bluuebunny/crossref_metadata_2025 huggingface dataset:
Preferential distribution of Vβ 8.2‐positive T cells in the central nervous system of rats with myelin basic protein‐induced autoimmune encephalomyelitis
Authors: Tsuchida Masanori , Matsumoto Yoh , Hirahara Hiroyuki , Hanawa Haruo , Tomiyama Katsuhiro , Abo Toru
Published: March 2007

To determine the role of encephalitogenic T cells in the formation of lesions in the central nervous system (CNS), experimental autoimmune encephalomyelitis (EAE) was induced in Lewis rats by immunization with either myelin basic protein (MBP) or the synthetic peptide which corresponds to the 87–100 sequence of guinea pig MBP, and T cells expressing T cell receptor (TcR) Vβ8.2, Vβ8.5, Vβ10 and Vβ16 in the lymphoid organs and CNS were localized and quantified by flow cytometry (FCM) and immunohistochemistry. In normal rats, the percentage of T cells expressing these Vβ phenotypes to the total number of TcR αβ+ T cells, as determined by FCM, ranged from 5% to 10% in the lymph node. Vβi6+ T cells were the most predominant population among the four Vβ subsets tested. Essentially the same findings were obtained from the analysis of the lymphoid organs of rats with EAE which had been induced by immunization with the same two antigens. In sharp contrast, 15–20% of the T cells isolated from lesions of MBP‐induced EAE expressed Vβ8.2+. Thus, the percentage of Vβ8.2+ T cells in the EAE lesions was threefold higher than that in the lymph node, while the proportions of Vβ8.5+, Vβ10+ and Vβ16+ T cells were about the same in both organs. The predominance of Vβ58.2+ T cells in EAE lesions was confirmed by counts of immunohistochemically stained T cells in the spinal cord. Moreover, it was revealed that (i) the predominance of Vβ8.2+ T cells was greatest during the development of EAE and became less obvious at the recovery stage, and (ii) at the peak stage of EAE, approximately 85% of Vβ8.2+ T cells were distributed in the parenchyma while 15% were in the perivascular space of the CNS vessels. These findings indicate that encephalitogenic T cells which express Vβ8.2 infiltrate the CNS at a very early stage of EAE and become the predominant population in infiltrating T cells, and further suggest that encephalitogenic T cells, not only recruit inflammatory cells in the CNS, but also cause neural tissue damage, such as demyelination.
Murine CD8+ T suppressors against mycobacterial 65‐kDa antigen compete for IL‐2 and show lack of major histocompatibility complex‐imposed restriction specificity in antigen recognition
Authors: Khetan Smita , Sainis Krishna Balaji , Rath Satyajit , Kamat Ramesh
Published: March 2007

The mechanism of antigen‐specific suppression and reasons for aberrant major histocompatibility complex (MHC) class II restriction mediated by CD8+ T cells was investigated in a previously reported murine model of immunosuppression, generated by intraperitoneal priming with Mycobacterium vaccae. Both the CD4+ T helper cells (Th) and CD8+ T supressor cell (Ts) of M.vaccae‐primed mice recognized the 65‐kDa antigen of the bacillus, presented by I‐A and I‐E, respectively. The CD8+ Ts could inhibit non‐antigen‐specific proliferation of primed CD4+ T cells induced by the exogenously added interleukin (IL)‐2 (concanvalin A‐stimulated culture supernatant). For inhibition, the Ts had to be activated by the 65‐kDa antigen. The degree of inhibition was dependent upon the amount of added IL‐2 and the relative numbers of primed CD8+ and CD4+ T cells. On incubation with antigen‐presenting cells, and the 65‐kDa antigen, the primed CD8+ T cells absorbed IL‐2 as efficiently as primed CD4+ T cells. Based on this, it was concluded that the primed CD8+ T cells induced suppression by competition for IL‐2.

Employing the same model, the MHC restriction of recognition of the suppressor epitope of the 65‐kDa antigen by the CD8+ Ts was investigated. The epitopes presented by diverse MHC class II molecules, such as self I‐A, I‐E and even allogeneic I‐E were similar, because they were recognized by the same population of primed CD8+ Ts. Further, immunization of C57BL/6 mice with Ltk‐cells expressing H‐2 DkKk alloantigens, stimulated CD8+ T cells capable of recognizing M.vaccae 65‐kDa antigen. Based on these data, it was proposed that recognition of the suppressor epitope of the 65‐kDa antigen by the primed CD8+ Ts exhibits lack of restriction specificity imposed by MHC diversity.
Polymorphism of the functional immunoglobulin variable region genes in the chicken by exchange of sequence with donor pseudogenes
Authors: Benatar Tania , Ratcliffe Michael J. H.
Published: March 2007

We have isolated a number of new allelic variants of the unique functional genes encoding chicken immunoglobulin heavy and light chain variable regions (VH1 and VL1, respectively). The distribution and nature of nucleotide variation among these and previously identified VH1 and VL1 alleles demonstrates that random point mutations are likely not the predominant cause of allelic variation at these loci. Comparison of the variant nucleotides with sequences from the pseudo‐VH and pseudo‐VL gene families, which lie 5′ to VH1 and VL1, respectively, suggests that the great majority of allelic variants can be accounted for by segmental transfer of sequence from donor pseudogenes into the germ‐line VH1 and VL1 genes. These results demonstrate that the chicken VH1 and VL1 genes are susceptible to sequence replacement at the germ‐line level as well as somatically during antibody diversification. The limited repertoire of Bcell specificities produced by gene rearrangement in the chicken has led to speculation that these specificities may play a critical role in the progression of chicken B cell development. The results presented here do not support this hypothesis since many of the allelic variant nucleotides described here encode non‐conservative amino acid substitutions within the antigen‐binding sites of the Ig molecule.
CD45‐mediated regulation of LFA1 function in human natural killer cells. Anti‐CD45 monoclonal antibodies inhibit the calcium mobilization induced via LFA1 molecules
Authors: Poggi Alessandro , Pardi Ruggero , Pella Nicoletta , Morelli Luigia , Sivori Simona , Vitale Massimo , Revello Valentino , Moretta Alessandro , Moretta Lorenzo
Published: March 2007

The TA218 and T205 monoclonal antibodies (mAb) were selected on the basis of their ability to inhibit the non‐major histocompatibility complex‐restricted lysis of the murine mastocytoma P815 cell line mediated by CD3−CD16+ natural killer (NK) cells. Both mAb were found to react with CD45 molecules, as demonstrated by immunoprecipitation after surface iodination and western blot analysis. A panel of tumor target cells susceptible to lysis by polyclonal or clonal CD3−CD16+ NK cells was used to study the mAb‐mediated inhibitory effect. The inhibition of cytolysis mediated by TA218 and T205 mAb was found to consistently parralel the inhibition mediated (with the same tumor target cells) by the anti‐LFAlα mAb TS.1.22 or by the anti‐LFA1β mAb TS.1.18. However, different from the anti‐LFAl mAb, T205 or TA218 mAb did not inhibit the binding of activated CD3−CD16+ effector NK cells to the same tumor target cells. This finding supported the concept that the anti‐CD45 mAb‐mediated inhibition could occur at a post‐binding stage. In polyclonal or clonal CD3‐CD16+ NK cellsT205 orTA218 mAb were found to reduce by 50–70 % the intracellular Ca++ ([Ca++]i) mobilization induced by anti‐LFAlα or anti‐LFA1β mAb. On the other hand, TA218 and T205 mAb did not inhibit the Ca++ mobilization induced by anti‐CD 16 mAb or phytohemagglutinin, thus suggesting that, in NK cells, CD45 molecules may exert a selective inhibitory effect on the signal transduction mediated by LFA1 molecules. In line with this hypothesis, the cytolytic activity of human NK clones was triggered in the presence of the hybridoma cells secreting either anti‐CD16 or anti‐LFAla mAb (as “triggering targets”). This effect of anti‐LFAlα, but not of anti‐CD16 hybridoma was susceptible to inhibition by the anti‐CD45 mAb T205 or TA218. Further, experiments on cloned NK cells indicated that T205 or TA218 mAb induced a strong decrease in the constitutive phosphorylation of the LFAlα chain (but not of HLA class I antigens). Taken together, these studies suggest that in human NK lymphocytes, CD45 molecule may regulate both the activation state and the function of the LFA1 molecule.
Murine platelet endothelial cell adhesion molecule (PECAM‐1)/CD31 modulates β2 integrins on lymphokine‐activated killer cells
Authors: Piali Luca , Albelda Steven M. , Scott Baldwin H. , Hammel Philippe , Gisler Roland H. , Imhof Beat A.
Published: March 2007

Lymphokine‐activated killer (LAK) cells are able to colonize sites of tumor lesions in mouse and man. The molecular mechanisms of homing in on tumors are largely unknown. However, before LAK cells can reach the tumor, they must adhere to the vascular endothelia within the lesion and then extravasate. We developed a novel mAb, EA‐3, which recognizes the murine homologue of the human adhesion molecule CD31. It is present on a subpopulation of murine LAK cells and all endothelial cells. CD31 was also involved in the adhesion of LAK cells to endothelium. Since CD31 can initiate integrin activation by inside‐out signaling after binding to its ligand, EA‐3 was used to minimic this in adhesion assays. It induces modifications in the β2 integrin LFA‐1, leading to increased binding capacities of the cells to endothelium. In contrast, β1 integrins and RGD‐binding integrins were not affected. These results suggest that expression of CD31 might confer adhesive advantages for LAK cells prone to tumor infiltration.
Regulation of CD59 expression on the human endothelial cell line EA.hy 926
Authors: Meri Seppo , Mattila Pirkko , Renkonen Risto
Published: March 2007

CD59 (protectin) is an 18–20‐kDa inhibitor of the membrane attack complex of complement. It protects homologous cells from complement‐mediated damage and has been shown to be present on the endothelial cell membranes both in vitro and in vivo. In this study we observed that the surface expression of CD59 on the cultured EA.hy 926 endothelial cell line can be up‐regulated to an approximately threefold higher level after a 72‐h stimulation by the protein kinase C inducers phorbol‐12‐myristate‐13 acetate (PMA; 10 nM) and calcium ionophore, A23187 (100 nM). Similarly, an increase in the level of CD59 expression was seen by the protein kinase A inducer dibutyryl‐cyclic adenosine monophosphate. In Northern blot analysis increases were observed in CD59 mRNA expression, particularly in the level of the longest 1.9‐kb, 2.1‐kb and 5.8‐kb transcripts. A functional significance for the increased CD59 expression was implied by an observed increased resistance of the PMA‐stimulated EA.hy 926 cells to complement‐mediated cell lysis.
The association of the protein tyrosine kinases p56lck and p60fyn with the glycosyl phosphatidylinositol‐anchored proteins Thy‐1 and CD48 in rat thymocytes is dependent on the state of cellular activation
Authors: Garnett David , Neil Barclay A. , Carmo Alexandre M. , Beyers Albertus D.
Published: March 2007

Cell surface glycoproteins anchored to the plasma membrane via glycosyl‐phosphatidylinositol (GPI) structures, and hence having no cytoplasmic domains, can nevertheless transmit activation signals in lymphocytes. By immunoprecipitation from detergent lysates and in vitro immune complex kinase reactions the GPI‐anchored molecules Thy‐1 and CD48 are shown to be associated with multimolecular complexes of phosphoproteins including the protein tyrosine kinases p56lck: and p60fyn in both rat and mouse thymocytes. Moreover, the kinase activity associated with Thy‐1 on rat thymocytes is shown to be dependent on the activation state of the cells, with stimulation by the lectin, concanavalin A, producing a marked decrease in Thy‐1‐associated kinase activity. In such activated cells, there is an increased association of kinase activity with CD48, but this may be explained in terms of increased surface expression of CD48 and of increased total kinase activity. Additional phosphoproteins of 85, 36 and 32 kDa were consistently seen as components of the complexes.
Dynamic response of murine gut intraepithelial T cells after infection by the coccidian parasite Eimeria
Authors: Craig Findly R. , Roberts Scott J. , Hayday Adrian C.
Published: March 2007

The response of murine intraepithelial lymphocyte (IEL) populations to challenge by Eimeria vermiformis, a naturally occurring protozoan parasite of the gut epithelium, has been studied. The number of recoverable IEL increased within 3 days post infection, was depleted by day 7 post infection, but was significantly increased again by about day 14 post infection. Special attention was paid to γ δ+ IEL T cells, because they are of unknown functions. These cells showed changes in numbers similar to the total IEL population. Moreover, by day 3, increased expression was detected among γ δ + IEL T cells, of T cell receptor genes not constitutively associated with the intestine. These results demonstrate that the IEL repertoire, and within that, the γ δ + T cell repertoire, can be extremely dynamic post infection with a naturally occurring epithelial‐tropic pathogen. In considering the potential benefits of such IEL changes, we hypothesize that they may be relevant to the transient protection of the host's epithelium, both from parasitic re‐infection, and from potentially damaging inflammation.
Endogenous retroviral superantigen presentation by B cells induces the development of type 1 CD4+ T helper lymphocytes
Authors: Gollob Kenneth J. , Nagelkerken Lex , Coffman Robert L.
Published: March 2007

The endogenous retroviral superantigen, minor lymphocyte stimulating antigen (Mls 1a, encoded by Mtv‐7), when presented by highly purified B cells induced the development of a highly polarized population of T helper (Th)1 cells from naive peripheral CD4+ T cells in vitro. Immobilized anti‐Vβb6+ antibodies similarly generated highly polarized, largely Vβ6+, Th 1 populations in vitro. In the presence of exogenous interleukin‐4, both stimuli were capable of generating Th 2, rather than Th 1 populations. Mls 1a presentation by B cells in vivo led to the development of an equally polarized Th 1 population. Using monoclonal antibodies against interferon‐γ and transforming growth factor‐β it was demonstrated that maximal Th 1 development with either stimulus in vitro was dependent on the endogenous production of these two cytokines. Thus, our results demonstrate that the retroviral encoded superantigen, Mls 1a, drives the development of Th 1 cells both in vitro and in vivo, and they suggest that B cell presentation does not, in itself, lead to the generation of Th 2 cells.
Ligation of CD28 receptor by B7 induces formation of D‐3 phosphoinositides in T lymphocytes independently of T cell receptor/CD3 activation
Authors: Ward Stephen G. , Westwick John , Hall Nicolas D. , Sansom David M.
Published: March 2007

The co‐stimulatory role of B7/CD28 interactions is important in promoting T cell activation. Very little is known about the intracellular events that follow CD28 engagement although recent evidence has implicated coupling of CD28 to a protein tyrosine kinase signal transduction pathway. In this study we have investigated the putative role of D‐3 phosphoinositides as mediators of CD28 receptor signaling, since phosphoinositide (PI) 3‐kinase, the enzyme responsible for D‐3 phosphoinositide formation, is a known substrate for protein tyrosine kinases associated with certain T cell surface receptors such as CD4 and interleukin‐2 receptor. The lipid products of PI 3‐kinase activity have been suggested to play a role in mitogenic signaling and growth regulation in other cells. Chinese hamster ovary cells (CHO) previously transfected with B7 cDNA, induced time‐dependent elevation above basal levels of phosphatidylinositol(3,4)‐bisphosphate (PtdIns(3,4)P2) and PtdIns(3,4,5)P3, while parental CHO cells that did not express B7 had no effect on these lipids. Moreover, the elevation of these same lipids by CD3 ligation was potentiated in an additive manner by CHO‐B7+ but not by CHO‐B7− cells. CHO‐B7+ and CHO‐B7− cells did not activate phospholipase C as evidenced by their inability to modulate basal or CD3‐induced changes in the levels of phosphatidic acid or D‐4 and D‐5 phosphoinositides. These data imply that PI 3‐kinase but not phospholipase C, may be an important signal transduction molecule with respect to CD28‐mediated co‐stimulation and T cell activation following ligation by B7.
T cell development in a major histocompatibility complex class II‐deficient patient
Authors: Van Eggermond Marja C. J. A. , Rijkers Ger T. , Kuis Wietse , Zegers Ben J. M. , van den Elsen Peter J.
Published: March 2007

In this report we show that the major histocompatibility complex (MHC) class II‐negative thymus of a bare lymphocyte syndrome (BLS) patient contains a reduced CD4+ CD8− T cell population when compared to thymocytes derived from a MHC class II‐expressing thymus. Of these CD4+ CD8− BLS thymocytes, approximately only one third co‐expressed the CD3 antigen, moreover at a lower expression level when compared to control thymocytes. This suggests a partial maturation of the CD4+ CD8− T cells in the absence of MHC class II expression. Among the BLS thymocytes, CD4+ CD8+ thymocytes could easily be detected. Noteworthy, the number of CD4− CD8+ thymocytes was significantly increased. CD4+ CD8− T cells could also be found among the BLS peripheral blood mononuclear cells, albeit at reduced numbers. Despite the absence of peripheral MHC class II expression, the majority of these CD4+ CD8− T cells co‐expressed the CD45RO marker. In the BLS patient, thymocytes as well as peripheral CD4+ CD8− T cells were not restricted in the use of the available T cell receptor (TcR) V gene family pool. However, the lack of detectable levels of thymic and peripheral MHC class II antigen expression in the BLS patient had altered the CD4−skewing patterns of TcR V gene families which were present in normal individuals. In conclusion, the lack of MHC class II expression in the BLS patient does not completely inhibit the CD4+ CD8− T cell development.
Viral hemagglutinin augments peptide‐specific cytotoxic T cell responses
Authors: Ertel Christian , Millar Neil S. , Emmerson Peter T. , Schirrmacher Volker , Von Hoegen Paul
Published: March 2007

In attempt to increase the induction of peptide‐specific cytolytic T cells (CTL) we investigated the effect of the Newcastle disease virus (NDV) hemagglutinin‐neuraminidase (HN) gene product on the activation of peptide‐specific CTL. Spleen cells of CH3 mice immunized against the influenza nucleoprotein peptide 50–63 (NP 50–63) were restimulated in vitro (i) with peptide‐pulsed syngeneic fibroblast cells (Ltk−) as antigen‐presenting cells, which were in addition (ii) infected with NDV or (iii) stably transfected with the HN cDN A of NDV. A greater than sixfold increase in peptide‐specific CTL responses was observed in cultures restimulated with peptide‐pulsed Ltk− cells which co‐expressed viral hemagglutinin due to either infection or transfection.

A similar augmentation was seen in CTL responses against other types of antigen (major histocompatibility complex alloantigens, minor histocompatibility antigens or tumor antigens) when suboptimal cultures were stimulated with the respective antigen‐presenting cells modified by NDV infection. These findings suggest that NDV or viral HN expressed on antigen‐presenting cells or tumor cells can exert a T cell co‐stimulatory function.
An age‐related γδ T cell suppressor activity correlates with the outcome of autoimmunity in experimental Trypanosoma cruzi infection
Authors: Cardillo Fabíola , Falcao Roberto P. , Rossi Marcos A. , Mengel José
Published: March 2007

In this work the suppressive activity of splenic T cells from young and aged BALB/c mice infected with Trypanosoma cruzi were compared and correlated with the development of autoimmune myocarditis. The T cells from young adult BALB/c mice with acute T. cruzi infection exhibit suppressor activity when added to full allogeneic or Mis‐disparate mixed lymphocyte cultures. This suppression could not be reverted by exogenous interleukin (IL)‐2 and was not directly dependent on the presence of IL‐4, IL‐10 or transforming growth factor‐β. Further characterization of the T cell lineage responsible for the suppressor activity by in vitro and/or in vivo depletion with monoclonal antibody to αβ or γδ T cell receptor revealed that splenic γδ T cells function as suppressor lymphocytes in young T. cruzi‐infected mice. In addition, these young adult BALB/c mice do not develop autoimmune myocarditis and showed a low incidence of syngeneic heart graft rejection in the early chronic phase of the infection. In contrast, T cells from acutely infected aged BALB/c mice lacked demonstrable T suppressor activity. Furthermore, these mice developed a severe autoimmune myocarditis as early as 2 months after the onset of the infection, when the majority of them reject syngeneic heart grafts. These findings suggest that a γδ T cell‐mediated suppressor mechanism may operate in the avoidance of the breaking of tissue‐specific tolerance during the acute infection. Moreover, such a mechanism is likely related to the immune system chronobiology.
Molecular cloning of a ligand for the inducible T cell gene 4‐1BB: a member of an emerging family of cytokines with homology to tumor necrosis factor
Authors: Goodwin Raymond G. , Din Wenie S. , Davis‐Smith Terri , Anderson Dirk M. , Gimpel Steven D. , Sato Timothy A. , Maliszewski Charles R. , Brannan Camilynn I. , Copeland Neal G. , Jenkins Nancy A. , Farrah Terry , Armitage Richard J. , Fanslow William C. , Smith Craig A.
Published: March 2007

4‐1BB is an inducible T cell antigen that shows sequence homology to members of an emerging family of cytokine receptors, including those for tumor necrosis factor and nerve growth factor. To aid in the analysis of the function of 4‐1BB we have utilized a soluble form of the molecule as a probe to identify and clone the gene which encodes its ligand. The ligand for 4‐1BB is a type II membrane glycoprotein that has homology to tumor necrosis factor, lymphotoxin, and the ligands for CD40 and CD27, all of which are themselves ligands to receptors in this superfamily. The gene for 4‐IBB is on mouse chromosome 4 and maps close to the p80 form of the tumor necrosis factor receptor as well as the gene for CD30. The gene for 4‐IBB ligand maps to mouse chromosome 17, but considerably distal to the tumor necrosis factor and lymphotoxin genes. Interaction of 4‐1BB with its ligand induces the proliferation of activated thymocytes and splenic T cells, a response which is mimicked on similar cell populations stimulated with an antibody to 4‐1BB.
Clonal anergy to staphylococcal enterotoxin B in vivo: selective effects on T cell subsets and lymphokines
Authors: Baschieri Selene , Lees Rosemary K. , Lussow Alexander R. , Robson MacDonald H.
Published: March 2007

Injection of bacterial superantigens such as staphylococcal enterotoxin B (SEB) in adult mice results in initial proliferation of SEB‐responsive Vβ8+ T cells followed by induction of a state of non‐responsiveness frequently referred to as clonal anergy. We show here that SEB‐induced anergy involves selective changes in lymphokine production and that it affects CD4+ Vβ 8+ and CD8+ Vβ 8+ T cells in different fashions. Whereas both CD4+ Vβ 8+ and CD8+ Vβ 8+ cells from anergic mice exhibit strongly reduced proliferative capacity and interleukin(IL)‐2 production upon restimulation with SEB either in vivo or in vitro the CD8+ subset from SEB‐injected mice produces other lymphokines (such as interferon(IFN)‐γ) at normal or slightly increased levels in response to SEB. Changes in the levels of production of IL‐2 and IFN‐γ protein correlated well with mRNA accumulation both in vivo and in vitro. Collectively these data suggest that superantigen‐induced anergy involves selective changes in signal transduction and/or gene regulation in T lymphocytes.
Chimaeric anti‐CD4 monoclonal antibody cross‐linked by monocyte Fcγ receptor mediates apoptosis of human CD4 lymphocytes
Authors: Choy Ernest H. S. , Adjaye James , Forrest Lesley , Kingsley Gabrielle H. , Panayi Gabriel S.
Published: March 2007

Previous studies have shown that murine anti‐CD4 monoclonal antibody, cross‐linked by rabbit anti‐mouse immunoglobulin, could mediate apoptosis of murine CD4+ lymphocytes when they were stimulated by T cell receptor antibody. In this study, we have shown that the murine anti‐CD4 monoclonal antibody, OKT4, can induce apoptosis in human CD4+ T cells stimulated by the recall antigen tuberculin purified protein derivative (PPD) only when cross‐linked by rabbit anti‐mouse immunoglobulin. The chimeric anti‐CD4 monoclonal antibody, cM‐T412 whose Fc fragment is human, was able to cause apoptosis without cross‐linking by a second antibody. Similarly, abolition of PPD‐induced proliferation of peripheral blood mononuclear cells by cM‐T412 did not require cross‐linking with rabbit anti‐human immunoglobulin. Inhibition of proliferation by cM‐T412 could be reduced by pre‐treating monocytes with heat‐aggregated human IgG. This suggested that monocyte Fcγ receptors might be cross‐linking the human Fc of cM‐T412. Propidium iodide staining together with immunofluorescence showed that the apoptotic cells were indeed CD4+ lymphocytes. It is proposed that during treatment with cM‐T412 in autoimmune disease such as rheumatoid arthritis, cM‐T412‐coated CD4 T cells, when they are subsequently stimulated by the unknown arthritogenic antigen, may undergo apoptotic cell death through cross‐linking of cM‐T412 on Fey receptor‐positive cells within the joint.
Regional sublocalization of the human CD69 gene to chromosome bands 12p12.3‐p13.2, the predicted region of the human natural killer cell gene complex
Authors: Schnittger Susanne , Hamann Jörg , Dannenberg Christine , Fiebig Helmut , Strauss Michael , Fonatsch Christa
Published: March 2007

The early activation antigen CD69 is a member of a supergene family of type II integral membrane proteins with a C‐type lectin domain. In recent reports the genes encoding the natural killer (NK) cell‐related molecules of this supergene family, NKR‐P1, NK1.1 and Ly‐49, were shown to be clustered in a chromosomal region in mouse, termed the NK gene complex. The human homologue of this complex is likely to reside on chromosome 12 near the PRP locus (12p13.2). By analyzing T cell hybrids, the CD69 gene was previously mapped to human chromosome 12. Here we report the regional sublocalization of the human CD69 gene to chromosome bands 12p12.3‐p13.2, suggesting that CD69 belongs to one linkage group together with different cell surface molecules on NK cells.
Mouse intestinal epithelial cells express the self superantigen Mis1
Authors: Kaiserlian Dominique , Vidal Karine , Robson MacDonald H. , Grosjean Isabelle
Published: March 2007

In previous studies, we demonstrated that intestinal epithelial cells of the mouse small intestine could present exogenous antigen to specific CD4+ T cell hybridomas. We now report on the abililty of normal enterocytes to present the self superantigen M1sla. Enterocytes from M1sla but not from M1slb strains stimulated interleukin‐2 production through aVβ6+ T cell hybridoma specific for M1sla determinants. Antibody inhibition experiments showed that enterocytes presented M is determinants via a major histocompatibility complex class II‐dependent mechanism. Furthermore, the ability of enterocytes to activate Vβ6+ M1sla‐specific T cells was inhibited by monoclonal antibodies against the Orf protein encoded by an Mtv‐7 provirus which is associated with M1sla expression. These findings provide evidence for the first time that M is determinants are expressed on normal enterocytes and support the theory of a possible role of these cells in extrathymic selection of T cell receptor Vβ repertoire of intraepithelial T lymphocytes.
CD21 expressed on basophilic cells is involved in histamine release triggered by CD23 and anti‐CD21 antibodies
Authors: Bacon Kevin , Gauchat Jean‐Francois , Aubry Jean‐Pierre , Pochon Sibylle , Graber Pierre , Henchoz Sybille , Bonnefoy Jean‐Yves
Published: March 2007

Recombinant full‐length human CD23 incorporated into fluorescent liposomes was used to detect a ligand for CD23 on the basophilic leukemia cell line, KU 812. Based on our recent finding that CD23 interacts with CD21 on subsets of B and T cells, we investigated if the same ligand was involved on KU 812 cells. An anti‐CD21 monoclonal antibody (mAb) BU‐33, was able to totally block CD23‐liposome binding to KU 812 cells. Moreover, KU 812 cells express CD21 mRNA and have a cell surface molecule that reacts with anti‐CD21 mAb. The CD23/CD21 interaction was not merely physical but was also associated with an increase in histamine release by KU 812 cells. Both recombinant soluble CD23 and an anti‐CD21 mAb increased histamine release by KU 812 cells. The CD23 and anti‐CD21 mAb‐mediated effect on histamine release was not restricted to the leukemic cell line, but was also observed with normal human blood basophils. These data demonstrate that CD21 is expressed on basophilic cells and that CD21 controls histamine production upon ligand‐induced stimulation (CD23 or anti‐CD21 mAb).
Graft rejection by T cells not restricted by conventional major histocompatibility complex molecules
Authors: Dierich Andrée , Chan Susan H. , Benoist Christophe , Mathis Diane
Published: March 2007

The appropriate crosses of mice lacking conventional major histocompatibility complex (MHC) class I or class II molecules generate single‐ and double‐deficient offspring. These were used as donors for skin grafts across major plus minor, or just minor, histocompatibility differences. Surprisingly, in the two circumstances, there was a rapid rejection of grafts lacking both MHC class I and class II molecules. Rejection was mediated by thymically derived CD4+ T cells of the host. We provide evidence that these T cells recognize an unconventional ligand, capable of activating a pre‐formed T cell compartment but incapable of positively selecting it. The existence of this unexpected rejection phenomenon should serve to caution those aiming to engineer “universal donor” cells by simply abrogating expression of MHC class I and class II molecules.
Contrasting efficacy of presentation by major histocompatibility complex class I and class II products when peptides are administered within a common protein carrier, self immunoglobulin
Authors: Zaghouani Habib , Kuzu Yukiko , Kuzu Hiroshi , Brumeanu Teodor D. , Swiggard William J. , Steinman Ralph M. , Bona Constantin A.
Published: March 2007

Major histocompatibility complex (MHC) class I and II products are specialized to present antigens via different intracellular processing routes. Peptides originating from proteins in the cytoplasm can gain access to class I peptide‐binding grooves, most likely in the rough endoplasmic reticulum. Peptides from proteins in acidic endocytic vacuoles gain access to class II. It has been proposed that MHC class I products also can capture peptides from “exogenous” or noninfectious sources, and this assumption underlies the use of intact proteins as vaccines for CD8+ cytotoxic T lymphocytes. Here we describe quantitative information comparing the efficacy of peptide presentation from exogenous proteins by administering a class I‐ and II‐restricted peptide within the same context, the CDR3 loop of the VH domain of a self immunoglobulin. Antigen‐presenting cells (APC), including primary dendritic cells, efficiently present an influenza hemagglutinin peptide from the immunoglobulin (Ig) carrier (50% maximal response at 10 nM Ig‐HA) to an MHC class II‐restricted T cell. In contrast, these same APC are unable to present an influenza nucleoprotein (NP) peptide from the same context (1 μM Ig‐NP) to an MHC class I‐restricted T cell. Ig‐NP DNA transfectants do present the nucleoprotein viral peptide on class I. Thus, peptides within the complementarity‐determining region loops of Ig carriers can be presented on class I or II MHC products, but the endocytic compartment, when offered MHC class I‐ and II‐restricted peptides within the same carrier protein context, favors presentation by class II by at least 1000‐fold.
Identification of an endogenous mammary tumor virus involved in the clonal deletion of Vβ2 T cells
Authors: Jouvin‐Marche Evelyne , Marche Patrice N. , Six Adrien , Liebe‐Gris Carine , Voegtle Danielle , Cazenave Pierre‐André
Published: March 2007

Expression of Vβ (β ‐chain variable region) gene segments was investigated in the Mus m. domesticus DDO strain, which possesses a large genomic deletion encompassing 20 of the 29 Vβ gene segments known in BALB/c. Stainings using Vβ ‐specific monoclonal antibodies revealed that up to 60% of the peripheral T cells use 3 Vβ gene segments. Variable frequencies of Vβ 2 T cells were observed among DDO individuals. Segregation analyses of F2 crosses between Vβ 2‐deletor mice and mammary tumor virus (Mtv)‐free mice led to the identification of a new endogenous Mtv, named Mtv‐DDO, mediating Vβ 2 Tcell clonal deletion. Mtv‐DDO structure is conserved with the exception of the carboxy‐terminal region as compared to other Mtv. Comparison between Mtv sharing the same Vβ specificity and isolated from laboratory or wild mice confirms that a stretch of 11 amino acids, defined as the Vβ ‐specific region, is required for the Vβ ‐specific interaction. Limited substitutions in this region account for the shift of the Mtv specificity towards different Vβ.
A new infectious mammary tumor virus in the milk of mice implanted with C4 hyperplastic alveolar nodules
Authors: Shakhov Alexander N. , Wang Hong , Acha‐Orbea Hans , Pauley Robert J. , Wei Wei‐Zen
Published: March 2007

We have previously characterized an infectious mouse mammary tumor virus [(MMTV(SW)] which induces a strong superantigen response in vivo. Here we describe the isolation and characterization of MMTV(C4) which was derived from milk of mice implanted with hyperplastic alveolar nodules. MMTV(C4) stimulates Vβ2 expressing T cells after local injection in vivo. Comparison with known open reading frame (orf) sequences revealed high homology to Mtv‐6, an endogenous virus interacting with Vβ3‐expressing T cells. The carboxyl‐terminal amino acids were, however, altered. High homology including the carboxyl‐terminal orf amino acids were found with MMTV(C3H‐K). We show here that MMTV(C3H‐K) has lost its superantigen function. Sequence comparisons permitted the characterization of few key amino acids which could be important for T cell receptor interaction and superantigen processing.
Tumors with reduced expression of a cytotoxic T lymphocyte recognized antigen lack immunogenicity but retain sensitivity to lysis by cytotoxic T lymphocytes
Authors: Koeppen Hartmut , Acena Maria , Drolet Aimee , Rowley Donald A. , Schreiber Hans
Published: March 2007

A murine solid tumor was transfected to express various levels of an allogeneic major histocompatibility complex class I gene (K216), in order to test the effect of the level of antigen expression on immunogenicity and sensitivity to lysis by cytotoxic T lymphocytes (CTL). The growth rates of clones of tumor cells expressing different levels of the transfected gene were similar in vitro and in nude mice. Although all tumor cells, including cells freshly isolated from growing tumors, were equally sensitive to lysis by specific CTL, only tumor cells expressing the highest level of the K216 antigen stimulated CTL and were rejected by normal mice. In contrast, tumor cells expressing lower levels of antigen failed to immunize for CTL and grew progressively in normal mice, despite retaining expression of the transfected gene and remaining fully sensitive to CTL‐mediated lysis; thus, the threshold of antigen needed to stimulate CTL responses was considerably higher than that needed to lyse tumor cells. Reduction of K216 antigen expression from 100‐fold to 40‐fold above background, impaired significantly the ability of the tumor cells to induce a K216‐specific immune response, while tumor cells expressing K216 at levels 2‐fold above background were as susceptible to CTL‐mediated lysis as tumor cells expressing 50‐fold more antigen. The important implication of these findings is that some tumors occurring in nature may not be immunogenic but nevertheless express antigens which are potential targets for immune therapy.
Activation of human monocytes and granulocytes by monoclonal antibodies to glycosylphosphatidelinositol‐anchored antigens
Authors: Lund‐Johansen Fridtjof , Olweus Johanna , Symington Frank W. , Arrli Aanen , Thompson John S. , Vilella Ramon , Skubitz Keith , Horejsi Vaclav
Published: March 2007

The present study investigated possible receptor‐like characteristics of glycosyl‐phosphatidylinositol (GPI)‐linked antigens on human monocytes and granulocytes by measuring cytoplasmic calcium fluxes and the oxidative burst in cells following cross‐linking of GPI‐linked antigens. Cross‐linking of cell‐bound anti‐CD14, ‐CDw52 and ‐CD55 induced cytoplasmic calcium fluxes and oxidative bursts in unprimed human monocytes similar to those observed following FcyR cross‐linking. In granulocytes primed with 200 nM N‐formyl‐Met‐Leu‐Phe (FMLP), cross‐linking of cell‐bound anti‐CD16, ‐CD24, ‐CD59 and ‐CD67 led to calcium fluxes and activation of the oxidative burst. The oxidative bursts mediated by GPI‐linked antigens were stronger than those induced by 200 nM FMLP, even though FMLP induced a larger increase in cytoplasmic calcium concentration. The responses were likely to be independent of FcyR interactions as F(ab′)2 fragments of IgG or IgM antibodies were used in the experiments. Activating effects of monoclonal antibody to GPI‐linked antigens were not observed in cells from patients with paroxysmal nocturnal hemoglobinuria, which are deficient in GPI‐linked antigens. In addition, treatment with GPI‐specific phospholipase C led to inhibition of cell activation through GPI‐linked antigens but not through transmembrane receptors. Cross‐linking of a number of non‐GPI‐linked antigens (CD11a, CD18, CD31, CD35, CD43, and CD45) neither induced calcium fluxes, nor activated the oxidative burst. The results indicate that most, if not all, GPI‐linked surface glycoproteins on myeloid cells are capable of mediating cell activation and suggest that the GPI anchor is a structure facilitating signal transduction.
Somatic diversification and affinity maturation of IgM and IgG anti‐DNA antibodies in murine lupus
Authors: Hirose Sachiko , Wakiya Midori , Kawano‐Nishi Yasuko , Yi Jiang , Sanokawa Reiko , Taki Shinsuke , Shimamura Toshiroh , Kishimoto Toshihiko , Tsurui Hiromichi , Nishimura Hiroyuki , Shirai Toshikazu
Published: March 2007

Molecular events occurring during the process of generation of pathogenic immunoglobulin (Ig)G anti‐DNA antibodies in systemic lupus erythematosus (SLE) were studied using a newly established method. We analyzed the Ig variable (V) region gene sequence and DNA‐binding activity of IgM and IgG anti‐DNA monoclonal antibodies (mAb) from individual SLE‐prone (NZB × NZW) F1 mice. The first event appeared to be clonal selection and expansion of IgM anti‐DNA clones, in which several clones had intraclonal V gene mutations. Although the number of mutations was small, the mutated IgM clones were associated with an increase in DNA‐binding activity. The somatic mutations located in complementarity‐determining regions (CDR) and in framework regions (FR) of V genes were apparently related to changes in DNA‐binding activity. IgG anti‐DNA clones that progressively increased in number with aging had numerous somatic mutations in the V region genes and there was a pair of clones which showed an intraclonal accumulation of mutations, in association with increase in the DNA‐binding activity. All these findings show that somatic mutations associated with affinity maturation of the V region begin immediately before isotype‐switching from IgM to IgG of the clones that have been selected and expanded, in an antigen‐driven manner and/or by other forces. We propose that further accumulations of intraclonal somatic hypermutation, in association with selection and expansion of high affinity IgG clones, may lead to formation of highly pathogenic anti‐DNA antibodies.
The lack of CD8α cytoplasmic domain resulted in a dramatic decrease in efficiency in thymic maturation but only a moderate reduction in cytotoxic function of CD8+ T lymphocytes
Authors: Fung‐Leung Wai‐Ping , Louie Marjorie C. , Limmer Andreas , Ohashi Pamela S. , Ngo Karen , Chen Lei , Kawai Kazuhiro , Lacy Elizabeth , Loh Dennis Y. , Mak Tak W.
Published: March 2007

The glycoprotein CD8 is believed to play an important role in the maturation and function of MHC class I‐restricted T lymphocytes. CD8 has been proposed to function as a co‐receptor of the TcR to participate in signal transduction, possibly through its cytoplasmic domain that binds to protein tyrosine kinase p56lck. A T cell‐specific transgene encoding CD8α truncated at the cytoplasmic domain (“tailless CD8α”), was introduced into CD8α‐deficient mice. This animal model was used to study the role of the CD8 cytoplasmic domain in T cell ontogeny and function. “Tailless CD8α” was expressed on the cell surface of thymocytes and peripheral T cells. A small population of peripheral CD4− T cells (6% of T lymphocytes) was found to have cell surface expression of “tailless CD8α” and endogenous CD8β indicating that these cells may belong to the CD8+ T cell lineage. A consistent result was obtained from CD8α‐deficient mice bearing the “tailless CD8α” and the MHC class I‐restricted 2C TcR transgenes. A small population of CD4 T cells expressing CD8β the “tailless CD8α” and the 2C TcR transgenes was present in the periphery of these mice in a selecting background, but was absent in a deleting background. When “tailless CD8α” mice were infected with lymphocytic choriomeningitis virus (LCMV), the peripheral CD8+ CD4− T cell subset expanded dramatically and a significant LCMV‐specific cytolytic activity was detected. The results suggest that the cytoplasmic portion of CD8α is not absolutely required but dramatically enhances the eficiency of thymic maturation of CD8+ T cells. The lack of CD8α cytoplasmic domain in peripheral CD8+ T cells does not abolish the generation of cytotoxicity in response to an in vivo LCMV infection, although the cytolytic activity is slightly reduced compared to that in control mice.
The Vχ genes of the L regions and the repertoire of Vχ gene sequences in the human germ line
Authors: Huber Christian , Schäble Karlheinz F. , Huber Erwin , Klein Ralph , Meindl Alfons , Thiebe Rainer , Lamm Rosemarie , Zachau Hans G.
Published: March 2007

Only 14 of the 25 Vχ genes and pseudogenes had been found before as parts of the L regions. The cloning and linking described in the accompanying report allowed us now to assign to Lp or Ld some Vχ genes which had been found before on scattered clones. In addition the sequences of several still unknown genes are reported here, thus completing the publication of the Vχ genes of the x locus as far as they are potentially functional or have only one or two 1‐bp defects. Of the Vχ genes of the K locus, 32 are potentially functional, 16 have minor defects, 3 have both potentially functional and slightly defective alleles and 25 are pseudogenes which amounts to a repertoire of 76 Vχ‐related gene sequences. The Vχ genes of the L regions are, within the subgroups, particularly similar to each other, which is in part due to common evolutionary origins and in part caused by gene conversion‐like events. One donor‐acceptor pair could be clearly identified, since converted and not‐converted alleles of the acceptor gene were found. In other cases the duplicates of the converted genes served as non‐converted controls.
Antisense oligonucleotides specific for regulatory factor RFX‐1 inhibit inducible but not constitutive expression of all major histocompatibility complex class II genes
Authors: Siegrist Claire‐Anne , Mach Bernard
Published: March 2007

The regulation of major histocompatibility complex (MHC) class II genes expression, which can be constitutive, inducible or both, is a crucial aspect of the control of an immune response. It involves binding of various regulatory factors to cis‐acting sequences of MHC class II promoters. Antisense oligonucleotides specific for RFX‐1, a regulatory factor binding to the functionally essential X box motive of MHC class II promoters, were designed to study the role of RFX‐1 in the various modes of MHC class II regulation and explore the possibility of experimentally modulating the level of expression of MHC class II genes by transcriptional intervention. RFX‐1 antisense oligonucleotides were first tested in cell‐free translation, selected for an inhibitory effect on RFX‐1 in vitro translation and then assayed in cell cultures for an effect on human histocompatibility leukocyte antigen (HLA) class II expression. We show that an RFX‐1 specific antisense oligonucleotide drastically inhibits induction of HLA‐DR, ‐DQ and ‐DP molecules by interferon γ in monocytic cells. Unexpectedly, the same agent has no effect on the constitutive expression of the same genes either in these cells or in B lymphocytes, indicating an uncoupling of the constitutive and inducible modes of class II regulation. This transient and reversible experimental modulation of MHC class II expression in live cells by transcriptional intervention provides a new tool to study the function of class II molecules in various biological models.
Production and characterization of two murine monoclonal antibodies directed against epitopes exclusive to soluble fragments of FcεRII/CD23
Authors: Ghaderi Abbas A. , Stanworth Denis R.
Published: March 2007

Two murine monoclonal antibodies (mAb) designated as SU1 and SU3 directed against soluble FcεRII/CD23 have been generated by fusing X.63.AG.8653 (a mouse myeloma cell line) with spleen cells from mice immunized with an Epstein Barr virus (EBV)‐transformed B cell line (RPMI‐8866). The antibodies have been shown to be capable of detecting affinity purified soluble FcεRII/CD23 in an enzyme‐linked immunosorbent assay. Indirect immunofluorescence has shown that the SU1 and SU3 mAb do not stain RPMI‐8866, a FcεRII/CD23+ B cell line. By studying the migration profiles of affinity purified SU1‐ and SU3‐reactive molecules on sodium dodecylsulfate‐polyacrylamide gel electrophoresis it has been shown that SU1 mAb immunoprecipitates 33‐ and 12‐kDa components, while the SU3 mAb recognized 25‐ and 45‐kDa proteins from culture supernatants of RPMI‐8866 cells. Moreover, affinity purified SU1‐ and SU3‐reactive proteins have been shown to be recognized by human IgE but not by the human IgG molecule. These results provide evidence that SU1 and SU3 mAb may recognize some putative post‐cleavage epitopes on the N‐terminal end of the low affinity receptor which appear, perhaps, following the process of fragmentation. In addition, the effect of these antibodies on continuous growth of a panel of lymphoblastoid cell lines indicates that SU1 mAb was found incapable of influencing the spontaneous proliferation of EBV‐immortalized B cell lines; whereas SU3 mAb completely blocked the spontaneous growth and proliferation of all B cell lines tested. The results are discussed in relation to the appearance of a functional post‐cleavage epitope on soluble FcεRII/CD23.