摘要:

AbstractBacterial 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)‐α, interleukin (IL)‐2, IL‐4, IL‐5, IL‐10, IL‐12, interferon (IFN)‐γ, granulocyte‐macrophage colony‐stimulating factor, and transforming growth factor‐β, whereas IL‐1‐α, IL‐1ra, IL‐1β, TNF‐β, IL‐3, IL‐6, and IL‐8 were undetectable. Most of the TNF‐α, IL‐2, IL‐12, and IFN‐γ were made by the infiltrating human leukocytes, while the colon carcinoma cells were induced to produce IL‐4, IL‐10, and TNF‐α. Up‐regulation of IFN‐γ 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 th

ISSN:0014-2980    

DOI:10.1002/eji.1830260102

出版商:WILEY‐VCH Verlag GmbH    

年代:1996

数据来源: WILEY

摘要:

AbstractB cell development is dependent on both direct interactions with stromal cells and their secreted cytokines. The precise mechanisms by which these interactions regulate B cell differentiation are currently unknown. We report here that a novel growth factor thymic stromal‐derived lymphopoietin (TSLP) can replace the activity of interleukin‐7 (IL‐7) in supporting B cell developmentin vitro.TSLP was found to promote the proliferation and differentiation of committed B220+B cell progenitors from day 15 fetal liver. Phenotypic analysis of these cells revealed that they are at the pro‐B cell stage of differentiation and express cell surface markers characteristic of pro‐B cells cultured in IL‐7. TSLP can replace the activity of IL‐7 in supporting the progression of B lymphocytes from uncommitted bipotential precursors. In the absence of either TSLP or IL‐7, the progeny of cells that give rise to mature B lymphocytes fail to develop from these bipotential precursors. Moreover, TSLP can substitute for IL‐7 in supporting the sustained proliferative response exhibited by B cell progenitors from CBA/N mice. Together these results show that TSLP can replace the requirement for IL‐7 duringin vitr

ISSN:0014-2980    

DOI:10.1002/eji.1830260103

出版商:WILEY‐VCH Verlag GmbH    

年代:1996

数据来源: WILEY

摘要:

AbstractPeripheral lymphoid tissues contain a fibroblastic cell type referred to as stromal cells or reticulum cells which interact with lymphocytes as part of the lymphoid microenvironment. After isolation from human tonsils and expansionin vitrowe analyzed the surface phenotype, extracellular matrix components, cytoskeletal products, cytokine production, binding and functional interaction with B lymphocytes ofin vitrocultured stromal cells (HTSC) both in resting condition and after activation with tumor necrosis factor (TNF)‐α and interferon (IFN)‐γ. Our results show that HTSC do not express specific myeloid, lymphoid, endothelial or epithelial markers. HTSC express CD54 (ICAM‐1), CD49a (VLA‐1), CD49b (VLA‐2), CD49c (VLA‐3), CD49e (VLA‐5), CD49f (VLA‐6), CD29, CD51, CD44 and produce vinculin, β‐tubulin, α‐actin, vimentin, fibronectin, laminin and collagen types I, III and IV. Activation of HTSC up‐regulated CD54 (ICAM‐1) and induced HLA‐DR and CD106 (VCAM‐1).HTSC constitutively produce interleukin (IL)‐6 which is enhanced upon activation with TNF‐α. IL‐8 and granulocyte/macrophage colony‐stimulating factor are detected only in the supernatants of activated HTSC. Reverse transcriptase polymerase chain reaction analysis revealed that HTSC display mRNA for IL‐1α, leukemia inhibitory factor and IL‐7. The adhesion of tonsillar B lymphocytes to activated HTSC is mediated by CD11a/CD18 and CD54. Furthermore, HTSC can induce maximal proliferation of IL‐2‐activated B lymphocytes cocultured in direct cell‐cell contact with HTSC. These results clearly distinguishin vitrocultured HTSC from common fibroblasts and other non‐lymphoid elements present in the lymphoid parenchyma, such as follicular dendritic cells, and show that HTSC actively participate in the lymphoid microenvironment.In vitrocultures of HTSC could therefore be a useful model system for detailed analysis of the interactions between stromal cells and lympho

ISSN:0014-2980    

DOI:10.1002/eji.1830260104

出版商:WILEY‐VCH Verlag GmbH    

年代:1996

数据来源: WILEY

摘要:

AbstractTransgenic mice in which mouse interleukin (IL)‐7 cDNA is expressed under the control of the mouse major histocompatibility complex (MHC) class II (Eα) promoter develop a lymphoproliferative disease characterized by the early polyclonal expansion of T cells followed in many cases by the development of lymphomas of immature B cells. Here, we have analyzed B cell development in these transgenic mice. Phenotypic analysis using monoclonal antibodies to B220, IgM, IgD,c‐kit, IL‐7 receptor, MHC class II, AA4.1, CD19, CD23, CD25, CD40 and CD43 shows that B lymphopoiesis in the bone marrow is dramatically altered and the number of pro/pre‐B and immature B cells is significantly increased. Interestingly, pro/pre‐B and immature B cells persist in the spleens of adult transgenic mice and are also present in lymph nodes and blood. Cell cycle analysis of lymph node cells shows that subpopulations of developing B cells retain the cell cycle profiles of their bone marrow counterparts. Limiting dilution analysis shows that the number of clonable pre‐B cells is significantly increased and that at limiting dilution, growth of transgenic pre‐B cells is still dependent on exogenous IL‐7. Using semiquantitative polymerase chain reaction (PCR) andin situhybridization, the level of IL‐7 transcripts in the spleen was found to decrease between 2 and 4 weeks in control mice with levels in transgenics mice being approximately 50 times greater. These transgenic mice represent an interesting model with which to study the effects of IL‐7 overexpression in the bonemarrow and raise interesting questions regarding the regulation of B lymphopo

ISSN:0014-2980    

DOI:10.1002/eji.1830260105

出版商:WILEY‐VCH Verlag GmbH    

年代:1996

数据来源: WILEY

摘要:

AbstractCD28 is a 44‐kDa homodimer present on T cells providing an important costimulatory signal for T cell proliferation, cytokine production and cytokine receptor expression. CD28 activation is mediated by interaction with its counter‐receptors, B7.1/CD80 and B7.2/B70/CD86. The biochemical basis of these costimulatory signals are still poorly understood, particularly in resting T cells. However, various biochemical pathways such as tyrosine phosphorylation, phospholipase C, sphingomyelinase and phosphatidylinositol 3‐kinase (PI3‐K) activation have been reported to play a role in CD28 signaling in tumor T cell lines and CD28‐transfected cells or pre‐activated T cells. In addition, recent reports propose that CD28‐B7.1 and B7.2 interaction could be involved in the production of Th1 and Th2 cytokines, respectively, but the putative biochemical basis for these different functions is still unknown. We have analyzed the functional and molecular consequences of CD28 activation by B7.1 and B7.2 in human resting T cells. We demonstrate in this report that both CD28‐B7.1 and CD28‐B7.2 interactions induce the association of PI3‐K to CD28 in the CD4 subpopulation, whereas it was barely detectable in CD8 cells. This association involves the binding of the src homology domain 2 (SH2) of p85 to tyrosine‐phosphorylated CD28 and does not require pre‐activation by CD3‐T cell receptor. Worthmannin, a specific inhibitor of PI3‐K enzymatic activity within the nanomolar range also inhibits the interleukin‐2 production induced by costimulation mediated by either the B7.1‐ and B7.2‐transfected cells or CD28 monoclonal antibodies. The only slight difference between B7.1 and B7.2 costimulation is the IC50of wortmannin being 25 and 110 nM, respectively, which could suggest differences in the

ISSN:0014-2980    

DOI:10.1002/eji.1830260106

出版商:WILEY‐VCH Verlag GmbH    

年代:1996

数据来源: WILEY

摘要:

AbstractPenicillin G (Pen G) and other beta‐lactam antibiotics frequently induce allergic reactions constituting typical examples of human immune responses to haptens. In fact, penicillins represent a unique set of haptens with outstanding structural variability on the basis of an identical protein‐reactive beta‐lactam containing backbone. Although both cellular and humoral responses are involved in drug‐induced allergies, little is known about the T cell reactivity to penicillins. To understand which structural features determine antigenic specificity, we isolated a panel of MHC‐restricted, Pen G‐reactive T cell clones from different penicillin‐allergic patients and tested them for their capacity to proliferate in the presence of other penicillin derivatives. We found that the antigenic epitope consists of both the amide‐linked side chain, which is different in every member of the penicillin family, as well as the thiazolidine ring common to all penicillin derivatives. We also demonstrated the presence of two different types of penicillin‐specific T cells, one dependent, and the other independent of antigen processing by autologous antigen‐presenting cells. Our data strongly suggest that penicillins form part of the epitopes contacting the antigen r

ISSN:0014-2980    

DOI:10.1002/eji.1830260107

出版商:WILEY‐VCH Verlag GmbH    

年代:1996

数据来源: WILEY

摘要:

AbstractInterleukin (IL)‐4, IL‐10 and IL‐13 are cytokines with potent anti‐inflammatory activities. Prevention of pathological inflammation at mucosal surfaces appears to be due, in part, to the presence of these cytokines. One potential source for these cytokines is the mast cell which resides at mucosal surfaces. Demonstrated in this report are the findings that bone marrow‐derived mucosal‐like mast cells constitutively expressed IL‐13 whereas bone marrow‐derived connective tissue‐like mast cells demonstrated IL‐13 transcription only after FcεRI‐mediated activation or the addition of exogenous IL‐3. A similar pattern of expression of IL‐10 by these mast cell types was also evident and matches that of IL‐4 previously reported. Intracellular cytokine staining indicated that IL‐10 protein is constitutively expressed by the bone marrow‐derived mucosal‐like mast cells but is only evident in the bone marrow‐derived connective tissue‐like mast cells after induction with IL‐3. The increase of IL‐13 and IL‐10 transcripts in the connective tissue‐like mast cells following IL‐3 treatment is not mast cell specific, in that splenic and bone marrow cells also demonstrated the same phenomenon. These data suggest that mucosal mast cells may have a constitutive repertoire of Th2 cytokines with potential anti‐inflammatory activity, while connective tissue mast cells may not. However, production of such cytokines can be induced in the connective tissue mast cell and other cell ty

ISSN:0014-2980    

DOI:10.1002/eji.1830260108

出版商:WILEY‐VCH Verlag GmbH    

年代:1996

数据来源: WILEY

摘要:

AbstractThe CD5 T cell glycoprotein which is expressed by a subset of B cells has been shown to be involved in T cell activation and proliferation. No similar studies, to date, have addressed the role of CD5 on the B cell subset. CD5+and CD5−B cells were sorted and stimulated with anti‐CD5 monoclonal antibody (mAb)in vitro.The activation and proliferative responses of these two populations, as measured by analysis of proliferation marker, did not differ following anti‐μ and interleukin (IL)‐2 stimulation. The addition of anti‐CD5 did not change the responsiveness of such activated CD5+B cells but resulted in a decrease in CD25 expression. Pre‐activation of B cells with phorbol 12‐myristate 13‐acetate, which increased CD5 expression, failed to alter the proliferative response of CD5+B cells to anti‐μ and IL‐2 with or without addition of anti‐CD5 mAb. Anti‐μ and IL‐2 treatment of CD5+cells resulted in optimal proliferation measured at day 3 which decreased by day 6. However, addition of anti‐CD5 mAb at day 3 prevented this decline in proliferative response. This dose‐dependent effect was observed only when the anti‐CD5 mAb was presented to the B cells in cross‐linked form. Co‐stimulation of CD5 did not lower the threshold of antigen to which the B cells responded. Taken together, these data support a functional role for CD5 on B cells acting as an accessory signal, following their primary activation through the B cell receptor complex and highlight differences in the role of CD5 assoc

ISSN:0014-2980    

DOI:10.1002/eji.1830260109

出版商:WILEY‐VCH Verlag GmbH    

年代:1996

数据来源: WILEY

摘要:

AbstractIn early steps of B cell differentiation, μ chains are transiently expressed in association with a surrogate light chain (ΨL) composed of the λ‐like and VpreB monomorphic polypeptides, thus forming a putative preB receptor. Using a monoclonal anti‐VpreB antibody, preB cells were isolated from two adult human bone marrow samples and their VDJ repertoire analyzed at the transcription level. All VH families were identified and further analysis focused on VH3 sequence analysis of 37 distinct VDJ cDNA clones. The VH3 genes expressed in the two bone marrow samples were also encountered in fetal liver and adult peripheral blood lymphocytes with a roughly similar contribution of 3.30, 3.23, 3.9 and 3.53. The characteristic features of the preB repertoire as compared to the activation B repertoire include the quasi absence of somatic mutations, limited N diversity and a shorter third complementarity‐determining region (CDR3). It also significantly differs from the fetal repertoire, which makes higher usage of DQ52 and has CDR3 of even shorter lengths. The almost constant presence of glycine residues in the CDR3 and predominance of JH4 with a low level of DQ52 DH usage, suggest that preB cell clones are submitted to an initial selective pressure which should be antigen independent. Thebona fideheavy chains would be merely selected for their ability to interact with the surrogate light chains, thus shaping the repertoire that will be co‐expressed with immunoglobulin light chains in IgM

ISSN:0014-2980    

DOI:10.1002/eji.1830260110

出版商:WILEY‐VCH Verlag GmbH    

年代:1996

数据来源: WILEY

摘要:

AbstractEosinophils synthesize and store various cytokines with potential autocrine activity. We hypothesized that eosinophils synthesize and store RANTES, a CC chemokine with potent eosinophil chemotactic activity. Expression of RANTES mRNA in highly purified eosinophil populations was detected by reverse transcription followed by polymerase chain reaction analysis.In situhybridization (ISH) with35S‐labeled RANTES‐specific riboprobes showed that 6.8–10% of peripheral blood eosinophils obtained from atopic subjects expressed RANTES mRNA, increasing to 25% after incubation (16 h) with interferon(IFN)‐γ, but not ionomycinin vitro.Peripheral blood eosinophils also showed specific immunoreactivity with an anti‐RANTES monoclonal antibody, consistent with translation of the mRNA. By enzyme‐linked immunosorbent assay, blood eosinophils were shown to contain a median of 7300 pg (range 5200–8800) RANTES per 106cells, of which a mean of 24% was released into culture supernatants after stimulation of the cells with serum‐coated particlesin vitro.These culture supernatants exhibited eosinophil chemotactic activity which was inhibited (mean 68%) by a specific anti‐RANTES antibody. Sequential immunocytochemistry and ISH on biopsies obtained from allergen‐induced late‐phase cutaneous reactions showed that 55–75% of the infiltrating RANTES mRNA+cells were EG2+eosinophils. Allergen, but not diluent challenge, was also associated with a time‐dependent increase in the number of cells showing RANTES immunoreactivity. Of these cells, 55% were identified as eosinophils by morphological criteria. Thus, human eosinophils have the capacity to synthesize, store and secrete physiologically relevant quantities of RANTES, and may therefore be an important source of this chemokin

ISSN:0014-2980    

DOI:10.1002/eji.1830260111

出版商:WILEY‐VCH Verlag GmbH    

年代:1996

数据来源: WILEY