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H G Ljunggren

Publications and source records attributed to H G Ljunggren.

At least 73 records · Page 4Linked to original sources
PubMed1997

Constitutive macropinocytosis allows TAP-dependent major histocompatibility complex class I presentation of exogenous soluble antigen by bone marrow-derived dendritic cells.

Dendritic cells expanded from mouse bone marrow (BMDC) with granulocyte/macrophage-colony-stimulating factor have potent T cell-stimulatory properties both in vitro and in vivo. This has been well documented for major histocompatibility complex (MHC) class II-restricted responses, and more recently using peptide-loaded and protein-pulsed DC for CD8 responses following adoptive transfer in mice. An unresolved question concerns the capacity of BMDC to present exogenous antigen on MHC class I molecules, an unconventional mode of MHC class I loading for which there is now considerable evidence, particularly in macrophages. Here, we show that BMDC exhibit high levels of macropinocytosis driven by constitutive membrane ruffling activity. Up to one-third of actively ruffling and macropinocytosing BMDC transferred pinocytosed horseradish peroxidase into the cytosol following a 15-min pulse, suggesting that they might be capable of presenting exogenous soluble antigen on MHC class I molecules. We show that BMDC presented exogenous ovalbumin to a T cell hybridoma more effectively, more rapidly, and at lower exogenous antigen concentrations than BM macrophages on a cell-for-cell basis. Presentation was TAP dependent, brefeldin A sensitive, and blocked by inhibitors of proteasomal processing, demonstrating use of the classical MHC class I pathway. Although effective presentation of exogenous antigen by BMDC occurred in the absence of agents which stimulate macropinocytosis, treatment with phorbol myristate acetate (PMA) enhanced both pinocytosis and MHC class I presentation by BMDC. Finally, PMA-stimulated BMDC exposed to exogenous ovalbumin in vitro were able to prime an antigen-specific cytotoxic T lymphocyte response following adoptive transfer in vivo.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1997

MHC class I presentation of live and heat-inactivated Sendai virus antigen in T2Kb cells depends on an intracellular compartment with endosomal characteristics.

T2Kb cells, which do not express TAP1/2 peptide transporters or the low molecular weight protein 2/7 (LMP2/7) proteasomal subunits, can still process and present both live and heat-inactivated Sendai virus (SV). As this operation may also reflect the existence of an alternative processing pathway in normal antigen-presenting cells (APC), the authors have characterized it using intracellular inhibitors and anti-Kb monoclonal antibodies (MoAbs). From the results with lipophilic amines (ammonium chloride, methylamine and chloroquine), cytoskeletal inhibitors (cytochalasin B and vinblastine), and an endoprotease inhibitor (phenylmethylsulfonyl fluoride, PMSF), the authors conclude that the processing of SV antigen in T2Kb cells has endosomal characteristics depending on cellular activities such as uptake, vesicular transport and intracellular-vesicular proteolysis. In addition, internalized 'empty' Kb molecules derived from the T2Kb cell surface appeared to be involved in the presentation of SV antigen, as demonstrated by a protocol using a combination of the Golgi inhibitor brefeldin A(BFA) and anti-Kb antibodies. The results thus indicate that T2Kb cells process SV antigen in an endosomal-like compartment which contain recycling 'empty' Kb molecules.

Amines↗
PubMed1997

Host MHC class I gene control of NK-cell specificity in the mouse.

The missing self model predicts that NK cells adapt somatically to the type as well as levels of MHC class I products expressed by their host. Transgenic and gene knock-out mice have provided conclusive evidence that MHC class I genes control specificity and tolerance of NK cells. The article describes this control and discusses the possible mechanisms behind it, starting from a genetic model to study how natural resistance to tumors is influenced by MHC class I expression in the host as well as in the target cells. Data on host gene regulation of NK-cell functional specificity as well as Ly49 receptor expression are reviewed, leading up to the central question: how does the system develop and maintain "useful" NK cells, while avoiding "harmful" and "useless" ones? The available data can be fitted within each of two mutually none-exclusive models: cellular adaptation and clonal selection. Recent studies supporting cellular adaptation bring the focus on different possibilities within this general mechanism, such as anergy, receptor calibration and, most importantly, whether the specificity of each NK cell is permanently fixed or subject to continuous regulation.

Animals↗
PubMed1996

External glycopeptide binding to MHC class-I in relation to expression of TAP transporters, beta 2-microglobulin and to pH.

MHC class-I binding glycopeptides are easily visualized on the cell surface by carbohydrate specific monoclonal antibodies. By comparing the staining intensity between anti-carbohydrate and anti-MHC class-I specific monoclonal antibodies, an estimation of the fraction of peptide accessible 'empty' sites on the cell surface of MHC class-I molecules can be made. This system was used to analyze glycopeptide binding to MHC class-I molecules in relation to transporter associated with antigen processing (TAP) peptide transporters and beta 2-M expression, using gene targeted mice, and in relation to pH. Approximately 15, 40, and 95% 'empty' Db molecules were found on activated T cells from normal, beta 2-M-/- and TAP -/- mice, respectively. The ASN9-6h-Gal2 glycopeptide also bound to transfected 'empty' Db molecules on T1-Db, T2-Db and T3-Db cells with a preference for T2-Db cells, lacking TAP peptide transporters. The stability of glycopeptide binding to H-2Db is also highest on T2-Db cells. pH was found to influence binding either positively or negatively, using four different glycopeptides, binding either to Db or Kb. We conclude that external glycopeptide binding may reflect important functional properties in the MHC class-I system and that pH in different processing compartments might influence the expressed peptide repertoire.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1996

Binding of H-2Kb-specific peptides to TAP and major histocompatibility complex class I in microsomes from wild-type, TAP1, and beta2-microglobulin mutant mice.

Major histocompatibility complex (MHC) class I molecules are trimolecular complexes consisting of a heavy chain (HC), beta2-microglobulin (beta2m), and a short peptide. Assembly of MHC class I molecules is thought to take place early during biosynthesis. Deficiency in either beta2m or the transporter associated with antigen processing (TAP) results in accumulation of class I molecules in the endoplasmic reticulum (ER). In this study, we have assessed peptide binding to TAP and MHC class I in purified microsomes derived from wild-type, TAP1(-/-), beta2m-/-, and TAP1/beta2m-/- mice using a cross-linkable H-2Kb-binding peptide. This enabled us to study the influence of an intact TAP complex and beta2m on peptide binding to MHC class I and to analyze the stepwise interaction of peptide with TAP and MHC class I molecules. Peptide bound both immature and mature (terminally glycosylated) class I molecules in intact as well as permeabilized microsomes from wild-type mice. Efficient peptide binding to immature class I molecules was also detected in permeabilized microsomes from TAP1(-/-) mice. In contrast, no peptide binding to beta2m-free HC was detected in permeabilized microsomes from beta2m-/- and TAP1/beta2m-/- mice. However, the addition of exogenous beta2m allowed peptide binding to class I in permeabilized beta2m-/- and TAP1/beta2m-/- microsomes. These results demonstrate that a preformed class I HC middle dotbeta2m heterodimer is necessary for efficient peptide binding under physiological conditions. The observed peptide binding to class I in permeabilized TAP1(-/-) microsomes further suggests that TAP1 is not required for peptide binding to class I in the ER. Finally, kinetic studies allowed the demonstration of a stepwise binding of peptide to TAP, subsequent translocation across the ER membrane, a step that required ATP hydrolysis, and binding of peptide to preformed class I HC.beta2m heterodimers.

Adenosine Triphosphate↗
PubMed1996

Natural resistance against tumors grafted into the brain in association with histocompatibility-class-I-antigen expression.

The role of MHC-class-I-antigen expression in intracerebral anti-tumor natural resistance was examined using MHC-positive Lym+ and MHC-negative Lym- lymphoma cell lines. Lym+ was sensitive to MHC-class-I-restricted CTL-mediated lysis, while lym- was resistant. Both lines were susceptible to NK-cell-mediated lysis. There was no difference in in vitro growth rate of in vivo intraperitoneal tumorigenicity between them. Inoculation of Lym+ cells into the brain caused upregulation of the intracellular MHC mRNA to the same level as after treatment with interferon-gamma, resulting in an increase in cell-surface MHC expression. Although inoculated Lym- cells also underwent an increase in cytosolic MHC mRNA, the cell-surface MHC expression remained negative. Immunoprecipitation revealed that the terminal glycosylation did not occur normally in Lym-. An in vivo intracerebral tumorigenicity assay, using 2 groups of untreated and NK-cell-depleted syngeneic mice, showed that Lym+ was less tumorigenic than Lym-. In T-cell-depleted mice, however, no difference was detected between them. In addition, when Lym+ and Lym- cells were inoculated into the brain of allogeneic or syngeneic preimmunized mice (immunized with tumor cells), Lym+ was rejected, while Lym- was accepted. When allogeneic mice had received treatment for T-cell depletion before intracerebral inoculation, no rejection was observed in Lym+. On the other hand, Lym- cells, when injected i.p. into NK-depleted mice, had greater killing activity than Lym+ cells, while in T-cell-depleted mice Lym- was less tumorigenic than Lym+. These results suggest that MHC-positive tumor cells grafted into the brain may be rejected by CTL in an MHC-dependent manner, whereas MHC-negative tumor cells can escape from T-cell-mediated immunosurveillance and grow progressively in the brain, due to absence of intracerebral natural resistance mediated by NK cells.

Animals↗
PubMed1996

Xenograft rejection of porcine islet-like cell clusters in normal and natural killer cell-depleted mice.

Fetal porcine islet-like cell clusters (ICC) were transplanted under the renal capsule of normoglycemic normal or athymic (nu/nu) C57BL/6 mice. Control animals were implanted with allogeneic minced kidney tissue from C57BL/Ks mice. The animals were killed 6 or 14 days after transplantation and the grafts were processed for flow cytometric analyses or immunohistochemistry. Xenograft destruction was evident in normal mice on day 6 after transplantation. The majority of infiltrating cells were macrophage-like cells expressing the F4/80 antigen. Lymphocytes expressing the CD3 antigen were in minority and mainly located in the peripheral parts of the ICC xenograft. The frequency and distribution of CD4+ cells were found to resemble those of the CD3+ cells. A large number of infiltrating cells, including several macrophage-like cells, expressed the Thy 1.2 antigen. Flow cytometry of infiltrating cells in the ICC xenograft revealed that approximately half of the cells expressing the F4/80 antigen also expressed Thy 1.2 and/or CD4. No cells were found expressing both the F4/80 and CD8 antigens. Both the F4/80 single-positive and the F4/80, CD4 double-positive cells were found to be larger and more granular than the CD4 single-positive cells. No co-expression of CD4 or Thy 1.2 with the F4/80 antigen was detected on cells infiltrating allogeneic tissue grafts. Moreover, a relative large number of cells (approximately 15%) in the xenograft expressed the NK 1.1 antigen as determined by flow cytometry. The role of natural killer (NK) cells in islet xenograft rejection was further evaluated in mice depleted of NK cells, using intraperitoneal injections of the monoclonal antibody NK 1.1. The simultaneous inoculation and subsequent growth of the NK cell-sensitive beta 2-microglobulin-deficient mutant, C4.4-25-, lymphoma cell line EL-4 served as an in vivo control of NK cell depletion. However, all NK cell-depleted mice rejected the ICC xenograft. In contrast, athymic mice permanently accepted the porcine ICC xenograft but, readily rejected the NK cell-sensitive lymphoma cell line. Taken together, ICC xenograft rejection in mice seems to be T cell dependent, as evidenced in the nude mice model, while the main effector cell appears to be a macrophage with a unique phenotype.

Animals↗
PubMed1996

TAP1-deficient mice select a CD8+ T cell repertoire that displays both diversity and peptide specificity.

Mice deficient in the gene encoding the transporter associated with antigen processing 1 (TAP1) are defective in providing major histocompatibility complex (MHC) class I molecules with cytosolic peptides. Consequently, these mice express reduced levels of MHC class I glycoproteins on the cell surface, and have reduced numbers of CD8+ T cells in the periphery. In the present study, we have addressed the diversity and specificity of the peripheral CD8+ T cell population in TAP1 -/- mice. CD8+ T cells were polyclonal with regard to T cell receptor (TCR) V beta expression. Overall, V beta usage in TAP1 -/- mice appear to be very similar to that in wild-type mice, with significantly reduced levels of V beta 5.1/5.2-expressing CD8+ T cells as the only clear exception. This polyclonal population of CD8+ T cells readily mounted epitope-specific CTL responses against four out of five well-defined MHC class I-restricted peptides. In contrast to allospecific CTL, peptide-specific CTL from TAP1 -/- mice did not cross-react on cells expressing normal levels of H-2b class I. The present results demonstrate that a polyclonal CD8+ T cell repertoire, displaying both diversity and peptide specificity, is positively selected in mice devoid of a functional peptide transporter. These observations imply that TAP-dependent peptides are not absolutely required for positive selection of a functionally diverse repertoire of CD8+ T cells.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1996

Chemically induced sarcomas from nude mice are more immunogenic than similar sarcomas from congenic normal mice.

To detect possible differences in immunogenicity between tumors induced in T cell-deficient mice and phenotypically normal congenic mice, 16 sarcomas, 8 having developed in nude BALB/c mice and 8 having developed in congenic normal (nu/+) mice, were transplanted to normal BALB/c recipients and the rates of rejection or acceptance were registered. The 16 tumors were chosen randomly from a panel of 39 sarcomas induced with 0.5% or 0.1% 3-methylcholanthrene and maintained as cell lines in culture. Out of the tumors originating from nude mice, 66% were rejected by the normal BALB/c recipients, while only 30% of the tumors originating from normal mice were rejected. Tumors with short induction times from normal mice were more readily accepted than tumors with long induction times. Tumors originating from nude mice had significantly longer mean latency times after transplantation to both normal and nude recipients than tumors originating from normal mice. Contrary to what has been reported by others, there was no correlation between the rejection rates of the individual tumors and their Kd, Dd or Ld major histocompatibility complex (MHC) class I surface expression as measured by flow cytometric analysis of cultured tumor cells. The Kd, Dd and Ld proteins of the transplanted tumor lines were analyzed by isoelectric focusing for the occurrence of mutations resulting in altered charge of the MHC protein. No such mutations were found, ruling out MHC mutations of that kind as the source of immunogenicity in the cell lines used in these experiments. Our results suggest the existence of a T cell-mediated selection in the original tumor cell mass of tumors induced in normal mice, adapting the tumor to growth in a host with a functional T cell system, but apparently there is no connection between this loss of immunogenicity and loss of MHC class I expression.

Animals↗
PubMed1996

Triggering of natural killer cells by the costimulatory molecule CD80 (B7-1)

NK cell-mediated cytotoxicity is influenced by triggering as well as inhibitory signals. The identification of inhibitory signals provided by MHC class I molecules has recently attracted significant attention. Much less is known about putative triggering signals. Using purified populations of mouse NK cells, we demonstrate that the CD80 (B7-1) gene product functions as a triggering signal for NK cell-mediated cytotoxicity. The strength of this response is such that it overrides the protection mediated by MHC class I molecules. Triggering of mouse NK cells by B7-1 occurred even in the absence of CD28 and could not be blocked by either anti-CD28 or anti-CTLA-4 antibodies. NK cells may thus, at least in part, use receptors other than CD28 and CTLA-4 in their interaction with B7-1. Furthermore, we demonstrate that bone marrow-derived macrophages and dendritic cells are highly susceptible to lysis by autologous NK cells.

Animals↗
PubMed1996

Principles of MHC class I-mediated antigen presentation and T cell selection.

Class I molecules of the major histocompatibility complex (MHC) are expressed on the cell surface of almost all nucleated mammalian cells. Their main function is to transport and present peptides, derived from intracellularly degraded proteins, to cytotoxic T cells (CTL). They are also directly involved in the process leading to maturation and selection of a functional CD8+ T cell repertoire. MHC class I molecules consist of a highly polymorphic membrane spanning heavy chain of approximately 45 kD that is non-covalently associated with a light chain, beta 2-microglobulin (beta 2m). Class I molecules bind peptides, usually 8-11 amino acids in length. The majority of the class I-bound peptides are generated in the cytosol and are subsequently translocated into the lumen of the endoplasmic reticulum (ER) through the ATP-dependent transporter associated with antigen processing 1/2 (TAP1/2). Here, we provide an up-to-date review summarizing the most essential parts relating to MHC class I-mediated antigen processing, presentation and T cell selection. A particular emphasis is devoted to the structure of MHC class I molecule, and MHC class I-bound peptides.

Animals↗
PubMed1995

Natural killer cell interaction with murine allogeneic MHC class I molecules.

Class I molecules of the MHC affect target cell sensitivity to NK cell repertoire. In the mouse, absence of MHC class I molecules on target cells is associated with an increased susceptibility to NK cell-mediated lysis, while syngeneic class I molecules confer protection. In contrast to the protective role of syngeneic class I molecules, less is known about the interaction between murine NK cells and allogenic class I MHC molecules. In theory, such could either be triggering, inert, or inhibitory. To directly address the role of allogeneic class I in interaction with NK cells of the mouse, a panel of polyclonal allogeneic murine NK cells were exposed to H-2b class I positive or negative target cells, and susceptibility to lysis was assessed. For all effectors studied, regardless of H-2 haplotype or genetic background, a preferential killing of class I-deficient targets was observed. This pattern was observed in vitro with tumor as well as lymphoblast targets, and in vivo in rapid elimination studies of radiolabeled tumor cells. The results demonstrate that protection from murine NK cell-mediated lysis can be conferred by the expression of allogeneic class I molecules. No evidence for a triggering effect caused by the expression of allogeneic class I molecules was observed. The data are discussed in relation to current models on NK cell/MHC class I interactions, alloreactivity mediated by NK cells, and the role of NK cells in allogeneic graft rejection responses.

Animals↗
PubMed1995

Rejection of tumors in mice with severe combined immunodeficiency syndrome determined by the major histocompatibility complex. Class I expression on the graft.

This study addresses the role of MHC class I molecules in the rejection of tumor grafts by SCID mice. Tumor cell lines, their corresponding MHC class I transfectants, and MHC class I-deficient mutants were inoculated to SCID mice. This allowed a study of tumor rejection responses in an environment with normal numbers of natural killer cells but largely devoid of functional T and B cells. C.B-17 (H-2d) SCID mice were found to reject low (10(2)) but not high (10(4)) doses of allogeneic (H-2b) tumor cells. The introduction of H-2Dd into such allogeneic tumor cells abrogated the rejection response with progressive tumor growth as a consequence. Introduction of H-2Kd or Ld had no or only marginal effects. The protective ability of H-2Dd was mapped to the alpha 1/alpha 2 domains of the molecule. H-2Dd protected allogeneic tumors from rejection also in C3H SCID mice of the H-2k haplotype, demonstrating that this ability was not dependent on H-2Dd expression in the host. Expression of endogenous H-2Kb and/or Db molecules partially protected wild-type allogeneic tumor cells from rejection since mutant allogeneic cells, devoid of class I expression, were rejected even after high-dose inoculation. Introduction of either allogeneic or xenogeneic class I molecules did not lead to rejection of otherwise MHC class I syngeneic (H-2d) tumor cells. The observed tumor cell rejection in SCID mice was dependent on natural killer cells. After depletion of asialo-GM1+ cells, all inoculated tumor cell lines grew progressively, independently of MHC class I expression. These results are compatible with a model where expression of certain, but not all, class I molecules protect from natural killer cell-mediated rejection. There was no evidence for rejection occurring as a consequence of the expression of allogeneic or xenogeneic class I molecules on the grafted cells. MHC class I expression may thus influence tumor cell recognition in mice lacking T-cell receptor expression.

Animals↗
PubMed1995

Heat-inactivated Sendai virus can enter multiple MHC class I processing pathways and generate cytotoxic T lymphocyte responses in vivo.

We have earlier described an alternative MHC class I processing pathway for Sendai virus (SV) in H-2Kb-transfected T2 cells (T2Kb). These cells have deleted genes for transporters associated with Ag processing (TAP1/2) and proteasome subunits LMP2/7 but can still process SV for the presentation of an immunodominant nucleoprotein CTL epitope (nucleoprotein peptide 324-332, FAPGNYPAL, SV9), even in the presence of the fungal metabolite brefeldin A (BFA). Presently we have compared live and heat-inactivated SV to investigate whether infectious virus, including early events such as binding and fusion at the host cell membrane, is important for nonclassical MHC class I processing and immunogenicity. We have found that heated virus (56 degrees C, boiled or autoclaved) with no fusion and hemagglutinin-neuraminidase activities, behaves similar to live SV in T2kb cells by entering a TAP-independent and BFA-resistant pathway. In EL-4 cells, which do not express this nonclassical TAP-independent and BFA-resistant pathway, heat-treated SV is processed in a BFA-sensitive way. In T1Kb- and TAP1/2-transfected T2Kb cells, as in T2Kb cells, processing of heat-inactivated SV was completely BFA resistant. Heat-inactivated SV was also found to prime CTLs in vivo. We conclude that heat-inactivated SV can enter both BFA-sensitive and -resistant MHC class I processing pathways and that SV in this respect may be particularly efficient. What property in the SV that is important for this characteristic is presently not clear but might be useful for the deliberate generation of CTL responses in vivo.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1995

Peptide influences the folding and intracellular transport of free major histocompatibility complex class I heavy chains.

Class I major histocompatibility complex molecules require both beta 2-microglobulin (beta 2m) and peptide for efficient intracellular transport. With the exception of H-2Db and Ld, class I heavy chains have not been detectable at the surface of cells lacking beta 2m. We show that properly conformed class I heavy chains can be detected in a terminally glycosylated form indicative of cell surface expression in H-2b, H-2d, and H-2s beta 2m-/- concanavalin A (Con A)-stimulated splenocytes incubated at reduced temperature. Furthermore, we demonstrate the presence of Kb molecules at the surface of beta 2m-/- cells cultured at 37 degrees C. The mode of assembly of class I molecules encompasses two major pathways: binding of peptide to preformed "empty" heterodimers, and binding of peptide to free heavy chains, followed by recruitment of beta 2m. In support of the existence of the latter pathway, we provide evidence for a role of peptide in intracellular transport of free class I heavy chains, through analysis of Con A-stimulated splenocytes from transporter associated with antigen processing 1 (TAP1)-/-, beta 2m-/-, and double-mutant TAP1/beta 2m-/- mice.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1995

Differential reactivity of residual CD8+ T lymphocytes in TAP1 and beta 2-microglobulin mutant mice.

TAP1 -/- and beta 2-microglobulin (beta 2m) -/- mice (H-2b background) express very low levels of major histocompatibility complex (MHC) class I molecules on the cell surface. Consequently these mice have low numbers of mature CD8+ T lymphocytes. However, TAP1 -/- mice have significantly higher numbers of CD8+ T cells than beta 2m -/- mice. Alloreactive CD8+ cytotoxic T lymphocyte (CTL) responses were also stronger in TAP1 -/- mice than in beta 2m -/- mice. Alloreactive CTL generated in TAP1 -/- and beta 2m -/- mice cross-react with H-2b-expressing cells. Surprisingly, such cross-reactivity was stronger with alloreactive CTL from beta 2m -/- mice than with similar cells from TAP1 -/- mice. The beta 2m -/- mice also responded more strongly when primed with and tested against cells expressing normal levels of H-2b MHC class I molecules. Such H-2b-reactive CD8+ CTL from beta 2m -/- mice but not from TAP1 -/- mice also reacted with TAP1 -/- and TAP2-deficient RMA-S cells. In contrast, H-2b-reactive CD8+ CTL from neither beta 2m -/- mice nor TAP1 -/- mice killed beta 2m -/- cells. In line with these results, beta 2m -/- mice also responded when primed and tested against TAP1 -/- cells. We conclude that the reactivity of residual CD8+ T cells differs between TAP1 -/- and beta 2m -/- mice. The MHC class I-deficient phenotype of TAP1 -/- and beta 2m -/- mice is not equivalent: class I expression differs between the two mouse lines with regard to quality as well as quantity. We propose that the differences observed in numbers of CD8+ T cells, their ability to react with alloantigens and their cross-reactivity with normal H-2b class I are caused by differences in the expression of MHC class I ligands on selecting cells in the thymus.

ATP Binding Cassette Transporter, Subfamily B, Mem↗