bluuebunny/crossref_metadata_2025 huggingface dataset:
Complement regulation on the surface of cultured schistosomula and adult worms of Schistosoma mansoni
Authors: Marikovsky Moshe , Parizade Miriam , Arnon Ruth , Fishelson Zvi
Published: March 2007

Cercaria and freshly prepared schistosomula of Schistosoma mansoni are highly sensitive to complement. However, early in their maturation, the schistosomula become resistant to complement killing. This conversion is preceded by a rapid and massive release of several acetabular proteases and of the glycocalyx coat. Thus, shedding of the glycocalyx which is a major immunogen and a strong activator of the alternative pathway of complement permits the parasite to escape immune damage. Mechanically transformed schistosomula, which were cultured in a defined synthetic medium and developed complement resistance, could be converted by proteolysis to complement sensitivity. Trypsin and pronase markedly increased the susceptibility of cultured schistosomula to complement. The trypsin‐induced complement sensitivity persisted for at least 19 h without recovery of resistance. Similar treatment with trypsin produced complete killing of adult worms by complement in absence of antibodies. Efficient killing was obtained with normal human serum (NHS), with normal guinea pig serum (GpS), and with C4‐depleted HS and C4‐deficient GpS indicating that the killing was mediated by the cytolytic alternative pathway of complement. Larger quantities of C3b with intact α′ chain could be demonstrated on trypsin‐treated than on non‐treated schistosomula. Antibodies which were raised in rabbits by immunization with the trypsin‐released material bound to cultured (non‐treated) schistosomula and to adult worms, and induced their killing in GpS and C4‐deficient GpS. These results suggest that following release of the glycocalyx, the transforming schistosomula of S. mansoni spontaneously express a complement regulatory protein(s). A similar regulator is postulated to be present on the surface of adult worms. Such regulatory molecules may serve as good targets for immunotherapy, since antibodies directed to them will inhibit their regulatory activity and thus potentiate in vivo the lytic action of complement.
A new HLA‐DRB1 allele within the DRw52 supertypic specificity (DRw13‐DwHAG): Sequencing and direct identification by oligonucleotide typing
Authors: Tiercy Jean‐Marie , Jeannet Michel , Mach Bernard
Published: March 2007

Molecular analysis of HLA class II polymorphism represents a crucial parameter for HLA matching in transplantation immunology, for the study of HLA‐disease association and for the understanding of major histocompatibility complex (MHC)‐restricted antigen presentation. We report here the DNA sequence and the deduced amino acid sequence of the polymorphic first domain exon of the DRB1 and DRB3 alleles of the homozygous cell line HAG (DRw13‐DwHAG‐DQw7). The DRB1 sequence represents a new DRB allele, which clearly shows a close relationship to other DRB1 genes from the DRw52 group and is now officially named DRB1*1303. The DRB1*1303 allele is very similar to the two DRw13 alleles we have described earlier, with only five amino acid differences at positions 32, 37, 47, 57 and 71. Furthermore, its sequence in the third hypervariable region is unique among all known DRB1 and DRB3 alleles. The sequence of the DRB3 gene of HAG shows that it corresponds to the previously described DRB3*0101 (DRw52a) allele. In addition we present analyses of a panel of healthy blood donors and leukemic patients by oligonucleotide typing showing that this new HLA‐DR specificity can now be unequivocally identified in routine oligotyping with an allele‐specific oligonucleotide probe.
Regulation of human IgE synthesis. I. Human IgE synthesis in vitro is determined by the reciprocal antagonistic effects of interleukin 4 and interferon‐γ
Authors: Chrétien Isabelle , Pène Jérǒme , Brière Francine , Malefijt René De Waal , Rousset Françoise , de Vries Jan E.
Published: March 2007

In the present study culture conditions resulting in optimal IgE synthesis by mononuclear cells (MNC) isolated from peripheral blood, tonsils or spleens from healthy nonallergic donors were investigated. The highest rate of IgE synthesis was obtained in a two‐step culture system in which the MNC were preincubated with interleukin 4 (IL4; 200 U/ml) for 48 h, washed and subsequently incubated with IL 4 (200 U/ml) for 9 days. Despite these culture conditions, IL4‐induced IgE synthesis varied considerably (1‐150 ng/ml) and MNC from 16/70 donors failed to produce IgE. Kinetic studies indicated that IL4 was required at the onset of the incubation phase. IgE synthesis was reduced by > 95% when addition of IL 4 in the incubation period was delayed 24 h or more. IL 4‐induced IgE synthesis was blocked by interferon‐γ (IFN‐γ). This inhibition is most effective when IFN‐γ was added in the 48‐h preincubation step or during the first 48 h of the incubation period. Interestingly, IL 4 was found to block spontaneous and lectin‐ or factor‐induced IFN‐γ production by MNC, purified CD3+, CD4+ or CD8+ Tcells. This down‐regulatory effect of IL 4 on IFN‐γ production occurred at the mRNA transcription level. Furthermore, it is shown that IL 4 induced the release of soluble CD23 and that recombinant soluble CD23 enhanced IL4‐ induced IgE synthesis, but only when IL 4 was present at suboptimal concentrations. Collectively, our data indicate that IL4 and IFN‐γ regulate the level of IgE synthesis by influencing each other's activities reciprocally during the first 3 days of the culture.
Structure of rearranged and germ‐line rabbit Vϰ genes indicated that the CDR3 is encoded by the Vϰ gene*
Authors: Ayadi Hammadi , Cazenave Pierre‐André , Marche Patrice N.
Published: March 2007

The third complementary‐determining regions (CDR3) of rabbit kappa chains are unusually long and the length is more heterogeneous when compared to those of the mouse and the human ϰ chains. To study how the rabbit ϰ light (L) chain genes create diversity and generate CDR3, we analyzed the structure of a rearranged variable ϰ gene (Vr) and the variable (Vg) and joining (Jg) regions of the putative precursor genes. Alignment of the Vr gene sequence with that of the Vg and Jg regions allowed precise determination of the recombination event. Five nucleotides between the recombination point and the J2 heptamer were deleted, indicating flexibility in the recombination producing rabbit ϰ chains. The entire Vg is contained in the rearranged product demonstrating that neither a D element nor an N sequence addition are required for the CDR3 formation. Comparison of the Vr and the Vg gene sequences show base substitutions suggesting that somatic mutations may contribute to rabbit ϰ L chain diversity.
Functional analysis of intercellular adhesion molecule‐1‐expressing human thyroid cells
Authors: Weetman Anthony P. , Freeman Mark , Borysiewicz Leszeck K. , Makgoba M. William
Published: March 2007

We have tested the potential role of thyroid cell intercellular adhesion molecule‐1 (ICAM‐1) expression by in vitro assays of cell clustering and cytotoxicity. Increased ICAM‐1 appeared within 24 h of thyroid cell stimulation with cytokines and was not inhibited by the antithyroid drug methimazole. Autologous and allogeneic lymphocyte‐thyroid cell cluster formation, assessed by flow cytometry, was reduced by about one‐third in the presence of a monoclonal antibody against ICAM‐1, regardless of whether thyroid cells were expressing basal levels of ICAM‐1 or had been stimulated with interferon‐γ. The cytotoxicity produced by interleukin 2‐stimulated allogeneic lymphocytes was not consistently inhibited by anti‐ICAM‐1 antibody, but phytohemagglutinin‐stimulated lymphocytes showed a reduction of 23% – 28% in cytotoxicity against untreated or interferon‐γ stimulated thyroid cells when the anti‐ICAM‐1 monoclonal antibody was present. Finally, thyroid cells could be infected by rhinovirus, confirming the presence of fully functional ligand. These results show that ICAM‐1 expression by thyroid cells may enhance immune cell recognition and play some role in cytotoxicity, features which could be important in the initiation or perpetuation of autoimmune thyroiditis.
Complement activation by immunoglobulin does not depend solely on C1q binding
Authors: Bindon Carol I. , Hale Geoffrey , Waldmann Herman
Published: March 2007

A matched set of rat chimeric antibodies has been studied for their ability to activate various key stages of the complement cascade. Rat IgM and IgG2b were efficient at all stages from C1q binding to cell lysis. However, for other isotypes, a direct correlation between C1q binding and cell lysis did not apply. IgG2a, which was only modestly efficient at C1q binding, was relatively more so for binding and activation of whole C1, and was by far the most effective isotype after IgG2b, and IgM for C4 and C3 binding. IgG2c was relatively efficient at binding C1q and C1, but less so for the binding of C4 or for later stages. IgA was efficient at binding C1, but again, this was not reflected in activation of later stages. The results suggest that properties of different isotypes, as well as influencing binding of C1q, may regulate attachment of the C1r2C1s2 tetramer. In addition, distinct features of certain isotypes may favor C4 activation and binding, independent of their ability to activate C1.
The implications of subunit interactions for the structure of the T cell receptor‐CD3 complex
Authors: Koning Frits , Lee Maloy W. , Coligan John E.
Published: March 2007

Cell surface‐expressed receptors are often multichain complexes. One of these, the T cell receptor (TcR)αβ‐CD3 complex, is known to contain at least seven chains: the α and β TcR chains plus the γ, δ, ϵ and two ζ chains from the CD3 complex (αβγδϵζ2). To gain insight into the structure of the complex we have used anti‐peptide antisera specific for the individual subunits of the complex, and nonionic and ionic detergents to determine subunit interactions within the complex. Four closely associated pairs of chains could be identified: αβ, ζ2, γϵ and δϵ. Interactions between the TcR αβ and either γϵ or δϵ could be observed in the apparent absence of other CD3 chains. Furthermore, a hierarchy in the strength of the association between the TcR and the individual CD3 chains could be distinguished: TcR ϵ > TcR δ > TcR γ. The ζ2 dimer could only be detected in “intact” TcR‐CD3 complexes shedding no light on possible interactions with either the TcR or CD3‐γ, δ and ϵ chains. Finally, cross‐linking experiments suggest a close spatial relationship between the TcR αβ and both the CD3‐γ and CD3‐ϵ chains. The results demonstrate that the methods used give valuable information on subunit interactions in a cell surface‐expressed receptor complex and suggest a TcR‐CD3 complex in which two ϵ chains are present, one linked to γ and the other to δ. The data further indicate that γϵ and δϵ complexes interact directly with the TcR chains. Based on the observations a model for the structure of the TcR‐CD3 is presented and discussed.
Polyclonal in vitro proliferative responses from nonimmune donors to Plasmodium falciparum malaria antigens require UCHL1+ (memory) T cells*
Authors: Rebecca Jones K. , Hickling Julian K. , Targett Geoffrey A. T. , Playfair John H. L.
Published: March 2007

The in vitro polyclonal proliferative responses of peripheral blood mononuclear cells to whole blood stage parasites or fractionated antigens from the human malaria parasite Plasmodium falciparum were studied. Cells from healthy laboratory donors who had never been exposed to malaria antigens in vivo consistently proliferated to P. falciparum antigens, as did cord blood mononuclear cells. This response was only observed in sheep rosette‐positive cells in the presence of adherent cells and was inhibited by NH4Cl, indicating a requirement for antigen processing. The proliferative response was strongest at day 6 and was dependent on the presence of cells expressing high levels of CD45 180‐kD isomer (UCHL1 monoclonal antibody), a marker for activated or memory cells, but not for CD45R (SN130 monoclonal antibody) a marker for naive or unprimed Tcells. This suggests a similarity to the recall response to tuberculin antigen. These results suggest that the proliferative response to malaria antigens observed previously and described as a nonspecific mitogenic response may be a cross‐reactive response to epitopes shared between P. falciparum and other common immunogens. This would explain the establishment of T cell clones to malaria antigens from such donors, but might suggest that the epitopes to which such clones are specific may be of questionable protective or diagnostic use.
Human memory T cells express intercellular adhesion molecule‐1 which can be increased by interleukin 2 and interferon‐γ
Authors: Buckle Anne‐Marie , Hogg Nancy
Published: March 2007

We have shown that low levels of intercellular adhesion molecule‐1 (ICAM‐1) expression can be detected on approximately 40% of the resting peripheral blood T cell population. The ICAM‐1+ T cells have the phenotypic markers of memory cells which can be distinguished functionally from naive T cells by their ability to respond rapidly to previously experienced antigen. These cells appear to be in a state of marginal activation in that they also express low levels of the interleukin 2 receptor (Tac antigen) and have increased cell size as compared to the naive T cells. In addition we have shown that the cytokines interleukin 2 and interferon‐γ, both of which are products of activated T cells, are able to increase the expression of ICAM‐1 on T cells. Finally, pretreating T cells with an anti‐ICAM‐1 monoclonal antibody inhibits their response to recall antigens, strongly suggesting a functional role for this protein on the memory T cell.
Induction of antigen‐specific CD4+ HLA‐DR‐restricted cytotoxic T lymphocytes as well as nonspecific nonrestricted killer cells by the recombinant mycobacterial 65‐kDa heat‐shock protein
Authors: Ab Birhane Kale , Kiessling Rolf , Van Embden Jan D. A. , Thole Jelle E. R. , Kumararatne Dinakantha S. , Pisa Pavel , Wondimu Assefa , Ottenhoff Tom H. M.
Published: March 2007

Acquired cell‐mediated immunity to intracellular parasites like mycobacteria is dependent on antigen‐specific T lymphocytes. We have recently found that mycobacteria not only induce helper T cells but also cytotoxic CD4+ and/or CD8+ T cells as well as nonspecific killer cells that lyse human macrophages in vitro. In addition, we have described that the recombinant heat‐shock protein (hsp) 65 of Mycobacterium bovis BCG/M. tuberculosis is an important target antigen for CD4+CD8− cytotoxic T cells.

We have now further investigated the cytotoxic effector cells that are induced by the hsp65 of BCG. Purified protein derivative of tuberculin (PPD)‐ or hsp65‐specific cytotoxic T cells specifically lysed PPD, hsp65 of BCG and hsp65 of M. leprae‐pulsed macrophages in an HLA‐DR‐restricted manner. Nonpulsed macrophages were lysed to a much lower but still significant extent. hsp65‐induced effector cells expressed CD3, CD5, CD4, CD8 and CD56 markers. Depletion experiments showed that the antigen‐specific HLA‐DR‐restricted killer cell was of the CD5+CD4+CD8−CD56− phenotype. Experiments using N‐terminal truncated hsp65 fusion (cro‐lacZ) proteins suggested that the N‐terminal 65 amino acid residues of the 540 amino acid molecule are critical for the expression of the cytotoxic target epitope(s) in two individuals tested.

In addition to inducing antigen‐specific cytotoxic effector cells, the hsp65 also triggered nonspecific nonrestricted effector cells with lytic activity against nonpulsed autologous or allogeneic macrophages as well as K‐562 and Daudi tumor cells. hsp65‐stimulated effector cells produced both interferon and tumor necrosis factor‐α. An important finding was that hsp65‐stimulated effector cells strongly inhibited colony‐forming unit formation from live BCG‐infected autologous macrophages.
Adipogenesis in a murine bone marrow stromal cell line capable of supporting B lineage lymphocyte growth and proliferation: Biochemical and molecular characterization
Authors: Gimble Jeffrey Martin , Dorheim Marie‐Annette , Cheng Qi , Medina Kay , Wang Chi‐Sun , Jones Robert , Koren Eugen , Pietrangeli Carolynn , Kincade Paul W.
Published: March 2007

Recent advances in long‐term bone marrow (BM) culture techniques have allowed investigators to dissect cellular components responsible for lympho‐hematopoiesis. Consequently, a number of “stromal” cell clones have been developed which are capable of supporting B lineage lymphocyte growth and proliferation in vitro by direct cell‐cell interactions and the release of cytokines. While much work has focused on the support function of these cells, questions remain regarding their own differentiation potential. We have examined adipo‐genesis in the cloned BM stromal cell, BMS2. The presence of hydrocortisone, methylisobutylxanthine, or 30% fetal calf serum each accelerated adipocyte differentiation. This process was accompanied by the accumulation of triglycerides and cholesterol esters along with the induction of adipocyte‐specific enzymes. Likewise, the steady‐state level of mRNA transcripts increased for genes related to lipid metabolism. However, the pattern of mRNA expression in BMS2 adipocytes differed from that of a well‐established, pre‐adipocyte cell line, 3T3‐L1, with respect to the following genes: glycerol phosphate dehydrogenase, CAAT/enhancer binding protein and angiotensinogen. Adipocyte BMS2 cells retailed the ability to support stromal cell‐dependent B lineage lymphocytes in methylcellulose assays. The adipocytes continued to express macrophagecolony‐stimulating factor mRNA constitutively and interleukin 6 mRNA in an inducible manner, similar to the BMS2 pre‐adipocytes. Together, these data document a close developmental relationship between a specialized fibroblasts and adipocytes in the BM and suggest that adipocyte stromal cells may play an active role in lympho‐hematopoiesis.
Interleukin 2, 4 and 5 are sequentially produced in mitogen‐stimulated murine spleen cell cultures
Authors: Cardell Susanna , Sander Birgitta
Published: March 2007

Lymphokine production was analyzed in murine spleen lymphocytes stimulated with different T cell mitogens. Using in situ hybridization, frequencies of cells and the kinetics of production of interleukin (IL) 2, 4 and 5 were analyzed. The different mitogens varied in their ability to induce the three interleukins. IL 2 was most successfully induced with a high dose of the calcium ionophore A23187 combined with phorbol 12‐myristate 13‐acetate (PMA). Significant frequencies of cells containing IL 4 or IL 5 mRNA were found among cells stimulated with an anti‐CD3 antibody together with PMA, or pokeweed mitogen. The combination of anti‐CD3 and PMA induced relatively high frequencies of all three cytokines. The production was sequential with the highest levels of IL 2 mRNA present during the first 24 h, IL 4 mRNA reaching a peak on day 2 and finally IL 5 peaking on day 3. When cells that had been stimulated with mitogens in vitro were restimulated, the lymphokines were produced more rapidly. The order of production was maintained with IL 2 mRNA reaching a maximum already at 3 h of culture, IL 4 mRNA at 8 h and IL 5 mRNA at 24 h.
Stable expression and function of EBV/C3d receptor following genomic transfection into murine fibroblast L cells
Authors: Cantaloube Jean François , Piechaczyk Marc , Calender Alain , Lenoir Gilbert , Minty Adrian , Carrière Dominique , Fischer Elizabeth , Poncelet Philippe
Published: March 2007

The Epstein‐Barr virus (EBV) and the C3d component of complement bind to the same cell surface receptor (EBVR/CR2) which is part of the B lymphocyte differentiation antigen recognized by the monoclonal antibodies (mAb) of the cluster of differentiation 21 (CD21). To analyze EBV and C3d binding to this receptor, mouse fibroblasts were transfected with human genomic DNA and rare CD21‐positive cells were selected and cloned by cell sorting. The presence of the human gene in host cell DNA as well as its transcription product were assayed with a cloned EBVR/CR2 cDNA by Southern and Northern blotting analysis, respectively. A glycoprotein of apparent molecular mass of 140 kDa, similar to that found in human B lymphocytes, was immunoprecipitated with anti‐CD21 mAb and proved to be functional since both C3d and EBV bound efficiently and specifically to mouse cells expressing EBVR/CR2. However, no expression of EBV nuclear antigens, early antigens and viral capsid antigens was detected in cells exposed to EBV. This indicates that the presence of EBVR/CR2 is not sufficient to allow full infection of mouse fibroblasts.
Effect of interferon‐gamma and tumor necrosis factor‐alpha on lymphoepithelial interactions within thymic nurse cells
Authors: Defresne Marie Paule , Humblet Chantal , Rongy Anne Michel , Greimers Roland , Boniver Jacques
Published: March 2007

Isolated thymic nurse cells (TNC) represent a specialized microenvironment in vivo where thymocytes interact specifically with subcapsular epithelial cells. They are thought to play a critical role in the process of T cell differentiation. We demonstrate that recombinant murine interferon‐γ and recombinant human tumor necrosis factor‐α can act on these interactions: they stimulate TNC‐derived epithelial cells to establish interactions with thymocytes in vitro and to form new lymphoepithelial complexes. This phenomenon is partially inhibited by anti‐la monoclonal antibodies. Implications of these findings for normal intrathymic differentiation are discussed.
Autoantibody production against the U small nuclear ribonucleoprotein particle proteins E, F and G in patients with connective tissue diseases
Authors: Reuter Rolf , Rothe Susanne , Habets Winand , van Venrooij Walter J. , Lührmann Reinhard
Published: March 2007

The nucleoplasmic U small nuclear ribonucleoprotein particles (snRNP) have a set of seven proteins in common which are designated B′, B, D, D′, E, F and G. Patients suffering from rheumatoid autoimmune diseases such as systemic lupus erythematosus often develop autoantibodies against the proteins B′, B, and D. Here we describe a sensitive immunoassay which allows the specific detection of autoantibodies reacting with the E, F or G snRNP proteins. We were able to identify several patient sera containing autoantibodies against one or more of these proteins. This demonstrates that all snRNP proteins described so far are potentially antigenic in systemic rheumatoid diseases. The characterization of the antibodies showed an immunological cross‐reactivity between the snRNP protein G and the 70‐kDa protein of U1 snRNP. Several sera contained autoantibodies which were specific for the F snRNP protein.
Induction of transplantation tolerance by intravenous injection of allogeneic lymphocytes across an H‐2 class II mismatch. Different mechanisms operate in tolerization across an H‐2 class I vs. H‐2 class II disparity
Authors: Van Twuyver Esther , Martin Kast W. , Mooijaart Rob J. D. , Melief Cornelis J. M. , De Waal Leo P.
Published: March 2007

Previously, we have shown that the intravenous (i.v.) injection of allogeneic lymphocytes across an H‐2 class I‐mutant disparity leads to specific skin allograft tolerance caused by irradiation‐sensitive donor T cells, which function as veto cells. In the present study, we show that the i.v. injection of H‐2 class II‐incompatible spleen cells also results in specific skin allograft tolerance. However, tolerance induction depends on the presence of irradiation‐resistant non‐T cells in the donor cell inoculum. Thus, different mechanisms operate in tolerance induction across an H‐2 class I vs. H‐2 class II mismatch. I.v. injection of allogeneic spleen cells across an H‐2 class I plus class II disparity does not result in skin allograft tolerance. Finally, our data show that transfusion‐induced suppression of the delayed‐type hypersensitivity response against alloantigens does not correlate with skin allograft tolerance induced by i.v. injected allogeneic lymphocytes. In conclusion, the type of H‐2 mismatch between transfusion donor and recipient not only determines the occurrence of allograft tolerance but also the mechanism leading to tolerization.
Exit of recirculating lymphocytes from lymph nodes is directed by specific exit signals
Authors: Binns Richard M. , Licence Stephen T.
Published: March 2007

During recirculation, lymphocytes leave the peculiar structurally inverted lymph nodes (LN) of pigs via blood vessels instead of via efferent lymphatics, as in sheep and other mammals. This functional difference provided an opportunity to show the existence of signals directing lymphocyte exit from LN. The recirculation of pig peripheral blood lymphocytes was traced through fetal sheep LN and of sheep PBL into and out of unsuckled newborn piglet LN, using the lack of natural antibody or natural killer cell function in these immunologically mature young to compare foreign and homologous lymphocyte behavior. In spite of some 50 million years of evolutionary divergence, the detailed kinetics and route of recirculation of the xenogeneic PBL were essentially the same as those of the host species. Thus determinants guiding the anomalous blood exit from pig LN must involve conserved “exit” signals in a new site and not changes in pig lymphocyte homing receptors.
Increased expression of Ia antigens on B cells after neonatal induction of lymphoid chimerism in mice: Role of interleukin 4
Authors: Abramowicz Daniel , Doutrelepont Jean‐Marc , Lambert Philippe , Vorst Philippe Van Der , Bruyns Catherine , Goldman Michel
Published: March 2007

BALB/c mice rendered chimeric at birth by injection of 108 (A/J × BALB/c)F1 spleen cells develop a lupus‐like autoimmune disease linked to the activation of donor B cells by host T cells. As in vitro studies previously indicated that interleukin 4 (IL4) was a mediator of the interactions between T and B cells, we analyzed the intensity of Ia antigen expression on B cells of chimeric mice. Flow cytometric analysis with anti‐Ia monoclonal antibodies (mAb) revealed that B cells from spleens and lymph nodes of 2‐week‐old chimeric BALB/c mice displayed a two‐ to threefold increase in membrane Ia antigen expression, this increase still being present in spleens of 30‐week‐old animals. An increase in Ia antigen expression was also found in the small number of donor B cells detected in spleens and lymph nodes of chimeric mice. IL4 was the major stimulus leading to increased B cell Ia antigen expression, as this phenomenon was substantially prevented by in vivo treatment of chimeric mice with the anti‐IL4 11B11 mAb.

In vitro experiments revealed that host splenic T cells of chimeric mice, while unable to generate anti‐donor cytotoxic T lymphocytes, secreted significant amounts of IL4 when stimulated in mixed lymphocyte cultures (MLC) with donor alloantigens. This IL4 secretion led to an increased expression of Ia antigens on donor‐type F1 B cells present in MLC. No significant increase in Ia antigen expression was found on syngeneic BALB/c B cells co‐cultured with T cells from chimeric mice unless A/J B cells were added to the cultures.

Taken together, these findings indicate that increased Ia antigen expression on donor B cells is induced by IL4 secreted by anti‐donor Tcells. IL 4 released in this setting also leads to increased Ia antigen expression on host B cells through a bystander effect.
Continuous cell activation is necessary for stable interaction of complement receptor type 3 with its counter‐structure in the aggregation response of human neutrophils
Authors: Kuypers Taco W. , Koenderman Leo , Weening Ron S. , Verhoeven Arthur J. , Roos Dirk
Published: March 2007

Human neutrophils aggregate after stimulation with various stimuli; this response is completely absent in neutrophils from patients with leukocyte adhesion defiency (LAD). To investigate the cellular requirements of this process a method was used in which neutrophils are separately loaded with hydroethidine (HE) and sulfofluorescein (SFDA), to give them either red fluorescence or green fluorescence. After mixing HE‐ and SFDA‐labeled cells in a ratio of 1 : 1, the number of double‐colored aggregates formed after activation was determined by analysis on a fluorescence‐activated cell sorter. In this way, essential information is obtained when cells of different origin are used. The formation of aggregates between neutrophils of an LAD patient and control neutrophils was thus quantified. Because neutrophil aggregation is dependent mostly on the presence of complement receptor type 3 (CR3), which is not present on LAD neutrophils, this result revealed the presence of a counter‐structure for CR3 on LAD neutrophils (and hence on normal human neutrophils). In addition to the presence of these proteins on the cell surface, aggregation required continuous cell triggering as indicated by the transient aggregation induced by short‐term activation of protein kinase C. This phenomenon was substantiated by the fact that energy depletion caused profound disaggregation. The present study reveals that neutrophil aggregation is a well‐controlled process, which needs constant activation of CR3 for a stable interaction with a constitutively expressed counter‐structure.
Preferential usage of Vα4 and Vβ10 T cell receptor genes by lymphocytic choriomeningitis virus glycoprotein‐specific H‐2Db‐restricted cytotoxic T cells
Authors: Aebischer Toni , Oehen Stephan , Hengartner Hans
Published: March 2007

Correlations between the T cell receptor (TcR) V gene usage and the specificity of T cells have been primarily described for major histocompatibility complex (MHC) class II‐restricted helper T cell responses. In the present study the TcR genes expressed by MHC class I‐restricted murine cytotoxic T cells (CTL) specific for a major epitope of the lymphocytic choriomeningitis virus (LCMV), LCMV‐GP2275–289, were investigated. The TcR primary structure of an LCMV‐GP2275–289 specific H‐2Db‐restricted CTL clone has been determined. It uses a member of the Vα4 family joined to JαAN14.4 for the α chain and Vβ10 rearranged to Dβ2.1 and Jβ2.4 for its β chain. Four other independent LCMV‐GP2275–289 specific H‐2Db‐restricted CTL clones also expressed Vα4 and Vβ10 gene elements. Furthermore, Vα4 and Vβ10 were preferentially expressed by polyclonal CTL of C57BL/6 origin specific for LCMV. These results suggest that both TcR Vα and Vβ regions are important for the recognition of the LCMV‐GP2275‐289 epitope on H‐2Db molecules.
Evidence that the level of the p55 component of the interleukin (IL) 2 receptor can control IL 2 responsiveness in a murine IL 3‐dependent cell
Authors: Collins Mary K. L. , Malde Prupti , Miyajima Atsushi , Arai Ken‐Ichi , Smith Kendall A. , Mulligan Richard C.
Published: March 2007

Although the role of interleukin 2 (IL 2) in mature T lymphocyte function is well documented, its effect on hematopoietic progenitor cells is less well characterized. Here we have used recombinant retroviruses to transduce and express a cDNA clone encoding the p55 component of the human IL 2 receptor (h‐p55), in a murine IL 3‐dependent cell line, BAF3. While the parental cells do not respond to IL 2, the h‐p55‐expressing cells proliferate upon treatment with recombinant IL 2 after an initial lag period. The responsiveness of individual cell clones is correlated with their level of h‐p55 expression, and can be inhibited by Tac monoclonal antibody. Furthermore, growth at limiting IL 2 concentrations selects a subset of cells expressing higher h‐p55 levels from a bulk population. Detailed 125I‐labeled IL 2 binding analysis on the highest h‐p55‐expressing clone detects the presence of 200 high‐affinity (KD = 25 pM) IL 2 receptors. We therefore propose that the level of h‐p55 expression governs the formation of high‐affinity receptors, and hence IL 2 responsiveness in BAF3 cells.
Stimulation of all T cells bearing Vβ1, Vβ3, Vβ11 and Vβ12 by staphylococcal enterotoxin A
Authors: Takimoto Hiroaki , Yoshikai Yasunobu , Kishihara Kenji , Matsuzaki Goro , Kuga Hiroshi , Otani Tsuyoshi , Nomoto Kikuo
Published: March 2007

To determine the molecular mechanisms of T cell stimulation by staphylococcal enterotoxin A (SEA), we examined the expression of T cell receptor (TcR) Vβ on the T cells from four strains of mice stimulated in vitro with SEA, using flow cytometric analysis for the number of T cells bearing Vβ3, Vβ6, Vβ8, Vβ11 and RNA blotting analysis for the amount of transcripts of Vβ1, Vβ5 and Vβ12. The number of T cell blasts bearing Vβ1, Vβ3, Vβ1 or Vβ12 were increased in the T cell blasts proliferating in vitro in response to SEA in C57BL/6 mice. In AKR/J mice, which contain few Vβ11‐ or Vβ12‐bearing T cells due to a tolerance to the self‐MHC class II IE‐antigens, T cells bearing Vβ1 or Vβ3 responded to SEA. SEA enriched only Vβ1‐bearing T cells in BALB/c mice carrying Mls‐2a which lack Mls‐1a‐reactive Vβ3‐bearing T cells as well as Vβ11‐ and Vβ12‐bearing T cells. In spite of the presence of Vβ1‐bearing T cells, C3H/He T cells exhibited a very low responsiveness to SEA. T cell repertoires skewed by clonal deletion of self‐reactive T cells may in part account for the different sensitivity to SEA among the different strains. A tolerance to SEA can be established in C57BL/6 mice which have been primed i.v. with SEA and treated i.p. with 200 mg/kg of cyclophosphamide 2 days later. All mature T cells bearing Vβ3 or Vβ11 were virtually abolished in the periphery of tolerant mice. These results suggest that most T cells reactive to SEA bear Vβ1, Vβ3, Vβ11 or Vβ12 and that clonal deletion of mature T cells reactive to SEA may account for the cellular mechanisms for cyclophosphamide‐induced tolerance to SEA.
Several epitopes on native human complement C9 are involved in interaction with the C5b‐8 complex and other C9 molecules
Authors: Kontermann Roland , Deppisch Reinhold , Rauterberg Ernst W.
Published: March 2007

Ten monoclonal antibodies (mAb) against native human C9 exhibiting various inhibitory effects on the hemolytic activity of C9 (Bausback, J., Kontermann, R. and Rauterberg, E. W., Immunobiology 1988. 178: 58) were further analyzed regarding their reactivities with monomeric C9 (mC9), polymerized C9 (pC9), and the non‐lytic SC5b‐9 complex in enzyme‐linked immunosorbent assay and with the membrane attack complex (MAC) generated on rabbit erythrocytes analyzed by flow cytometry. In addition, the inhibitory effects of mAb on zinc‐induced C9 polymerization were investigated. One epitope of the C‐terminal half of C9b exposed on the surface of pC9 and the MAC seems not to participate directly in lytic function or polymerization since no inhibitory effect of the respective mAb was observed. The nine other mAb directed against epitopes of the C9a part exhibit various inhibitory potentials. The mAb inhibit either hemolysis or polymerization, or both processes. Due to the reactivity with the tested antigens the mAb can be divided into two groups. mAb of the first group bind with nearly the same affinity to all four antigens, whereas mAb of the second group react preferentially with mC9 while their affinity to pC9, SC5b‐9 and the MAC is reduced. Comparison of reaction patterns and inhibitory effects strongly suggest that different epitopes on the surface of native C9 are involved in interaction of C9 with C5b‐8 and/or in C9‐C9 interaction. The finding that mAb inhibiting polymerization of C9 in vitro have no inhibitory effect on hemolysis confirms that C9 polymers are no prerequisite for lysis.
Regulation of T helper‐B lymphocyte adhesion through CD4‐HLA class II interaction
Authors: Mazerolles Fabienne , Amblard François , Lumbroso Catherine , Lecomte Olivia , van de Moortele Pierre‐François , Barbat Christiane , Piatier‐Tonneau Dominique , Auffray Charles , Fischer Alain
Published: March 2007

Antigen‐independent adhesion of CD4+ T lymphocytes to Epstein‐Barr virus (EBV)‐transformed B cells is mediated by CD2/lymphocyte function‐associated antigen (LFA)‐3 and LFA‐1/intracellular adhesion molecule (ICAM)‐1. Although some anti‐CD4 antibodies block the antigen‐independent adhesion of CD4+ T lymphocytes, the CD4‐HLA class II interaction does not appear to significantly contribute to the forces of cell adhesion since CD4+ T cells equally bind HLA class II+ and HLA class II− mutant B cells. In addition, conjugates formed between CD4+ T cells and HLA class II− B cells remain stable for at least 1 h while CD4+T/HLA class II+ B cell conjugate percentages promptly drop off. Down‐regulation of CD4 or spontaneous low expression of CD4 also results in a persistance of conjugates formed with B cells. The role of the CD4‐HLA class II interaction has been further studied by investigating the inhibitory effect of synthetic 12‐mer peptides analogous to HLA class II and containing the Arg‐Phe‐Asp‐Ser sequence conserved in the β1 domain. These peptides were previously found to inhibit HLA class II‐restricted T cell responses, this sequence being thought to be involved in CD4‐HLA class II interaction. These peptides block conjugate formation of CD4+ resting T cells or clones but not of CD8+ T cells, by interacting with the T cells as shown by preincubation experiments. Down‐regulation of CD4 or spontaneous low expression results in the loss of the inhibitory activity. The peptide‐mediated inhibition is neutralized by a soluble dimeric CD4 molecule. Alteration within the Arg‐Phe‐Asp‐Ser sequence results in a significant loss of inhibition. It is thus proposed that the CD4‐HLA class II interaction negatively regulates antigen‐independent adhesion of T cells, this interaction involving the highly conserved Arg‐Phe‐Asp‐Ser sequence in the HLA class II β1 sequence as a CD4‐binding site.
Presence of intestinal intraepithelial lymphocytes in mice with severe combined immunodeficiency disease
Authors: Croitoru Kenneth , Stead Ron H. , Bienenstock John , Fulop Gabrielle , Harnish Delsworth G. , Shultz Leonard D. , Jeffery Peter K. , Ernst Peter B.
Published: March 2007

The murine intestinal epithelium contains a heterogeneous population of intraepithelial leukocytes (IEL) most of which are granulated, Thy‐1‐CD5−CD8+. In order to assess the lineage relationship of this subgroup of IEL to peripheral T cells, we examined IEL in mice with the severe combined immunodeficiency (scid/scid) mutation, which lack T and B cells in peripheral lymphoid tissues. Electron and light microscopy showed that the intestine from scid/scid mice had granulated IEL similar to IEL in normal C.B‐17 mice. Flow cytometry of isolated IEL stained with monoclonal antibodies against Thy‐1, CD3, CD4, CD5 and CD8 showed that scid/scid mice IEL contained cells with the Thy‐1−CD4−CD5−CD8+ phenotype. Immunohistochemical staining of IEL in tissue sections with antibodies to Thy‐1 and CD8 confirmed that the Thy‐1−CD8+ cells were in the intestinal epithelium. These scid/scid IEL also lacked CD3 expression and mRNA for the Vγ7 V region gene of the γ T cell receptor. We conclude that scid/scid mice contain precursors for IEL that can differentiate into a granulated Thy‐1−CD5−CD8+ IEL in the intestine. The absence of CD8+ peripheral T cells in these mice suggests that these IEL differ from classical T cells in their ability to differentiate and express CD8 and do not require T cell receptor expression for their localization to the intestine.
Induction of interleukin 6 by human and murine recombinant interleukin 1 in mice
Authors: Libert Claude , Brouckaert Peter , Shaw Alan , Fiers Walter
Published: March 2007

Interleukin (IL) 6 is a pleistropic cytokine with activities, among others, on immune cells, hematopoietic precursor cells and hepatocytes. We have investigated the kinetics and amplitude of its in vivo induction in mice after injection of four different IL 1 species as well as murine (m) and human (h) tumor necrosis factor (TNF) and bacterial lipopolysaccharide (LPS) using a sensitive bioassay on 7TD1 cells to measure the IL 6 concentrations. Recombinant mIL 1β, administered as a single i.v. injection in mice, induced the appearance of IL 6 in the plasma with peak levels observed after 2 h. A dose‐response correlation was found between serum IL 6 levels and injected IL 1α concentrations at 3 and 8 h after the injection. We then compared the ability of h/mIL 1α, h/mIL 1β, h/mTNF and LPS to induce IL 6 in mice. We found: (a) LPS is the most potent inducer of IL 6; (b) 3 h after injection, the four IL 1 preparations had induced IL 6 levels comparable with the IL 6 levels observed after TNF injection; (c) high doses of mIL 1, α or β, but not hIL 1, resulted in a high IL 6 level persisting for over 8 h. We conclude that IL 1 is a potent inducer of IL 6 in vivo and that no major differences are observed between the four IL 1 preparations, as evaluated at 3 h after the injection. However, mIL 1α and mIL 1β, in contrast to hIL 1α and hIL 1β, induced a sustained IL 6 level over a longer time period. This pattern of prolonged IL 6 induction is even much more pronounced after mTNF injection, but not after hTNF injection.
CD1c as a target recognition structure for human T lymphocytes: Analysis with peripheral blood γ/δ cells
Authors: Faure Florence , Jitsukawa Setsuko , Miossec Christine , Hercend Thierry
Published: March 2007

It has been shown recently that one γ/δ cell line, termed IDP2, derived from a immunodeficient patient recognizes the CD1c molecule on the surface of target cells. In light of these data, we have tested 43 cloned and 11 polyclonal γ/δ cell lines derived from peripheral blood of 19 donors following nonspecific mitogenic stimulation. In this panel, which included lymphocytes expressing various combinations of γ and δ chains, only one clone, termed J2B7, was found to interact with target cells via a CD1c‐dependent recognition pathway. These J2B7 lymphocytes have, like IDP2, a δ chain which results from the frequent V1/J1 rearrangement while they use a distinct Vγ gene segment. The data support the view that the CD1c major histocompatibility complex “class I‐like” gene product does not have a pivotal contribution to the repertoire of peripheral blood γ/δ cells in adult individuals.
Origin and selection of peripheral CD4−CD8− T cells bearing α/β T cell antigen receptors in autoimmune gld mice
Authors: Wadsworth Scott , Yui Katsuyuki , Siegel Richard M. , Tenenholz Drew E. , Hirsch Joshua A. , Greene Mark I.
Published: March 2007

We have analyzed the origin and development of unusual CD4−CD8− α/β T cell receptor‐positive peripheral T cells produced in large numbers by mice homozygous for the gld mutation (C3H‐gld/gld). These mice may be an important model for investigating processes controlling T cell development. Bone marrow transfers demonstrated that the gld defect was intrinsic to bone marrow‐derived cells. Clonal deletion of potentially autoreactive cells was observed in peripheral gld CD4−CD8−, CD4+CD8−, and CD4−CD8+ T cells, as well as mature thymocytes. This suggests that gld CD4−CD8− T cells have passed through the thymus in ontogeny and that gld autoimmunity does not result from a general defect in elimination of self‐reactive thymocytes. These observations, combined with demethylation of the CD8 gene in the CD4−CD8− population, support prior expression of CD4 and/or CD8 in gld CD4−CD8− T cell ontogeny, perhaps at a CD4+CD8+ stage. Steroid sensitivity of gld thymocytes and CD4−CD8− T cells was normal. Therefore, we found no gross abnormalities in two major mechanisms of inducible cell death in the gld thymus, the clonal deletion process associated with tolerance and the steroid‐inducible endogenous endonuclease thought to be involved in apoptosis of unselected thymocytes. The data suggest that if gld CD4−CD8− T cells arise via escape from normal elimination in the thymus, they must do so by a novel defect in thymic selection (perhaps related to aberrant positive signals) and/or are expanded by an extrathymic process which allows clonal deletion to occur.
Regulation of interleukin 1 generation in immune‐activated fibroblasts
Authors: Huleihel Mahmoud , Douvdevani Amos , Segal Shraga , Apte Ron N.
Published: March 2007

In the present study we have demonstrated that fibroblasts can generate the inflammatory cytokine interleukin 1 (IL 1) under conditions similar to those abundant in cellular immune responses. Thus, induction of IL 1 requires a sequential two‐step protocol which consists of preactivation of mouse embryo fibroblasts (MEF) with crude preparations of T cell or macrophage‐derived conditioned media (CM; 72 h), followed by a challenge with lipopolysaccharide (LPS; 24 h). Unstimulated fibroblasts or such cells activated by either CM or LPS produced only low levels of IL 1, while a synergism between both signals was observed for obtaining maximal IL 1‐like activity in MEF. Each of a series of individual recombinant lymphokines and cytokines (IL 2, granulocyte/macrophage‐colony‐stimulating factor, tumor necrosis factor, IL 1β and interferons‐α, β and γ) was shown to serve as an efficient priming signal for the induction of IL 1. IL 1‐like activity in fibroblasts was detected in cell lysates or associated with the producing‐cell membrane but not in culture fluids. Immune‐stimulated fibroblasts, activated under such experimental conditions, were shown to actively transcribe mRNA of both IL 1 genes (α and β). For the expression of IL 1‐specific mRNA in fibroblasts a single stimulus, provided by either LPS or a lymphokine/cytokine, was sufficient; however, a more intense signal was observed when both stimuli were applied. The IL 1‐like biological activity of fibroblast origin was significantly reduced by anti‐IL 1α antibodies. Thus, fibroblasts, when activated by immune and bacterial products, generate IL 1 which in turn possibly amplifies cellular immune responses or inflammatory processes in connective tissues.
Variable region sequences of pathogenic anti‐mouse red blood cell autoantibodies from autoimmune NZB mice
Authors: Reininger Luc , Shibata Takanori , Ozaki Shoichi , Shirai Toshikazu , Jaton Jean‐Claude , Izui Shozo
Published: March 2007

New Zealand Black (NZB) mice spontaneously develop a severe autoimmune hemolytic anemia due to the production of anti‐mouse red blood cell (MRBC) autoantibodies. The contribution of variable region genes and somatic mutations in the pathogenicity of anti‐MRBC autoantibodies was investigated by mRNA sequencing of eight NZB anti‐MRBC monoclonal autoantibodies, among which five are capable of inducing anemia in BALB/c mice. Here we report that at least three VH gene families (J558, J606 and 3609) and five Vϰ, subgroups (Vϰ8, 9, 19, 21 and 28), in combination with several D, JH and Jϰ, gene segments, encode anti‐MRBC autoantibodies. Thus, the NZB anti‐MRBC autoantibodies, whether pathogenic or not, are encoded by a large number of immunoglobulin gene elements and by members of known VH and Vϰ, gene families with preferential usage of VH gene families most distal to the D regions. The presence of several mutations in the JH gene segments of both IgM and IgG anti‐MRBC autoantibodies, whether pathogenic or not, strongly suggests that their VH regions may be highly mutated and that the mechanism of somatic diversification might be important in the generation of anti‐MRBC autoantibodies. Our results support the idea that anti‐MRBC autoimmune responses are likely to be generated by an antigen‐driven mechanism.