摘要:

AbstractThe effect of the guanosine triphosphate‐binding protein (G‐protein) inhibitors cholera toxin (Ctx) and pertussis toxin (Ptx) has been analyzed on lymphocyte function‐associated antigen 1 (LFA‐1)‐dependent adhesion and signal transduction in human natural killer (NK) cells. Ctx, but not Ptx, inhibited the LFA‐1‐dependent adhesion of NK cells to tumor target cells which constitutively express the intercellular cell adhesion molecule‐1 (ICAM‐1) and to NIH/3T3 mouse fibroblasts stably transfected with human ICAM‐1. This effect was detectable only by the use of the entire Ctx but not of the Ctx B subunit. In addition, Ctx could inhibit both NK cell binding and spreading to purified ICAM‐1 protein. NK cell treatment with Ctx modified neither the surface expression of LFA‐1 nor its Mg2+binding site. These findings, together with the absence of any detectable effect of Ctx on the constitutive phosphorylation of LFA‐1α, suggests that this toxin modifies the avidity of LFA‐1 for ICAM‐1 by acting on LFA‐1‐cytoskeletal protein association. Unlike Ctx, Ptx did not affect NK cell adhesion. The effects of Ctx and Ptx are unlikely to depend on intracellular levels of cyclic adenosine 3′,5′‐monophosphate (cAMP), since a strong increase of cAMP was induced by both toxins. Moreover, this was confirmed by the observation that the LFA‐1‐dependent adhesion was not inhibited by the adenylate cyclase activator forskolin (FSK), the phosphodiesterase inhibitor isobutyl‐1‐methylxantine (IBMX), or both, which increase intracellular cAMP levels. Unlike the differential effect on cell adhesion, both the intracellular calcium [Ca2+]iincrease and phosphoinositide breakdown mediated via LFA‐1 were consistently inhibited in a dose‐dependent manner by both Ctx and Ptx. Also in this case, the inhibitory effect did not depend on an increase of intracellular cAMP as indicated by NK cell treatment with FSK, IBMX, or both. Further evidence of the involvement of G‐proteins in LFA‐1‐mediated signal transduction was the inhibitory effect of the GDP analog guanosine‐5′‐O‐2‐thiodiphosphate (GDPßS) on LFA‐1‐mediated calcium mobilization. Taken together, our data provide evidence that the LFA‐1‐mediated NK cell adhesion and signal transduction are p

ISSN:0014-2980    

DOI:10.1002/eji.1830260502

出版商:WILEY‐VCH Verlag GmbH    

年代:1996

数据来源: WILEY

摘要:

AbstractIn a previous study, we identified T cell receptor and major histocompatibility complex (MHC) contact sites on the pigeon cytochromecp43‐58 peptide. Positions 46 and 54 of p43‐58 were shown to be the MHC‐binding sites. Specific amino acids were identified on the MHC‐binding sites which bound to the relevant I‐A molecule. In the present study, using NOD (I‐Ag7) mice, we established a T cell hybridoma, NOE33‐1‐2, specific for a p43‐58 analog 46R50E54A with arginine (R) and alanine (A) at positions 46 and 54, respectively. Interestingly, NOE 33‐1‐2 recognized 46R50E54A in the presence of not only I‐Ag7, but also I‐Ad, s, u and v. In contrast to previous reports that promiscuous T cells were able to recognize peptide antigens with various HLA‐DR or I‐E molecules consist of monomorphic α and polymorphic β chains, the promiscuous T cell clone NOE33‐1‐2 recognized peptides with various I‐A mo

ISSN:0014-2980    

DOI:10.1002/eji.1830260503

出版商:WILEY‐VCH Verlag GmbH    

年代:1996

数据来源: WILEY

摘要:

AbstractIn the Lewis rat, myelin basic protein (MBP)‐specific, encephalitogenic T cells preferentially recognize sequence 68–88, and use the Vβ8.2 gene to encode their T cell receptors. To analyze the structural prerequisites for the development of the MBP‐specific T cell repertoire, we reconstituted severe‐combined immunodeficient (SCID) mice with fetal (embryonic day 15–16) Lewis rat lymphoid tissue, and then isolated MBP‐specific T cell lines from the adult chimeras after immunization. Two types of chimera were constructed: SCID mice reconstituted with rat fetal liver cells only, allowing T cell maturation within a chimeric SCID thymus consisting of mouse thymic epithelium and rat interdigitating dendritic cells, and SCID mice reconstituted with rat fetal liver cells and rat fetal thymus grafts, allowing T cell maturation within the chimeric SCID and the intact Lewis rat thymic microenvironment. Without exception, the T cell lines isolated from MBP‐immunized SCID chimeras were restricted by MHC class II of the Lewis rat (RT1.B1), and none by I‐Adof the SCID mouse. Most of the T cell lines recognized the immunodominant MBP epitope 68–88. In striking contrast to intact Lewis rats, in SCID mice reconstituted by rat fetal liver only, MBP‐specific T cell clones used a seemingly random repertoire of Vβ genes without a bias for Vβ8.2. In chimeras containing fetal Lewis liver plus fetal thymus grafted under the kidney capsule, however, dominant utilization of Vβ8.2 was restored. The migration of liver‐derived stem cells through rat thymus grafts was documented by combining fetal tissues from wild‐type and transgenic Lewis rats. The results confirm that the recognition of the immunodominant epitope 68–88 by MBP‐specific encephalitogenic T cells is a genetically determined feature of the Lewis rat T cell repertoire. They further suggest that the formation of the repertoire requires T cell differentiation in a sy

ISSN:0014-2980    

DOI:10.1002/eji.1830260504

出版商:WILEY‐VCH Verlag GmbH    

年代:1996

数据来源: WILEY

摘要:

AbstractFas is a cell surface molecule that is expressed on a wide array of cell types and triggers apoptosis. While in most situations Fas ligation activates programmed cell death, on resting T lymphocytes it can co‐stimulate proliferation with the T cell receptor (TCR)/CD3 complex. This incongruity suggests that Fas may elicit signaling events that overlap with those used by proliferation cues. We observe that in the human T cell line Jurkat and in human peripheral blood lymphocytes, Fas stimulation does not signal by the Ras/Raf‐1/mitogen‐activated protein kinase (MAPK) pathway or by increased intracellular calcium. Rather, Fas ligation strongly activates Jun kinase (JNK). This activity, as well as Fas‐induced apoptosis, is blocked by increased levels of cAMP. The balance between proliferation and apoptosis by Fas triggering of T lymphocytes may therefore reflect a signaling ratio between TCR activation of the Ras/Raf‐1/MAPK pathway versus JNK activati

ISSN:0014-2980    

DOI:10.1002/eji.1830260505

出版商:WILEY‐VCH Verlag GmbH    

年代:1996

数据来源: WILEY

摘要:

AbstractInterleukin‐12 (IL‐12) is a heterodimeric cytokine that induces interferon (IFN)‐γ production and an increased generation of Th1 cells. Both IL‐12 and IL‐12 antagonists are being studied for the treatment of allergic reactions, autoimmune disease and malignancy. The goal of the present experiments was to examine the importance of IL‐12 in endotoxin‐induced ocular inflammation. The number of inflammatory cells infiltrating eyes with endotoxin‐induced uveitis (EIU) was significantly increased in animals treated with intraperitoneal anti‐IL‐12 antibody when compared to control animals, but there was no difference in infiltrating inflammatory cells in the eyes of animals treated with IL‐12 when compared to controls. In contrast, intraocular injection of IL‐12 significantly inhibited the development of endotoxin‐induced intraocular inflammation. The infiltrating inflammatory cells were reduced in the eyes of animals receiving intraocular IL‐12 when compared to controls. Cytokine analysis of the aqueous humor obtained from eyes with EIU showed increased levels of IFN‐γ and decreased levels of IL‐6 in eyes receiving intraocular IL‐12. These data show that IL‐12 has an inhibitory effect on endotoxin‐induced inflammation in the eye and suggest that IL‐12 can have an immunoregulatory funct

ISSN:0014-2980    

DOI:10.1002/eji.1830260506

出版商:WILEY‐VCH Verlag GmbH    

年代:1996

数据来源: WILEY

摘要:

AbstractThe superantigen (SAg) expressed by mouse mammary tumor virus (MMTV) has been shown to play an essential role in the course of the viral life cycle. In the present study, we describe a Vβ4‐specific SAg encoded by a new exogenous MMTV carried by the SIM mouse strain. This is the first report of a viral or bacterial SAg reacting with mouse Vβ4+T cells. Injection of MMTV(SIM) into adult BALB/c mice leads to a rapid and strong stimulation of Vβ4+CD4+T cells, followed by a slow deletion of these cells. Neonatal exposure to the virus also leads to a progressive deletion of Vβ4+T cells. In contrast to other strong MMTV SAg, this new SAg requires the presence of major histocompatibility complex class II I‐E molecules to be presented efficiently to T cells. Sequence analysis revealed a new predicted amino acid sequence in the C‐terminal polymorphic region of this SAg. Furthermore, sequence comparisons to the most closely related SAg with different Vβ specificities hint at the specific residues involved in the interaction with the T cel

ISSN:0014-2980    

DOI:10.1002/eji.1830260507

出版商:WILEY‐VCH Verlag GmbH    

年代:1996

数据来源: WILEY

摘要:

AbstractCD27, which belongs to the tumor necrosis‐factor receptor family, is expressed on germinal center (GC) but not on naive B cells, suggesting an important function of this molecule in the regulation of the GC reaction. We described here the expression of CD70, which is the ligand for CD27. We observed that in most tonsils, CD70 is only expressed on part of the IgD−, CD38−B cell population, which have been described as memory B cells. However, in 10 % of the tonsils tested, CD70+IgD+GC were found. The CD70+GC B cells were small cells that also expressed CD44 and CD39, but were CD10−and CD38−, suggesting that they represent very recent immigrants that are in the process of forming a GC. The concordant expression of CD27 and its ligand CD70 on this primordial subset of GC B cells suggests an important role for CD27/CD70 interaction at this stage of GC

ISSN:0014-2980    

DOI:10.1002/eji.1830260508

出版商:WILEY‐VCH Verlag GmbH    

年代:1996

数据来源: WILEY

摘要:

AbstractClonal selection of T cells mediated through the T cell antigen receptor (TCR) mostly occurs at the CD4+CD8+double positive thymocyte stage. Immature CD4+CD8+thymocytes expressing self‐reactive TCR are induced to die upon clonotypic engagement of TCR by self antigens. CD3 engagement by antibody of the surface TCR‐CD3 complex is known to induce apoptosis of CD4+CD8+thymocytes, a process that is generally thought to represent antigen‐induced negative selection in the thymus. The present study shows that the CD3‐induced apoptosis of CD4+CD8+thymocytes can occur even in TCRα−mutant mice which do not express the TCRαβ/CD3 antigen receptor. Anti‐CD3 antibody induces death of CD4+CD8+thymocytes in TCRα−mice either in cell cultures or upon administrationin vivo. Interestingly, most surface CD3 chains expressed on CD4+CD8+thymocytes from TCRα−mice are not associated with clonotypic TCR chains, including TCRβ. Thus, apoptosis of CD4+CD8+thymocytes appear to be induced through the CD3 complex even in the absence of clonotypic antigen receptor chains. These results shed light on previously unknown functions of the clonotype‐independent CD3 complex expressed on CD4+CD8+thymocytes, and suggest its function as an apoptotic receptor inducing elimination o

ISSN:0014-2980    

DOI:10.1002/eji.1830260509

出版商:WILEY‐VCH Verlag GmbH    

年代:1996

数据来源: WILEY

摘要:

AbstractRecent studies demonstrated that administration of a p55‐tumor necrosis factor (TNF) receptor IgG‐fusion protein (TNFR‐IgG) prevented the clinical onset of experimental autoimmune encephalomyelitis but did not alter the number or tissue distribution of autoantigen‐specific CD4+effector T cells which trafficked into the central nervous system. To determine whether specific target tissues of autoimmune damage remain intact after TNFR‐IgG treatment despite the presence of inflammatory cells within the tissues, we examined rats with experimental autoimmune uveoretinitis (EAU), as in this model, the main target of autoreactive CD4+T cells, the retinal rod outer segments (ROS), can be examined readily by light microscopy. As judged by direct ophthalmoscopy, the onset of inflammation in the anterior chamber of the eye in EAU following administration of TNFR‐IgG was delayed by 6 days compared to untreated controls, but the magnitude of the response was only slightly less than controls. Histological examination of the retinae and direct assessment of retinal inflammation revealed a disproportionate sparing of ROS in the TNFR‐IgG‐treated animals despite a level of retinal inflammation not substantially less than controls in which ROS damage was marked. Analysis of retinal leukocytes by immunofluo‐rescence microscopy and flow cytometry indicated that approximately equal numbers of CD4+αβTCR+lymphocytes were present in treated and control retinae, more than 30% of CD4+cells in both experimental groups expressed the CD25 or MRC OX40 activation markers and most cells, which would include the CD4+T lymphocytes, were activated as evidenced by MHC class II expression. Fewer activated macrophages and granulocytes were present in the treated retinae, possibly reflecting the lower level of tissue damage and subsequent accumulation of these inflammatory cells. The results demonstrate directly that a tissue specifically targeted for autoimmune destruction can be protected despite the influx of fully ac

ISSN:0014-2980    

DOI:10.1002/eji.1830260510

出版商:WILEY‐VCH Verlag GmbH    

年代:1996

数据来源: WILEY

摘要:

AbstractWe have utilized synthetic ribozymes to modulate the lipopolysaccharide (LPS)‐induced production of tumor necrosis factor‐α (TNF‐α) by peritoneal cells. Two hammerhead ribozymes (mRz1 and mRz2) were prepared by transcriptionin vitroand their activitiesin vitroandin vivowere investigated. Both ribozymes cleaved their RNA target with an apparent turnover number (kcat) of 2 min−1, and inhibited TNF‐α gene expressionin vitroby 50% and 70%, respectively. When mRz1 and mRz2, entrapped in liposomes, were delivered into mice by intraperitoneal injection, they inhibited LPS‐induced TNF‐α gene expressionin vivowith mRz2 being the most effective. This enhanced activity could result from the facilitation of catalysis by cellular endogenous proteins, since they specifically bind to mRz2 as compared to mRz1. Furthermore, a significant mRz2 activity can be recovered from peritoneal cells 2 days post‐administrationin vivo. The anti‐TNF‐α ribozyme treatmentin vivoresulted in a more significant reduction of LPS‐induced IFN‐γ protein secretion compared to IL‐10. In contrast to this pleiotropic effect, the anti‐TNF‐α ribozyme treatment did not affect the heterogenous expression of Fas ligand by peritoneal cells, indicating the specificity of the treatment. Taken together, the present data indicate that the biological effects of TNF‐α can be modulated by ribozymes. In addition, the data suggest that ribozymes can be administered in a drug‐like manner, and therefore indicate

ISSN:0014-2980    

DOI:10.1002/eji.1830260511

出版商:WILEY‐VCH Verlag GmbH    

年代:1996

数据来源: WILEY