bluuebunny/crossref_metadata_2025 huggingface dataset:
Cell components required for deletion of an autoreactive T cell repertoire
Authors: Fukushi Noriko , Wang Bingyan , Arase Hisashi , Ogasawara Kazumasa , Good Robert A. , Onoé Kazunori
Published: March 2007

T cells become tolerant to self antigens during their development in the thymus. Clonal deletion of thymocytes bearing T cell receptor (TcR) which recognize self antigens is a major mechanism for generating tolerance. In the present study we have used allogeneic bone marrow (BM) chimeras, prepared with various combinations of mouse strains and focusing especially on expressions of I‐E molecules and Mls‐1a antigens on the cell surface, to investigate both immuno‐histochemically and by flow cytometry the cell components that contribute to the clonal deletion of T cells positive for Vβ6 TcR. The Vβ6 TcR expression is strongly associated with T cell recognition of both I‐E and Mls‐1a antigens. We found that I‐E+ cells derived from donor BM (and thus not of recipient lineage) represented a primary requirement for deletion of Mls‐1a−reactive thymocytes which bear Vβ6 TcR. Immunohistochemical analysis revealed that the donor‐derived I‐E+ cells were distributed mainly to the thymic medulla and that the Vβ6+ cells were eliminated from the thymic medulla between 2 and 3 weeks following BM transplantation. In contrast, Mls‐1a+ cells of either donor or recipient origin might be responsible for the deletion, even though cortical epithelial cells appeared not to express Mls‐1a antigens.
Primary responses of human T cells to mycobacteria: a frequent set of γ/δ T cells are stimulated by protease‐resistant ligands
Authors: Pfeffer Klaus , Schoel Bernd , Gulle Heinz , Kaufmann Stefan H. E. , Wagner Hermann
Published: March 2007

T lymphocyte subsets expressing either T cell receptor α/β or γ/δ were selected from human peripheral blood T cells and proliferative responsesto molecular mass‐fractionated mycobacterial lysates were determined. α/β T cells primarily responded to fractions > 30 kDa whereas γ/δ Tcells preferentially reacted to fractions < 3 kDa. Protease digestion abolished the stimulating activities for α/β T cells, confirming that α/β Tcells respond to protein components. In contrast, components recognized by γ/δ T cells proved resistant to protease digestion. In limiting dilution studies, frequencies of proliferating γ/β T cells remained virtually unaltered by protease treatment of stimulating lysates, while those of α/β T cells became almost undetectable. Furthermore, only few γ/δ T cells responded to the 65‐kDa heat‐shock protein. Our data indicate that, unlike α/β T cells, γ/δ T cells respond to mycobacterial components which are resistant to vigorous protease digestion.
The activation of protein kinase C is not necessary for the monoclonalantibody‐induced modulation of CD3 and CD4 antigens
Authors: Thuillier Laure , Selz Françoise , Métézeau Philippe , Pérignon Jean‐louis
Published: March 2007

We studied the effect of staurosporine, a potent inhibitor of protein kinase C (PKC) activity, on the phorbol ester‐ or monoclonal antibody (mAb)‐induced modulation of CD3 and CD4 surface antigens. Staurosporine (10−5 M) completely inhibited phorbol ester‐induced modulation but had no effect on that induced by mAb. These results indicate that the down‐regulation of CD3 and CD4 observed after activation of the cells by the corresponding mAb is independent from PKC‐mediated phosphorylations, and thus that the activation of PKC is sufficient but not necessary to induce the modulation of CD3 and CD4 antigens.
Immune responses to defined epitopes of the circumsporozoite protein of the murine malaria parasite, Plasmodium yoelii
Authors: Grillot Didier , Michel Martine , Müller Ingrid , Tougne Chantal , Rènia Laurent , Mazier Dominique , Corradin Giampietro , Lambert Paul‐henri , Louis Jacques A. , del Giudice Giuseppe
Published: March 2007

We have investigated the immunogenicity of defined sequences of the circumsporozoite (CS) protein of the murine malaria parasite, Plasmodium yoelii. A 21‐mer synthetic peptide from the nonrepetitive region of the CS protein (position 59—79, referred to as Py1) induced T cell proliferative responses in H2d and, to a lesser extent, in H2b mice. Conversely, a synthetic peptide (referred to as Py4) consisting of four (QGPGAP) repeats of the P. yoelii CS protein, induced anantibody response only in H2b mice. No antibody response was observed whenthe Py3 peptide, consisting of three (QGPGAP) repeats, was used as an immunogen. When cross‐linked to the Py4 repetitive peptide, the Py1 sequence behaved as a T helper epitope allowing the production of anti‐Py4 antibodies in H2d mice. Several long‐term T cell lines and clones specific for the nonrepetitive Py1 peptide were originated in vitro from both H‐2d and H2b mice. These lines and clones were CD4+ and proliferated in a major histocompatibility complex‐restricted fashion. Furthermore, Py1‐specific T cell lines and clones did not proliferate in the presence of synthetic peptides from an analogous region of another rodent malaria parasite, P. berghei, despite the high degree of homology existing in this sequence of the two CS proteins. Finally, supernatants from 7 out of 13 clones (from BALB/c mice) produced detectable amounts of interleukin 2 and interferon‐γ; whereas supernatants from the 4 clones from C57BL/6 and 2 from BALB/c mice contained detectable amounts of interleukin 5. These results show that functionally heterogenous CD4+ T cell populations, belonging to either TH1 or TH2 subset, are activated upon immunization of mice with the P. yoelii Py1 synthetic peptide. It is not yet known what differential role these CD4+ subsets play during the malaria infection or after immunization with different malaria T cell epitopes. This knowledge may have a particular impact in the design of effective subunit vaccines against malaria.
Cell and type specificity of interferon action. Unusual characteristics of the transcriptional control of gene expression by interferon‐γ in T cells*
Authors: Gribaudo Giorgio , Gariglio Marisa , Cavallo Giorgio , Landolfo Santo
Published: March 2007

We have examined the mechanisms by which interferon (IFN)‐γ and IFN‐α regulate the expression of 2′‐5′‐oligoadenylate synthetase (2–5A synthetase) and class I major histocompatibility complex antigens in murine T cells and in cell types of other histological origin. When treated with IFN‐α both fibroblasts and T cell lines displayed a marked increase of the 2–5A synthetase activity and of the corresponding mRNA. The augmentation of the enzyme activityin T cells was induced by IFN‐α at the transcriptional level, as determined by nuclear run‐on analysis. In contrast IFN‐γ was capable of increasing 2–5A synthetase activity only in fibroblasts, but not in T cells. Nuclear run‐on assays revealed that the 2–5A synthetase gene in T cells is not transcriptionally activated by IFN‐γ. After IFN‐α and ‐γ treatment we also observed a significant increase in class I gene expression in fibroblasts and T cell lines as measured both on the cell surface and by cytoplasmic RNA accumulation. In the case of the T cell line, DO1110, the observed increase in the steady‐state levels of class I transcripts was a consequence of a high rate of H‐2 gene transcription as demonstrated by run‐on analysis. However, the molecular mechanisms involved in this IFN‐dependent H‐2 gene transcriptional activation are differentbetween IFN‐α and IFN‐γ. When the T cell lines DO1110, L12‐R4 and EL4 were transfected with a plasmid containing a reporter gene (chloramphenicol acetyltransferase) under the control of a regulatory IFN‐responsive DNA element of 237 bp or 1.4 kb, IFN‐α was able to activate the transcription of these constructs. In contrast, IFN‐γ did not recognize the IFN‐responsive element which, by itself, activated transcription of the reporter gene in response to IFN‐γ in other cellular types of non‐T cell origin. Therefore, in the T cell linesexamined, IFN‐γ increases the H‐2 gene expression by acting on DNA elements located upstream of the regulatory segment used in this study or downstream of thecap site. This suggests a possible cell specificity in the activation of an IFN‐responsive element, that in turn may regulate the IFN‐inducible gene expression in acell‐specific fashion. Thus, the differential biological activities of IFN‐γ on T cells could be generated by a differential gene activation at the transcriptional level.
Adoptive transfer of fluorescence‐labeled cells shows that resident peritonealmacrophages are able to migrate into specialized lymphoid organs and inflammatory sites in the mouse
Authors: Rosen Hugh , Gordon Siamon
Published: March 2007

We have examined the migration of murine macrophages from the vascular compartment tonormal and inflammatory tissues by the adoptive transfer of resident peritoneal macrophages (RPMΦ) fluorescently labeled with the hydrophobic dye 1,1′‐dioctadecyl3,3,3′,3′‐tetramethylindocarbocyanine perchlorate (DiI). After initial labeling of the plasma membrane of RPMΦ, the dye accumulated stably in intracellular vesicles of low density (Q = 1.042—1.045 kg/l) and cells remained viable in culture for 4 weeks. Like the normal monocyte, DiI‐RPMΦ, but not exudate‐derived or fixed cells, migrated to peritoneal exudates, following i.v. adoptive transfer, by a mechanism inhibitable by an antibody to the type 3 complement receptor. In the absence of an inflammatory stimulus there was no migration to the peritoneal cavity, and DiI‐RPMΦ, accumulated within 4 h in the red pulp and marginal zone of the spleen. By day 6 these cells still formed a tight ring of fluorescence in the marginal zonealone, outside the marginal metallophil cells. DiI‐RPMΦ injected into the peritoneal cavity migrated to the parathymic lymph nodes where they were found in the subcapsular sinus and in the medullary cords, whereas very few fluorescent cells could be found in the T cell areas. The migration of RPMΦ to lymphoid organs required viable cells but, unlike the recruitment of cells to peritoneal exudates, was not inhibitable by antibodies to CR3. We conclude that the RPMΦ is a useful surrogate for the analysis of constitutive and induced monocyte migration to secondary lymphoid and inflammatory sites, respectively.
Human suppressor T cell clones lack CD28
Authors: Li Shu Guang , Ottenhoff Tom H. M. , van den Elsen Peter , Koning Frits , Zhang Li , Mak Tak , De Vries René R. P.
Published: March 2007

Previously we showed that certain T cell lines and clones from a lepromatous leprosy patient displayed a dose‐dependent suppression of the proliferation of autologous T cells to Mycobacterium leprae (M. leprae) but not mitogen or an unrelatedantigen. The latter cells were also cloned and did not display this suppressive activity, were CD4+ and proliferated vigorously to M. leprae presented by autologous HLA‐DR molecules. We shall refer to these cells as T helper (Th) cells. Most of the suppressive T cell clones (Ts) were also CD4+ and also proliferated to M. leprae presented by HLA‐DR, but much less strongly than Th cells.

In this study we report on our search for (a) the mechanism of this apparently antigen‐specific suppression by T cells, and (b) a possible phenotypic difference between Th and Ts clones. The two main conclusions are that Ts clones possess a lytic machinery, but that M. leprae‐specific suppression and cytotoxicity can be clearly dissociated, and that the only phenotypic difference between Th and Ts is the presence of the CD28 marker on Th and its absence on Ts clones.
Structure of class II genes in wild mouse Mus saxicola: functional and evolutionary implications
Authors: Cam Philippe , Jouvin‐marche Evelyne , Leguern Christian , Marche Patrice N.
Published: March 2007

Class II β genes have been characterized from Mus saxicola, a wild mouse which diverged five millions years ago from the house mouse. The nucleotide sequences of the exons 2 and 3 of Eβ1, Aβ2 and Eβ2 genes have been determined in order to define the molecular mechanisms of interspecies variability. The Eβ1 and Aβ1 (J. X. She, personal communication) genes from Mus saxicola display extensive sequence variability in exon 2 when compared tothose of the house mouse, in association with the variability of residues probably involved in antigen binding. On the other hand, most putative T cell contact residues from the I‐Eβ chain, and to a lesser extent from the I‐Aβ chain, were found conserved in mice. T cell contact residues from Mus saxicola I‐Aβ and I‐Eβ chains are identical to those of the b and q, and of the k haplotypes of the house mouse, respectively. Comparisons performed withAβ2 and Eβ2 genes revealed that both are highly conserved and, in contrast to Aβ1 and Eβ1 genes, that a strong selective pressure for conservation occurs in the Aβ2 second exon. The different aspects of Aβ2 and Eβ2 genes evolution may correlate with the original function of their products. Comparisons of the intervening sequences revealed that the repeated motifs of the hotspot region present in the Eβ1b gene are also conserved in Mus saxicola. Recombinations may have occurred, in some cases in association with χ‐like sequences.
Human antigen‐specific memory T cells express the homing receptor (LAM‐1) necessary for lymphocyte recirculation
Authors: Tedder Thomas F. , Matsuyama Takami , Rothstein David , Schlossman Stuart F. , Morimoto Chikao
Published: March 2007

Lymphocytes must circulate from blood into lymphoid tissues and sites of infection and inflammation to function efficiently in vivo. This process of “homing” is in part directed by the expression of the leukocyte adhesion molecule (LAM‐1, also known as TQ1 and Leu‐8) in humans and the homologous MEL‐14 antigen in mice. In this report, we demonstrate that the LAM‐1 molecule is a 74‐kDaprotein and that only half of the CD4+ T cells in humans which have a memory phenotype (CD45RA −CD29hi) express the LAM‐1 molecule. Functionally, these two phenotypically distinct subpopulations of memory cells were quite different. The LAM‐1+ memory cells proliferated better to recall antigen and induced three to seven times higher levels of B cell immunoglobulin secretion than their LAM‐1− counterparts. Thus, antigen‐specific memory T cellswithin the helper lineage express the homing receptor appropriate for regulating their migration to secondary lymphoid tissues and sites of inflammation.
Lack of somatic mutation in a χ light chain transgene
Authors: Sharpe Melanie J. , Neuberger Michael , Pannell Richard , Azim Surani M. , Milstein César
Published: March 2007

Analysis of mice transgenic for immunoglobulin genes should allow definition of the cis‐acting DNA sequences required to target somatic mutation to antibody V genes. We have looked for mutations in a chimeric χ transgene encoding a V region specific for the hapten 2‐phenyloxazolone (phOx) linked to a rat Cχ gene. Two independent lines of transgenic mice were hyperimmunized with phOx and splenic hybridomas established. In B cells that had been selected by antigen and which used mouse anti‐phOx genes, the endogenous sequences were found to be mutated whereas the transgene remained unchanged. These results suggest either that (a) if the transgene is a “passenger” gene expressed at a low level, transgene mutation is a rare event, or that (b)sequences far from the χ coding region are necessary to direct somatic mutation.
Human T cell proliferative responses to Plasmodium vivax antigens: evidence ofimmunosuppression following prolonged exposure to endemic malaria
Authors: Goonewardene Renu , Carter Richard , Priyntha Gamage C. , del Giudice Giuseppe , David Peter H. , Howie Sarah , Mendis Kamini N.
Published: March 2007

Human T cell proliferative responses, of 33 adult Sri Lankans convalescing from Plasmodium vivax infections, to several P. vivax antigens (i.e. a soluble extract of asexual erythrocytic stage parasites and two cloned antigens that are potential vaccine candidates PV200 and GAM‐1) were assessed. The peripheral blood mononuclear cell proliferative responses to the soluble extract of P. vivax, as assessed by studying both the proportion of responders and the degree of the response, were significantly lower in a group of individuals resident in a malaria endemic area in Sri Lanka than in another group that did not have a life‐long exposure to malaria but had acquired the disease on a visit to an endemic region. Individuals of both groups responded equally well tomitogen. The responses to a non‐malarial antigen such as purified protein derivative of tuberculin were only marginally lower in residents of the malaria‐endemic region. These findings suggest that exposure to endemic P. vivax malaria leads to a specific immuno‐suppression to P. vivax antigens. Immunosuppression of a much lower degree was evident to a non‐malarial antigen.
The cytoplasmic domain of CD4 is required for stable association with the lymphocyte‐specific tyrosine protein kinase p56lck
Authors: Veillette André , Sleckman Barry P. , Ratnofsky Sheldon , Bolen Joseph B. , Burakoff Steven J.
Published: March 2007

The CD4 T cell surface molecule binds MHC class II determinants expressed on antigen‐presenting cells. CD4 is thought to enhance T cell activation by serving as an adhesion molecule as well as possibly by transducing an independent intracellular signal during the process of antigen stimulation. The recent observation that CD4 is physically associated with the Src‐related tyrosine protein kinase p56lck suggests thattyrosine phosphorylation might be involved in these CD4 “signaling” events. The results presented in this report demonstrate that deletion of the cytoplasmic domain of CD4 significantly diminishes its ability to stably associate with p56lck. Thisobservation provides a biochemical basis for the decreased ability of this mutant CD4 molecule to enhance T cell activation during suboptimal antigen stimulation.
Anti‐immunoglobulins induce death by apoptosis in WEHI‐231 B lymphoma cells
Authors: Benhamou Laure E. , Cazenave Pierre‐andré , Sarthou Pierre
Published: March 2007

Anti‐membrane immunoglobulin (anti‐mIg) antibodies exert inhibitory effects in immature B lymphocytes such as WEHI‐231 cells. We show here that anti‐mIg treatment causes DNA fragmentation (apoptosis) in these cells. We also report thatco‐treatment with the protein kinase C activator phorbol 12‐myristate 13‐acetate prevents apoptosis induced by anti‐mIg. These results are in agreement with our initial proposal that sensitivity to the toxic effects of anti‐mIg reflects, at least partially, altered signal transduction in immature B lymphocytes. Variations in signal transduction pathways during B lymphocyte ontogeny may, therefore, play a critical role in determining whether B cells should be activated or inhibited via their mIg.
A physical map and analysis of the murine Cχ‐RS region show the presence of a conserved element
Authors: Müller Brigitte , Stappert Heike , Reth Michael
Published: March 2007

Lambda‐producing B lymphocytes have frequently deleted one or, more often, both Igχ loci. This deletion is mediated by the rearrangement of an element which lies 3′ of Cχ and which is called RS (recombining sequence) in the mouse and Kde (kappa‐deleting element) in the human. The tight correlation between Vλ to Jλ rearrangements and an RS‐mediated deletion may indicate that sequences in the Cχ‐RS region are controlling the activation of the Igλ locus. We have linked the Cχ exon and the RS element byphage cloning and compared the Cχ‐RS region to the previously cloned human Cχ‐Kde region. The distance between Cχ and RS is 25 kb and is thus similar to the distance of 24 kb separating the human Cχ exon and Kde element. Both mouse and man carry a conserved sequence of 470 bp (Rx) which lies 9 kb 3′ of the mouse Cχ and 12 kb 3′ of the human Cχ exon. The conserved mouse Rx sequence contains part of the χ3′ enhancer.
CD43 (leukosialin, sialophorin, large sialoglycoprotein) can be expressed in both normal and Wiskott‐Aldrich fibroblasts via transfection of a leukosialin cDNA
Authors: Pallant Adam , Fukuda Minoru , Frelinger John G.
Published: March 2007

Human leukosialin is among the most abundant sialoglycoproteins found on the surface of cells of the lympho‐hematopoietic system. Leukosialin, also known as sialophorin, is involved in T cell proliferation, and its molecular isoform changes upon cellular activation. We show that human leukosialin is identical to the antigens described by the monoclonal antibodies (mAb) G10‐2, G19‐1 (CD43) and B1B6 (large sialoglycoprotein). This identity was suggested by immunoblot analysis of transformed cell lysates. Further, fibroblasts transfected with the human leukosialin cDNA gain reactivity to these mAb, showing conclusively that molecules recognized by these mAb are determined by the same cDNA. Expression of the leukosialin gene is readily detected on the surface of transfected human and mouse fibroblasts. Immunoblot analysis of the transfectants indicates that processing of the human protein occurs in both species. Alterations of leukosialin expression have been reported in patients with the Wiskott‐Aldrich Syndrome (WAS), an X‐chromosome‐linked immunodeficiency disease. While essentially all of the transfected tumor and primary fibroblasts from normal individuals express the transfected gene on the cell surface, only half of the transfected Wiskott‐Aldrich fibroblasts express CD43. Nonetheless, the antigenic pattern by immunoblot analysis of both normal and WAS‐transfected fibroblasts appears identical. These results indicate that WAS‐derived cells can express leukosialin and that the product of WAS X‐chromosome mutation may not be expressed in fibroblasts.
A malaria heat‐shock‐like determinant expressed on the infected hepatocyte surface is the target of antibody‐dependent cell‐mediated cytotoxic mechanisms by nonparenchymal liver cells
Authors: Rénia Laurent , Mattei Denise , Goma Joseph , Pied Sylviane , Dubois Philippe , Miltgen François , Nüssler Andreas , Matile Hugues , Menégaux Fabrice , Gentilini Marc , Mazier Dominique
Published: March 2007

Cultured hepatic stages of Plasmodium falciparum and P. yoelii and with a monoclonal antibody recognizing a C‐terminal fragment of the P. falciparum heat‐shock‐like protein (Pfhsp70) revealed that synthesis of this antigen first occurs during intrahepatic development of the parasite, at the two nuclei stage. Using a variety of techniques, including scanning electron microscopy, we observed that this antigenic determinant was expressed on the infected hepatocyte membrane. Its participation in antibody‐dependent cell‐mediated cytotoxicity was investigated. While no effect was obtained with peripheral blood cells, we found that 25% of the schizonts were specifically lysed when using spleen cells at a killer/target ratio of 30/1. More interestingly, with nonparenchymal liver cells, up to 50% of the hepatic parasites disappeared with a killer/target ratio of 10/1.
Modulation of immune responses with monoclonal antibodies. I. Effects on regional lymph node morphology and on anti‐hapten responses to haptenized monoclonal antibodies
Authors: Mjaaland Siri , Fossum Sigbjøm
Published: March 2007

Repeated injections of monoclonal antibody (mAb) culture supernatants into rat footpads increased the weights of the draining lymph nodes. Immunostained freeze sections showed that injection of MRC OX2, a mAb reacting with rat follicular dendritic cells and MRC OX7 (anti‐Thy‐1.1), led to gross hypertrophy primarily of the follicular areas, whereas MRC OX6 (anti‐rat major histocompatibility complex class II molecules) resulted in selective stimulation of the paracortex. These findings indicate that mAb, when conjugated to certain antigens, would modulate the immune response to these antigens. Consequencly, the mAb were conjugated with fluorescein isothiocyanate (FITC) and the humoral response against the hapten measured. The primary anti‐FITC antibody response was tenfold stronger than after stimulation with FITC conjugated to a conventional carrier such as ovalbumin, and had some characteristics of a secondary response: a fast increase of IgG level to very high titers and a long duration without further amplification at later antigen challenges.
Clonal anergy in self‐reactive α/β T cells is abrogated by heat‐shock protein‐reactive γ/δ T cells in aged athymic nude mice
Authors: Yuuki Hiroyuki , Yoshikai Yasunobu , Kishihara Kenji , Iwasaki Akinori , Matsuzaki Goto , Takimoto Hiroaki , Nomoto Kikuo
Published: March 2007

Although T cells proliferate and differentiate primarily in the thymus, athymic nude mice contain an appreciable level of T cell receptor α/β and γ/δ T cells, suggesting the existence of the extrathymic pathway in the development of both T cells. Recent studies with nude mice indicate that clonal deletion of self‐reactive T cells does not occur extrathymically. In the present study, we have investigated the responsiveness of self‐reactive T cells differentiating along an extrathymic pathway in aged BALB/c (H‐2d, Mls‐1b2a, I‐E+, 7–8 month old) nude mice. Consistent with recent reports, T cells bearing Vβ3 or Vβ11, which are important for recognizing proteins encoded by the Mls‐2a or the I‐E allele, respectively, are readily detected in age nude mice. The Vβ3‐ or Vβ11‐bearing T cells, however, do not proliferate in response to staphylococcal enterotoxin A which specifically stimulates Vβ3‐ or Vβ11‐bearing T cells. When exogenous recombinant interleukin 2 was added to the culture, the Vβ3‐bearing T cells in aged nude mice significantly proliferated in response to staphylococcal enterotoxin A. Aged nude mice also contained a substantial level of γ/δ T cells which account for 15.6% of all Thy‐1.2+ cells. The γ/δ T cells proliferated and produced a significant level of interleukin 2 in response to the 65‐kDa mycobacterial heat‐shock protein, which is highly homologous to its eukaryotic counterpart. These results suggest that unresponsiveness of self‐reactive T cells may be reversed by T cells responding to stress proteins expressed by the invading microbes and/or the stressed autologous cells.
Thy‐1‐specific helper T cells can provide intramolecular help, but can provide intermolecular help only if the phosphatidylinositol anchor of Thy‐1 is replaced by a membrane insert
Authors: Stauss Hans , Mitchison N. Avrion , Yao Michelle
Published: March 2007

We have investigated the ability of Thy‐1 to serve as a helper antigen in delivering intermolecular help. The parental Thy‐1 molecule, which is attached to the cell surface via a phosphatidylinositol anchor, failed to deliver intermolecular help for an antibody response to H‐2 alloantigens. This failure appears to be due to the phosphatidylinositol anchor since a Thy‐1 hybrid construct, which is expressed as a transmembrane cell surface molecule, can serve as helper antigen and deliver intermolecular help.
The rat B cell system: The anatomical localization of flow cytometry‐defined B cell subpopulations
Authors: Kroese Frans G. M. , Butcher Eugene C. , Lalor Paul A. , Stall Alan M. , Herzenberg Leonore A.
Published: March 2007

Two‐color flow cytometrical (FCM) analysis of rat peripheral lymphoid organs shows two distinct IgM/IgD‐defined B cell subpopulations, similar to those of the mouse: a major population of cells expressing little IgM and high levels of IgD (population I) and a minor population of cells expressing high levels of IgM but little IgD (population III). In peripheral lymphoid organs population III cells are mainly found in spleen where they represent about 25% of the B cells; population III cells are almost absent from lymph nodes and Peyer's patches. In adult bone marrow and in neonatal spleen the majority of IgM/IgD‐defined B cells (> 70%) are population III cells, similar to what is observed in the mouse. In contrast with mice, only a low proportion of the cells (1%) recovered from the peritoneal cavity are B cells, and most of them belong to population I.

Previously defined monoclonal antibodies (HIS22 and HIS24) to B cell forms of the leukocyte common antigen (CD45R) in combination with staining for surface IgM and surface IgD demonstrates a further heterogeneity of rat B cells by three‐color FCM analyses. HIS22 labels most population I cells; population III cells and a small subset (about one third) of population I express only very low levels of the HIS22 determinant. HIS24 reacts with population I cells and subdivides population III into two subsets: about one third of splenic population III cells are brightly stained with this antibody whereas fluorescence of the remaining two‐thirds is lower. The HIS24bright population III cells likely are newly formed B cells since cells with this phenotype are the predominant surface Ig population found in adult bone marrow and neonatal spleen.

In tissue sections of lymphoid organs, HIS22‐ and HIS24‐positive cells are mainly found in lymphoid follicles; splenic marginal zones are almost unstained. Combining immunohistological analysis with the FCM data, we therefore conclude that the small follicular B cells are in population I and marginal zone B cells are found in the HIS24dull population III. The in situ localization of HIS24bright population III cells and the HIS22dull population I cells is not clear.

Based upon the relative expression of surface markers on B cells we propose a model of B cell differentiation in the rat in which a common precursor cell (the HIS24bright population III cell), produced in the bone marrow, enters the spleen and leads to two distinct B cell differentiation pathways: one leading to marginal zone B cells and the other giving rise (through an intermediate cell type) to the majority of B cells, the small resting follicular B cells.
Cytotoxic α/β+ and γ/δ+ T cells in murine intestinal epithelium
Authors: Viney Joanne L. , Kilshaw Peter J. , MacDonald Thomas T.
Published: March 2007

Murine intraepithelial lymphocytes (IEL) consist of two main populations. Approximately half are Thy‐1+, most of which are CD3+, T cell receptor (TcR) α/β+, and the remainder are Thy‐1−, most of which are CD3+, TcR α/β (presumably TcR γ/δ+). In redirected cytotoxicity assays, TcR α/β+ IEL are potent cytotoxic effectors. Thy‐1−, CD3+, TcR α/β− IEL isolated from athymic mice are also cytotoxic. Thus, regardless of TcR usage or Thy‐1 expression, IEL are cytotoxic effectors.
Monoclonal antibody against α(1 → 3) dextran transfers suppression of the immune response to the acetylcholine receptor
Authors: Tong Zhou , Dwyer Donard S.
Published: March 2007

Previous studies have demonstrated that immunization of BALB/c mice with α(1 → 3) dextran (Dex) is accompanied by a reduction in the subsequent immune response to the acetylcholine receptor (AChR), depending on the timing of the Dex administration relative to AChR challenge. Here, we report that suppression of the anti‐AChR response can be transferred by a monoclonal antibody, known as DX2, which is specific for Dex. Serum transfer experiments have also supported the notion that antibody is important for this effect. In addition, two new idiotypic markers have been defined that are expressed mainly by antibodies against Dex, including DX2. The anti‐idiotypic reagents (Sh135 and EB5) are derived from the immune response to the AChR. A human monoclonal antibody which binds to Dex (SR 11) resembles the BALB/c antibodies that are involved in the suppression of the anti‐AChR response. These findings emphasize the functional relevance of the AChR‐Dex network not only for the BALB/c immune response to the AChR, but also for humans with the autoimmune disease, myasthenia gravis.
Persistence of anti‐donor allohelper T cells after neonatal induction of allotolerance in mice
Authors: Abramowicz Daniel , Vandervorst Philippe , Bruyns Catherine , Doutrelepont Jean‐Marc , Vandenabeele Peter , Goldman Michel
Published: March 2007

BALB/c mice rendered tolerant to A/J alloantigens by neonatal injection of 108 (A/J × BALB/c)F1 spleen cells develop an autoimmune disease associated with a polyclonal activation of donor B cells. To study the mechanisms leading to donor B cell activation in tolerant mice, we prepared mixed lymphocyte cultures (MLC) between splenic T cells from neonatally injected mice and donor‐type (A/J × BALB/c)F1 or third‐party (C57BL/6 × BALB/c)F1 B cells. T cells from tolerized mice were unable to generate cytotoxic T lymphocytes, to proliferate or to secrete interleukin (IL)2 after stimulation with donor alloantigens in MLC. These T cell responses were present after MLC with third‐party antigens, but were of lower intensity than those generated by control BALB/c T cells. In contrast. T cells from tolerized mice stimulated immunoglobulin production by donor‐type (A/J × BALB/c)F1 B cells much more powerfully than T cells from control BALB/c mice. The stimulation of donor‐type (A/J × BALB/c)F1 B cells was polyclonal, as attested by the levels of anti‐hapten and anti‐DNA antibodies in the MLC supernatants. IgM was the dominant isotype secreted in vitro, but IgG1 and IgG3 were also produced in significant amounts. Lysis experiments indicated that the T cells responsible for F1 B cell stimulation in MLC were CD4+ host T cells. These T helper cells were alloreactive since they did not stimulate syngeneic BALB/c B cells, and their effect on donor B cells was specifically blocked by anti‐donor Ia monoclonal antibodies. Addition of anti‐IL 4 monoclonal antibody to MLC between T cells from tolerant mice and (A/J × BALB/c)F1 B cells almost completely abolished the production of IgG1, but not that of IgM or IgG3. Taken together, these findings indicate that neonatal injection of alloantigens in BALB/c mice induces a state of dissociated tolerance, with unresponsiveness of anti‐donor T cells secreting IL 2 on the one hand, and persistence of T cells responsible for B cell help and IL 4 secretion on the other hand.
Cytokine‐induced generation of multinucleated giant cells in vitro requires interferon‐γ and expression of LFA‐1
Authors: Möst Johannes , Neumayer Heinz P. , Dierich Manfred P.
Published: March 2007

Multinucleated giant cells (MGC), which are a common feature of various pathologic states, were generated in vitro by cytokine‐stimulation of human peripheral blood monocytes. As expected, conditioned medium, i.e. the supernatant of concanavalin A‐stimulated peripheral blood mononuclear cells, readily caused generation of MGC. Addition of a monoclonal antibody (mAb) against interferon‐γ (IFN‐γ) completely abrogated this effect. IFN‐γ alone, however, had a much smaller effect than the conditioned medium. All other cytokines tested [including interleukin (IL) 2, IL 4 and tumor necrosis factor‐α, which are known to activate monocytes] did not induce MGC nor did they enhance the effect of IFN‐δ.

Formation of MGC could almost entirely be inhibited by mAb to the α or β chain of LFA‐1 and to a lesser extent by relatively high concentrations of a mAb against ICAM‐1, one of the ligands of LFA‐1. In contrast to the anti‐IFN‐γ mAb that had no significant effect on the formation of monocyte clusters, mAb against LFA‐1 inhibited clustering very efficiently.

Antibodies directed to a number of different antigens present on the surface of monocytes (α chains of CR3 and CR4, HLA class I and II molecules, CD14 and CD16 antigens) had little or no effect on the generation of MGC. IFN‐γ, but not the concanavalin A‐induced supernatant clearly enhanced expression of LFA‐1 and ICAM‐1 on monocytes.

The results indicate that cytokine‐induced generation of MGC is not possible without IFN‐γ, but most probably additional factor(s) enhance this effect. The mechanism(s) by which IFN‐γ promotes monocyte fusion apparently includes, among others, up‐regulation of LFA‐1 whose expression seems to be necessary but not sufficient for fusion.
Expression of complement alternative pathway proteins by endothelial cells. Differential regulation by interleukin 1 and glucocorticoids
Authors: Dauchel Hélène , Julen Nathalie , Lemercier Claudie , Daveau Maryvonne , Ozanne Denyse , Fontaine Marc , Ripoche Jean
Published: March 2007

We have studied the secretion of proteins of the alternative pathway of complement C3, factor B and factor H by human umbilical vein endothelial cells (HUVEC). Results showed that factor H and factor B are quantitatively secreted in abundance whereas C3 could only be detected when the cells are maintained in culture during long periods of time. Interferon‐γ stimulated factor H, factor B and, to a lesser extent, C3 secretions. Interleukin (IL) 1 had a differential effect on spontaneous C3, factor B and factor H secretions. In the presence of IL 1, there was a significant secretion of C3 occurring within a short period of culture. IL 1 also stimulated factor B secretion. There was a synergistic stimulating effect between IL 1 and interferon‐γ to bring C3 and factor B productions by HUVEC to very high levels. In contrast, factor H secretion was consistently inhibited by IL 1. Local increase in C3 and factor B secretions by endothelial cells in the presence of IL 1 may have important implications in the inflammatory reaction. In striking contrast, the glucocorticoid dexamethasone (DXM) had modulatory effects which are consistent with its anti‐inflammatory properties. DXM, at therapeutic concentrations, decreased C3 and factor B secretions and increased factor H secretion. Local modulation of complement protein secretion by DXM appears to be a new mechanism by which this glucocorticoid may control inflammation.
Non‐tolerance and differential susceptibility to diabetes in transgenic mice expressing major histocompatibility class II genes on pancreatic β cells
Authors: Götz Jürgen , Eibel Hermann , Köhler Georges
Published: March 2007

In insulin‐dependent diabetes mellitus aberrant expression of major histocompatibility complex (MHC) class II antigens might induce autoimmune destruction of insulin‐producing pancreatic β cells. Here we demonstrate that already the expression of single MHC class II chains (Eαk and Eβb) is sufficient to cause diabetes without affecting β cell morphology and cell numbers. Our transgenes interfere at least in two points of insulin production leading to a severely diabetic phenotype. In one case Eβb expression led to a 10‐30‐fold decrease of mouse insulin mRNA. In another case Eαk expression reduced insulin secretion to background levels. In addition, we also found a patchy distribution of both insulin and Eαk expression, indicating heterogeneity of the β cell population, without the concommittant development of diabetes. Although the transgenic Eαk Eβb MHC class II molecules were expressed on the surface of pancreatic β cells, the transgenic mice did not prove to be tolerant for I‐E antigen. Autoimmune reactions remained absent showing that aberrant MHC class II expression on pancreatic β cells alone is not sufficient for the development of autoimmune diabetes in mice.
Immunoglobulin‐specific T‐B cell interaction. IV. B cell presentation of idiotypic determinant(s) of monoclonal anti‐surface immunoglobulin antibody to idiotope‐recognizing helper T clones
Authors: Rudensky Alexander Y. , Mazel Svetlana M. , Blechman Janna M. , Yurin Vitalij L.
Published: March 2007

Previously, we have demonstrated T‐B cell interactions mediated by T cell recognition of immunoglobulin (Ig) peptide/major histocompatibility complex (MHC) class II complexes derived by the B cell processing of endogenously synthesized Ig molecules. In this report Ig‐specific T‐B cell interaction mediated by B cell presentation of idiotopes (Id) of anti‐sIg antibodies to Id‐specific T cell clones has been studied in Igϰ‐1‐congenic rat strains. A panel of August (RT‐1c; Igϰ‐1a) rat T helper clones specific for Id of syngeneic anti‐Igϰ‐1b C3B9 monoclonal antibody (mAb) has been developed to study IdC3B9 presentation by Igϰ‐1b‐bearing B cells from congenic August.1b (RT‐1c; Igϰ‐1b) rats. Five of seven IdC3B9‐specific T clones responded even at very low concentrations (100‐200 pg/ml) of C3B9 mAb presented by Igϰ‐1b+ B cells. In contrast, the presentation of intact C3B9 mAb by nonspecific antigen‐presenting cells (macrophages, Igϰ‐1a+ B cells, etc.) to IdC3B9‐specific T cells was of low efficiency. The IdC3B9‐specific T cell response to idiotopes of anti‐Igϰ‐1b C3B9 mAb was found to be restricted by RT‐1B molecule and required the processing of intact C3B9 molecule. IdC3B9 epitope recognized by C31 and C5 clones was mapped to the heavy chain of C3B9 mAb. Thus, B cells are able to present peptides related to the V region of anti‐sIg Ab, i.e. Id peptide/MHC class II complexes, to Id‐recognizing T cells. IdC3B9‐presenting B cells are specifically activated both to proliferation and Ig production upon interaction with IdC3B9‐specific T clones. Based on the results of our studies on B cell presnetation of Ig epitopes to T cells a hypothetical model of Ig peptide‐driven T‐B cell interaction has been proposed.
Corynebacterium parvum (Propionibacterium acnes): An inducer of tumor necrosis factor‐α in human peripheral blood mononuclear cells and monocytes in vitro
Authors: Rossol Siegbert , Voth Rita , Brunner Silvia , Müller Werner E. G. , Büttner Matthias , Gallati Harald , zum Büschenfelde Karl‐Hermann Meyer , Hess Georg
Published: March 2007

The present study investigates the potential capacity of the immunostimulant Corynebacterium parvum (C.p.) to induce tumor necrosis factor‐α (TNF‐α) in human peripheral blood mononuclear cells (PBMC) and blood monocytes (BMo) in vitro. Both at the mRNA and protein level, stimulation of PBMC and BMo upon C.p. induces TNF‐α. Compared to the hitherto used TNF‐α inducers in vitro such as Sendai virus, phytohemagglutinin or lipopolysaccharide the C.p. stimulus displayed a threefold stronger induction of TNF‐α production (p < 0.001). Using C.p. as an inducer it was possible to demonstrate that TNF‐α production is regulated by prostaglandin E2; preincubation of the cells with prostaglandin E2 resulted in a reduced C.p.‐mediated TNF‐α production (p < 0.001). Coincubation of interferon‐γ (IFN‐γ) together with C.p. led to an enhanced release of TNF‐α, supporting the assumption that C.p. is a potent TNF‐α inducer. The additive effect of IFN‐γ and TNF‐α on the receptor level was demonstrated by addition of IFN‐γ antibodies to the PBMC cultures. Under these conditions TNF‐α production, stimulated by C.p. and IFN‐γ, was decreased by 30%, compared to the production in assays supplemented with C.p. alone. From these data we conclude that C.p. is a new inducer of TNF‐α in vitro and a useful tool to study TNF‐α production of PBMC and BMo from either healthy donors or from patients.
Cellular requirements for anti‐DNA production induced in mice by immunization with bacterial DNA
Authors: Gilkeson Gary S. , Pritchard Anita J. , Pisetsky David S.
Published: March 2007

To further define DNA immunization as a model for anti‐DNA production, we investigated the cellular requirements for this response in mice immunized with single‐stranded DNA from E. coli. The anti‐DNA responses of genetically immune‐deficient mice and congenic controls were measured by ELISA after immunization with E. coli DNA as complexes with methylated bovine serum albumin in complete Freund's adjuvant. T cell‐deficient BALB/c‐nulnu mice failed to produce IgG anti‐DNA by this protocol despite high backgrounds of IgM anti‐DNA. In contrast, CBA/N mice expressing the xid defect displayed IgG anti‐DNA responses comparable to those of CBA/J mice despite a reduced IgM response; the specificity of CBA/N and CBA/J anti‐DNA antibodies was similar as determined by binding to synthetic DNA and RNA antigens. These results suggest that the anti‐DNA response stimulated by DNA immunization is dependent on T cells but not the B cell population affected by xid. The intact IgG response of immunized xid mice differs from that of lupus mice bearing xid where this gene defect leads to significant reduction of spontaneous anti‐DNA production.
Molecular cloning of sheep T cell receptor γ and δ chain constant regions: unusual primary structure of γ chain hinge segments
Authors: Hein Wayne R. , Dudler Lisbeth , Marcuz Anne , Grossberger Dario
Published: March 2007

The primary structure of sheep T cell receptor (TcR) γ and δ chain constant (C) regions has been determined by cDNA cloning. A comparison of the nucleotide and deduced amino acid sequences of the sheep chains with known human and mouse sequences shows that the primary structure of the immunoglobulin, transmembrane and cytoplasmic Cγ domains and all of the Cδ region has been substantially conserved. However, the hinge or connector region of sheep γ chains differs significantly from all known TcR chains. Clones representing two different sheep Cγ genes were isolated and both contain additional sequence in this region, making them the longest TcR chains so far identified. The hinge region of both sheep Cγ sequences contains two additional cysteine residues and a motif of five amino acids (TTESP or TTEPP) which has been triplicated in one of the clones. Other repetitive segments of 13–17 amino acids could also be identified suggesting that, as in the human Cγ2 gene, this region of the sheep genes could have arisen from an exon duplication or triplication event. Southern blot analysis of sheep DNA confirmed the presence of one Cδ gene and at least two Cγgenes. A restriction fragment length polymorphism that is probably associated with allelic sequence variation in the sheep Cδ gene was detected in DNA from different animals. Although the essential structure of the γ/δ TcR appears well conserved through evolution, the marked heterogeneity evident in the hinge region of γ chains both within and between species, and particularly the presence of additional cysteine residues in the sheep sequences, may be of structural and functional importance.
Differential peripheral expansion and in vivo antigen reactivity of α/β and γ/δ T cells emigrating from the early fetal lamb thymus
Authors: Hein Wayne R. , Dudler Lisbeth , Morris Bede
Published: March 2007

The concentrations of different lymphocyte subsets in the blood of lambs which had been thymectomized (Tx) in utero between days 67‐75 of fetal gestation were measured at birth and at various intervals during the first year of life. Compared to thymus‐intact (Ti) controls, Tx lambs were severely depleted of both α/β and γ/δ T cells at birth (< 10% of control levels). The majority of the residual α/β T cells present in Tx lambs at birth were CD4+CD8−. As the Tx lambs aged, the concentration of α/β T cells in blood increased steadily to reach levels around 50% of control values. In contrast, the circulating γ/δ T cells did not expand in Tx animals and remained barely detectable throughout the observation period, although these cells accounted for 30%‐60% of the T cells in the blood of Ti lambs. The expansion of α/β but not γ/δ T cells was also reflected in changes in the cellular composition of solid lymphoid organs in Tx lambs. B cell numbers were similar in both groups at birth but Tx lambs were persistently B lymphopenic from 3 weeks of age onwards. The α/β T cells that had expanded in Tx lambs responded to stimulation with bacterial antigens in a way that was qualitatively similar to the response in Ti lambs. By contrast, the few γ/δ T cells in Tx lambs responded abnormally. Our results show that although sheep α/δ and γ/δ T cells are equally thymus dependent during ontogeny, the early fetal thymic emigrants which establish the two T cell lineages in the periphery have strikingly different antigen reactivities and capacities for self‐renewal and expansion.