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R Schwinzer

Publications and source records attributed to R Schwinzer.

At least 37 records · Page 2Linked to original sources
PubMed1994

Human CD45RA+ and CD45R0+ T cells exhibit similar CD3/T cell receptor-mediated transmembrane signaling capacities but differ in response to co-stimulatory signals.

CD45RA+ cells have been described to be less responsive to CD3/T cell receptor (TcR)-mediated activation than CD45R0+ T cells. To analyze the underlying mechanism of the differential responses we compared CD3/TcR-triggered tyrosine phosphorylation in the two subsets and studied the role of co-stimulatory signals provided either by accessory cells or pharmacologic activation of protein kinase C by phorbol ester. Stimulation of purified CD45RA+ and CD45R0+ T cells with CD3/TcR antibodies induced similar patterns and intensities of tyrosine phosphorylation in the two subsets, but no proliferation. If accessory cells were used as the source of co-stimulatory signals, strong expression of the 55-kDa chain of the interleukin-2 (IL-2) receptor (CD25), significant IL-2 production and vigorous proliferation were observed in CD45R0+ cells, whereas CD45RA+ cells responded weakly. However, when CD3/TcR-mediated triggering was combined with activation of protein kinase C by phorbol ester, CD45RA+ cells responded strongly. These data indicate that the transmembrane signaling capacity of the T cell receptor expressed by CD45RA+ and CD45R0+ cells is similar and, therefore, is presumably not responsible for the differential reactivities of the two subsets. It is more likely that co-stimulatory signals determine whether CD3/TcR-initiated activation results in strong or weak responses.

Antigen-Presenting Cells↗
PubMed1993

Characterization of the progression signal for human T-cell proliferation provided by monoclonal antibodies to the CD3-TCR complex.

Following T-cell activation, two distinct stages in the proliferative response can be identified. The first stage or state of competence (G0 to G1 transition) is defined by the acquisition of the ability of activated T cells to proliferate in response to growth factors. The second or progression phase relates to the progression of cells to DNA synthesis in response to exogenous growth factors (e.g., IL-2) or inducers of growth factor synthesis (e.g., phorbol esters). In the present manuscript we provide evidence that soluble monoclonal antibodies (mAbs) to CD3 or the T-cell receptor (TCR), similar to the phorbol esters, can induce IL-2 production and cell proliferation in competent but not resting T cells. The data indicate the role of CD3-TCR complex triggering not only as an "activating or competence signal" but demonstrate its capacity to signal in the progression phase as well. However, there were distinct differences when competent T cells were stimulated by phorbol ester compared to soluble antibodies to CD3 or TCR. Antibodies to the CD3-TCR complex, when compared to phorbol ester, induced less proliferation in competent, memory T cells (CD45RO) than in competent, naive T cells (CD45RA). In contrast to phorbol ester, the induction of IL-2 transcription and cell proliferation in competent cells by soluble mAbs BMA 031 (anti-alpha/beta T-cell receptor) or OKT3 (anti-CD3) was inhibited by cyclosporin A (CsA). This implies that in activated T cells there is a step between activation of the CD3-TCR complex and the point(s) of action of phorbol ester that is susceptible to CsA in activated cells. Alternatively, the data may indicate involvement of different regulatory elements with different sensitivities to CsA for the induction of IL-2 transcription by phorbol ester or antibodies to the CD3-TCR complex.

Antibodies, Monoclonal↗
PubMed1993

Responsiveness for mouse IgG2b antibodies to the human alpha/beta T-cell receptor/CD3 complex: evidence for genetic determination and low grade antibody cross-linking not mediated by known Fc gamma receptors.

Mouse IgG2a and IgG3 antibodies directed to the human T-cell receptor/CD3 complex stimulate peripheral blood mononuclear cells in almost all individuals. By contrast, responder and non-responder individuals exist for the stimulatory effect of mouse IgG1 and IgG2b antibodies. Whereas responsiveness to IgG1 antibodies is rather frequent (60-70%) and is known to be determined by an Fc gamma RII polymorphism on the accessory cells, little is known about the underlying factors of the rare (6%) IgG2b responsiveness. In this study it is shown that (1) IgG2b responsiveness is genetically determined with a dominant pattern of inheritance; (2) IgG2b responsiveness is determined by a radioresistant feature of responder accessory cells which can be substituted by artificial cross-linking, but not by IL-1 beta or IL-2; (3) stimulation of peripheral blood mononuclear cells of an IgG2b responder by IgG2b antibodies requires rather high antibody concentration compared with stimulation by IgG2a antibodies; (4) the stimulation leads to proliferation and IL-2 receptor expression, but no measurable IL-2 production; (5) antibody binding to known Fc gamma receptors (Fc gamma RI, Fc gamma RII, Fc gamma RIII) is not involved in the stimulatory effect of the IgG2b antibodies in responders. These results demonstrate that responsiveness to IgG2b antibodies against the TcR/CD3 complex depends on a genetically determined feature of accessory cells that is not identical with any of the known Fc gamma receptors. Most likely, the stimulatory effect observed in IgG2b responders is explained by a low grade cross-linking of the antibody by a not yet identified polymorphic structure on the surface of accessory cells.

Animals↗
PubMed1992

Monoclonal antibodies directed to different epitopes in the CD3-TCR complex induce different states of competence in resting human T cells.

After the initial stages of activation, T cells are not able to proliferate on their own but become competent to proliferate in response to exogenously added lymphokines. In the present study we compared the capacity of mAb directed to CD3 (OKT3, Leu4, UCHT1) or to common epitopes on the alpha/beta T-cell receptor (BMA 031, BMA 032) to induce competence in purified resting T cells. Stimulation with either soluble anti-CD3 or anti-alpha/beta TCR mAb rendered cells competent to progress to DNA synthesis in response to exogenous IL-2. In contrast, only soluble BMA 031 and BMA 032 were able to induce responsiveness to IL-4; anti-CD3 mAb had either to be immobilized or used in combination with anti-CD28 mAb to induce responsiveness to IL-4. Further, BMA 031-induced IL-4 responsiveness was selectively found in the CD45RA+ T cell subset. Analysis of early activation events revealed that the capacity of soluble BMA 031 and BMA 032 to induce responsiveness to IL-4 did not correlate with the ability of these mAb to increase the level of cytosolic Ca2+ or to induce detectable tyrosine phosphorylation. On the other hand, soluble Leu4 (anti-CD3) triggered an increase in both intracellular Ca2+ and tyrosine phosphorylation but was unable to induce IL-4 responsiveness. These data indicate that the induction of IL-2 and IL-4 responsiveness requires different sets of activation signals which can be induced by stimulating different epitopes in the CD3-TCR complex. This supports the concept that distinct activation pathways are coupled to the CD3-TCR complex.

Antibodies, Monoclonal↗
PubMed1992

Phenotypical and biochemical characterization of a variant CD45R expression pattern in human leukocytes.

As a result of a regulatory polymorphism a variant pattern of CD45R expression can be found in 8% of healthy individuals. We have previously reported that all resting T cells of these individuals react with CD45RA monoclonal antibodies (mAb) that are directed to the high-molecular mass isoforms (220 and 205 kDa) of the leukocyte common antigen (CD45). In addition, their T cells remain CD45RA+ after in vitro activation. In this report we studied the regulatory CD45R polymorphism in more detail. Flow cytometry studies revealed that CD45RA antigens are weakly expressed on normal monocytes but are absent from granulocytes. In individuals displaying the variant CD45R pattern both monocytes and granulocytes were CD45RA+. In controls and variant T cells CD45RA mAb precipitated two chains with molecular masses of about 220 and 205 kDa. In activated T cells from controls (CD45RA-) neither p220 nor the p205 chain of CD45RA could be detected. However, activated T cells from variant individuals (CD45RA+) did not express p220 but expressed p205 of CD45RA. These data indicate that there is a failure to down-regulate p205 CD45RA in individuals displaying the variant CD45R expression pattern. Since their T cells lost p220 CD45RA during activation the data also suggest that the two isoforms detected by CD45RA mAb can be regulated independently.

Antigens, CD↗
PubMed1992

Monoclonal antibodies to common epitopes of the human alpha/beta T-cell receptor preferentially activate CD45RA+ T-cells.

The murine monoclonal antibody BMA 031 (IgG2b) is directed to a monomorphic epitope on the human alpha/beta T-cell receptor. In contrast to anti-CD3 antibodies of the IgG2b isotype, BMA 031 is able to induce a proliferative response in T-cells from IgG2b low responders. This response occurs independently of cross-linking conditions indicating that the mode of activation differs from stimulation by the anti-CD3 antibody OKT3 (IgG2a) which strictly depends on cross-linking conditions. to further characterize the stimulatory potential of the two antibodies we studied the lymphocyte subsets responsive to stimulation by BMA 031 and OKT3. In CD45RA+ cells both antibodies exhibited similar effects. They induced weak expression of the 55-kDa chain of the interleukin-2 receptor (CD25), virtually no interleukin-2 secretion, but nevertheless strong proliferation. In CD45R0+ cells OKT3 and BMA 031 showed markedly different effects. OKT3 stimulated strong CD25 expression, strong interleukin-2 production, and marked proliferation. In contrast, CD45R0+ cells stimulated by BMA 031 showed only weak CD25 expression but neither interleukin-2 production nor proliferation. These data suggest that CD45RA+ and CD45R0+ cells differ in their capability to produce interleukin-2 upon stimulation via the CD3/T-cell receptor complex and also in the requirement for interleukin-2 to mount a proliferative response. The differential effect of OKT3 and BMA 031 in CD45R0+ cells probably results from the failure of BMA 031 to trigger interleukin-2 production which may be a consequence of its inability to induce CD3/T-cell receptor cross-linking in IgG2b low responders BMA 031 is therefore a useful tool for the selective activation of CD45RA+ cells in these individuals.

Animals↗
PubMed1991

Unusual phenotype of intestinal intraepithelial lymphocytes in the rat: predominance of T cell receptor alpha/beta+/CD2- cells and high expression of the RT6 alloantigen.

The phenotype of intraepithelial lymphocytes (IEL) from the small intestine of adult rats was studied by flow cytometry. Using appropriate monoclonal antibodies the expression patterns of the T cell receptor alpha/beta (TcR2), CD2, the alloantigen RT6 and several other T cell antigens were analyzed. The vast majority of rat IEL expressed TcR2 which is in contrast with data reported for the mouse. The comparison of IEL with lymph node cells revealed major phenotypic differences. Whereas CD2 was present on virtually all lymph node T cells it was found on only less than 5% of IEL. The T cell-specific differentiation antigen RT6 present on only a fraction of lymph node cells was found on about 99% of IEL demonstrating uniform expression with an approximately tenfold higher density. Identity of the detected molecule with RT6 was proven by using congenic controls and by the demonstration of phosphatidylinositol linkage to the IEL membrane. About 86% of IEL expressed CD8 but a substantial proportion of these cells co-expressed the CD4 molecule (34%). Two-color analysis revealed that the CD4+CD8+ double-positive subset completely lacked CD45RB suggesting that they represent memory cells. In the CD4-CD8+TcR2+ subset there was a remarkable heterogeneity of CD5 expression. A substantial number of these cells did not express CD5 despite high density of TcR2. Phenotypic peculiarities found on all or most IEL such as the lack of CD2 and the increased expression of RT6 indicate that the intestinal epithelial environment exerts strong effects on the development and maturation of these cells.

Animals↗
PubMed1991

Diabetes-prone BB rats express the RT6 alloantigen on intestinal intraepithelial lymphocytes.

The RT6 alloantigen of the rat is expressed on most peripheral T cells but not on thymocytes and thus represents a marker for postthymic T lymphocyte maturation in this species. Diabetes-prone (DP) BB rats exhibit a genetically determined T cell lymphopenia associated with a deficiency of RT6+ T cells. In this study we have analyzed the expression of RT6 on lymph node (LN) cells and intestinal intraepithelial lymphocytes (IEL) in two DP BB strains (BB/OK and BB/Mol) and two control strains (non-lymphopenic BB/PhiK and LEW) by flow cytometry. In the DP BB rats the number of LN T cells was substantially reduced (less than 25% TcR2+ cells) and completely lacked RT6 expression. The IEL population was also reduced in number and in marked contrast to normal rats consisted predominantly of CD4+ cells. The majority of IEL, however clearly expressed RT6. Treatment with a phosphatidylinositol (PI)-specific phospholipase C markedly reduced the RT6 density showing that PI-mediated anchoring of RT6 in the cell membrane also applies to IEL of DP BB rats. The results demonstrate that the DP BB strains possess a functional RT6 gene and are also able to generate the PI anchor. The defect in RT6 expression is thus unlikely to be the primary cause of the T cell lymphopenia.

Animals↗
PubMed1991

Activation of human thymocytes by antibodies to the CD3/T-cell receptor complex: triggering of different epitopes results in different signals.

Monoclonal antibodies (mAb) against the CD3/T cell receptor (TcR) complex were analyzed for their ability to activate human thymocytes. In addition to mAb detecting epitopes on the CD3 complex (OKT3, BMA 030) the activation potential of recently developed mAb against common epitopes on the alpha/beta T-cell receptor (anti-TcR mAb: BMA 031, BMA 032) was evaluated. Several differences were observed between the two types of mAb: (a) Binding of the tested anti-CD3 mAb to thymocytes resulted in a rapid increase in the level of cytoplasmic free calcium ions [Ca2+]i, whereas no significant changes in [Ca2+]i were detected in thymocytes stimulated with BMA 031 or BMA 032. (b) Induction of effective proliferation induced by mAb OKT3 depended on exogenous IL-2 and in addition on the presence of accessory cells or phorbol-ester. Proliferation induced by BMA 031 only required exogenous IL-2. (c) OKT3 but not BMA 031 inhibited proliferation of thymocytes induced via the CD2 molecule. These studies indicate that anti-CD3 and anti-TcR mAb transduce different signals in thymocytes. Since the two types of mAb are directed to the same molecular complex the observed differences also support the idea that there are functionally different compartments in the CD3/TcR complex which may activate different signaling pathways.

Antibodies, Monoclonal↗
PubMed1991

Cholera toxin-mediated inhibition of signalling in Jurkat cells is followed by, but not due to a loss of T cell receptor complex.

Cholera toxin treatment of the human T cell lymphoma Jurkat resulted in inhibition of signalling via the T cell antigen receptor complex (TcR/CD3-complex). Cholera toxin specifically ADP-ribosylated the alpha-subunit of the stimulatory G-protein of the adenylate cyclase (Gs alpha), no other proteins were modified in the intact cells. ADP-ribosylation of Gs alpha and its subsequent activation led to an increase of the cyclic AMP level and in addition, to a drastic reduction of the cell-surface density of the TcR/CD3-complex. Recently, we demonstrated that the effect of cholera toxin at the receptor level is not due to an increased cAMP level (4). As inhibition of signalling is also not cAMP-mediated (8), we examined whether the modulation of the TcR/CD3-complex could be the reason for the interruption of the signalling cascade. Analyzing the time courses of the multiple cholera toxin effects in Jurkat cells at 37 degrees C, the following sequence was found: ADP-ribosylation of Gs alpha--increase of cyclic AMP level--inhibition of signalling via the TcR/CD3-complex--decrease of cell-surface density of the TcR/CD3-complex. Treatment of Jurkat cells at 20 degrees C with cholera toxin resulted in an increase of cyclic AMP and inhibition of signal transduction, while no decrease of TcR/CD3-complex density could be observed. These data imply that receptor loss from the cell-surface is not causative for the inhibition of signalling. More likely, activation of Gs uncouples signal transduction in Jurkat cells via the TcR, which by a so far unknown mechanism is followed by a loss of the receptor from the cell surface.

Antigens, Differentiation, T-Lymphocyte↗
PubMed1990

Genetically determined lack of CD45R- T cells in healthy individuals. Evidence for a regulatory polymorphism of CD45R antigen expression.

A genetic polymorphism of CD45R expression was identified resulting in a lack of CD45R- lymphocytes in approximately 8% of healthy individuals. Family studies revealed an autosomal dominant mode of inheritance of the variant CD45R expression pattern. PBMC from donors possessing the variant type did not lose the CD45R antigen after in vitro activation, whereas a decrease of CD45R molecules was readily detected in individuals with the normal pattern. The expression of CD45RO antigens, as well as memory cell function, did not differ between both groups. These data show that activation and in vivo priming of T cells is not necessarily associated with a loss of CD45R antigen expression.

Antibodies, Monoclonal↗
PubMed1990

The G protein coupling T cell antigen receptor/CD3-complex and phospholipase C in the human T cell lymphoma Jurkat is not a target for cholera toxin.

Intact Jurkat cells could be stimulated by monoclonal antibodies against the Tcell antigen receptor complex (OKT3 directed against the CD3 complex, BMA031 directed against constant framework epitopes in the alpha/beta heterodimer). The accumulation of inositol phosphates was inhibited by prior incubation of the cells with cholera holotoxin. The inhibitory effect of cholera toxin (CT) was not cAMP mediated because forskolin (a direct activator of adenylate cyclase) did not mimic the inhibitory effect. When measuring phospholipase C (PLC) in a cell-free assay system by using [3H]inositol-labeled membranes, the enzyme could be stimulated by the poorly hydrolyzable GTP analogue guanosine 5'-O-(thiotriphosphate (GTP gamma S). Both anti-receptor antibodies augmented the GTP gamma S stimulatory effect, while the antibodies alone had no stimulatory capacity. In membranes from CT-pretreated cells, whereas the antibodies lost their stimulatory effect on PLC as in untreated cells, whereas the antibodies lost their stimulatory capacity in the presence of GTP gamma S. These data imply that CT exerts its inhibitory effect on signaling by acting at the receptor level while the PLC regulating G protein is not a target for CT-mediated alterations. This assumption is supported by the finding that in intact Jurkat cells CT, which ADP ribosylated only the alpha-subunit of the stimulatory G protein of the adenylate cyclase, led to a loss of the T cell antigen receptor complex from the cell surface as demonstrated by a decrease of receptor density using flow cytometry analysis. Receptor loss could not be achieved by forskolin treatment or incubation of the cells with the binding subunit of the toxin alone.

Adenosine Diphosphate Ribose↗
PubMed1990

Lymphocyte subsets in the blood. The influence of splenectomy, splenic autotransplantation, ageing, and the site of blood sampling on the number of B, T, CD4+, and CD8+ lymphocytes in the rat.

Removal of the largest single lymphoid organ, the spleen, leads to an increase in severe infections. To prevent this, transplantation of splenic fragments can be performed, which may, however, cause an increase in CD8+ lymphocytes in the blood of these patients. This is controversial since in the clinical situation it is often difficult to account for the different age of the patients, the time point after the operation and many other factors known to influence the number of lymphocyte subsets. Using a well-defined animal model, B, T, CD4+, and CD8+ lymphocytes were determined preoperatively in adult rats. Then, either sham splenectomy, splenectomy, or splenic autotransplantation was performed and the animals were followed up for 15 months after the operation. The surgical procedure itself, the site of blood sampling and ageing all influenced the number of lymphocyte subsets profoundly. Furthermore, giving the data as relative or absolute numbers leads to different results. Splenectomy caused lymphocytosis, due to a significant increase in B and CD8+ lymphocytes, as did splenic autotransplantation, which indicates that the number of lymphocyte subsets in the blood should not be used to argue in favour of or against splenic autotransplantation. This study demonstrates that the number of lymphocyte subsets in the blood is influenced by many factors and therefore should be determined in a highly standardized fashion.

Aging↗