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L D'Adamio

Publications and source records attributed to L D'Adamio.

At least 37 records · Page 2Linked to original sources
PubMed1993

Interleukin-4 protects double-negative and CD4 single-positive thymocytes from dexamethasone-induced apoptosis.

Glucocorticoid hormones (GCH) and anti-CD3 monoclonal antibodies (MoAbs) induce in mouse thymocytes and T-cell tumor lines an active process of cell death called apoptosis. Interleukins (IL), including IL-1 and IL-2, have been reported to inhibit such apoptosis. In this study we show that IL-4 also reduced the DNA fragmentation characteristic of dexamethasone (DEX)-induced apoptosis in thymocytes. This effect, studied in both time-course and dose-response experiments, was also detected at low IL-4 concentrations (1 U/mL) and against high DEX levels (10(-7) mol/L). The effect of IL-4 was blocked by an anti-IL-4 but not by an anti-IL-1 alpha MoAb, and was thus both specific and direct. Phenotypic analysis showed that IL-4 protects predominantly CD4-CD8- and CD4+CD8- cells. Our findings suggest that intrathymic T-cell development may be influenced by IL-4.

Animals↗
PubMed1993

Overexpression of CD3 eta during thymic development does not alter the negative selection process.

To elucidate the role of CD3 eta in thymic development and to determine whether CD3 eta is involved in the negative selection process, CD3 eta was overexpressed > 100 fold in transgenic (tg) mice using a Thy-1 promoter and regulatory elements. CD3 eta was readily observed in the majority of cortical thymocytes and in a fraction of medullary thymocytes in tg mice by immunohistochemical staining with an anti-CD3 eta-specific mAb. In contrast, endogenous CD3 eta levels were too low to detect in normal littermates. Flow cytometric analysis demonstrated an increased level of TCR on thymocytes with intermediate TCR density in tg animals and parallel biochemical studies showed a marked increased in TCR-associated CD3 zeta-eta heterodimers and CD3 eta-eta homodimers relative to controls. Despite this change in surface TCR phenotype, there was no significant alteration in the total numbers or proportion of CD4+CD8+ double-positive or CD4+CD8- or CD4-CD8+ single-positive thymocytes or peripheral T cells. Percentages of SP V beta 5, V beta 6, and V beta 8 thymocytes in tg animals were unaltered compared to normal littermates when backcrossed either to C57BL/6 (H-2b) or DBA/2 (H-2d) backgrounds. Furthermore, induction of DNA fragmentation with anti-CD3 epsilon mAb treatment in vivo was not significantly different for tg and normal littermates. Collectively, these data imply that CD3 eta is not a limiting component of the negative selection process.

Animals↗
PubMed1993

Thymic and peripheral apoptosis of antigen-specific T cells might cooperate in establishing self tolerance.

Aside from CD4+CD8+ double-positive (DP) thymocytes, the subpopulations of T lineage cells affected by negative selection are unknown. To address whether this process occurs in more mature cell types, we have compared the responses of purified single-positive (SP) murine thymocytes and peripheral T cells to the superantigen staphylococcal enterotoxin B (SEB) utilizing as antigen-presenting cells (APC) a fibroblast cell line expressing transfected I-Ek class II molecules. Whereas approximately 70% of SEB-reactive SP thymocytes, either CD4+ or CD8+, undergo programmed cell death (apoptosis) and, therefore, negative selection, CD4+ and CD8+ antigen-specific peripheral T cells are predominantly activated and proliferate to APC+SEB. Thus, mature thymocytes and peripheral T cells, with identical patterns and levels of expression of CD4, CD8 and T cell receptor (TCR), are programmed to elicit different responses following TCR stimulation. Unexpectedly, however activation of peripheral T cells was preceded by deletion of a large fraction of V beta 8+ T lymphocytes (SEB specific). This surprising phenomenon was also observed in in vivo studies: in fact, administration of SEB to adult mice resulted in depletion of the majority of antigen-specific T cells from the peripheral lymphoid tissues analyzed (lymph nodes and spleen). This depletion is the consequence of deletion as indicated by program cell death of V beta 8+ T cells and is followed by proliferation of the remaining SEB-reactive T cells. Clonal elimination of peripheral T cells may represent a mechanism by which tolerance to self antigens never expressed in and/or exported to the thymus is achieved.

Animals↗
PubMed1993

Interleukin-2 induces apoptosis in mouse thymocytes.

Interleukins play a role in the process of T-cell development and, like other cytokines, seem able to modulate apoptosis. Interleukin-2 has been reported to inhibit apoptotic cell death of thymocytes induced in vitro by either activation of CD3/TCR complex or treatment with glucocorticoid hormone. We demonstrate here that IL-2 can provoke DNA fragmentation and cell death of CD4+ CD8+ mouse thymocytes by activating an endogenous apoptotic pathway. Thymocytes, incubated with high IL-2 concentrations in vitro, showed the morphological characteristics of apoptotic cells, including reduction in nuclear size, derangement in chromatin structure, and DNA fragmentation in oligonucleosomal subunits. Inhibition of mRNA and protein synthesis and addition of the PKC-inhibitor H-7, Zn2+ ions, and IL-4 counteracted the IL-2 effect. These data suggest that high IL-2 concentrations may induce an active process of cell death on mouse thymocytes in vitro.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1992

Negative selection of thymocytes. A novel polymerase chain reaction-based molecular analysis detects requirements for macromolecular synthesis.

Self-tolerance is mainly established through clonal deletion of autoreactive T cells during thymic differentiation. The mechanisms by which deletion is achieved are poorly understood. Here we use a specific polymerase chain reaction-based system to characterize DNA fragmentation and show that after in vivo treatment of neonatal mice with staphylococcus enterotoxin B, selective apoptosis of V beta 8+ thymocytes occurs. This process precedes detectable deletion of V beta 8+ cells as determined by phenotypic analysis. Moreover, in vivo administration of cycloheximide and, to a lesser extent, actinomycin D, inhibits apoptosis of staphylococcus enterotoxin B specific thymocytes. Thus, macromolecular synthesis is a requirement for negative selection.

Animals↗
PubMed1992

T cell receptor complexes containing Fc epsilon RI gamma homodimers in lieu of CD3 zeta and CD3 eta components: a novel isoform expressed on large granular lymphocytes.

CD3 zeta and CD3 eta form disulfide-linked homo- or heterodimers important in targeting partially assembled Ti alpha-beta/CD3 gamma delta epsilon T cell receptor (TCR) complexes to the cell surface and transducing stimulatory signals after antigen recognition. Here we identify a new TCR isoform expressed on splenic CD2+, CD3/Ti alpha-beta+, CD4-, CD8-, CD16+, NK1.1+ mouse large granular lymphocytes (LGL), which are devoid of CD3 zeta and CD3 eta proteins. The TCRs of this subset contain homodimers of the gamma subunit of the high affinity receptor for IgE (Fc epsilon RI gamma) in lieu of CD3 zeta and/or CD3 eta proteins. The LGL display natural killer-like activity and are cytotoxic for B cell hybridomas producing anti-CD3 epsilon and anti-CD16 monoclonal antibodies, demonstrating the signaling capacity of both TCR and CD16 in this cell type. These findings provide evidence for an additional level of complexity of TCR signal transduction isoforms in naturally occurring T cell subsets.

Animals↗
PubMed1992

Pidotimod stimulates natural killer cell activity and inhibits thymocyte cell death.

Experiments were performed to analyze the possible effect of the immunomodulating agent Pidotimod (3-L-pyroglutamyl-L-thiazolidine-4-carboxylic acid) on mouse Natural Killer (NK) cell activity and glucocorticoid hormone(GCH)-induced thymocyte apoptosis. The results indicate that in vivo treatment with Pidotimod (200 mg/Kg ip for 5 days) causes a significant increase in NK activity and in vitro treatment produces a significant reduction of dexamethasone-induced thymocyte apoptosis. This inhibition appears to be dose-dependent and is also evident against TPA or Ca(++)ionophore-induced apoptosis.

Adjuvants, Immunologic↗
PubMed1991

CD3 eta and CD3 zeta are alternatively spliced products of a common genetic locus and are transcriptionally and/or post-transcriptionally regulated during T-cell development.

The CD3 eta subunit of the T-cell receptor is thought to subserve an important role in signal transduction and possibly T-cell development. Herein we characterize the organization of the mouse CD3 eta gene and show that it is part of one gene locus that also encodes CD3 zeta on chromosome 1. The NH2-terminal sequence of CD3 zeta and CD3 eta, which share the same leader peptide and are identical through amino acid 122 of each mature protein, is encoded by exons 1-7. However, exons 8 and 9 are differentially spliced to give rise to CD3 zeta and CD3 eta: exons 1-8 encode CD3 zeta and exons 1-7 plus 9 encode CD3 eta. RNase protection analysis with RNA from a variety of fetal, neonatal, and adult cell types indicates that expression of both gene products is T-lineage-restricted. Importantly, expression of CD3 zeta and CD3 eta mRNA appears before or on day 16 of fetal gestation. Expression is apparently coordinate since no cell types tested express CD3 zeta or CD3 eta alone. The steady-state level of CD3 zeta mRNA is greater than or equal to 40-60 times that of CD3 eta mRNA. In immature CD4+CD8+CD3low double-positive thymocytes and CD4+CD8-CD3high or CD4-CD8+CD3high single-positive thymocytes, the respective steady-state CD3 zeta and CD3 eta mRNA levels are equivalent, whereas the amount of receptor-associated CD3 zeta and CD3 eta proteins in double-positive thymocytes is approximately 10 times less than in single-positive thymocytes. Nevertheless, the CD3 zeta/CD3 eta protein ratio remains constant in all populations (40-60:1). Furthermore, discordance between mRNA and protein levels for CD3 zeta and CD3 eta is also observed in splenic T cells. Thus, posttranscriptional and/or transcriptional regulatory mechanisms control CD3 zeta and CD3 eta expression during T-cell development.

Amino Acid Sequence↗
PubMed1991

T-cell receptor isoforms and signal transduction.

Recent cDNA and genomic cloning have identified CD3 eta as an alternatively spliced product of the same gene locus that encodes CD3 zeta. Three distinct T-cell receptor isoforms have now been identified. A current view of the signal transduction function of these isoforms in thymocytes and T cells is discussed.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1990

Characterization of thymus-derived lymphocytes expressing Ti alpha-beta CD3 gamma delta epsilon zeta-zeta, Ti alpha-beta CD3 gamma delta epsilon eta-eta or Ti alpha-beta CD3 gamma delta epsilon zeta-zeta/zeta-eta antigen receptor isoforms: analysis by gene transfection.

To characterize the function of the CD3 eta subunit of the T cell receptor (TCR), we have used cDNAs encoding CD3 zeta, CD3 eta, or both to reconstitute a variant of a cytochrome c-specific, I-Ek-restricted murine T cell hybridoma, termed MA5.8, which lacks CD3 zeta and CD3 eta proteins. We provide direct evidence that assembly and surface expression of TCRs can be mediated by either of these subunits separately or together. However, the level of TCR expression on zeta transfectants is up to one order of magnitude greater than that on eta transfectants, implying that CD3 eta is weakly associated with the pentameric Ti alpha-beta CD3 gamma delta epsilon complex and/or inefficient at salvaging the incomplete TCR from lysosomal degradation. As a component of the TCR, the CD3 eta subunit preferentially forms a heterodimer with CD3 zeta, but is also able to form a CD3 eta-eta homodimer. Crosslinking of Ti alpha-beta CD3 gamma delta epsilon zeta-zeta, Ti alpha-beta CD3 gamma delta epsilon eta-eta, or Ti alpha-beta CD3 gamma delta epsilon zeta-zeta/zeta-eta TCR isotypes with anti-CD3 epsilon monoclonal antibody or a cytochrome c peptide epitope on I-Ek antigen-presenting cells mediates signal transduction resulting in reversible cell-cycle arrest of transfected clones. Given the potential for diversity of signals generated by these functional TCR isotypes and the expression of the CD3 eta gene product in the thymus, CD3 eta is likely to play a role in selection and/or activation of thymocytes during development.

Animals↗
PubMed1990

Downregulation of enkephalin-mediated inflammatory responses by CD10/neutral endopeptidase 24.11.

The antigen CD10 (common acute lymphoblastic leukaemia antigen), which is the zinc metalloprotease, neutral endopeptidase 24.11 (also known as NEP or 'enkephalinase'), is expressed by acute lymphoblastic leukaemias, normal lymphoid progenitors, mature polymorphonuclear leukocytes and certain nonhaematopoietic cells. CD10/NEP hydrolyses several naturally occurring peptides, including the endogenous opioid pentapeptides Met- and Leu-enkephalin. In invertebrate organisms such as the mollusc Mytilus edulis, Met-enkephalin triggers inflammatory responses by inducing morphological changes, directed migration and aggregation of haemocytes. We report here that a structure related to CD10/NEP is expressed by M. edulis haemocytes and that abrogation of CD10/NEP enzymatic activity reduces the amount of Met-enkephalin required for haemocyte activation by five orders of magnitude. Similar results are obtained with CD10+ human polymorphonuclear leukocytes, indicating that CD10/NEP related structures regulate enkephalin-mediated inflammatory responses in organisms whose ancestors diverged approximately 500 million years ago.

Animals↗
PubMed1990

Human natural killer cells and mature T lymphocytes express identical CD3 zeta subunits as defined by cDNA cloning and sequence analysis.

In order to characterize the CD3 zeta-related protein found in human natural killer (NK) cells and compare it with CD3 zeta expressed in T lymphocytes, the present study was performed. A polyclonal CD3-CD16+NK population displaying a strong non-major histocompatibility complex-restricted cytotoxic activity against the NK target K-562 was isolated and a product corresponding to CD3 zeta amplified using the polymerase chain reaction method. This 0.6-kb product was present in similar amounts in NK cells and T cells. In contrast, a product corresponding to CD3 delta was amplified from T lymphocytes exclusively. Thus, the CD3 zeta product detected in NK cells did not originate from contaminating T cells. DNA sequence analysis of two independent polymerase chain reaction products from the NK cells demonstrates that human NK cells and mature T cells share a CD3 zeta subunit with an identical primary amino acid sequence. The nucleotide sequence of a third NK-derived cDNA revealed an insertion of a CAG triplet encoding an additional glutamine residue in the cytoplasmic domain. Since this residue is encoded by nucleotides at a putative RNA splice junction, it possibly results from a difference in pre-mRNA splicing. Taken together, these data show that CD3 zeta is not structurally distinct in NK cells and in T lymphocytes.

Amino Acid Sequence↗
PubMed1989

The common acute lymphoblastic leukemia antigen gene maps to chromosomal region 3 (q21-q27).

Complementary DNA and genomic clones corresponding to the gene for the common acute lymphoblastic leukemia antigen (CALLA) were used to investigate the genetic structure and location of the CALLA locus. The gene, which encodes a 100-kDa type II transmembrane glycoprotein, appears to be a single copy locus of greater than 45 kb which is not rearranged in malignancies expressing cell surface CALLA. Cell hybrid analysis indicates that the CALLA-related DNA sequences are found on human chromosome 3. In situ hybridization studies reveal the regional location of the CALLA locus to be 3q21-27.

Animals↗
PubMed1989

Common acute lymphoblastic leukemia antigen (CALLA) is active neutral endopeptidase 24.11 ("enkephalinase"): direct evidence by cDNA transfection analysis.

The common acute lymphoblastic leukemia antigen (CALLA) is a 749-amino acid type II integral membrane protein expressed by most acute lymphoblastic leukemias, certain other lymphoid malignancies with an immature phenotype, and normal lymphoid progenitors. A computer search against the most recent GenBank release (no. 56) indicates that human CALLA cDNA encodes a protein nearly identical to the rat and rabbit neutral endopeptidase 24.11 ("enkephalinase;" EC 3.4.24.11). This zinc metalloendopeptidase, which has been shown to inactivate a variety of peptide hormones including enkephalin, chemotactic peptide, substance P, neurotensin, oxytocin, bradykinin, and angiotensins I and II, had not been identified in lymphoid cells. To determine whether CALLA cDNA derived from human acute lymphoblastic leukemia cells (Nalm-6 cell line) encodes functional neutral endopeptidase activity, we generated CALLA+ stable transfectants in the CALLA- murine myeloma cell line J558 and analyzed them for enzymatic activity in a fluorometric assay based upon cleavage of the substrate glutaryl-Ala-Ala-Phe 4-methoxy-2-naphthylamide at the Ala-Phe bond. Total lysates as well as whole-cell suspensions of the Nalm-6 line and of the CALLA+ transfectants, but not of the CALLA- J558 cells, possessed neutral endopeptidase activity. This enzymatic activity was associated with the cellular membrane fraction and was abrogated by the specific neutral endopeptidase inhibitor phosphoramidon. The unequivocal identification of CALLA as a functional neutral endopeptidase provides insight into its potential role in both normal and malignant lymphoid function.

Animals↗
PubMed1989

Organization of the gene encoding common acute lymphoblastic leukemia antigen (neutral endopeptidase 24.11): multiple miniexons and separate 5' untranslated regions.

The common acute lymphoblastic leukemia antigen (CALLA) is a 749-amino acid type II integral membrane protein that has been identified recently as the neutral endopeptidase 24.11 [NEP (EC 3.4.24.11)]. Herein, we characterize the organization of the human CALLA/NEP gene and show that it spans more than 80 kilobases (kb) and is composed of 24 exons. Exons 1 and 2 encode 5' untranslated sequences; exon 3 [170 base pairs (bp)] encodes the initiation codon and transmembrane and cytoplasmic domain; 20 short exons (exons 4-23), ranging in size from 36 to 162 bp, encode most of the extracellular portion of the enzyme; and exon 24 (approximately 3400 bp) encodes the COOH-terminal 32 amino acids of the protein and contains the entire 3' untranslated region (UTR). Of note, the pentapeptide sequence (His-Glu-Ile-Thr-His) associated with metalloprotease zinc binding and substrate catalysis is encoded within a single exon (exon 19). Three types of CALLA/NEP cDNAs have been identified: these clones contain 5' UTR sequences differing from one another upstream of exon 3. These human 5' sequences are homologous to those found in rat brain and rabbit kidney NEP cDNAs. The three human CALLA cDNA types result from alternative splicing of exons 1, 2a, or 2b to the common exon 3. Moreover, exons 2a and 2b share the same 5' sequence but differ from each other by the use of two distinct donor splice sites 171 bp apart in the gene. The substantial conservation of 5' untranslated sequences among species and the existence of 5' alternative splicing suggest that CALLA gene expression may be differentially controlled in a tissue-specific and/or developmentally regulated fashion.

Amino Acid Sequence↗
PubMed1987

Interleukin-1 augments the interleukin-2-dependent generation of natural killer cells from bone marrow precursors.

We have studied the possible role of interleukin-1 (IL-1) on the interleukin-2 (IL-2) dependent development of mouse natural killer (NK) cells from primitive bone marrow (BM) precursors. Results indicate that both IL-1 alpha and IL-1 beta (1-10 U/ml) are able to stimulate the generation of NK cells from 5-fluorouracil (5-FU) resistant BM progenitors. These precursor cells are asialoGM1-, Thy-1+, Lyt-1- and Lyt-2-. Effector cells generated by culturing with IL-2 (40 U/ml) and IL-1 (5 U/ml) are Thy-1+, asialoGM1+, Lyt-5+, Lyt-1-, Lyt-2- and lyse only NK-susceptible targets. Generation of NK cells is blocked by addition of anti-IL-2/r. These data indicate that IL-1 may play a role in the generation of mature NK cells from undifferentiated BM precursors.

Animals↗
PubMed1986

Generation of mouse natural killer (NK) cell activity: effect of interleukin-2 (IL-2) and interferon (IFN) on the in vivo development of natural killer cells from bone marrow (BM) progenitor cells.

The possible effect of IL-2, alpha,beta-IFN and poly I:C (an IFN inducer) administration on the generation of NK cells of LI and BM-reconstituted animals was investigated. B6D2F1 mice were LI (9.5 Gy) by total-body irradiation and reconstituted by i.v. injection of different doses (ranging from 10(6) to 2 X 10(7)) of syngeneic BM, after which the levels of splenic NK activity were evaluated on days 4, 7, 9, 12 and 14 after LI and BM graft. After a marked decline on day 4 (no detectable NK activity at any effector to target ratio tested), NK activity gradually returned, reaching the levels of untreated controls on day 9. Groups of LI and BM-reconstituted mice were also treated i.p. with mouse or human recombinant IL-2 from day 0 through day 3 (15-50 U/day/mouse) after BM transplantation. It appears that an earlier reconstitution of NK activity occurs in IL-2-treated animals as compared to medium-injected controls. LI and BM-reconstituted animals were also treated i.p. with alpha,beta-IFN (10(4) U/mouse) or Poly I:C (1 mg/kg/mouse) from day 0 through day 3, and the splenic NK activity was evaluated at 4, 7, 9, 12 and 14 days after LI and BM graft. Our data indicate that in vivo administration of IFN or Poly I:C was able to cause an earlier maturation of NK activity as measured on days 7 and 9 after LI. In contrast, when the NK activity of IFN-treated animals was compared with that of controls 14 days after LI and BM graft, a significant inhibition was found due to the induction of suppressor cells. Pre-treatment of donor BM with Poly I:C or IFN was also able to induce a more rapid reconstitution of NK activity of recipient mice. The NK activity reconstitution paralleled the increase in the number of splenic LGL. A synergistic effect was obtained when LI mice were transplanted with Poly I:C-pre-treated BM and then treated with IL-2. The effector cell in the IFN and IL-2 treated chimeras is a typical NK cell: asialo GM-1+, Thy 1 +/-, Lyt 1-, Lyt 2- and reactive only against NK-susceptible targets. These data suggest that IL-2 as well as IFN may represent maturational signals in the in vivo physiological regulation of growth and differentiation of BM NK stem-cells.

Animals↗