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T Hünig

Publications and source records attributed to T Hünig.

At least 73 records · Page 4Linked to original sources
PubMed1991

Development and function of T cells in mice rendered interleukin-2 deficient by gene targeting.

Interleukin-2 (IL-2) is a lymphocytotropic hormone which is thought to have a key role in the immune response of mammalian cells. It is produced by a subpopulation of activated T-lymphocytes and acts in vitro as the principal auto- and paracrine T-cell growth factor (for reviews see refs 1-3). IL-2 is, however, not the sole T-cell growth factor, nor does it act exclusively on T cells, also promoting growth of NK cells and differentiation of B cells. A role for IL-2 in T-cell development has been postulated but remains controversial. Here we test the requirement for IL-2 in vivo using IL-2-deficient mice generated by targeted recombination. We find that mice homozygous for the IL-2 gene mutation are normal with regard to thymocyte and peripheral T-cell subset composition, but that a dysregulation of the immune system is manifested by reduced polyclonal in vitro T-cell responses and by dramatic changes in the isotype levels of serum immunoglobulins.

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PubMed1991

T cell receptor-mediated selection of functional rat CD8 T cells from defined immature thymocyte precursors in short-term suspension culture.

Recent results have indicated that positive and negative repertoire selection act on the major population of CD4,8 double-positive (DP) thymocytes that express 5-10-fold less T cell receptor (TCR) than mature T cells (i.e., they are TCRlow). Since DP cells obtained ex vivo are heterogeneous with regard to their stage within thymic selection, a homogeneous population of virgin DP cells suitable for selection studies was generated in vitro from their immediate precursors, the CD8 single-positive (SP) immature blast cells. To mimic TCR-mediated selection signals, these virgin DP cells were then cultured for another 2 d in the presence of immobilized anti-TCR monoclonal antibodies with or without interleukin 2 (IL-2). Daily monitoring of recovery and phenotype showed that without TCR stimulation, the cells remained DP and became small, TCRlow cells that were lost with a half-life of 1 d, regardless of the presence of IL-2. TCR stimulation resulted in rapid downregulation of CD4 and CD8, maintenance of a larger cell size, and induction of the CD53 antigen that marks mature and CD4,8 double-negative rat thymocytes. In the absence of IL-2, viability decreased as rapidly as without TCR stimulation. Addition of IL-2 rescued TCR-stimulated virgin DP cells and prevented CD8 downregulation, so that 50-80% of input DP cells were recovered after 2 d as CD4-8+53+ cells. After release from modulation, these in vitro generated CD8 SP cells quantitatively upregulated the TCR to the TCRhigh phenotype and were readily induced to proliferate and exhibit cytotoxic T lymphocyte (CTL) activity in a polyclonal readout. Evidence is presented implicating an IL-2 receptor (IL-2R) not containing the p55 chain (i.e., most likely the p70 intermediate affinity IL-2R) in the TCR plus IL-2-driven in vitro differentiation of virgin DP cells towards the mature CD8 SP phenotype.

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PubMed1991

Rat "first-wave" mature thymocytes: cycling lymphoblasts that are sensitive to activation-induced cell death but rescued by interleukin 2.

The sequential appearance of thymocyte subsets in rat ontogeny was studied using the surface markers CD4, CD8, and the alpha/beta T cell receptor (here referred to as TcR). It was noted that the first TcRhigh thymocytes, appearing around birth, are not resting lymphocytes but cycling blast cells. These "first wave" TcRhigh cells are medullary in location and predominantly of the CD4/CD8 "single-positive" phenotype. Only about 5% express the light chain of the interleukin (IL)2 receptor, indicating that binding of IL 2 to high-affinity receptors is not driving proliferation of these blasts. Newborn TcR high blast cells were purified and analyzed in vitro. When cultured without further additions, they rapidly stopped cycling. Stimulation with cross-linked anti-TcR monoclonal antibody plus IL 2 resulted in vigorous and rapid proliferation that exhibited accelerated kinetics as compared to peripheral resting T cells. In contrast, TcR cross-linking without exogenous IL 2 induced cell death. This TcR-induced cell death involved fragmentation of DNA characteristic of apoptosis that was readily detectable within 18 h of culture. Addition of IL 2 to these short-term cultures prevented TcR-induced DNA fragmentation. Together, these results suggest that in newly formed TcRhigh thymocytes, TcR engagement results in clonal deletion if the IL 2 receptor remains unoccupied but allows clonal expansion if IL 2 is provided. This mechanism may be operative in the establishment of self-tolerance during T cell development.

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PubMed1990

Identification and characterization of rat T cell subpopulations expressing T cell receptors alpha/beta and gamma/delta.

Peripheral lymphoid organs of the rat were investigated for the presence of lymphocytes that expressed the pan-T cell markers CD5 and OX-52 but not the T cell receptor (TcR) alpha/beta. Two such populations were identified: 2% to 4% of lymphocytes in adult spleen, lymph nodes and peripheral blood are CD5+ TcR alpha/beta- and express the OX-52 antigen at the same density as TcR alpha/beta+ T-cells. About 90% of these cells are CD8+. A second population is CD5-, CD8+ and OX-52low. Radioimmunoprecipitation from digitonin lysates of surface-labeled cells with an anti-CD3 antiserum showed that the CD5+, but not the CD5- population of TcR alpha/beta- cells expresses a CD3-associated disulfide-linked cell surface molecule of about 100 kDa apparent mol. mass. Upon reduction, one major band, migrating with 48 kDa was observed. A band of the same size was obtained with an anti-human delta chain peptide antiserum, indicating that the CD3-associated non-TcR alpha/beta molecule is the rat TcR gamma/delta. Functional assays showed that most, if not all natural killer (NK) cell activity is present in the CD5(-)-OX-52low population. Reactivity to foreign major histocompatibility complex (MHC) antigens in mixed lymphocyte reaction was exclusively found in TcR alpha/beta+ splenic T cells. It is concluded that rat gamma/delta T cells in the spleen do not contain a high frequency of cells with specificity for foreign MHC antigens. The seeding of the periphery with alpha/beta and the presumptive gamma/delta T cells was followed from birth. Most prominently in the spleen, alpha/beta T cells reached adult levels much later than gamma/delta T cells. Taken together, these findings demonstrate the expression of the TcR gamma/delta on a minor population of peripheral rat T cells with the predominant phenotype CD4-CD8+ that has no NK cell activity when freshly isolated and does not contain a high frequency of alloreactive cells.

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PubMed1990

The neonatally T cell receptor 2-suppressed rat: lymphocyte subset composition and immune reactivity.

Rats were injected intraperitoneally from birth on with a mouse monoclonal antibody (R73) to a constant determinant of the rat T cell receptor (TcR)2. Throughout the observation period (6 months), TcR2+ cells in peripheral lymphoid organs and blood were absent in treated animals with the exception of few (less than 10%) cells with a tenfold reduced TcR2 density; peripheral TcR2-CD3+ cells, i.e. most likely TcR1+ T cells, were increased in frequency. Among thymocyte subpopulations, only those expressing the TcR2 at a high level were reduced in number. The lack of a visible effect on immature thymocytes may, however, be due to the fact that despite high serum levels, thymic R73 determinants were incompletely saturated. Spleen and lymph node cells from TcR2-suppressed rats were completely unresponsive in mixed lymphocyte reaction (two fully allogeneic haplotypes tested) even in the presence of interleukin 2. Reactivity to the T cell mitogen concanavalin A was, in contrast, only partially reduced. Since rat TcR1 cells are activated by concanavalin A, these results suggest that the TcR1 cells present in TcR2-suppressed rats are functional, but do not respond to foreign major histocompatibility complex antigens at a high frequency, a finding of possible importance for immunosuppression with anti-TcR2 monoclonal antibody in human allografting. Neonatally TcR2-suppressed rats were unable to respond to the strong T-dependent antigen keyhole limpet hemocyanin administered intraperitoneally in alum with B. pertussis. Thus, in the absence of peripheral TcR2 cells, the numerically expanded TcR1 T cells are not capable of providing help for B lymphocytes.

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PubMed1990

Suppression and prevention of adjuvant arthritis in rats by a monoclonal antibody to the alpha/beta T cell receptor.

Adjuvant arthritis (AA) in rats is an experimentally induced autoimmune disease mediated by T lymphocytes specific for Mycobacterium tuberculosis. We raised the question whether T cells carrying the gamma/delta T cell receptor (TcR), reactive or not to mycobacterial antigens, are involved in the pathogenesis of AA. For this purpose, T cells bearing the TcR alpha/beta were depleted from circulation by treatment with a monoclonal antibody against the rat TcR alpha/beta (R73). This treatment efficiently suppressed existing disease. Even more efficient was pretreatment with R73 from birth, which prevented AA induction completely. In these alpha/beta+ T cell-depleted animals an elevated level of alpha/beta- T cells (about 15% vs. 1% in normal rats) was evident, which was not significantly increased by Mycobacterium tuberculosis injection. We found no positive evidence that gamma/delta + T cells do contribute to AA induction. Moreover, treatment with an anti-TcR alpha/beta monoclonal antibody may be very efficient treatment of T cell-mediated autoimmune diseases.

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PubMed1990

The suppressive effect of an antibody to the alpha beta cell receptor in rat adjuvant arthritis: studies on optimal treatment protocols.

An antibody (R73) to the alpha beta T cell receptor (TCR) was able to dramatically suppress adjuvant arthritis (AA) in rats. The efficacy of R73 treatment was investigated with regard to antibody dosage, injection route and timing of injections. R73 was equally effective in reducing joint swelling throughout a wide dose range (80 to 2000 microgram/dose) when given on day 15, 18, and 21 after arthritis induction. Both i.p. and i.v. injection were able to suppress the pre-existing joint swelling to the same extent. However, R73 was only effective when given before or at the peak of joint swelling which occurred between day 18 to 24. Synovial membrane hyperplasia, mononuclear cell infiltration as well as cartilage and bone destruction were markedly reduced after therapy. The effect of R73 was associated with depletion of alpha beta+ T cells from the circulation even at low antibody doses. Only few alpha beta+ T cells were found in the pannus tissue. Late treatment on day 27, 30 and 33 after arthritis induction did not influence clinical scoring of the disease and histological examination thereafter did not show any difference between treated animals and controls. We conclude that antibody therapy directed at the TCR seems to be very effective even in pre-existing autoimmune diseases if the relevant T cell population is affected. However, inflammation and joint destruction may reach a state at which anti-TCR treatment is no longer effective.

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PubMed1989

A monoclonal antibody to a constant determinant of the rat T cell antigen receptor that induces T cell activation. Differential reactivity with subsets of immature and mature T lymphocytes.

mAb R73 detects a T cell-specific surface molecule consisting of two disulfide-linked subunits of 40 and 46 kD, respectively, on 97% of peripheral rat T cells, as defined by the OX-52 marker. Of the few OX-52+ R73- cells, none are CD4+ but many express the CD8 antigen known to be present on rat NK cells. mAb R73 is mitogenic for unseparated spleen cells and for purified T cells. In the absence of non-T "accessory cells", stimulation by R73 requires artificial crosslinking of the mAb and is largely dependent on exogenous IL-2. Overnight incubation of purified T cells with crosslinked R73 mAb induces blastoid transformation, IL-2-R expression, and modulation of the R73 antigen. In the rat thymus, mature medullary cells express the R73 determinant at the same level as peripheral T cells, whereas 94% of CD4-CD8- thymocytes are R73-. The major CD4+8+ thymocyte population contains 25% R73- and 70% R73low cells. Thymocytes of the CD4-CD8+OX-44- subpopulation that are the direct precursors of CD4+CD8+ cells display a continuum of R73 antigen density from undetectable to very low levels. We conclude that R73 is most likely directed at a constant determinant of the rat alpha/beta heterodimeric TCR and suggest that CD8+ immature thymocytes are the first cells in the T cell differentiation pathway to express this molecule at their surface.

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PubMed1989

The role of CD2/LFA-3 interaction in antigen- and mitogen-induced activation of human T cells.

Binding of LFA-3 to the T cell surface receptor CD2 promotes intercellular adhesion and is costimulatory with anti-CD2 mAbs in 'alternative pathway' activation of T cells. Since all AC-dependent systems of T cell activation are inhibited by anti-LFA-3 mAb, it was asked whether in mitogen- and antigen-induced activation of human T cells, the function of CD2/LFA-3 interaction involves signalling beyond its function in promoting intercellular adhesion. In order to selectively block and reconstitute CD2/LFA-3 interaction while leaving other AC functions available, the response of unseparated PBMC to various T cell mitogens and to allogeneic cells was blocked by a newly developed mAb (G26) to human LFA-3. Addition of purified T11TS, the sheep form of LFA-3 that binds to human CD2 but is not recognized by mAb G26, restored the T cell response to PHA-P but not to ConA, surface aldehydes, anti-CD3 mAb, or allogeneic cells. In addition, purified resting human T cells which were unresponsive to stimulation by lectins or anti-CD3 mAbs were activated by PHA-P in the presence of purified T11TS, demonstrating that provision of LFA-3 is a sufficient accessory cell function in the activation of human T cells by this mitogen. Again, the responses to ConA, cell surface aldehydes, or soluble anti-CD3 mAb were not restored by T11TS. T cell activation by PHA-P, but not by the other polyclonal T-cell activators studied thus seems to be mechanistically similar to 'alternative pathway' activation induced by anti-CD2 mAb in that the costimulatory effect of LFA-3 is independent of its prescence on an accessory cell membrane.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

Restricted expression of CD2 among subsets of sheep thymocytes and T lymphocytes.

A monoclonal antibody (mAb) generated against sheep T-cell blasts, called I/35 A, blocks sheep autologous E rosetting and competes with purified T11 target structure (TS), the sheep form of LFA3, for binding sites on the sheep T-cell surface. Immunoprecipitation from lysates of surface iodinated sheep T cells identifies the cell surface molecule recognized by mAb I/35 A as a single chain polypeptide migrating as a diffuse band of MW 55,000. From its binding properties and the biochemical nature of the target antigen, we conclude that mAb I/35 A is directed at sheep CD2. This finding makes sheep the first animal model in which the CD2-LFA3 (T11TS) system is defined by mAbs to both receptor and ligand. When analysed by two-colour flow cytometry and by immunohistochemistry, the cellular expression of CD2 in sheep differs significantly to that reported in humans. In peripheral blood, CD2 is found exclusively on CD4+8- and CD4-8+ T cells, while the third, CD4-8- (predominantly SBU-T19+) subset of sheep T cells (around 20% in peripheral blood) is CD2-. In thymus, only low to moderate levels of CD2 expression occurs on 80% of cells. Among these, medullary 'single positive' thymocytes express the highest level of CD2, whereas the CD4-8- 'double negative' population (which in contrast to peripheral CD4-8- T cells contains only very few SBU-T19+ cells) consists of CD2- and weakly positive cells. In peripheral lymphoid organs, CD2+ lymphocytes occur in the T-cell regions of spleen, lymph nodes and jejunal Peyer's patches (JPP). Tissue macrophages found in B-cell follicles of lymph nodes and JPP are also CD2+. The implications of these findings are discussed in terms of the role CD2 plays in the proliferation of immature thymocytes and of the possible importance of CD2/LFA3 interactions in lymphocyte recirculation.

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PubMed1988

Analysis of CD3 and antigen receptor expression on T cell subpopulations of aged athymic mice.

The expression of the T cell antigen receptor on subpopulations of extrathymically generated T cells from athymic mice was investigated and compared to antigen receptor expression in normal mice. To this end, spleen and lymph node cells from 18 individual athymic BALB/c nu/nu mice between 6 and 12 months of age and from normal controls were enriched for T cells by nylon wool filtration. Expression of the following cell surface markers was analyzed by two-color flow cytometry: Thy-1, CD4, CD8, V beta 8 and CD3. The distribution of subpopulations as defined by these markers varied much more among athymic than among normal mice. Some recurrent patterns did, however, occur that may be characteristic of the extrathymic pathway of T cell differentiation in nu/nu mice. Among Thy-1.2+ cells, CD8+ cells predominated over CD4+ cells. No CD4+8+ "double positive" cells were found, but CD4-8- "double negative" cell constituted 16% on average. All nu/nu nylon wool-nonadherent cells expressing Thy-1 at a normal level also expressed CD3, whereas Thy-1low and Thy-1- cells were CD3-. The fraction of V beta 8 T cells among the CD4+ and CD8+ subsets was near to normal in the majority of these animals. Most interestingly, the density of V beta 8 and CD3 expression was lower in athymic than in euthymic animals. This level of T cell antigen receptor expression was, however, higher than on immature CD4+8+ thymocytes. A fraction of the nude T cells presently characterized responded with proliferation to both anti-T cell receptor V beta 8 monoclonal antibody and to concanavalin A. Despite their apparently normal phenotype (with the exception of reduced T cell receptor expression), this response was, however, 4 to 10 times smaller than that of normal control cells. The presently described Thy-1+ T cell receptor intermediate cells may either be a(n) aberrant lineage(s) only generated extrathymically, or represent the accumulation of an immature intermediate stage of normal (i.e. thymic) T cell differentiation.

Animals↗
PubMed1988

Cross-linking of the T cell antigen receptor interferes with the generation of CD4+8+ thymocytes from their immediate CD4-8+ precursors.

The majority of thymocytes are immature cells co-expressing the surface markers CD4 and CD8. About two thirds of these cells also express the T cell antigen receptor (TcR), though at a level distinctly lower than found on mature T cells. The direct precursors of these "double-positive" thymocytes are cycling cortical blast cells of the CD4-8+ phenotype. Using a new monoclonal antibody to a constant determinant of the rat TcR alpha/beta, it is shown here that (a) about 50% of these CD8 "single-positive" committed precursor cells already express the TcR alpha/beta, though at very low levels, (b) during short-term suspension culture in medium supplemented only with fetal calf serum they not only acquire CD4 but also TcR alpha/beta levels characteristic of CD4,8 "double-positive" thymocytes, and (c) cross-linking of the TcR during culture inhibits the acquisition of the CD4 antigen in the majority of these cells.

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PubMed1987

Serologic cross-reactivity of T11 target structure and lymphocyte function-associated antigen 3. Evidence for structural homology of the sheep and human ligands of CD2.

T11 target structure (T11TS) and lymphocyte function-associated antigen (LFA) 3 are the cell-surface glycoproteins on sheep and human erythrocytes (E) binding to cluster differentiation 2 (the E-receptor) on T cells in E rosette formation. Here we show that this functional cross-reactivity is most likely due to a structural homology of these molecules. A rabbit antiserum to sheep T11TS is shown to cross-react with LFA-3 in several independent assays: (a) rabbit anti-T11TS antiserum blocks the formation of E rosettes by human T cells with both autologous and xenogeneic (sheep) E by binding to the respective E; (b) the antiserum blocks the binding of anti-LFA-3 monoclonal antibody to human E; and (c) it reacts with purified LFA-3 in Western blotting. Together, these findings demonstrate that T11TS on sheep E and LFA-3 on human E are serologically related, providing further support for the notion that T11TS and LFA-3 are the sheep and human forms of the same cell interaction molecule.

Animals↗
PubMed1987

Rosetting of human T lymphocytes with sheep and human erythrocytes. Comparison of human and sheep ligand binding using purified E receptor.

Previous studies have shown that the purified T lymphocyte glycoprotein, cluster differentiation 2 (CD2) (also known as T11, lymphocyte function-associated antigen (LFA)-2, and the erythrocyte (E) rosette receptor) interacts with the LFA-3 molecule on human E. We have examined the interaction of the purified CD2 molecule with the T11 target structure (T11TS) molecule on sheep E, and compared the two interactions. Purified, 125I-labeled CD2 bound to sheep E and the binding was inhibited by anti-T11TS monoclonal antibody (mAb). Reciprocally, the binding of T11TS mAb to sheep E was inhibited by pretreatment of sheep E with purified CD2. High concentrations of purified CD2 aggregated sheep E, possibly by inserting into the membrane, and the aggregation was inhibited by T11TS mAb. The affinity and number of binding sites for purified CD2 on sheep and human E was found to be similar, with Ka of 9 X 10(7)/M and 6 X 10(7)/M and 9800 and 8300 CD2 binding sites/E, respectively. Thus, the human T lymphocyte CD2 molecule is a receptor that cross-reacts between LFA-3 on human E and T11TS on sheep E, suggesting that LFA-3 and T11TS are functionally homologous ligands. As measured by saturation mAb binding, there are 8100 and 3900 ligand molecules/sheep and human E, respectively. Human and sheep E have surface areas of 145 and 54 micron 2, respectively. The 3.2- to 5.6-fold higher ligand density on sheep E appears to account for the ability of sheep but not human E to rosette with certain types of human T lymphocytes.

Animals↗
PubMed1987

Purified lymphocyte function-associated antigen-3 and T11 target structure are active in CD2-mediated T cell stimulation.

In this study we have used cells expressing LFA-3 or T11TS, the human and sheep forms of the ligand of CD2, as well as the purified LFA-3 and T11TS molecules themselves to study their effects on T cell activation via the CD2-mediated "alternative pathway". Sheep red blood cells, which bind to CD2 via T11TS in E-rosette formation, and human autologous monocytes, which express the LFA-3 molecule, both induce proliferation of resting T cells in the presence of per se submitogenic concentrations of anti-T11(2) plus anti-T11(3) monoclonal antibodies (mAb). This effect is blocked by mAb to LFA-3, T11TS and CD2 known to inhibit CD2-ligand interaction. In addition, purified LFA-3 and T11TS, when added at ng amounts to cultures containing submitogenic concentrations of anti-T11(2 + 3) mAb, are also strongly mitogenic for resting human T cells. Thus, both LFA-3 and T11TS are potent co-stimulators of the alternative pathway of T cell activation but by themselves do not provide a mitogenic signal. This finding is discussed with regard to a physiological role of CD2-LFA-3 interaction in T cell activation.

Animals↗
PubMed1987

The "erythrocyte receptor" of T-lymphocytes and T11 target structure (T11TS): complementary cell interaction molecules involved in T-cell activation.

The CD2 or T11 glycoprotein on T-lymphocytes is the receptor for both sheep and human erythrocytes in the formation of spontaneous ("E"-)rosettes. Recent evidence employing monoclonal anti-T11 antibodies suggested that T11 is also a signal transducing molecule with a function in T-cell activation. The present report summarizes the identification of T11 target structure (T11TS), a natural ligand of T11, and its biochemical and functional characterization. T11TS is defined by a mAb to sheep erythrocytes that completely blocks their binding to CD2. It is a glycoprotein of 42 kDa MW expressed on all types of blood cells and some other tissues. While the anti-T11TS mAb used is specific for sheep cells, an antiserum raised to purified T11TS also blocks human autologous E-rosetting. Evidence is presented that the human lymphocyte function associated antigen (LFA)-3, which had recently been shown to be the likely human ligand of CD2, is the structural and functional human homologue of T11TS. Functional studies on T-cell activation employing sheep erythrocytes as one ligand of CD2 indicate that binding of T11TS to the E-receptor provides one of the signals required for T-cell activation through the CD2 molecule.

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