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A M Kruisbeek

Publications and source records attributed to A M Kruisbeek.

At least 19 recordsLinked to original sources
PubMed1999

Costimulatory signals are required for induction of transcription factor Nur77 during negative selection of CD4(+)CD8(+) thymocytes.

A major question in end-stage T cell development is how T cell receptor(TCR) ligation on immature CD4(+)CD8(+) double positive thymocytes is translated into either survival (positive selection) or apoptotic (negative selection) signals. Because different types of antigen-presenting cells (APCs) induce positive or negative selection in the thymus and express different costimulatory molecules, involvement of such costimulatory molecules in determining cell fate of DP thymocytes is considered here. If TCR-generated signals are modulated by APCs, this should be reflected in the activation of distinct biochemical pathways. We here demonstrate that costimulatory signals involved in negative selection also are required for induction of protein expression of Nur77 and its family members. These transcription factors are critically involved in negative but not positive selection. In contrast, the signals that costimulate negative selection are not required for induction of several molecular events associated with positive selection. These include activation of the immediate early gene Egr-1, the mitogen-activated protein kinase ERK2, and surface expression of the CD69 marker. Thus, costimulation for negative selection selectively provides signals for activation of apoptotic mediators. These data provide molecular insights into how TCR-engagement by ligands on different thymic APCs can determine cell fate.

Animals↗
PubMed1999

Systemic T cell expansion during localized viral infection.

In a local immune response, the priming and expansion of the antigen-specific T cell population has been thought to largely take place in the draining lymphoid tissue. This model was primarily based on indirect enumeration of antigen-specific T cells by limiting dilution analyses. Here, tetrameric MHC class I complexes were used to evaluate the contribution of different secondary lymphoid organs in a local immune response by following the CD8+ T cell responses against the immunodominant epitopes of influenza A virus and herpes simplex virus-1. Mice were either intranasally infected with influenza A virus and developed pneumonia or were intradermally injected with herpes simplex virus-1. Remarkably, even though these viruses cause a local infection, the spleen of infected animals contains approximately 50-fold more antigen-specific cytotoxic T cells than the draining lymph nodes. Although antigen-specific T cells in spleen appear not to have experienced any recent encounter with antigen, this population is actively dividing, and over time, the formation of a memory T cell population is observed. These data reveal that there is a remarkably large and distinct population of antigen-specific T cells in spleen in the course of a local antigenic challenge. This T cell compartment may not only form the foundation of a memory T cell pool but could also provide a safeguard against systemic spreading of an infection.

Animals↗
PubMed1999

Cell-fate decisions in early T cell development: regulation by cytokine receptors and the pre-TCR.

During lymphocyte development, cell-fate decisions are determined by a myriad of signals produced by the micro- environment of the thymus and the bone marrow. These yet to be fully defined developmental cues regulate stage-specific gene expression, and the extraordinarily well-characterized stages of T and B cell development have provided attractive model systems for studying regulation of cellular differentiation. In particular, studies on the contribution of both antigen receptors and cytokine receptors to lymphoid development have illuminated essential signalling pathways in early T and B cells. Here, we review investigations supporting an obligatory role for the IL-7 receptor pathway in early T cell development. IL-7 is produced by both thymus and bone marrow stromal cells, and its potential contribution to survival, differentiation and proliferation of pro-T cells is discussed. We also address the contribution of the pre-T cell receptor (pre-TCR) to differentiation past the pro-T cell stage, and recent advances in deciphering the composition and function of the pre-TCR complex are discussed. Finally, we suggest future directions in this field that may serve to reveal whether and how signals initiated by the cytokine receptors and pre-TCR may intersect, and to define which down-stream molecular events are regulated by these receptors.

Animals↗
PubMed1998

The CD3gamma chain is essential for development of both the TCRalphabeta and TCRgammadelta lineages.

CD3gamma and CD3delta are the most closely related CD3 components, both of which participate in the TCRalphabeta-CD3 complex expressed on mature T cells. Interestingly, however, CD3delta does not appear to participate functionally in the pre-T-cell receptor (TCR) complex that is expressed on immature T cells: disruption of CD3delta gene expression has no effect on the developmental steps controlled by the pre-TCR. Here we report that in contrast with CD3delta, CD3gamma is an essential component of the pre-TCR. We generated mice selectively lacking expression of CD3gamma, in which expression of CD3delta, CD3epsilon, CD3zeta, pTalpha and TCRbeta remained undisturbed. Thus, all components for composing a pre-TCR are available, with the exception of CD3gamma. Nevertheless, T-cell development is severely inhibited in CD3gamma-deficient mice. The number of cells in the thymus is reduced to <1% of that in normal mice, and the large majority of thymocytes lack CD4 and CD8 and are arrested at the CD44-CD25+ double negative (DN) stage of development. Peripheral lymphoid organs are also practically devoid of T cells, with absolute numbers of peripheral T cells reduced to only 2-5% of those in normal mice. Both TCRalphabeta and TCRgammadelta lineages fail to develop effectively in CD3gamma-deficient mice, although absence of CD3gamma has no effect on gene rearrangements of the TCRbeta, delta and gamma loci. Furthermore, absence of CD3gamma results in a severe reduction in the level of TCR and CD3epsilon expression at the cell surface of thymocytes and peripheral T cells. The defect in the DN to double positive transition in mice lacking CD3gamma can be overcome by anti-CD3epsilon-mediated cross-linking. CD3gamma is thus essential for pre-TCR function.

Animals↗
PubMed1998

The TNF receptor family member CD27 signals to Jun N-terminal kinase via Traf-2.

CD27 is a lymphocyte-specific member of the TNF receptor (TNFR) family. It is a costimulatory molecule for peripheral T cells, as defined by its ability to enhance the TCR-induced proliferative response. We show here that CD27 augments TCR-induced Jun N-terminal kinase (JNK) activity in primary murine lymph node T cells. To investigate how CD27 couples to JNK, we performed a yeast two hybrid screen with the CD27 cytoplasmic tail. This revealed that CD27 directly associates with Traf-2. Transfection experiments using dominant negative Traf-2 indicated that CD27 communicates with JNK via Traf-2. These findings group CD27 together with other members of the TNFR family, TNFR-1, -2, CD30 and CD40, which have all been shown to couple to Traf proteins. Since Traf proteins have been reported to initiate an anti-apoptotic signaling pathway, our data suggest that CD27 not only regulates proliferation, but also survival of T lymphocytes.

Animals↗
PubMed1998

Non-responsiveness of antigen-experienced CD4 T cells reflects more stringent co-stimulatory requirements.

We recently reported that previously activated T cells, irrespective of the nature of the first stimulus they encountered, are unable to respond to Staphylococcal enterotoxin B (SEB), nor to soluble anti-CD3 monoclonal antibody (mAb) presented by splenic antigen-presenting cells (APC). Such previously activated T cells are, however, fully capable of responding to plate-bound anti-CD3 plus splenic APC. These data suggest differential integration of the T-cell receptor (TCR) and co-stimulatory signalling pathways in naive versus antigen-experienced T cells. Consistent with this hypothesis, anti-CD28 mAb restores the proliferative capacity of resting ex vivo CD45RBlo CD4+ T cells (representing previously activated T cells) to both soluble anti-CD3 mAb and SEB. Interestingly, mAb-mediated engagement of cytotoxic T-lymphocyte antigen-4 (CTLA-4) completely negates the rescue effects mediated by anti-CD28 mAb in CD45RBlo cells. Nevertheless, the non-responsiveness of CD45RBlo CD4+ T cells cannot be reversed by anti-CTLA-4 Fab fragments, indicating that it is not related to negative regulatory effects of CTLA-4 engagement itself. Interestingly, the addition of interleukin-2 (IL-2) restores the proliferative capacity of CD45RBlo CD4+ T cells to SEB and soluble anti-CD3 mAb. Moreover, when rescued by IL-2, the cells are less susceptible to the negative regulatory effects of CTLA-4 engagement. Together, these findings suggest that the non-responsiveness of CD45RBlo CD4+ T cells to certain stimuli may be related to inadequate TCR signalling, primarily affecting IL-2 production.

Abatacept↗
PubMed1998

CD40 and CD70 co-stimulate a potent in vivo antitumor T cell response.

In several studies, CD80, a potent co-stimulatory molecule, has been reported to be responsible for the induction of CD8+ antitumor T cell responses by CD80-transfected tumor cells. However, expression of CD80 by tumors not always ensures generation of a T cell-mediated antitumor response. Variables such as the inherent immunogenicity of a tumor and its major histocompatibility complex (MHC) expression status affect the efficacy of this approach. Therefore, in this study two other co-stimulatory ligands, CD40 and CD70, have been investigated for their ability to co-stimulate antitumor responses. The efficacy of CD40 and CD70 is compared with that of CD80, with respect to CD4 and CD8 T cell co-stimulatory capacity in vitro and their ability to induce in vivo antitumor responses. Furthermore, CD40 and CD70 are tested for their capacity to induce a long-lived memory response in vivo, as defined both by induction of tumor-specific cytotoxic T lymphocytes (CTLs) and rejection of wild-type tumor cells. It was found that, despite the fact that CD40 predominantly stimulates CD4 T cells, CD40-transfected MHC class II-negative P815 tumor cells become highly immunogenic and induce long-lasting memory tumor-specific CTLs in vivo. Furthermore, CD40 and CD70 emerge as powerful and even superior alternatives to CD80 for improving tumor immunogenicity in vivo. While the mechanisms by which they do so remain to be defined, these findings suggest additional strategies for immunotherapy.

Animals↗
PubMed1998

Thymocyte selection: not by TCR alone.

During development of T cells in the thymus, T-cell receptor (TCR)-mediated recognition of self-MHC/self-peptide complexes on thymic stroma dictates the developmental fate of immature CD4+CD8+ (double positive) thymocytes. Intriguingly, TCR-generated intracellular signals can elicit two entirely different cellular responses in such thymocytes: apoptosis or further differentiation. The critical issue in understanding end-stage T-cell development is how TCR occupancy can be perceived in such markedly different ways by the TCR. Here, we review the cytoplasmic and nuclear events that result from TCR signaling during thymocyte selection. Studies aimed at distinguishing molecular components involved in positive selection (resulting in signals for further differentiation) and negative selection (resulting in apoptosis) will help solve this fascinating feature of T-lymphocyte biology. We also discuss how non-TCR-derived signaling might serve to fine tune the TCR-driven selection events in thymocytes. Central to this aspect of the conceptual framework needed to explain thymocyte selection is the observation that thymic antigen-presenting cells appear to be specialized in the induction of either positive or negative selection. Finally, we suggest a hypothesis that integrates the facts currently available on developing thymocytes, and which may serve to refine our exploration of unresolved issues in thymocyte selection. This hypothesis expands our focus to include signals from receptors other than TCRs as modulating and amplifying factors in thymocyte signaling.

Animals↗
PubMed1997

Cytotoxic T lymphocyte antigen 4 (CTLA-4) interferes with extracellular signal-regulated kinase (ERK) and Jun NH2-terminal kinase (JNK) activation, but does not affect phosphorylation of T cell receptor zeta and ZAP70.

Cytotoxic T lymphocyte antigen 4 (CTLA-4) is an important regulator of T cell homeostasis. Ligation of this receptor leads to prominent downregulation of T cell proliferation, mainly as a consequence of interference with IL-2 production. We here report that CTLA-4 engagement strikingly selectively shuts off activation of downstream T cell receptor (TCR)/CD28 signaling events, i.e., activation of the microtubule-associated protein kinase (MAPKs) ERK and JNK. In sharp contrast, proximal TCR signaling events such as ZAP70 and TCR-zeta chain phosphorylation are not affected by CTLA-4 engagement on activated T cells. Since activation of the ERK and JNK kinases is required for stimulation of interleukin (IL)-2 transcription, these data provide a molecular explanation for the block in IL-2 production imposed by CTLA-4.

Abatacept↗
PubMed1997

Induction of TCR gene rearrangements in uncommitted stem cells by a subset of IL-7 producing, MHC class-II-expressing thymic stromal cells.

The embryonic thymic microenvironment provides the necessary elements for T cell lineage commitment, but the precise role of individual stromal cell components remains to be determined. Here we address the question of which stromal cell types are required for initiation of V-DJ rearrangements of the TCR-beta and TCR-delta locus in CD117+CD45+ uncommitted fetal liver progenitors. We show that fetal thymic stroma alone is necessary and sufficient for induction of TCR-beta and TCR-delta rearrangements. Furthermore, the ability to induce this T cell commitment step is confined to a subset of MHC class II-positive epithelial cells. Thymic stroma derived from mice with a targeted deletion in the IL-7 gene, however, lacks this ability. These findings set the stage for a further definition of the nature of the thymic stromal cell support in the regulation of T cell commitment.

Animals↗
PubMed1997

Nonresponsiveness and susceptibility to CTLA-4 of antigen-exposed CD4 T cells are not regulated by the Bcl-2 family of apoptotic mediators, but can be restored by IL-2.

Memory T cells are considered to be less dependent on costimulation and to respond more vigorously to TCR triggering compared to their naive counterparts. We and others, however, observed that memory CD4 T cells display nonresponsiveness to a variety of stimuli, including superantigens and soluble anti-CD3. We now report that CD28-derived costimulation can revert the nonresponsive state of antigen-exposed CD4 T cells. Interestingly, the rescuing effect of CD28 can be completely negated by CTLA-4 engagement. The malfunction of memory T cells is related to increased cell death; the viability can be restored by CD28 engagement and is negatively regulated by CTLA-4 engagement. Importantly, it has been reported that antigen-exposed T cells express lower levels of the anti-apoptotic mediator Bcl-2. In addition, CD28 costimulation was reported to upregulate the expression levels of Bcl-xL. We therefore examined the possible role of Bcl-2 family proteins in the nonresponsiveness of antigen-exposed CD4 T cells, and determined whether CTLA-4, in analogy to CD28, mediates its negative regulatory effects via the Bcl-2 family of apoptotic mediators. Our data indicate that neither the nonresponsiveness nor the susceptibility to CTLA-4 of antigen-experienced CD4 T cells are related to the expression levels of Bcl-2 or Bax. The rescuing effects of CD28, however, may be related to increased Bcl-xL levels. Addition of IL-2 normalizes the nonresponsiveness of memory CD4 T cells and renders these cells resistant to the negative effects of CTLA-4 engagement. Impaired IL-2 production is therefore likely to be the cause of the malfunction and CTLA-4 susceptibility of memory CD4 T cells.

Abatacept↗
PubMed1996

Isolation of an immunodominant viral peptide that is endogenously bound to the stress protein GP96/GRP94.

Heat shock protein gp96 primes class I restricted cytotoxic T cells against antigens present in the cells from which it was isolated. Moreover, gp96 derived from certain tumors functions as an effective vaccine, causing complete tumor regressions in in vivo tumor challenge protocols. Because tumor-derived gp96 did not differ from gp96 isolated from normal tissues, a role for gp96 as a peptide carrier has been proposed. To test this hypothesis, we analyzed whether such an association of antigenic peptides with gp96 occurs in a well-defined viral model system. Here we present the full characterization of an antigenic peptide that endogenously associates with the stress protein gp96 in cells infected with vesicular stomatitis virus (VSV). This peptide is identical to the immunodominant peptide of VSV, which is also naturally presented by H-2Kb major histocompatibility complex class I molecules. This peptide associates with gp96 in VSV-infected cells regardless of the major histocompatibility com- plex haplotype of the cell. Our observations provide a biochemical basis for the vaccine function of gp96.

Animals↗
PubMed1996

Defects in cardiac outflow tract formation and pro-B-lymphocyte expansion in mice lacking Sox-4.

A striking example of the relationship between regulation of transcription and phenotype is the central role of the Y-chromosomal gene Sry in mammalian sex determination. Sry is the founding member of a large family of so-called Sox genes. During murine embryogenesis, the transcriptional activator Sox-4 is expressed at several sites, but in adult mice expression is restricted to immature B and T lymphocytes. Using targeted gene distruption, we have found that SOX-4(-/-) embryos succumb to circulatory failure at day E14. This was a result of impaired development of the endocardial ridges (a specific site of Sox-4 expression) into the semilunar valves and the outlet portion of the muscular ventricular septum. The observed range of septation defects is known as 'common arterial trunk' in man. We studied haemopoiesis in lethally irradiated mice reconstituted with SOX-4(-/-) fetal liver cells and found that a specific block occurred in B-cell development at the pro-B cell stage. In line with this, the frequency and proliferative capacity of IL-7-responsive B cell progenitors in fetal liver were severely decreased in vitro.

Animals↗
PubMed1996

Identification and functional analysis of Ly-6A/E as a thymic and bone marrow stromal antigen.

We recently described an mAb (MTS23) reactive with a membrane Ag expressed on a subset of thymic medullary stromal cells. This Ag is also constitutively expressed at high levels on peripheral B cells, macrophages, and thymic and splenic dendritic cells of C57BL/6 mice. A number of stromal cell lines derived from thymus and bone marrow also stain with MTS23, but thymocytes and peripheral T cells only weakly express the Ag detected by MTS23. Here we show that the molecule detected by MTS23 is a member of the Ly-6 family of phosphatidylinositol-anchored membrane proteins. Treatment of stromal cells with phosphatidylinositol-phospholipase C before staining completely abolished expression. Using transient expression of 293T cells and a cDNA library of a stromal cell line cloned into the pEF-BOS vector, a cDNA encoding the MTS23-target Ag was isolated. Partial sequencing and restriction enzyme mapping revealed that it represents the Ly-6A/E protein. While the physiologic significance of the presence of Ly-6 molecules on stromal cells is not clear, it has been known for some time that, at least in lymphocytes, cellular activation events can be induced upon Ly-6 engagement. We now demonstrate that Ly-6 also functions as a signal transduction molecule on stromal cells, in that granulocyte-macrophage CSF can be produced by a variety of stromal cell lines upon mAb-mediated cross-linking of Ly-6. Together with the dramatic up-regulation of Ly-6 expression on stromal cells upon IFN-gamma treatment, this is the first indication of a biologic function of an Ly-6 gene product on nonhemopoietic cells. The results suggest that Ly-6 may play a role in the cross-talk between lymphocyte precursors and stromal cells.

Animals↗
PubMed1996

Mechanisms underlying T-cell tolerance.

To understand how the immune system manages to disarm potentially harmful T cells with reactivity against self-antigens, many immunologists have turned to model systems in which transgenic expression of TCRs, foreign antigens, or both, allows visualization of otherwise undetectable mechanisms. Recent data have offered several new insights into the variables that dictate the mechanism for tolerance employed by peripheral T cells. Some of these mechanisms can also dampen ongoing immune responses against foreign antigens and provide a strikingly powerful supplement to the primary mechanism of tolerance, thymic clonal deletion.

Animals↗
PubMed1996

Expression of tyrosine kinase gene in mouse thymic stromal cells.

Amongst the most important signal transduction molecules involved in regulating growth and differentiation are the protein tyrosine kinases (PTK). Since T cell development is a consequence of interactions between thymic stromal cells (TSC) and thymocytes, identification of the PTK in both compartments is required to dissect the mechanisms that control this process. Here we report a search for PTK in mouse TSC, using RT-PCR to survey the repertoire of PTK mRNAs expressed in a freshly isolated TSC preparation. We identified 10 different PTK cDNAs among the 216 cDNAs sequenced, and demonstrate that transcripts of three of those (ufo, fyn and fer) are widely expressed among a large panel of immortalized thymic epithelial cell lines (TEC) and in primary cultures of TSC. Of the other seven, none were expressed in established TEC lines but, instead, displayed distinct expression patterns in cell types likely to have contaminated the fresh TSC preparation, i.e., macrophages, B cells, T cells and fibroblasts. Among the three PTK expressed in TEC lines, only one, ufo, exhibited expression exclusively in cells of non-hemopoietic origin. Although expression of ufo (also known as tyro 7, axl or ark) is not thymic-specific, in that it is also expressed in cell types of mesodermal origin in other tissues, its presence in TEC suggests a role for ufo in differentiation of the TSC compartment. Consistent with this notion, high-level expression of this receptor PTK at the protein level could be documented in every TEC line investigated, as well as in fresh thymus tissue sections. These data provide the first example of a receptor PTK in TSC and open new approaches to study the regulation of TSC differentiation.

Amino Acid Sequence↗