing Ca(2+) influx through store-operated Ca(2+) channels. The threshold TCC concentration to increase intracellular Ca(2+) concentration in this study was lower than the maximum TCC concentrations reported in human blood samples. Therefore, we anticipate that TCC at concentrations reported in human blood samples might disturb intracellular Ca(2+) signaling in human lymphocytes. Copyright © 2014 Elsevier B.V. All rights reserved.

  17. Exposure of tumor-bearing mice to extremely high-frequency electromagnetic radiation modifies the composition of fatty acids in thymocytes and tumor tissue.

    PubMed

    Gapeyev, Andrew B; Kulagina, Tatiana P; Aripovsky, Alexander V

    2013-08-01

    To test the participation of fatty acids (FA) in antitumor effects of extremely high-frequency electromagnetic radiation (EHF EMR), the changes in the FA composition in the thymus, liver, blood plasma, muscle tissue, and tumor tissue in mice with Ehrlich solid carcinoma exposed to EHF EMR were studied. Normal and tumor-bearing mice were exposed to EHF EMR with effective parameters (42.2 GHz, 0.1 mW/cm2, 20 min daily during five consecutive days beginning the first day after the inoculation of tumor cells). Fatty acid composition of various organs and tissues of mice were determined using a gas chromatography. It was shown that the exposure of normal mice to EHF EMR or tumor growth significantly increased the content of monounsaturated FA (MUFA) and decreased the content of polyunsaturated FA (PUFA) in all tissues examined. Exposure of tumor-bearing mice to EHF EMR led to the recovery of FA composition in thymocytes to the state that is typical for normal animals. In other tissues of tumor-bearing mice, the exposure to EHF EMR did not induce considerable changes that would be significantly distinguished between disturbances caused by EHF EMR exposure or tumor growth separately. In tumor tissue which is characterized by elevated level of MUFA, the exposure to EHF EMR significantly decreased the summary content of MUFA and increased the summary content of PUFA. The recovery of the FA composition in thymocytes and the modification of the FA composition in the tumor under the influence of EHF EMR on tumor-bearing animals may have crucial importance for elucidating the mechanisms of antitumor effects of the electromagnetic radiation.

  18. Efficient adenovirus-mediated gene transfer into primary T cells and thymocytes in a new coxsackie/adenovirus receptor transgenic model

    PubMed Central

    Hurez, Vincent; Dzialo-Hatton, Robin; Oliver, James; Matthews, R James; Weaver, Casey T

    2002-01-01

    Background Gene transfer studies in primary T cells have suffered from the limitations of conventional viral transduction or transfection techniques. Replication-defective adenoviral vectors are an attractive alternative for gene delivery. However, naive lymphocytes are not readily susceptible to infection with adenoviruses due to insufficient expression of the coxsackie/adenovirus receptor. Results To render T cells susceptible to adenoviral gene transfer, we have developed three new murine transgenic lines in which expression of the human coxsackie/adenovirus receptor (hCAR) with a truncated cytoplasmic domain (hCARΔcyt) is limited to thymocytes and lymphocytes under direction of a human CD2 mini-gene. hCARΔcyt.CD2 transgenic mice were crossed with DO11.10 T cell receptor transgenic mice (DO11.hCARΔcyt) to allow developmental studies in a defined, clonal T cell population. Expression of hCARΔcyt enabled adenoviral transduction of resting primary CD4+ T cells, differentiated effector T cells and thymocytes from DO11.hCARΔcyt with high efficiency. Expression of hCARΔcyt transgene did not perturb T cell development in these mice and adenoviral transduction of DO11.hCARΔcyt T cells did not alter their activation status, functional responses or differentiative potential. Adoptive transfer of the transduced T cells into normal recipients did not modify their physiologic localization. Conclusion The DO11.hCARΔcyt transgenic model thus allows efficient gene transfer in primary T cell populations and will be valuable for novel studies of T cell activation and differentiation. PMID:12019030

  19. A Handbook of Double Stars

    NASA Astrophysics Data System (ADS)

    Crossley, Edward; Gledhill, Joseph; Wilson, James M.

    2011-11-01

    Preface; Part I. Historical, and Descriptive of Instruments and Methods: 1. Historical introduction; 2. The Equatorial: its construction and adjustments; 3. Some account of the Equatorials which have been used by double-star observers; 4. The micrometer; 5. Methods of observing double stars; Part II. On the Calculation of the Orbit of a Binary Star: 1. Introduction; 2. Example of an orbit worked by a graphical method; 3. Dr. Doberck's example of an orbit worked by analytical methods; 4. On relative rectilinear motion; 5. On the effect of proper motion and parallax on the observed position angles and distance of an optically double star; 6. On the errors of observation and the combination of observations; Part III. The Catalogue and Measures: Introductory remarks; A catalogue of binary and other double starts deserving of attention; Lists of measures, with historical notes, etc.; Supplementary list of measures; Appendix; Additional notes to measures; Binary stars classified; Note on systematic errors in the measures of angle and distance of double stars; Part IV. Bibliography: A. Some of the most important works and papers on double stars; B. Some papers on the micrometer; C. Some papers on the colours of double stars; Additional notes; Corrections 1880.

  20. Double inflation

    SciTech Connect

    Silk, J.; Turner, M.S.

    1986-04-01

    The Zel'dovich spectrum of adiabatic density perturbations is a generic prediction of inflation. There is increasing evidence that when the spectrum is normalized by observational data on small scales, there is not enough power on large scales to account for the observed large-scale structure in the Universe. Decoupling the spectrum on large and small scales could solve this problem. As a means of decoupling the large and small scales we propose double inflation (i.e., two episodes of inflation). In this scenario the spectrum on large scales is determined by the first episode of inflation and those on small scales by a second episode of inflation. We present three models for such a scenario. By nearly saturating the large angular-scale cosmic microwave anisotropy bound, we can easily account for the observed large-scale structure. We take the perturbations on small scales to be very large, deltarho/rho approx. = 0.1 to 0.01, which results in the production of primordial black holes (PBHs), early formation of structure, reionization of the Universe, and a rich array of astrophysical events. The ..cap omega..-problem is also addressed by our scenario. Allowing the density perturbations produced by the second episode of inflation to be large also lessens the fine-tuning required in the scalar potential and makes reheating much easier. We briefly speculate on the possibility that the second episode of inflation proceeds through the nucleation of bubbles, which today manifest themselves as empty bubbles whose surfaces are covered with galaxies. 37 refs., 1 fig.

  1. Seeing Double

    NASA Astrophysics Data System (ADS)

    Pesic, Peter

    2003-10-01

    The separateness and connection of individuals is perhaps the central question of human life: What, exactly, is my individuality? To what degree is it unique? To what degree can it be shared, and how? To the many philosophical and literary speculations about these topics over time, modern science has added the curious twist of quantum theory, which requires that the elementary particles of which everything consists have no individuality at all. All aspects of chemistry depend on this lack of individuality, as do many branches of physics. From where, then, does our individuality come? In Seeing Double, Peter Pesic invites readers to explore this intriguing set of questions. He draws on literary and historical examples that open the mind (from Homer to Martin Guerre to Kafka), philosophical analyses that have helped to make our thinking and speech more precise, and scientific work that has enabled us to characterize the phenomena of nature. Though he does not try to be all-inclusive, Pesic presents a broad range of ideas, building toward a specific point of view: that the crux of modern quantum theory is its clash with our ordinary concept of individuality. This represents a departure from the usual understanding of quantum theory. Pesic argues that what is bizarre about quantum theory becomes more intelligible as we reconsider what we mean by individuality and identity in ordinary experience. In turn, quantum identity opens a new perspective on us. Peter Pesic is a Tutor and Musician-in-Residence at St. John's College, Santa Fe, New Mexico. He has a Ph.D. in physics from Stanford University.

  2. Restrictions Limiting the Generation of DNA Double Strand Breaks during Chromosomal V(D)J Recombination

    PubMed Central

    Tillman, Robert E.; Wooley, Andrea L.; Hughes, Maureen M.; Wehrly, Tara D.; Swat, Wojciech; Sleckman, Barry P.

    2002-01-01

    Antigen receptor loci are composed of numerous variable (V), diversity (D), and joining (J) gene segments, each flanked by recombination signal sequences (RSSs). The V(D)J recombination reaction proceeds through RSS recognition and DNA cleavage steps making it possible for multiple DNA double strand breaks (DSBs) to be introduced at a single locus. Here we use ligation-mediated PCR to analyze DNA cleavage intermediates in thymocytes from mice with targeted RSS mutations at the endogenous TCRβ locus. We show that DNA cleavage does not occur at individual RSSs but rather must be coordinated between RSS pairs flanking gene segments that ultimately form coding joins. Coordination of the DNA cleavage step occurs over great distances in the chromosome and favors intra- over interchromosomal recombination. Furthermore, through several restrictions imposed on the generation of both nonpaired and paired DNA DSBs, this requirement promotes antigen receptor gene integrity and genomic stability in developing lymphocytes undergoing V(D)J recombination. PMID:11828005

  3. Transcriptomic analysis reveals sex-specific differences in the expression of Dcl1 and Fis1 genes in the radio-adaptive response of thymocytes to TRP53-mediated apoptosis.

    PubMed

    López-Nieva, Pilar; Malavé, Manuel; González-Sánchez, Laura; Fernández-Piqueras, José; Fernández-Navarro, Pablo; Santos, Javier

    2016-08-31

    Radio-Adaptive Response (RAR) is a biological defense mechanism whereby exposure to low dose ionizing radiation (IR) mitigates the detrimental effects of high dose irradiation. RAR has been widely observed in vivo using as endpoint less induction of apoptosis. However, sex differences associated with RAR and variations between males and females on global gene expression influenced by RAR have not been still investigated. In addition, the response to radiation-induced apoptosis is associated with phosphorylation of TRP53 at both the serine 15 (ser-18 in the mouse) and serine 392 (ser-389 in mice) residues, but the role of these two phosphorylated forms in male and female RAR remains to be elucidated. We analyzed the effect of administering priming low dose radiation (0.075 Gy of X-rays) prior to high dose radiation (1.75 Gy of γ-rays) on the level of caspase-3-mediated apoptosis and on global transcriptional expression in thymocytes of male and female mice. Here, we provide the first evidence of a differential sex effect of RAR on the reduction of thymocyte apoptosis with males showing lesser levels of caspase-3-mediated apoptosis than females. Analysis of transcriptomic profiles of 1944 genes involved in apoptosis signaling in radio-adapted thymocytes identified 17 transcripts exhibiting differential expression between both sexes. Among them, Dlc1 and Fis1 are closely related to the apoptosis mediated by the TRP53 protein. Our data demonstrate that overexpression of Dlc1 and Fis1 occur concomitantly with a highest accumulation of phosphoserine-18-TRP53 and caspase-3 in radio-adapted thymocytes of female mice. In an opposite way, both down-modulation of Fis1 and phosphoserine-389-TRP53 accumulation appear to be associated with protection from thymocyte apoptosis mediated by caspase-3 in males. Transcriptomic analysis performed in this work reveals for the first time sex-specific differences in gene expression influenced by RAR. Our results also suggest a sex

  4. Double Your Major, Double Your Return?

    ERIC Educational Resources Information Center

    Del Rossi, Alison F.; Hersch, Joni

    2008-01-01

    We use the 2003 National Survey of College Graduates to provide the first estimates of the effect on earnings of having a double major. Overall, double majoring increases earnings by 2.3% relative to having a single major among college graduates without graduate degrees. Most of the gains from having a double major come from choosing fields across…

  5. Buparlisib plus fulvestrant versus placebo plus fulvestrant in postmenopausal, hormone receptor-positive, HER2-negative, advanced breast cancer (BELLE-2): a randomised, double-blind, placebo-controlled, phase 3 trial.

    PubMed

    Baselga, José; Im, Seock-Ah; Iwata, Hiroji; Cortés, Javier; De Laurentiis, Michele; Jiang, Zefei; Arteaga, Carlos L; Jonat, Walter; Clemons, Mark; Ito, Yoshinori; Awada, Ahmad; Chia, Stephen; Jagiełło-Gruszfeld, Agnieszka; Pistilli, Barbara; Tseng, Ling-Ming; Hurvitz, Sara; Masuda, Norikazu; Takahashi, Masato; Vuylsteke, Peter; Hachemi, Soulef; Dharan, Bharani; Di Tomaso, Emmanuelle; Urban, Patrick; Massacesi, Cristian; Campone, Mario

    2017-07-01

    Phosphatidylinositol 3-kinase (PI3K) pathway activation is a hallmark of endocrine therapy-resistant, hormone receptor-positive breast cancer. This phase 3 study assessed the efficacy of the pan-PI3K inhibitor buparlisib plus fulvestrant in patients with advanced breast cancer, including an evaluation of the PI3K pathway activation status as a biomarker for clinical benefit. The BELLE-2 trial was a randomised, double-blind, placebo-controlled, multicentre study. Postmenopausal women aged 18 years or older with histologically confirmed, hormone receptor-positive and human epidermal growth factor (HER2)-negative inoperable locally advanced or metastatic breast cancer whose disease had progressed on or after aromatase inhibitor treatment and had received up to one previous line of chemotherapy for advanced disease were included. Eligible patients were randomly assigned (1:1) using interactive voice response technology (block size of 6) on day 15 of cycle 1 to receive oral buparlisib (100 mg/day) or matching placebo, starting on day 15 of cycle 1, plus intramuscular fulvestrant (500 mg) on days 1 and 15 of cycle 1, and on day 1 of subsequent 28-day cycles. Patients were assigned randomisation numbers with a validated interactive response technology; these numbers were linked to different treatment groups which in turn were linked to treatment numbers. PI3K status in tumour tissue was determined via central laboratory during a 14-day run-in phase. Randomisation was stratified by PI3K pathway activation status (activated vs non-activated vs and unknown) and visceral disease status (present vs absent). Patients, investigators, local radiologists, study team, and anyone involved in the study were masked to the identity of the treatment until unblinding. The primary endpoints were progression-free survival by local investigator assessment per Response Evaluation Criteria In Solid Tumors (version 1.1) in the total population, in patients with known (activated or non

  6. Tenofovir alafenamide versus tenofovir disoproxil fumarate for the treatment of HBeAg-positive chronic hepatitis B virus infection: a randomised, double-blind, phase 3, non-inferiority trial.

    PubMed

    Chan, Henry L Y; Fung, Scott; Seto, Wai Kay; Chuang, Wan-Long; Chen, Chi-Yi; Kim, Hyung Joon; Hui, Aric Josun; Janssen, Harry L A; Chowdhury, Abhijit; Tsang, Tak Yin Owen; Mehta, Rajiv; Gane, Edward; Flaherty, John F; Massetto, Benedetta; Gaggar, Anuj; Kitrinos, Kathryn M; Lin, Lanjia; Subramanian, G Mani; McHutchison, John G; Lim, Young-Suk; Acharya, Subrat K; Agarwal, Kosh

    2016-11-01

    Tenofovir alafenamide is a novel prodrug formulated to deliver the active metabolite to target cells more efficiently than tenofovir disoproxil fumarate at a lower dose, thereby reducing systemic exposure. In patients with HIV, tenofovir alafenamide was as efficacious as tenofovir disoproxil fumarate, with reduced bone and renal toxic effects. We compared the efficacy and safety of the two drugs in patients with HBeAg-positive chronic hepatitis B virus (HBV) infection in a non-inferiority study. We did this ongoing double-blind, non-inferiority study in 161 outpatient centres in 19 countries. Patients with chronic HBV infection who were positive for the hepatitis B e antigen (HBeAg) were randomly assigned (2:1) to receive either 25 mg tenofovir alafenamide or 300 mg tenofovir disoproxil fumarate with matching placebo. Randomisation was done by a computer-generated allocation sequence (block size six) stratified by plasma HBV DNA concentration and previous treatment experience. The primary efficacy endpoint was the proportion of patients with HBV DNA less than 29 IU/mL at week 48 in all patients who were randomly assigned and received at least one dose of study drug using a missing-equals-failed approach. The pre-specified non-inferiority margin was 10%. Key prespecified safety endpoints were bone and renal parameters at week 48. This study is registered with ClinicalTrials.gov, number NCT01940471. Of the 1473 patients screened from Sept 11, 2013, to Dec 20, 2014, 875 eligible patients were randomly assigned and 873 received treatment (581 with tenofovir alafenamide and 292 with tenofovir disoproxil fumarate). 371 (64%) patients receiving tenofovir alafenamide had HBV DNA less than 29 IU/mL at week 48, which was non-inferior to the 195 (67%) of patients receiving tenofovir disoproxil fumarate who had HBV DNA less than 29 IU/mL (adjusted difference -3·6% [95% CI -9·8 to 2·6]; p=0·25). Patients given tenofovir alafenamide had a significantly smaller decrease in

  7. Anti-thymocyte globulin as graft-versus-host disease prevention in the setting of allogeneic peripheral blood stem cell transplantation: a review from the Acute Leukemia Working Party of the European Society for Blood and Marrow Transplantation.

    PubMed

    Baron, Frédéric; Mohty, Mohamad; Blaise, Didier; Socié, Gérard; Labopin, Myriam; Esteve, Jordi; Ciceri, Fabio; Giebel, Sebastian; Gorin, Norbert Claude; Savani, Bipin N; Schmid, Christoph; Nagler, Arnon

    2017-02-01

    Allogeneic hematopoietic stem cell transplantation is increasingly used as treatment for patients with life-threatening blood diseases. Its curative potential is largely based on immune-mediated graft-versus-leukemia effects caused by donor T cells contained in the graft. Unfortunately, donor T cells are also the cause of graft-versus-host disease. The vast majority of human leukocyte antigen-matched allogeneic hematopoietic stem cell transplants are nowadays carried out with peripheral blood stem cells as the stem cell source. In comparison with bone marrows, peripheral blood stem cells contain more hematopoietic stem/progenitor cells but also one log more T cells. Consequently, the use of peripheral blood stem cells instead of bone marrow has been associated with faster hematologic recovery and a lower risk of relapse in patients with advanced disease, but also with a higher incidence of chronic graft-versus-host disease. These observations have been the basis for several studies aimed at assessing the impact of immunoregulation with anti-thymocyte globulin on transplantation outcomes in patients given human leukocyte antigen-matched peripheral blood stem cells from related or unrelated donors. After a brief introduction on anti-thymocyte globulin, this article reviews recent studies assessing the impact of anti-thymocyte globulin on transplantation outcomes in patients given peripheral blood stem cells from human leukocyte antigen-matched related or unrelated donors as well as in recipients of grafts from human leukocyte antigen haploidentical donors. Copyright© Ferrata Storti Foundation.

  8. Anti-thymocyte globulin as graft-versus-host disease prevention in the setting of allogeneic peripheral blood stem cell transplantation: a review from the Acute Leukemia Working Party of the European Society for Blood and Marrow Transplantation

    PubMed Central

    Baron, Frédéric; Mohty, Mohamad; Blaise, Didier; Socié, Gérard; Labopin, Myriam; Esteve, Jordi; Ciceri, Fabio; Giebel, Sebastian; Gorin, Norbert Claude; Savani, Bipin N; Schmid, Christoph; Nagler, Arnon

    2017-01-01

    Allogeneic hematopoietic stem cell transplantation is increasingly used as treatment for patients with life-threatening blood diseases. Its curative potential is largely based on immune-mediated graft-versus-leukemia effects caused by donor T cells contained in the graft. Unfortunately, donor T cells are also the cause of graft-versus-host disease. The vast majority of human leukocyte antigen-matched allogeneic hematopoietic stem cell transplants are nowadays carried out with peripheral blood stem cells as the stem cell source. In comparison with bone marrows, peripheral blood stem cells contain more hematopoietic stem/progenitor cells but also one log more T cells. Consequently, the use of peripheral blood stem cells instead of bone marrow has been associated with faster hematologic recovery and a lower risk of relapse in patients with advanced disease, but also with a higher incidence of chronic graft-versus-host disease. These observations have been the basis for several studies aimed at assessing the impact of immunoregulation with anti-thymocyte globulin on transplantation outcomes in patients given human leukocyte antigen-matched peripheral blood stem cells from related or unrelated donors. After a brief introduction on anti-thymocyte globulin, this article reviews recent studies assessing the impact of anti-thymocyte globulin on transplantation outcomes in patients given peripheral blood stem cells from human leukocyte antigen-matched related or unrelated donors as well as in recipients of grafts from human leukocyte antigen haploidentical donors. PMID:27927772

  9. Comparison of Different Rabbit Anti-Thymocyte Globulin Formulations in Allogeneic Stem Cell Transplantation: Systematic Literature Review and Network Meta-Analysis.

    PubMed

    Gagelmann, Nico; Ayuk, Francis; Wolschke, Christine; Kröger, Nicolaus

    2017-08-29

    Since 2000, phase III randomized controlled trials (RCT) investigated the efficacy of rabbit anti-thymocyte globulins (ATG) in patients following allogeneic stem cell transplantation (allo-SCT). However, comparisons of different ATG formulations are lacking. Our aim was to synthesize all efficacy evidence, enabling a comparison of all available formulations of rabbit ATG in the allo-SCT setting. We performed a systematic literature review to identify all available phase III RCT evidence. We searched the Cochrane Library, MEDLINE, MEDLINE In-Process and the website www.ClinicalTrials.gov. In addition, one trial presented at the Annual Meeting of the American Society of Hematology 2016 was added to include the most recent evidence. In total, six RCTs were identified, including two formulations: Anti-t-lymphocyte globulins (ATLG, Grafalon(®), Neovii Biotech) and polyclonal globulins immunized with human thymocytes (Thymo, Thymoglobulin(®), Genzyme-Sanofi). The evidence was synthesized using a conventional network meta-analysis (NMA). The best treatment option to prevent GVHD was ATLG with the most favorable hazard ratio compared to standard treatment regarding chronic graft-versus-host-disease (GVHD; 0.42, 95% CI 0.31 to 0.56), acute GVHD II- IV (0.54, 95% CI 0.39 to 0.73) and acute GVHD III+IV (0.50, 95% CI 0.29 to 0.86), whereas both ATLG and Thymo were at least similarly effective regarding transplant-related mortality (TRM; 0.90, 95% CI 0.61 to 1.32 and 0.90, 95% CI 0.56 to 1.44). Thymo tended to be the better treatment option regarding overall survival (OS; 0.86, 95% CI 0.59 to 1.26). Our NMA provides the first relative efficacy of all available rabbit ATG formulations in patients undergoing allo-SCT. Until additional data from randomized head to head comparisons are available, based on this analysis, ATLG seems to be the best option to prevent chronic and acute GVHD. Both formulations show similar efficacy in TRM while Thymo tends to be the better treatment

  10. Doubling an investment

    NASA Astrophysics Data System (ADS)

    Eliazar, Iddo

    2004-01-01

    We study the issue of optimal long-term portfolio management in continuous time multi-asset financial markets. Rather than following the abstract notion of ‘utility’ and its implied paradigm of ‘maximization of expected utility’ we suggest a different approach: The investor sets a goal-such as reaching a desired fortune level, or doubling the initial investment-and then operates to minimize the expected time-to-goal, i.e., achieving the goal as quick as possible. We assume the ‘standard model’ of multi-asset financial markets where assets are governed by correlated Geometric Brownian motion dynamics, and study optimality under the criteria of ‘minimization of the expected time-to-goal’. We explicitly compute: (i) the optimal holding strategies; (ii) the dynamics and behavior of the optimal investment portfolios; and, (iii) the statistics-mean, variance, and Laplace transform-of the time-to-goal (under the optimal investment strategy). Also, an investment paradox arising in this context-in which some portfolios have exponential mean growth but have a positive probability of never doubling their initial value-is discussed and explained.

  11. Expression of a gp33/27,000 MW activation inducer molecule (AIM) on human lymphoid tissues. Induction of cell proliferation on thymocytes and B lymphocytes by anti-AIM antibodies.

    PubMed Central

    Sánchez-Mateos, P; Cebrián, M; Acevedo, A; López-Botet, M; De Landázuri, M O; Sánchez-Madrid, F

    1989-01-01

    We have recently described several monoclonal antibodies (mAb) that recognize a heterodimeric structure (gp33/27,000 MW) expressed on the surface of human peripheral blood T lymphocytes upon activation with different mitogenic stimuli. Such mAb, when used in combination with submitogenic doses of phorbol ester, were capable of triggering T-cell proliferation. The antigen has been designated as activation inducer molecule (AIM). In the present study we have investigated the expression of the AIM in different lymphoid and non-lymphoid tissues. In addition, we have analysed the ability of lymphocyte subsets derived from thymus and tonsil to proliferate in response to anti-AIM mAb. The presence of AIM on subpopulations of lymphoid cells from thymus, tonsil, lymph node and spleen has been demonstrated by immunoprecipitation, flow cytometry and immunoperoxidase staining of tissue sections. By contrast, non-lymphoid cells from tissue such as brain, kidney, liver, lung or skin did not react with anti-AIM mAb. In thymus, the AIM expression was restricted to a subset of CD3+ medullary thymocytes, whereas CD1+ CD3- cortical thymocytes did not express this antigen. Nevertheless, the majority of both purified CD1- and CD3- thymocytes expressed AIM antigen after treatment with PMA. In tonsil and lymph node, a strong staining of a subset of CD3+ T lymphocytes located in the germinal centre was observed by immunohistochemical labelling with anti-AIM mAb. Certain T cells from the paracortical zone and CD19+ B lymphocytes from mantle region were also reactive. Both purified tonsillar T and B lymphocytes strongly expressed AIM after activation with PMA. The anti-AIM mAb was able to induce a strong proliferative response on purified CD1- thymocytes as well as on both purified tonsillar T and B lymphocytes in the presence of submitogenic doses of PMA. By contrast, no proliferative response was induced through the AIM in the CD3- immature thymocyte subset. Images Figure 1 Figure 3

  12. Intersubunit distances in full-length, dimeric, bacterial phytochrome Agp1, as measured by pulsed electron-electron double resonance (PELDOR) between different spin label positions, remain unchanged upon photoconversion.

    PubMed

    Kacprzak, Sylwia; Njimona, Ibrahim; Renz, Anja; Feng, Juan; Reijerse, Edward; Lubitz, Wolfgang; Krauss, Norbert; Scheerer, Patrick; Nagano, Soshichiro; Lamparter, Tilman; Weber, Stefan

    2017-05-05

    Bacterial phytochromes are dimeric light-regulated histidine kinases that convert red light into signaling events. Light absorption by the N-terminal photosensory core module (PCM) causes the proteins to switch between two spectrally distinct forms, Pr and Pfr, thus resulting in a conformational change that modulates the C-terminal histidine kinase region. To provide further insights into structural details of photoactivation, we investigated the full-length Agp1 bacteriophytochrome from the soil bacterium Agrobacterium fabrum using a combined spectroscopic and modeling approach. We generated seven mutants suitable for spin labeling to enable application of pulsed EPR techniques. The distances between attached spin labels were measured using pulsed electron-electron double resonance spectroscopy to probe the arrangement of the subunits within the dimer. We found very good agreement of experimental and calculated distances for the histidine-kinase region when both subunits are in a parallel orientation. However, experimental distance distributions surprisingly showed only limited agreement with either parallel- or antiparallel-arranged dimer structures when spin labels were placed into the PCM region. This observation indicates that the arrangements of the PCM subunits in the full-length protein dimer in solution differ significantly from that in the PCM crystals. The pulsed electron-electron double resonance data presented here revealed either no or only minor changes of distance distributions upon Pr-to-Pfr photoconversion. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  13. Differential depletion of total T cells and regulatory T cells and prolonged allotransplant survival in CD3Ɛ humanized mice treated with polyclonal anti human thymocyte globulin

    PubMed Central

    Buszko, Maja; Cardini, Benno; Oberhuber, Rupert; Oberhuber, Lukas; Jakic, Bojana; Beierfuss, Anja; Wick, Georg; Cappellano, Giuseppe

    2017-01-01

    Thymoglobulin (ATG) is a polyclonal rabbit antibody against human thymocytes used as a T cell-depleting agent to prevent or treat allotransplant rejection. The aim of the present study was to investigate the effect of low dose ATG treatment exclusively on T cells using a humanized BALB/c human CD3Ɛ transgenic mouse model expressing both human and murine T cell receptors (TCR). Mice received a single intravenous (i.v.) injection of ATG. Blood and peripheral lymphoid organs were obtained after different time points. We found a significant T cell depletion in this mouse model. In addition, regulatory T cells (Tregs) proved to be less sensitive to depletion than the rest of T cells and the Treg:non-Treg ratio was therefore increased. Finally, we also investigated the effect of ATG in a heterotopic allogenic murine model of heart transplantation. Survival and transplant function were significantly prolonged in ATG-treated mice. In conclusion, we showed (a) an immunosuppressive effect of ATG in this humanized mouse model which is exclusively mediated by reactivity against human CD3Ɛ; (b) provided evidence for a relative resistance of Tregs against this regimen; and (c) demonstrated the immunomodulatory effect of ATG under these experimental circumstances by prolongation of heart allograft survival. PMID:28257450

  14. Application of Natural Language Processing and Network Analysis Techniques to Post-market Reports for the Evaluation of Dose-related Anti-Thymocyte Globulin Safety Patterns.

    PubMed

    Botsis, Taxiarchis; Foster, Matthew; Arya, Nina; Kreimeyer, Kory; Pandey, Abhishek; Arya, Deepa

    2017-04-26

    To evaluate the feasibility of automated dose and adverse event information retrieval in supporting the identification of safety patterns. We extracted all rabbit Anti-Thymocyte Globulin (rATG) reports submitted to the United States Food and Drug Administration Adverse Event Reporting System (FAERS) from the product's initial licensure in April 16, 1984 through February 8, 2016. We processed the narratives using the Medication Extraction (MedEx) and the Event-based Text-mining of Health Electronic Records (ETHER) systems and retrieved the appropriate medication, clinical, and temporal information. When necessary, the extracted information was manually curated. This process resulted in a high quality dataset that was analyzed with the Pattern-based and Advanced Network Analyzer for Clinical Evaluation and Assessment (PANACEA) to explore the association of rATG dosing with post-transplant lymphoproliferative disorder (PTLD). Although manual curation was necessary to i