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T Kalland

Publications and source records attributed to T Kalland.

At least 37 records · Page 2Linked to original sources
PubMed1997

T-cell stimulation and cytokine release induced by staphylococcal enterotoxin A (SEA) and the SEAD227A mutant.

Previous work demonstrated that human cytotoxic T cells activated by superantigens can lyse major histocompatibility complex (MHC) class II-positive target cells as well as MHC class II-negative tumour cells coated with conjugates of monoclonal antibodies and superantigens. In order to decrease MHC class II affinity, and therefore unwanted binding of the superantigen staphylococcal enterotoxin A (SEA) to MHC class II molecules, a point mutation was introduced into the SEA gene. This mutation (SEAD227A) resulted in an approximately 3-log reduction of affinity to human leucocyte antigen (HLA)-DR, but cytotoxicity mediated by this mutant superantigen towards antibody-labelled tumour cells is as efficient as cytotoxicity mediated by the native superantigen. We therefore compared the T-cell activating potency of native and mutated SEA. Our data show that SEAD227A is 4- to 5-log less effective than native SEA when activation of resting T cells is assayed in terms of blast formation, expression of cell surface activation markers and cytokine release. Furthermore, presenting either SEA or SEAD227A to MHC class II-negative mononuclear cells by MHC class II-negative tumour cells did not result in significant blast formation of T cells, up-regulation of CD25 or cytokine release. This suggests that lysis of MHC class II-negative tumour cells is efficiently induced by monoclonal antibody targeted superantigen, while activation of resting T cells requires additional co-stimulatory signals.

Cell Culture Techniques↗
PubMed1997

The tumour associated cell surface antigen A6H is costimulatory for human CD4+ but not CD8+ T cells.

The A6H monoclonal antibody (mAb) recognizes a 120,000-140,000 MW antigen that is expressed at similar densities on 85-90% of human CD4+ and CD8+ T cells and on renal cell carcinomas. The binding of the A6H mAb induced a costimulatory signal in anti-CD3 activated T cells. In the present report, we show that A6H costimulated cell proliferation and cytokine production in purified CD4+ T cells. Unexpectedly, the CD8+ T-cell subpopulation failed to respond. CD4+ T cells costimulated with the A6H mAb upregulated CD80, CD86, CD71, interleukin-2 (IL-2)R alpha, IL-2R beta and IL-2R gamma, while no corresponding up-regulation of these cell surface molecules was seen in CD8+ T cells. In order to investigate the nature of the A6H mAb costimulus at the transcriptional level we have examined induction of the transcription factors OCT-1, AP-1 and NF-kappa B which are known to be transcriptional regulators of several cytokine and cytokine receptor genes, including the IL-2 and IL-2R genes. Co-ligation of the A6H antigen and the CD3 complex induced expression of the transcription factor AP-1 in CD4+ T cells, whereas no increase in NF-kappa B and octamer-binding (Oct) proteins was seen compared to T cells stimulated with anti-CD3 alone. Furthermore, no induction of AP-1 was seen in A6H costimulated CD8+ T cells. These results suggests that both proximal steps in CD8+ T-cell activation as well as the later phases are unresponsive to A6H ligation. Molecular differences of the A6H molecule or distinct regulation of the A6H transduced AP-1 activation pathway may exist in CD4+ and CD8+ T cell subpopulations.

Antibodies, Monoclonal↗
PubMed1997

Superantigen-induced lysis of melanoma cells.

Superantigens like the Staphylococcus enterotoxin A (SEA) can direct cytotoxic T lymphocytes expressing certain T cell receptor V beta regions to lyse MHC class II-positive target cells. This superantigen-dependent cellular cytotoxicity (SDCC) has been extended to MHC class II-negative tumour cells by targeting T cells via conjugates of a tumour-specific monoclonal antibody (moAb) and a superantigen. In the present study the MHC class II-negative human melanoma cell lines G361 and MaRI were tested for susceptibility to SDCC in vitro. Antibodies recognizing the disialoganglioside GD3 and the CD10 antigen were linked to SEA either by a recombinant protein A-SEA fusion protein or an anti-kappa moAb-SEA chemical conjugate. Specific lysis of melanoma cells was dose- and effector to target (E:T) cell ratio-dependent. Introduction of a point mutation into the SEA gene (producing SEAm9) in order to reduce MHC II affinity of the superantigen, which has already been shown to severely diminish superantigen-dependent binding and lysis of MHC class II-positive cells, did not influence antibody-targeted SDCC. Cytotoxicity was equal with both antibodies (anti-GD3 and anti-CD10) and independent of whether protein A-SEA, protein A-SEAm9 or anti-kappa-SEA were used.

Antigens, Neoplasm↗
PubMed1997

A mutation of F47 to A in staphylococcus enterotoxin A activates the T-cell receptor Vbeta repertoire in vivo.

The bacterial superantigen staphylococcal enterotoxin A (SEA) binds with high affinity to major histocompatibility complex (MHC) class II molecules and subsequently activates T cells bearing particular T-cell receptor (TCR) Vbeta chains. Structural and mutational studies have defined two distinct MHC class II binding sites located in the N-terminal and C-terminal domains of SEA. The N-terminal F47 amino acid is critically involved in a low-affinity interaction to the MHC class II alpha-chain, while the C-terminal residues H187, H225, and D227 coordinate a Zn2+ ion and bind with moderate affinity to the beta-chain. In order to analyze whether the SEA-MHC class II alpha-chain interaction plays a role in dictating the in vivo repertoire of T-cell subsets, we studied distinct Vbeta populations after stimulation with wild-type SEA [SEA(wt)] and SEA with an F47A mutation [SEA(F47A)]. Injections of SEA(wt) in C57BL/6 mice induced cytokine release in serum, strong cytotoxic T-lymphocyte activity, expansion of T-cell subsets, and modulated expression of the T-cell activation antigens CD25, CD11a, CD44, CD62L, and CD69. SEA-reactive TCR Vbeta3+ and Vbeta11+ T cells were activated, while TCR Vbeta8+ T cells remained unaffected. The SEA(F47A) mutant protein induced a weaker T-cell response and failed to induce substantial interleukin-6 production compared to SEA(wt). Notably, SEA(F47A) failed to activate TCR Vbeta11+ T cells, whereas in vivo expansion and modulation of T-cell activation markers on TCR Vbeta3+ T cells were similar to those for SEA(wt). A similar response to SEA(F47A) was seen among CD4+ and CD8+ T cells. Activation of TCR Vbeta3+ and TCR Vbeta11+ T-cell hybridomas confirmed that SEA(F47A) activates TCR Vbeta3+ but not TCR Vbeta11+ T cells. The data support the view that the SEA-N-terminal MHC class II alpha-chain interaction defines a topology that is required for engagement of certain TCR Vbeta chains in vivo.

Animals↗
PubMed1997

Costimulation by B7-1 and LFA-3 targets distinct nuclear factors that bind to the interleukin-2 promoter: B7-1 negatively regulates LFA-3-induced NF-AT DNA binding.

We have characterized the regulation of nuclear factors involved in transcriptional control of the interleukin-2 (IL-2) promoter-enhancer activity in Jurkat T cells stimulated with superantigen presented on HLA-DR transfectants combined with the ligands LFA-3 (CD58) and B7-1 (CD80). Gel shift analyses showed that NF-AT was strongly induced in LFA-3-costimulated Jurkat T cells, suggesting that NF-AT is a key target nuclear factor for the CD2-LFA-3 pathway. Studies using HLA-DR-B7-1-LFA-3 triple transfectants showed that the LFA-3-induced NF-AT DNA binding activity was negatively regulated by B7-1 costimulation. In contrast, induction of a CD28 response complex containing only c-Rel proteins was seen after B7-1 costimulation. Both LFA-3 costimulation and B7-1 costimulation induced the AP-1 and NF-kappaB nuclear factors. Distinct compositions of the NF-AT complexes were seen in B7-1- and LFA-3-costimulated cells. LFA-3 induced primarily Jun-D, Fra-1, and Fra-2, while B7-1 induced June-D-Fos complexes. In contrast, AP-1 and NF-kappaB complexes induced in B7-1- and LFA-3-costimulated T cells showed similar contents. Transient transfection of Jurkat T cells with a construct encoding the IL-2 enhancer-promoter region (position -500 to +60) linked to a luciferase reporter gene revealed that B7-1 costimulation was required to induce strong transcriptional activity. Combined B7-1-LFA-3 costimulation resulted in a synergistic increase in IL-2 transcriptional activity. Multimers of the AP-1, NF-AT, NF-kappaB, and CD28 response elements showed distinct kinetics and activity after LFA-3 and B7-1 costimulation and revealed that B7-1 and LFA-3 converge to superinduce transcriptional activity of the AP-1, NF-AT, and CD28 response elements. Transcriptional studies with an IL-2 enhancer-promoter carrying a mutation in the CD28 response element site revealed that the activity was reduced by 80% after B7-1 and B7-1-LFA-3 costimulation whereas the transcriptional activity induced by LFA-3 was unaffected. Our data strongly suggest a selectivity in induction of nuclear factors by the CD2-LFA-3 and CD28-B7-1 pathways. This selectivity may contribute to regulation of the levels of IL-2 induced by LFA-3 and B7-1 costimulation and favor autocrine and paracrine T-cell responses, respectively.

B7-1 Antigen↗
PubMed1997

Superantigen-based immunotherapy: a phase I trial of PNU-214565, a monoclonal antibody-staphylococcal enterotoxin A recombinant fusion protein, in advanced pancreatic and colorectal cancer.

PURPOSE: To establish the maximum-tolerated dose (MTD) and define the toxicities of a single-dose infusion of PNU-214565, a recombinant Escherichia coli-derived fusion protein of Staphylococcal enterotoxin A (SEA) and the Fab-fragment of the C242 monoclonal antibody in patients with advanced colorectal and pancreatic carcinomas. To investigate the capability of PNU-214565 to induce a superantigen (SAg) response resulting in cytokine production and tumor regression. PATIENTS AND METHODS: Twenty-one patients (age range, 39 to 76 years; median, 64; 12 men, nine women; 18 colorectal, three pancreatic cancers) were treated with a single 3-hour infusion of PNU-214565, with doses ranging from 0.01 to 1.5 ng/kg. All patients had prior chemotherapy and a good performance status Eastern Cooperative Oncology Group [ECOG] performance status [PS] = 0 [n = 10]; PS = 1 [n = 11]), 10 had prior radiation, and 18 had prior surgery. RESULTS: Fever and hypotension were the most common toxicities. Fever of any grade occurred in 16 of 21 patients (76%): four of 21 (19%) with grade 2 and two of 21 (9.5%) with grade 3. Hypotension of any grade occurred in 13 of 21 (62%): four of 21 with grade 2 and one of 21 (5%) with grade 3. Interleukin-2 (IL-2) and tumor necrosis factor alpha (TNF alpha) induction correlated with toxicity. In the two patients with grade 3 fever, peak IL-2 and TNF alpha levels were 2.9 IU/mL and 165 pg/mL, and 8.3 IU/mL and 245 pg/mL, respectively. Transient, > or = 50% decreases in circulating monocytes were observed in 17 of 21 patients as early as 0.5 hours (median time, 2 hours) from the start of infusion. Decreases (mean 33%) in circulating lymphocytes were observed in seven of 21 patients. All three patients with grade 3 toxicity were treated at the 0.5-ng/kg dose. The significance of baseline anti-SEA, human antimouse antibody (HAMA), CA242-soluble antigen levels, and T-cell receptor variable beta region (TCR V beta) subsets and histocompatibility leukocyte antigen-DR (HLA-DR) genotypes was assessed as possible predictors of toxicity. All toxicities were transient and easily managed. No grade 3 toxicity occurred at the higher dose levels. CONCLUSION: PNU-214565, a SAg-based tumor targeted therapy, is safe when given as a single 3-hour infusion at doses up to 1.5 ng/kg. The MTD for a single dose was not determined. The safety of a repeated dose schedule is currently under investigation, beginning with doses determined to be safe in this trial.

Adult↗
PubMed1997

Tumor therapy with an antibody-targeted superantigen generates a dichotomy between local and systemic immune responses.

Repeated injections of a fusion protein containing the superantigen staphylococcal enterotoxin A (SEA) combined with a Fab fragment of a tumor-specific antibody is a highly efficient immunotherapy for mice expressing lung melanoma micrometastasis. In the present study, the systemic and local immune responses generated by this therapy were analyzed at a cellular level. Two distinct but coupled immune reactions occurred after repeated therapy. Tumor necrosis factor and macrophage inflammatory protein-1 alpha and -1 beta were immediately synthesized, in the absence of T lymphocytes, at the local tumor site in the lung. This was followed by the induction of VCAM-1 adhesion molecule expression on pulmonary vascular endothelial cells. Concurrently, the early response in the spleen was characterized by the induction of selective T cells producing interleukin (IL)-2. The primed and expanded SEA-reactive V beta 3- and V beta 11-expressing T lymphocytes accumulated to the tumor area only after Fab-SEA therapy and were not present in the lung when SEA, Fab fragment, or recombinant IL-2 was injected. The tumor-infiltrating T cells produced large amounts of interferon-gamma, but no IL-2 or Th2 type of lymphokines were detected at the tumor site in the Fab-SEA-targeted antitumor immune response. These results emphasize the necessity to investigate several sites of antigen presentation to elucidate the effects of immunotherapy.

Animals↗
PubMed1996

Mutations in the MHC class II binding domains of staphylococcal enterotoxin A differentially affect T cell receptor Vbeta specificity.

C-terminal residues of staphylococcal enterotoxin A (SEA), including H187, D225, and D227, are involved in moderate affinity binding to MHC class II beta-chain, whereas N-terminal residues, including F47, are involved in low affinity binding to MHC class II alpha-chain. The effect of alanine substitutions at residues D227 or F47 on induction of T cell proliferation and the expansion of specific TCR Vbeta families was determined. SEA wild type specifically activated T cells expressing Vbeta1, Vbeta5.2, Vbeta6, Vbeta7, Vbeta9, Vbeta18, and Vbeta22. Although SEA-D227A exhibited substantially reduced mitogenicity compared with SEA wild type, it expanded the same Vbeta-bearing T cells, except those expressing Vbeta1. By contrast, SEA-F47A, which was slightly less mitogenic than SEA wild type, induced expansion only of T cells expressing Vbeta6, Vbeta7, and to a lesser extent Vbeta22. Therefore, specific mutations affecting either MHC class II alpha or beta binding sites differentially affect the Vbeta specificity of this superantigen. The lack of expansion in four of seven Vbeta families by SEA-F47A suggests that the class II alpha binding site may position SEA on the MHC class II molecules in an appropriate conformation for interaction with certain Vbeta elements.

Antigens, Bacterial↗
PubMed1996

Regulation of superantigen-induced T cell activation in the absence and the presence of MHC class II.

To study MHC class II-dependent and -independent SAg2 activation and the relative importance of CD80/CD28 costimulation, staphylococcal enterotoxin A (SEA) was presented to T cells as a fusion protein containing the Fab fragment of an mAb directed against the CA215 glycoprotein. Chinese hamster ovary (CHO) cells transfected with HLA-DR4, CA215, and CD80, individually or in combinations, were used as presenting cells. A strong T cell proliferation was obtained when C215Fab-SEA fusion proteins were presented by CHO-DR/CD80 or CHO-CA215/CD80 double transfectants, whereas only low levels of proliferation were seen in the absence of CD80. Large amounts of IL-2, IFN-gamma, and TNF were produced in addition to an increase in IL-2 mRNA as a result of CD80 costimulation. Only approximately 50% of the SEA-reactive T cells responded by expression of IL-2 receptor chains and by blast formation when activated with SEA in the absence of MHC class II. Reverse transcription-PCR-assisted repertoire analysis of SEA-reactive TCR V beta families showed that the CA215-dependent activation involved an expansion of fewer TCR V beta families compared with MHC class II-dependent activation. One-half of the six analyzed TCR V beta families were expanded independently of class II. This indicates that MHC class II has only a partial influence on the TCR V beta repertoire imprinted by SAg. This finding redefines the role of MHC class II in SAg presentation. It is suggested that MHC class II molecules are selected as SAg-binding molecules mainly as a suitable targeting receptor for professional APC expressing costimulatory molecules such as CD80 and CD86.

Animals↗
PubMed1996

Immune response during tumor therapy with antibody-superantigen fusion proteins.

To engineer superantigens (SAg) to express tumor reactivity, we genetically fused the Fab-part of the tumor-reactive MAb C215 and the bacterial SAg staphylococcal enterotoxin A (SEA). Treatment of mice carrying established lung micrometastases of the C215-transfected syngeneic B16 melanoma with 3-4 daily injections of C215Fab-SEA resulted in strong antitumor effects, while only moderate effects were seen when treatment was given every 4th day (intermittent treatment). High serum levels of IL-2, TNF-alpha, IFN-gamma and strong induction of CTLs (cytotoxic T lymphocytes) were noted after priming with the fusion protein. T cells responded well to 3 daily injections of C215Fab-SEA and then gradually entered a hyporesponsive state, characterized by a reduced ability to produce IL-2, TNF-alpha and IFN-gamma and failure to mediate cytotoxicity in vitro. Intermittent treatment was characterized by increased levels of IL-10, concomitant with accentuated loss of IL-2, TNF-alpha and IFN-gamma production. A 10-fold increase in SEA-reactive TCR V(beta)3+ CD4+ cells was observed in the spleen, while a loss of TCR V(beta)3+ CD8+ and V(beta)11+ CD8+ cells was noted. This is in striking contrast to injections of native SEA which induced a marked deletion of TCR V(beta)3+ CD4+ T cells, but not of CD8+ cells. Recovery of the TH1 cytokine profile occurred within 1-2 weeks, while restoration of cytotoxicity required several months and correlated with recovery of TCR V(beta)3+ CD8+ and TCR V(beta)11+ CD8+ T cells. These results show that the temporal relationship of SAg stimulations dictates the cytokine profile. Moreover, different mechanisms appear to regulate hyporesponsiveness in CD4+ and CD8+ T cells.

Animals↗
PubMed1996

Fusion of a signal sequence to the interleukin-1 beta gene directs the protein from cytoplasmic accumulation to extracellular release.

Interleukin (IL)-1 differs from most other cytokines by the lack of a signal sequence, which results in the retention of the immature proform intracellularly (i.c.). Several cell types have the capacity to produce IL-1, but release has been shown to be restricted predominantly to monocytes/macrophages and associated with apoptosis of the producer cell. These features have limited the studies on IL-1 in early T cell-APC interactions. To develop a model for studying the biological effects of IL-1 beta release during long-lasting immune responses, we have established cells transfected with IL-1 beta cDNA constructs. To construct a hybrid gene for IL-1 beta release, the signal sequence from the related IL-1 receptor antagonist was fused to the gene encoding the 17-kDa mature form of IL-1 beta. A murine fibroblast cell line was transduced with retroviral technique and analyzed for the expression of human IL-1 beta, with or without a signal sequence (ssIL-1 beta and IL-1 beta, respectively). The fibroblasts transduced with either IL-1 beta or ssIL-1 beta expressed similar levels of human IL-1 beta mRNA. High levels of IL-1 bioactivity were recorded in freeze-thaw extracts from cells expressing the IL-1 beta protein i.c., and in supernatants of ssIL-1 beta-transduced cells, which indicates that the initial formation of a proform of IL-1 beta is not required for correct folding of the protein. Treatment of ssIL-1 beta-transduced cells with Brefeldin A (BFA), an inhibitor of protein transport in the endoplasmatic reticulum, induced accumulation of the protein i.c. BFA treatment did not affect IL-1 beta-transduced cells, while lipopolysaccharide-activated human monocytes increased the secretion of IL-1 beta. Cytoplasmic staining of single cells demonstrated that expression of the ssIL-1 beta gene directed the protein to a perinuclear Golgi-like compartment, whereas cells transduced with IL-1 beta cDNA showed a diffuse cytoplasmic distribution pattern. Secretion of IL-1 beta from human monocytes was under certain conditions accompanied by cell death. In contrast, in the fibroblast cell line transduced to secrete IL-1 beta, no accompanying cell death could be detected. Gene targeting of IL-1 to the secretory or cytoplasmic pathway may be useful for elucidating the role of IL-1 in T cell-APC interactions, avoiding cell death of the producer cells.

3T3 Cells↗
PubMed1996

In vivo anergized CD4+ T cells express perturbed AP-1 and NF-kappa B transcription factors.

Anergy is a major mechanism to ensure antigen-specific tolerance in T lymphocytes in the adult. In vivo, anergy has mainly been studied at the cellular level. In this study, we used the T-cell-activating superantigen staphylococcal enterotoxin A (SEA) to investigate molecular mechanisms of T-lymphocyte anergy in vivo. Injection of SEA to adult mice activates CD4+ T cells expressing certain T-cell receptor (TCR) variable region beta-chain families and induces strong and rapid production of interleukin 2 (IL-2). In contrast, repeated injections of SEA cause CD4+ T-cell deletion and anergy in the remaining CD4+ T cells, characterized by reduced expression of IL-2 at mRNA and protein levels. We analyzed expression of AP-1, NF-kappa B, NF-AT, and octamer binding transcription factors, which are known to be involved in the regulation of IL-2 gene promoter activity. Large amounts of AP-1 and NF-kappa B and significant quantities of NF-AT were induced in SEA-activated CD4+ spleen T cells, whereas Oct-1 and Oct-2 DNA binding activity was similar in both resting and activated T cells. In contrast, anergic CD4+ T cells contained severely reduced levels of AP-1 and Fos/Jun-containing NF-AT complexes but expressed significant amounts of NF-kappa B and Oct binding proteins after SEA stimulation. Resolution of the NF-kappa B complex demonstrated predominant expression of p50-p65 heterodimers in activated CD4+ T cells, while anergic cells mainly expressed the transcriptionally inactive p50 homodimer. These alterations of transcription factors are likely to be responsible for repression of IL-2 in anergic T cells.

Adult↗
PubMed1996

Antibody-targeted superantigen therapy induces tumor-infiltrating lymphocytes, excessive cytokine production, and apoptosis in human colon carcinoma.

Bacterial superantigens are the most potent known activators of human T lymphocytes. To engineer superantigens for immunotherapy of human colon carcinoma, the superantigen, staphylococcal enterotoxin A (SEA) was genetically fused to the Fab region of the colon carcinoma-reactive monoclonal antibody C242. In the present study the effector mechanisms involved in the anti-tumor response to C242 Fab-SEA were characterized. Immunohistochemistry and computer-aided image analysis were used in studies of cryopreserved tumor tissue to evaluate the phenotype of infiltrating cells and their cytokine profiles in response to therapy. Human T cells and monocytes were recruited to the tumor area and penetrated the entire tumor mass within hours after injection of C242 Fab-SEA. The production of cytokines at the single-cell level was found to be dominated by tumor necrosis factor (TNF)-alpha, interleukin (IL)-2, IL-4, IL-5, IL-10, IL-12, interferon (IFN)-gamma, granulocyte-macrophage colony-stimulating factor, and transforming growth factor-beta, whereas IL-1-alpha, IL-1ra, IL-1 beta, TNF-beta, IL-3, IL-6, and IL-8 were undetectable. Most of the TNF-alpha, IL-2, IL-12, and IFN-gamma were made by the infiltrating human leukocytes, while the colon carcinoma cells were induced to produce IL-4, IL-10, and TNF-alpha. Up-regulation of IFN-gamma receptors and TNF R p60 receptors was found, while the TNF R p80 receptor was absent. The cytokine production, T cell infiltration, and CD95 Fas receptor expression concomitantly occurred to induce programmed cell death in the tumor cells. This was followed by a strong reduction of the tumor mass that was seen within 24 h after C242 Fab-SEA infusion. These findings demonstrate that antibody-superantigen proteins efficiently recruit tumor-infiltrating lymphocytes actively producing a variety of cytokines likely to be essential for the therapeutic effects observed in the model. Although the humanized SCID model has obvious limitations in its predictive value for treatment of human cancer, we believe that these results encourage clinical evaluation of antibody-targeted superantigens.

Animals↗
PubMed1996

Superantigen-mediated cellular cytotoxicity is dependent on antigen expression, but independent of the P-glycoprotein multidrug resistance phenotype.

Superantigen-activated T cells can be targeted by monoclonal antibodies (mAb) to lyse MHC class II negative tumour cells. In this study we determined the susceptibility of the T-lymphoblastoid leukaemic cell line CCRF-CEM and its multidrug resistant sublines CCRF VCR100, CCRF VCR1000 and CCRF ADR5000 to lysis by monoclonal antibody-targeted and superantigen-activated T cells (superantigen-dependent cellular cytotoxicity, SDCC). A recombinant fusion protein of protein A and the superantigen Staphylococcus enterotoxin A (SEA) was used together with the mAbs anti-CD7, anti-CD38, anti-CD45RA and 4E3 (anti-P-glycoprotein) to correlate susceptibility to SDCC with expression of the MDR1-gene product. Our results demonstrated SDCC to be independent of MDR1-gene expression. This was further confirmed by blocking the function of Pgp in the leukaemic cell lines with a cyclosporine A derivative, which had no influence on SDCC. As expected, expression of the respective cell surface antigens on target cells had a strong impact on SDCC, although other factors seem to influence efficiency of SDCC as well.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1996

Linomide increases plasma corticosterone in normal rats, but does not prevent the inhibitory action of IL-1 on beta-cells in vivo or ex vivo.

Recently, the synthetic immunomodulator Linomide (quinoline-3-carboxamide, LS 2616) was reported to prevent IDDM and insulitis in NOD mice. The mechanism for this protective effect is not known. The cytokine interleukin 1 (IL-1) may be a pathogenetic factor in the initial destruction of the beta-cells leading to IDDM. This study was undertaken to investigate the influence of Linomide on IL-1beta induced diabetogenic and hormonal changes in the rat in vivo, and on IL-1beta mediated synthesis of NO and inhibition of insulin secretion in isolated islets of Langerhans ex vivo. Normal male Wistar Kyoto rats received 4.0 microg/kg of recombinant human IL-1beta (rhIL-1beta) i.p. daily for 5 days with or without Linomide (8-9 mg/kg/day) in the drinking water. Litters of neonatal Wistar rats were pretreated for 3 days with injections of 10 mg/kg of Linomide i.p., and pancreatic islets of Langerhans were isolated for ex vivo studies. Linomide alone caused significant hypercorticosteronemia, hypoglucagonemia, lymphopenia and neutrophilia. Linomide had no effect on IL-1beta induced hyperglycemia, hyperglucagonemia, lymphopenia, neutrocytosis, or hypercorticosteronemia on day three and hypocorticosteronemia on day five. Further, Linomide did not prevent rhIL-1beta mediated reduction in insulin secretion or increase in NO synthesis ex vivo. In conclusion, Linomide does not seem to exert its protective effect on IDDM development via inhibition of interleukin 1 action on islet insulin release or NO production, but the increase in plasma corticosterone may contribute to the understanding of the immunomodulatory effects of Linomide.

Adjuvants, Immunologic↗
PubMed1995

Cross-linking of major histocompatibility complex class II molecules by staphylococcal enterotoxin A superantigen is a requirement for inflammatory cytokine gene expression.

Staphylococcal enterotoxin A (SEA) has two distinct binding sites for major histocompatibility complex (MHC) class II molecules. The aspartic acid located at position 227 (D227) in the COOH terminus of SEA is one of the three residues involved in its interaction with the DR beta chain, whereas the phenylalanine 47 (F47) of the NH2 terminus is critical for its binding to the DR alpha chain. Upon interaction with MHC class II molecules, SEA triggers several cellular events leading to cytokine gene expression. In the present study, we have demonstrated that, contrary to wild-type SEA, stimulation of the THP1 monocytic cell line with SEA mutated at position 47 (SEAF47A) or at position 227 (SEAD227A) failed to induce interleukin 1 beta and tumor necrosis factor-alpha messenger RNA expression. Pretreatment of the cells with a 10-fold excess of either SEAF47A or SEAD227A prevented the increase in cytokine messenger RNA induced by wild-type SEA. However, cross-linking of SEAF47A or SEAD227A bound to MHC class II molecules with F(ab')2 anti-SEA mAb leads to cytokine gene expression, whereas cross-linking with F(ab) fragments had no effect. Taken together, these results indicate that cross-linking of two MHC class II molecules by one single SEA molecule is a requirement for cytokine gene expression.

Antigens, Bacterial↗