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T Hünig

Publications and source records attributed to T Hünig.

At least 55 records · Page 3Linked to original sources
PubMed1995

Characterization of mouse CD53: epitope mapping, cellular distribution and induction by T cell receptor engagement during repertoire selection.

The pan-leukocyte antigen CD53 is a member of the poorly understood transmembrane 4 superfamily (TM4SF) of cell membrane glycoproteins. CD53 is proposed to play a role in thymopoiesis, since rat CD53 is expressed on immature CD4-8-thymocytes and the functionally mature single-positive subset, but is largely absent from the intermediate CD4+8+ cells. We have characterized CD53 in the mouse through the production of two new monoclonal antibodies, MRC OX-79 and OX-80, which were raised against the RAW 264 cell line and screened on recombinant CD53 fusion proteins. The epitopes recognized by both antibodies are dependent on disulfide bonding and map to the major extracellular region of CD53, requiring the presence of a single threonine residue at position 154. Mouse CD53 has a molecular mass of 35-45 kDa and is expressed on virtually all peripheral leukocytes, but not on cells outside the lymphoid or myeloid lineages. CD53 expression distinguishes subpopulations of thymocytes in the mouse and resembles the expression pattern of rat CD53. Amongst the immature CD4-8-thymocytes, mouse CD53 is clearly detectable on the earliest CD44high25- subset, but down-regulated on the later CD44high25+, CD44low25+ and CD44low25- stages. Also, the subsequent transient TcR-/low CD4-8+ cells and most CD4+8+ thymocytes express little or no CD53. This is consistent with the idea that cells which are committed to enter the selectable CD4+8+ compartment switch off CD53. The effect of T cell receptor (TcR) engagement on the re-expression of CD53 on CD4+8+ thymocytes was studied both ex vivo and in vitro using F5 mice, transgenic for the H-2b/influenza nucleoprotein-peptide-specific TcR, back-crossed onto an H-2q or H-2b background of RAG-2-deficient mice. CD4+8+ thymocytes from non-selecting H-2q F5 mice are CD53 negative, but in vitro stimulation through the TcR dramatically induces CD53 expression. In contrast, a fraction of CD4+8+ thymocytes from positively selecting H-2b F5 transgenic mice express CD53. Therefore TcR engagement by selecting major histocompatibility complex peptide complexes, or surrogate ligands, induces CD53 expression on otherwise CD53-negative, non-selected CD4+8+ thymocytes. Whether CD53 itself participates as a signaling molecule in further stages of thymic selection is still a matter of speculation.

Amino Acid Sequence↗
PubMed1995

Structural analysis of the rat T-cell receptor Tcra V4 gene family.

The rat Tcra V gene locus is only poorly characterized, although rats are widely used in a variety of T-cell-mediated experimental animal models. Recently, we described the first monoclonal antibody, G99, directed against a rat Tcra V4 segment. We examined cDNA transcripts of G99-positively sorted T cells and show that the monoclonal antibody G99 most likely recognizes at least two members of the Tcra V4 family. Moreover, we analyzed the genomic repertoire of this VA family and report 15 novel Tcra V4 DNA sequences. Based on sequence and Southern blot analysis, the Tcra V4 family could be divided into four subgroups, which were also detected in mice. These findings corroborate previous findings of a similar genetic organization of the Tcra V loci in both species.

Amino Acid Sequence↗
PubMed1995

Gamma/delta T cells in fetal, neonatal, and adult rat lymphoid organs.

In the present study, we have analyzed the appearance and maturation of gamma/delta T cells, recognized with a new mAb V65, in the central and peripheral lymphoid organs of fetal, neonatal, and adult Wistar rats. Cytofluorometrical analysis demonstrated the first V65+ gamma/delta T cells in the thymus of 16-17-day embryonic rats, although by immunohistology, they were identified only in 19-day rat embryos in both the cortico-medullary border and thymic medulla. Phenotypically, gamma/delta thymocytes from fetal and neonatal thymus expressed CD3, CD2, and CD5, but only 60-80% were CD8+ and approximately 40-50% expressed the alpha chain (p55) of the IL-2R. In the periphery, the immunohistological study identified for the first time gamma/delta T cells in the splenic white pulp and the gut of 21-day fetal rats, where they occurred within the epithelium as well as in the lamina propria. After birth, gamma/delta lymphocytes appeared in the skin, where they were present as dendritic epidermal T cells in increasing numbers during postnatal life. Whereas these gamma/delta T cells formed the predominant T-cell population in the rat skin, gamma/delta T cells in peripheral lymphoid organs, BALT, or the gut only represented a minor T-cell population. These results are discussed in comparison to gamma/delta T cells of other vertebrate species.

Animals↗
PubMed1994

Induction of interleukin 2 receptor beta chain expression by self-recognition in the thymus.

1-2% of adult mouse thymocytes express the T cell receptor alpha/beta (TCR-alpha/beta) together with the interleukin (IL) 2R beta (p70), but not the alpha (p 55) chain. We show that the previously described alpha/beta-TCR +CD4-8- and the partially overlapping Ly6C+ thymocytes are contained within this subset. Most IL-2R beta+ alpha/beta-TCR+ cells have a mature and activated (heat stable antigen [HSA]-, thymic shared antigen 1 [TSA-1]-, CD44high, CD69+) phenotype. Overrepresentation of V beta 8.2 in both CD4-8- and CD4 and/or CD8+ IL-2R beta+ thymocytes suggests that IL-2R beta expression is induced by a TCR-mediated activation event. In mice transgenic for an H-2Kb-specific TCR, IL-2R beta+ cells were abundant under conditions of mainstream negative selection, i.e., in the presence of Kb, but absent under conditions of mainstream positive selection or in a nonselecting environment. Together, these results show that in addition to clonal deletion, self-recognition by immature thymocytes leads to phenotypic maturation of a small subset of thymocytes expressing IL-2R beta. IL-2-deficient mice contain normal numbers of IL-2R beta+ alpha/beta-TCR+ thymocytes, indicating that like mainstream T cell development, this minor pathway of positive selection does not depend on IL-2. However, in the absence of IL-2, the CD4/CD8 subset composition of IL-2R beta+ thymocytes is skewed towards CD4-8+, mostly at the expense of CD4-8-. A possible relevance of this finding for the development of the immune pathology of IL-2-deficient mice is discussed.

Animals↗
PubMed1994

Identification and characterization of rat gamma/delta T lymphocytes in peripheral lymphoid organs, small intestine, and skin with a monoclonal antibody to a constant determinant of the gamma/delta T cell receptor.

A mAb called V65 was raised to a CD3+, TCR-alpha/beta- rat/mouse T cell hybrid that selectively reacts with all CD3+, TCR-alpha/beta- rat lymphocytes. Both anti-CD3 and V65 precipitate a 48- to 50-kDa heterodimeric protein from digitonin-lysed surface-iodinated cells. V65+ but not V65- T cells and T cell hybridoma cells express TCR-gamma mRNA. Together, these results show that V65 detects a constant determinant of the rat TCR-gamma/delta. In the presence of either IL-2 or IL-4, V65 stimulates proliferation in peripheral rat gamma/delta T cells. Approximately 90% of gamma/delta T cells from peripheral lymphoid organs have the same cell surface phenotype as thymus-derived MHC class I-restricted alpha/beta T cells, i.e., they are CD4- but express the CD8 alpha/beta heterodimer together with CD2 and CD5. In contrast, gamma/delta T cells from the epithelium of the small intestine lack CD2, CD4, and CD5 and express CD8 alpha only. Finally, V65 directly identifies a dense network of dendritic cells in the epidermis as gamma/delta T cells. These dendritic epidermal T cells are absent from athymic rats, indicating that like their mouse counterparts, they are thymus dependent.

Animals↗
PubMed1994

Control of the rat T cell response to retroviral and bacterial superantigens by class II MHC products and Tcrb-V8.2 alleles.

The in vitro response of unprimed rat T cells to retroviral and bacterial superantigens (SAg) was analyzed with TCR V beta 8.2-, 8.5-, 10-, and 16-specific mAbs. Specific stimulation of V beta 8.2 and 8.5 CD4 cells was observed in the response to Mls1a, the retroviral SAg encoded by integrated provirus Mtv-7 (Mtv-7 SAg), which was presented by mouse B cells or mouse fibroblasts transfected with DR1 genes and the Mtv-7 SAg. Additionally, a strong response of V beta 16 CD4 cells to an as yet unidentified mouse SAg was found. Only some of the bacterial SAg known to stimulate mouse and human T cells also activated rat lymph node cells. SEA, SEE, and TSST-1 stimulated rat T cells well; SEB, SEC1, and SED did not. This defect was apparently a result of weak binding to rat MHC class II molecules because presentation by human MHC class II molecules restored T cell activation. Under these conditions, SEB stimulated V beta 8.2+ and 8.5+ CD4 and CD8 cells from Lewis rats. A comparison of several rat strains revealed an unresponsiveness to SEB or Mtv-7 SAg for V beta 8.2 cells from F344 and DA rats. Determination of the nucleotide sequences of the Tcrb-V8.2 of these strains revealed differences between SAg-responsive and SAg-unresponsive Tcrb-V8.2 in seven amino acids, four of them located in the putative SAg contact site. The significance of these findings for the evolution of TCR-SAg interactions is discussed.

Alleles↗
PubMed1994

Preferential TCR V usage in rat repertoire selection: V alpha 8 imparts both positive thymic selection by and alloreactivity to RT1f.

Using a panel of newly developed mAb to two rat TCR V alpha and four TCR V beta segments, TCR V usage in CD4 and CD8 T cells of eight RT1 congenic strains sharing the LEW background was analyzed by flow cytometry. While no striking effects on V beta 8.2 and 8.5 usage were observed, a 3- to 4-fold over-representation of V beta 10 in the CD4 as compared with the CD8 subset in all strains suggested a preference of V beta 10 for MHC class II products. The degree of 'overselection' was mapped to the RT1.B/D region. In addition, an allele-specific overselection of V alpha 4+ CD4 T cells was mapped to RT1.B/Du and of V beta 16+ CD8 T cells to RT1.Au. Finally, a dramatic overselection of V alpha 8+ CD8 T cells by RT1f (14% in RT1f versus 1-2% in other haplotypes) provides the most striking case yet for an intrinsic affinity of a TCR V segment for an MHC product. V alpha 8+ CD8 T cells are not only overselected by RT1f in the thymus, but also during the alloreactive response of peripheral CD8 T cells to RT1f. The implications of these findings for the contribution of TCR V segments to TCR-MHC interactions in repertoire selection and alloreactivity are discussed.

Animals↗
PubMed1994

T-cell receptor diversity in dendritic epidermal T cells in the rat.

The rat epidermis contains a population of dendritic CD3+ cells. For a better characterization of these cells and to investigate their relationship to epidermal lymphocytes of other species, we stained rat epidermal sheets using a variety of monoclonal antibodies against rat leukocyte differentiation antigens in an indirect immunofluorescence procedure. Additionally, we attempted to define their T-cell receptor (TCR) isotype at both the nucleic acid and protein level. Results obtained showed that the majority of the CD3+ dendritic epidermal cells are CD45+, CD2+, TCR alpha beta-, major histocompatibility complex class II-, Thy-1-, asialo GM1-, CD4-, CD5-, and CD8- lymphocytes. We further observed that, in contrast to the mouse system, the rat epidermis additionally harbors a small but distinctive portion of dendritic CD3+ cells that exhibit reactivity with an anti-pan TCR alpha beta monoclonal antibody. Our further finding that rat epidermal cells enriched for CD3+ lymphocytes express full-length C delta mRNA suggests that the vast majority of rat epidermal T cells carry surface-bound TCR gamma delta moieties. On the basis of these findings, one may speculate that the indigenous T-cell population of the epidermis is not necessarily programmed to uniformly express monomorphic TCR gamma delta molecules but, to effectively fulfill its role in host defense, is capable of adaptation to the specific challenges encountered by a given species.

Animals↗
PubMed1993

T cell receptor ligation induces interleukin (IL) 2R beta chain expression in rat CD4,8 double positive thymocytes, initiating an IL-2-dependent differentiation pathway of CD8 alpha+/beta- T cells.

The role of interleukin (IL)2 in intrathymic T cell development is highly controversial, and nothing is known about IL-2R expression on thymocytes of the T cell receptor (TCR) alpha/beta lineage undergoing TCR-driven differentiation events. We analyze here IL-2R alpha and beta mRNA expression in an in vitro system where newly generated rat CD4,8 double positive (DP) thymocytes respond to TCR ligation plus IL-2 (but not to either stimulus alone) with rapid differentiation to functional CD8 single positive T cells (Hünig, T., and R. Mitnacht. 1991. J. Exp. Med. 173:561). TCR ligation induced expression of IL-2R beta (but not alpha) chain mRNA in DP thymocytes. Addition of IL-2 then lead to functional maturation and expression of the IL-2R alpha chain. To investigate if the CD8 T cells generated via this IL-2R beta-driven pathway in vitro correspond to the bulk of CD8 T cells seeding peripheral lymphoid organs in vivo, we compared their phenotype to that of lymph node CD8 T cells. Surprisingly, analysis of CD8 cell surface expression using a novel anti-CD8 monoclonal antibody specific for the alpha/beta heterodimeric isoform, and of CD8 alpha and beta chain mRNA revealed that T cells generated by TCR ligation plus IL-2 resemble thymus-independent rather than thymus-derived CD8 cells in that they express CD8 alpha without beta chains. These findings demonstrate that TCR crosslinking induces functional IL-2R on immature DP rat thymocytes. In addition, they show that at least in vitro, CD8 alpha/alpha T cells are generated from TCR-stimulated DP thymocytes (which express the CD8 alpha/beta in the heterodimeric isoform) along an IL-2-driven pathway of T cell differentiation.

Animals↗
PubMed1993

Early phenotypic and functional alterations in lymphocytes from simian immunodeficiency virus infected macaques.

Phenotypic and functional changes in lymphocytes from rhesus monkeys (Macaca mulatta) were investigated during the first 6 months after infection with SIV mac 32H. Animals preimmunized with keyhole limpet hemocyanin (KLH) were sacrificed 1, 3, 6, 12, and 24 weeks post infection. Subset composition and function of lymphocytes from blood, spleen, lymph node and thymus were analysed. In addition to a rapid decline in CD4/CD8 ratios, a massive reduction in CD29+ CD4+ cells was seen in the periphery. Although depletion of this subset was observed throughout the course of this experiment, the loss of proliferative T cell responses was most pronounced very early after infection and partially recovered after Month 3. Polyclonal cytotoxic responses were only slightly affected. In the thymus, a gradual, but moderate loss of CD4+CD8+ immature thymocytes, and a relative increase in both CD4+ and CD8+ mature subsets was observed. Infectious virus was readily recovered from homogenates of lymph node and spleen, but not of thymus tissue. Interestingly, however, virus was detected in thymocytes from all infected animals by cocultivation with a simian immunodeficiency virus (SIV) susceptible cell line.

Animals↗
PubMed1992

Prevention and therapy of experimental autoimmune neuritis by an antibody against T cell receptors-alpha/beta.

The mAb R73 directed to the TCR-alpha/beta of rat lymphocytes was tested for its therapeutic potential during the effector phase of experimental autoimmune neuritis (EAN) in Lewis rats. EAN can be actively induced by immunization with bovine peripheral nerve myelin, bovine P2 protein, or a peptide containing its neuritogenic epitope and serves as a model of the human Guilain-Barré syndrome. Adoptive transfer of activated P2-specific T lymphocytes also produces the monophasic disease (AT-EAN) characterized by inflammation and demyelination of peripheral nerves and highlights the central role of T lymphocytes in the pathogenesis of EAN. A single administration of the mAb R73 immediately after injection of activated P2-specific T line cells completely prevented the development of clinical and electrophysiologic signs of EAN in most animals and greatly alleviated the disease in the others. In further experiments mAb R73 was applied after the appearance of first clinical signs of EAN actively induced by immunization with a neuritogenic peptide or bovine peripheral nerve myelin. In both cases the anti-TCR-alpha/beta mAb reversed clinical signs of EAN and prevented the development of peripheral nerve dysfunction. In vivo and in vitro data suggest that impairment of Ag recognition and T cell function by occupancy of the TCR and R73-induced TCR-modulation rather than depletion of TCR-alpha/beta-bearing lymphocytes is the decisive mechanism underlying suppression of EAN that is apparent already within 48 h of the first R73 injection.

Animals↗
PubMed1992

Differential thymus dependence of rat CD8 isoform expression.

Expression of the rat CD8 molecule was studied using five novel monoclonal antibodies (mAb), four of which are specific for the V-like domain of CD8 alpha, whereas one reacts either with the beta chain or with a determinant only expressed on the CD8 alpha/beta heterodimer. mAb to both chains effectively blocked purified lymph node CD8 T cells in mixed lymphocyte reaction and in cell-mediated cytotoxicity. Flow cytometric analysis showed that CD8 T cells from lymph nodes or spleen of normal rats almost exclusively express the alpha/beta isoform, regardless of the T cell receptor isotype (alpha/beta or gamma/delta). In contrast, natural killer (NK) cells carry only CD8 alpha chains. This CD8 alpha + beta - phenotype was also prominent among CD8 T cells from athymic rats and from intestinal epithelium of normal rats. CD8 alpha homodimers can also be expressed as a result of activation, as shown by analysis of CD4 CD8 double-positive T cells obtained from highly purified lymph node CD4 T cells by in vitrok stimulation. Such CD4+CD8 alpha + beta - cells also represent a major subset among adult intestinal intraepithelial lymphocytes (IEL), suggesting local activation. Taken together, the difference in CD8 isoform expression among T cells from athymic rats, NK cells, and gut IEL versus CD8 T cells from peripheral lymphatic organs of euthymic animals suggests that like in mice, expression of the CD8 heterodimer is more dependent on intrathymic maturation than that of the homodimer. Since the more stringent thymus dependence of CD8 alpha + beta + T cells may be due to a requirement for thymic selection on self major histocompatibility complex class I antigens, the virtually exclusive CD8 alpha + beta + phenotype of peripheral rat gamma/delta T cells could mean that antigen recognition by this subset is also restricted by MHC class I molecules.

Animals↗
PubMed1992

Taking the thymus to pieces.

Complex in vitro and in vivo techniques are being combined to unlock the remaining secrets of the thymus. In this report from a recent thymus workshop*, Bruno Kyewski and Thomas Hünig describe the genetic manipulations aimed at clarifying the mechanisms of T-cell selection and lineage commitment, and the use of organ culture and immunohistology to identify the thymic microenvironments in which these events take place.

Animals↗
PubMed1992

Maturation of CD4-8- alpha/beta TCR+ T cells induced by CD2-stimulation in vivo and in vitro.

Newly generated ('virgin') rat thymocytes of the immature CD4+8+ double positive (DP) subset were treated in suspension culture for 2 days with the stimulatory pair of anti-CD2 monoclonal antibodies OX-54 and OX-55. Approximately 50% of the recovered cells had downregulated CD4 and CD8 and upregulated the T cell antigen receptor (TCR). CD2-stimulated, but not control thymocytes proliferated in response to TCR plus IL-2 stimulation. In vivo, postnatal injection of OX-54/55 led to a dramatic and selective increase in functionally mature CD4-CD8- double negative (DN) alpha/beta--TCR(high) thymocytes and peripheral T cells. These findings show that CD2 stimulation can promote T cell differentiation and suggest that DN TCR(high) thymocytes can be generated from DP thymocytes via alternative pathways of T cell maturation.

Animals↗