bluuebunny/crossref_metadata_2025 huggingface dataset:
Anti‐B cell autoantibodies encoded by VH 4–21 genes in human fetal spleen do not require in vivo somatic selection
Authors: Parr Tyler B. , Johnson Todd A. , Silberstein Leslie E. , Kipps Thomas J.
Published: March 2007

We isolated immunoglobulin (Ig) VH4 genes that were rearranged in the genomic DNA of 160 day human fetal spleen. Productively rearranged VH 4‐21 genes were cloned into pRTM1, a human IgM expression vector. This allowed us to generate IgMϰK‐expressing transfectomas by co‐transfecting each of these constructs with pSVG‐VϰK3, an Ig ϰK light‐chain expression vector that has a variable region encoded Humkv325, a conserved VϰK gene that is frequently expressed early B cell ontogeny. We find that all transfectomas expressing IgMϰK encoded by VH 4‐21 make IgM autoantibodies reactive with i, a linear poly‐N‐acetyllactosamine determinant present on neonatal red blood cells and a B cell‐restricted isoform of the CD45 surface molecule. In contrast, a transfectoma expressing pSVG‐VϰK3 and pRTM1 containing a rearranged VH4‐59 (V71‐4) gene isolated from a chronic lymphocytic leukemia B cell population, designated WIL, produced IgMϰK antibodies that had no detectable anti‐i binding activity. However, transfectomas expressing VH 4‐21 fused onto the Ig heavy‐chain third complementarity determining region (CDR3) of WIL are found to make anti‐B cell autoantibodies with anti‐i activity. These studies indicate that VH 4‐21 genes rearranged in human fetal B cell ontogeny can encode anti‐B cell autoantibodies with a binding specificity that does not require in vivo somatic selection.
A peptide encoded by human gene MAGE‐3 and presented by HLA‐A2 induces cytolytic T lymphocytes that recognize tumor cells expressing MAGE‐3
Authors: van der Bruggen Pierre , Bastin Judy , Gajewski Thomas , Coulie Pierre G. , Boël Pascale , De Smet Charles , Traversari Catia , Townsend Alain , Boon Thierry
Published: March 2007

The human MAGE‐3 gene is expressed in many tumors of several histological types but it is silent in normal tissues, with the exception of testis. Antigens encoded by MAGE‐3 may, therefore, be useful targets for specific anti‐tumor immunization of cancer patients. We reported previously that MAGE‐3 codes for an antigenic peptide recognized on a melanoma cell line by autologous cytolytic T lymphocytes (CTL) restricted by HLA‐A1. Here we report that the MAGE‐3 gene also codes for another antigenic peptide that is recognized by CTL restricted by HLA‐A2. MAGE‐3 peptides bearing consensus anchor residues for HLA‐A2 were synthesized and tested for binding. T lymphocytes from normal individuals were stimulated with autologous irradiated lymphoblasts pulsed with each of three peptides that showed strong binding to HLA‐A2. Peptide FLWGPRALV was able to induce CTL. We obtained CTL clones that recognized not only HLA‐A2 cells pulsed with this peptide but also HLA‐A2 tumor cell lines expressing the MAGE‐3 gene. The proportion of melanoma tumors expressing this antigen should be approximately 32 % in Caucasian populations, since 49 % of individuals carry the HLA‐A2 allele and 65 % of melanomas express MAGE‐3.
Increased susceptibility of mice infected with Schistosoma mansoni to recombinant vaccinia virus: Association of viral persistence with egg granuloma formation
Authors: Actor Jeffrey K. , Marshall Margaret A. , Eltoum Isam A. , Buller R. Mark L. , Berzofsky Jay A. , Sher Alan
Published: March 2007

BALB/c mice infected 7 weeks previously with Schistosoma mansoni and challenged with a recombinant vaccinia virus vPE16 expressing the human immunodeficiency virus envelope protein gp160 show a marked delay in hepatic viral clearance as compared to mice infected with vPE16 alone. This increase in viral persistence is accompanied by reduced gp120‐specific Th1‐associated cytokine responses as well as by impaired cytotoxic T lymphocyte (CTL) activity against targets expressing epitopes of the same antigen. To investigate the contribution of these defects to the observed delay in clearance of recombinant vaccinia virus, animals were challenged with vPE16 at different times following S. mansoni infection, and virus titers in tissues and viral‐specific immune responses were measured simultaneously in the same animals. While normal resolution of virus occurred in schistosome‐infected mice prior to parasite egg deposition, persistence within the liver was observed in animals challenged during the onset and peak phase of granuloma formation (6 to 8 weeks after S. mansoni infection). At later times, when schistosomiasis is in its chronic phase, normal viral clearance returned. This time course of viral resolution correlated in part with the observed pattern of decreased Th1 cytokine production toward viral antigens but was clearly less temporally related to the defect in virus‐specific CTL activity. Immunohistochemical staining of liver sections from vaccinia/S. mansoni co‐infected mice with polyclonal anti‐vaccinia antibodies revealed that viral epitopes are localized primarily within granulomas. These experiments suggest that egg granulomas, by providing a microenvironment for viral expansion, in combination with the cytokine imbalance present during schistosome infection, can promote the expansion of vaccinia virus and possibly other viral agents.
IgG isotype distribution of local and systemic immune responses induced by influenza virus infection
Authors: Fazekas György , Rosenwirth Brigitte , Dukor Peter , Gergely János , Rajnavölgyi Éva
Published: March 2007

The IgG isotype profile of the influenza virus‐specific immune response was studied by quantitation of serum antibody (Ab) levels in correlation with the enumeration of antibody‐secreting cells (ASC) detected in the lung, spleen, mediastinal lymph nodes (MLN), Peyer's patches and bone marrow (BM). Distincst isotypic patterns for serum Ab and Ab produced by cells present at or close to the site of infection were found after primary or repeated infections. An elevated number of IgM ASC was found after primary challenge in the spleen, lung and MLN. In contrast, the site of IgA and IgG production is restricted to the lung and lymph nodes draining the site of infection. In these organs IgA, IgG2a and IgG1 ASC are found as a result of primary virus infection while viral challenge induces mostly activation of IgA‐producing cells and secretion of IgA to the lung lavage. In contrast, the majority (80‐90%) of Ab detected in the serum belong to the IgG2a subclass and their serum level is maintained at a high level during the whole period of the response. The relative level of virus‐specific serum IgG2a in correlation with the production of IgG2a Ab found predominantly in MLN and lung is highly dependent on the viral dose used for priming or challenge. As IgG2a ASC can be detected at relatively low numbers in the spleen and BM these results suggest that the production of the dominant IgG2a isotype of serum Ab occurs close to the viral challenge site. These data, however, point to distinct isotypic regulation in systemic versus local virus‐specific Ab responses.
Inhibition of the development of immediate hypersensitivity by staphylococcal enterotoxin B
Authors: Saloga Joachim , Lack Gideon , Bradley Kathrine , Renz Harald , Larsen Gary , Leung Donald Y. M. , Gelfand Erwin W.
Published: March 2007

We investigated the ability of staphylococcal enterotoxin B (SEB) to modify the immediate hypersensitivity response induced in BALB/c mice following sensitization to ovalbumin (OVA), a response mediated by OVA‐reactive Vβ8 T cells. Mice were sensitized by skin painting with OVA every second day over a period of 2 weeks. SEB, a potent activator of Vβ8+ T cells, was administered at the same site where OVA was applied (skin of the lower abdomen) following two different protocols. In protocol (A) SEB was injected intradermally 1 day before painting with OVA and on day 7; in protocol B, SEB was injected each time OVA was applied to the skin (eight times). SEB (but not SEA) altered the development of immediate hypersensitivity to OVA, as demonstrated by the reduction in allergen‐specific IgE, decreased OVA‐specific immediate skin test responsiveness, and prevented the development of increased airways responsiveness after bronchial challenge with OVA. Injections of SEB did not alter the proliferative responses of local draining lymph node cells or spleen mononuclear cells to OVA, indicating that administration of SEB did not inhibit the sensitization to OVA, but shifted the immune response away from an immediate type response (IgE/IgG1) to IgG2a, IgG2b and IgG3. Although both protocols of SEB treatment did not lead to a major deletion of the Vβ8 T cell population, they did reduce the proliferative response of Vβ8+ T cells to OVA. These data indicate that the bacterial toxin SEB is capable of modifying the immediate hypersensitivity response induced by OVA by altering the functional capacity of antigen‐reactive Vβ8 T cells.
On the various manifestations of spontaneous autoimmune diabetes in rodent models
Authors: Degermann Sylvie , Reilly Christina , Scott Bernadette , Ogata Lynn , Boehmer Harald Von , Lo David
Published: March 2007

Autoimmune (type 1) diabetes mellitus in mouse, rat, and humans shares several features, including T lymphocyte infiltration into pancreatic islets and a dependence on permissive class II major histocompatibility complex (MHC) alleles. We report here on an experimental model involving mice that express influenza hemagglutinin (HA) under the control of the insulin promoter and, at the same time, a transgenic class II MHC‐restricted T cell receptor (TcR) specific for an HA peptide. These mice spontaneously develop islet infiltrates resembling those found in NOD mice and most animals become diabetic within 8 weeks of age. Because of the availability of a clonotypic TcR antibody, we can be confident that the Ins‐HA transgene does not induce any measurable alterations in the vast majority of T cells with the transgenic TcR in primary and secondary lymphoid organs. Continuous export of large numbers of HA‐specific lymphocytes from the thymus was not required for the manifestation of the disease since mice thymectomized at 3 days after birth still developed the disease albeit with smaller infiltrates.
Elongated peptides, not the predicted nonapeptide stimulate a major histocompatibility complex class I‐restricted cytotoxic T lymphocyte clone with specificity for a bacterial heat shock protein
Authors: Schoel Bernd , Zügel Ulrich , Ruppert Thomas , Kaufmann Stefan H. E.
Published: March 2007

The peptides recognized by an H‐2Db‐restricted CD8 cytotoxic T lymphocyte (CTL) clone which is specific for the 60‐kDa mycobacterial heat shock protein (hsp) and cross‐reacts with stressed host cells were characterized. None of the nonapeptides from hsp60 conforming to the H‐2Db binding motif were able to sensitize target cells for lysis by this CTL clone. Sequence analysis of the stimulatory fraction from a trypsin digest of hsp60, together with synthetic peptide studies, defined a cluster of overlapping epitopes. Carboxy‐terminal extension by at least one amino acid of the nonamer predicted to bind best to H‐2Db was essential for CTL recognition. Two such elongated peptides, a 10‐mer and a 12‐mer stimulated the clone at similarly low concentrations in the 100 pM range. We assume that these two peptides comply best with the natural epitope. In contrast, the 11‐mer was inactive. The stimulatory 10‐mer bound to H‐2Db with an efficacy similar to that of the nonapeptide corresponding to the H‐2Db motif, as revealed by peptide induced major histocompatibility complex (MHC) surface expression on RMA‐S cells and competitive blocking of epitope recognition by the nonamer. Binding of these carboxy‐terminally extended peptides to the MHC groove can be explained by anchoring through the amino acid residue Asn in position 5 of the peptide and by intrusion of the hydrophobic carboxy‐terminal Ala (10‐mer) or Leu (12‐mer), but not Gly (11‐mer), into the hydrophobic pocket of the H‐2Db cleft. Because the carboxy‐terminal part is thus larger than predicted this region of the peptide may arch up from the binding groove. We assume that recognition of steric components of the MHC/peptide complex broaden the range of epitope specificity for a single T cell receptor. This flexibility not only promotes recognition of several overlapping peptides from a single antigen, but may also increase the chance of cross‐reaction with similar peptides from unrelated proteins, including autoantigens. Consistent with this latter assumption, the T cell clone cross‐recognizes mycobacterial hsp60 and stressed host cells.
Expression and regulation of adhesion molecules by γδ T cells from lymphoid tissues and intestinal epithelium
Authors: Chao Cheng‐Chi , Sandor Matyas , Dailey Morris O.
Published: March 2007

T cells bearing the γδ T cell receptor localize largely in epithelial tissues, but are also present at low frequency in organized secondary lymphoid organs. To assess the role of cell surface adhesion molecules in the traffic and tissue localization of γδ T cells, we compared the expression of these molecules on both αβ and γδ T cells in several lymphoid and non‐lymphoid organs. In the gut epithelium, γδ cells express less LFA‐1 (CD11a), Pgp‐1 (CD44), and α4 integrin than the corresponding αβ cells. In lymph nodes (LN) and Peyer's patches (PP), adhesion molecule expression by γδ cells is heterogeneous, with some of the cells having a phenotype similar to that of intraepithelial γδ cells and the rest expressing high levels of CD44 and L‐selectin (CD62L) but lower β7 and αM290, a phenotype more like lymph node αβ cells. Therefore, the particular set of adhesion molecules expressed by a T cell is dependent, in part, on its anatomic location. Superimposed upon this, however, are differences in expression that are based on the type of T cell; LN and PP γδ T cells express less CD44 but much more β7, αM290 and ICAM‐1 (CD54) than αβ T cells in the same organ. The differences in adhesion molecules between αβ and γδ cells are not due simply to differences in their activation status, because these molecules are regulated differently after activation through the T cell receptor (TcR)/CD3 complex. The differential expression of adhesion molecules on cells bearing a particular TcR V region suggests that distinct adhesion phenotypes may arise from prior contact with specific antigen and resultant cell activation in vivo. Lastly, the presence of high level expression of α4β7 and αM290 on L‐selectinlo γδ cells in lymph nodes suggests that these γδ cells may be uniquely capable of migrating to the gut. The differences in adhesion molecule expression and regulation between γδ and αβ T cells could explain, in part, the distinct homing and tissue localization of these T cell subsets in vivo.
Induction by transforming growth factor‐β1 of the interleukin‐1 receptor antagonist and of its intracellular form in human polymorphonuclear cells
Authors: Muzio Marta , Sironi Marina , Polentarutti Nadia , Mantovani Alberto , Colotta Francesco
Published: March 2007

The aim of this study was to examine the expression of interleukin‐1 receptor antagonist (IL‐1ra) in human polymorphonuclear cells (PMN) treated with transforming growth factor‐β1 (TGFβ1). TGFβ1 induced IL‐1ra transcripts in human circulating PMN and the induction was not blocked by protein synthesis inhibitors. Actinomycin D blocked induction by TGFβ1 of IL‐1ra transcripts, suggesting the involvement of gene transcription. The half life of IL‐1ra transcripts was prolonged by TGFβ1. By reverse transcriptase‐polymerase chain reaction, TGFβ1 was found to augment the transcripts coding for both the intracellular (keratinocyte type) and the secreted form of IL‐1ra. TGFβ1 induced the production of IL‐1ra in PMN. Induction of IL‐1ra by TGFβ1 in PMN may represent a further mechanism by which this molecule can counteract the potent pro‐inflammatory properties of IL‐1.
Functional and molecular characterization of B cell‐responsive Vδ1+ γδ T cells
Authors: Orsini Daniela L. M. , van Gils Mariëlle , Kooy Yvonne M. C. , Struyk Linda , Klein George , Elsen Peter Van Den , Koning Frits
Published: March 2007

Cells expressing the Vδ1+ gene segment are a minor γδ T cell population in human peripheral blood but predominate in epithelia and (inflamed) tissues. The characteristic dendritic‐like morphology of these γδ T cells is consistent with their putative immune surveillance role in epithelia. Their function, however, remains unknown. We and others previously reported that a subset of Vδ1+ γδ T cells proliferates after stimulation with Epstein‐Barr virus (EBV)‐transformed B lymphoblastoid cell lines (LCL), but not with fresh peripheral blood‐derived B cells. These responses were independent of the type of T cell receptor (TcR) γ chain co‐expressed with the Vδ1 chain. The in vivo relevance of this LCL‐mediated activation as well as the nature of the stimulatory ligand on the LCL is not well established. In this study, we tested the proliferative response of Vδ1+ LCL‐responsive T cells against non‐EBV‐transformed B cells, activated through CD40 by murine EL4 B5 cells, and to a panel of B cell lines differing in the expression of EBV nuclear antigen proteins and adhesion/co‐stimulatory molecules. The role of the Epstein‐Barr virus‐derived antigen in the induction of this response could be excluded as the activated (non‐EBV‐transformed) peripheral blood B cells were also able to induce a proliferative response in the LCL‐responsive Vδ1+ T cells. Therefore, the stimulatory ligand on B cells is of cellular rather than of viral origin, and its expression is up‐regulated upon activation of B cells. The expression of B7 and CD39 molecules on the surface of activated B cells appeared to be crucial since antibodies to these structures could block the induction of proliferation of the Vδ1+ T cells. Finally, we investigated the diversity of the responding Vδ1+ γδ T cell clones by sequence analysis of the TcRδ junctional regions. No restricted V‐D‐J sequences were found among the LCL‐responsive Vδ1+ T cell clones, arguing strongly against a mono‐ or oligoclonal Vδ1+ γδ T cell response to LCL. These findings may explain the presence of polyclonally activated Vδ1+ T cells in inflamed tissues where activated B cells are often present.
Detachment and cytolysis of human endothelial cells by proteinase 3
Authors: Ballieux Bart E. P. B. , Hiemstra Pieter S. , Klar‐Mohamad Ngaisah , Hagen E. Christiaan , van Es Leendert A. , van der Woude Fokko J. , Daha Mohamed R.
Published: March 2007

Activation and degranulation of polymorphonuclear leukocytes (PMN) with release of proteolytic enzymes, such as proteinase 3 (PR3) and elastase, in the vessels of patients with Wegener's granulomatosis (WG) is thought to play an important role in the vascular endothelial cell damage. We have investigated the detachment and cytolysis of 51Cr‐labeled umbilical vein endothelial cells (HUVEC) induced by highly purified, enzymatically active, PR3 and elastase. Incubation of confluent monolayers of HUVEC with 100 mU/ml of PR3 for 3 h at 37°C generally resulted in 20% detachment and 30% cytolysis. Elastase (350 mU/ml) induced approximately 40% detachment and 15% cytolysis. Both PR3‐mediated and elastase‐mediated detachment and cytolysis were fully inhibited by alpha‐1‐proteinase inhibitor (α1PI), while anti‐leukoprotease (ALP) only inhibited elastase‐mediated endothelial damage. By selective inhibition of an azurophilic granule extract with either α1PI or ALP we calculated that PR3 is responsible for 23% of the total detachment and cytolysis induced by the extract. Elastase was responsible for 60% of the detachment and 19% of the cytolysis. Detachment induced by PR3 was inhibited by three out of five IgG preparations purified from c‐ANCA‐positive sera of WG patients. PR3‐mediated cytolysis was inhibited by each of the c‐ANCA + IgG preparations and also to a limited extent by control IgG, suggesting a partial nonspecific stabilization of the endothelial cells. These studies provide evidence that besides elastase, PR3 also plays an important role in the PMN‐mediated endothelial cell damage.
Interleukin‐4 but not interleukin‐10 inhibits the production of leukemia inhibitory factor by rheumatoid synovium and synoviocytes
Authors: Dechanet Julie , Taupin Jean‐Luc , Chomarat Pascale , Rissoan Marie‐Clotilde , Moreau Jean‐François , Banchereau Jacques , Miossec Pierre
Published: March 2007

The expression of the proinflammatory cytokine leukemia inhibitory factor (LIF) has been reported in the cartilage and synovium of rheumatoid arthritis (RA) patients. Here, we show that high levels of LIF were constitutively produced by cultures of synovium pieces. Low levels of LIF were produced spontaneously by isolated synoviocytes, but interleukin (IL)‐1β caused a fourfold enhancement of this secretion. The anti‐inflammatory cytokine IL‐4 reduced the production of LIF by synovium pieces by 75%, as observed earlier with IL‐6, IL‐1β and tumor necrosis factor (TNF)‐α. IL‐4 had a direct effect since it inhibited LIF production by unstimulated and IL‐1β‐ or TNF‐α‐stimulated synoviocytes. Conversely, IL‐4 enhanced the production of IL‐6, which shares with LIF biological activities and receptor components. The inhibitory effect of IL‐4 was dose dependent and was reversed using a blocking anti‐IL‐4 receptor antibody. Similar inhibitory action of IL‐4 on LIF production was observed on synovium pieces from patients with osteoarthritis and on normal synoviocytes. IL‐10, another anti‐inflammatory cytokine acting on monocytes, had no effect on LIF production by either synovium pieces or isolated synoviocytes. Thus, the production of LIF by synovium tissue was inhibited by IL‐4 through both a direct effect on synoviocytes and an indirect effect by inhibition of the production of LIF‐inducing cytokines.
Induction of natural killer cell migration by monocyte chemotactic protein−1, −2 and −3
Authors: Allavena Paola , Bianchi Giancarlo , Zhou Dan , Van Damme Jo , Jílek Petr , Sozzani Silvano , Mantovani Alberto
Published: March 2007

Under certain physiological and pathological conditions, naturall killer (NK) cells rapidly accumulate in tissues. Chemokines are an essential component of the current paradigm of leukocyte recruitment. The present study was designed to investigate the responsiveness of NK cells to the prototypic C‐C chemokine, monocyte chemotactic protein‐1 (MCP‐1). MCP‐1 induced migration across filters of interleukin (IL)‐2‐activated NK cells, whereas it was a weak attractant for unstimulated cells. Maximal induction of migration required a positive concentration gradient between the lower and the upper compartment of the chemotaxis chamber. Preliminary characterization of the MCP‐1 receptor on NK cells indicated that the chemotactic response to MCP‐1 was blocked by pre‐treatment of cells with Bordetella pertussis toxin, and MCP‐1 but not IL‐8 displaced 125I‐labeled MCP‐1 from IL‐2‐activated NK cells.

The related chemokines MCP‐2 and MCP‐3 were also active – though less potent – attractants for activated NK cells. Thus the spectrum of action of MCP‐1, ‐2 and ‐3 encompasses NK cells and chemokines are likely to play a role in regulating extravasation of these cells.
T cell receptor and peptide‐contacting residues in the HLA‐DR17(3) β1 chain
Authors: Geluk Annemieke , Fu Xin‐Ting , van Meijgaarden Krista E. , Jansen Yanourishka Y. R. , de Vries René R. P. , Karr Robert W. , Ottenhoff Tom H. M.
Published: March 2007

Previously, we have proposed that the β1 residues 9–13, 26, 28 and 86 in HLA‐DR17, the most common subtype of DR3, might be critical for the binding of an immunodominant, mycobacterial epitope (peptide 3–13 of the 65‐kDa heat shock protein). In order to examine directly (i) which DR17 residues are involved in peptide binding, (ii) whether the same or other DR17 residues are involved in the binding of different peptides, and (iii) whether subtle differences in the mode of peptide binding can influence T cell stimulation, we have now systematically mutated 15 highly polymorphic DR17β1 residues, located in the proposed peptide binding groove of DR17, and examined the effect thereof on binding and presentation of two peptides, hsp65 p3–13 and p56–65 of the 30/31‐kDa secreted mycobacterial protein. Mutations in residues 28 (D → H) and 86 (V → G) completely eliminated binding of p3–13 and significantly reduced binding of p56–65. A mutation in residue 26 (Y → F) decreased binding of p3–13 but did not affect binding of p56–65. Substitutions of amino acid residues 28, 67, 71 and 86 in the DR17β1 chain abrogated peptide‐specific stimulation of both the p3–13‐ and the p56–65‐specific T cell clones, while specific stimulation by only one peptide was eliminated by substitution at positions 26 and 74 (p3–13) and by substitution of residues 11 and 37 (p56–65). The observation that substitution of several other peptide‐contacting DR17β1 chain residues does not significantly affect peptide binding but does affect T cell stimulation, suggests that these substitutions alter the conformation of the bound peptide.
Experimental therapy of systemic lupus erythematosus: the treatment of NZB/W mice with mouse soluble interferon‐γ receptor inhibits the onset of glomerulonephritis
Authors: Ozmen Laurence , Roman Danièle , Fountoulakis Michael , Schmid Georges , Ryffel Bernhard , Garotta Gianni
Published: March 2007

Female NZB/W F1 mice develop an autoimmune disease similar to human systemic lupus erythematosus (SLE), and ultimately die of glomerulonephritis. Starting at the age of 16 weeks NZB/W F1 mice were treated for a period of 19 weeks with soluble interferon‐γ receptor (sIFN‐γR), anti‐IFN‐γ monoclonal antibody (mAb) or IFN‐γ. All mice treated with sIFN‐γR or anti‐IFN‐γ mAb were alive 4 weeks after the treatment was discontinued, whereas 50% of mice died in the placebo groups and 78% of the mice died in the IFN‐γ‐treated group. Histologically, there was severe membrano‐proliferative glomerulonephritis in IFN‐γ‐ and placebo‐treated mice, and minimal or no mesangioproliferative disease in mice receiving sIFN‐γR or anti‐IFN‐γ mAb. The renal mononuclear infiltrate (T lymphocytes and monocytes), expression of major histocompatibility complex class II antigen and glomerular immunoglobulin and complement deposition were reduced in those mice. These data suggest that an IFN‐γ inhibitor, such as the soluble IFN‐γR, can be used for SLE therapy in the early stages of the disease.
Regulation of non‐classical protein kinase C isoenzymes in a human T cell line
Authors: Keenan Carolyn , Kelleher Dermot , Long Aideen
Published: March 2007

We have examined the expression and responses to activation, of novel/atypical protein kinase C (PKC) isoforms ε,ζ, and δ in the T cell lymphoma cell line K‐4. The effects of 1‐h phorbol 12‐myristate 13‐acetate (PMA) and OKT3 activation of K‐4 cells on PKC isoform distribution were examined. In addition, the effects of PMA‐mediated down‐regulation on the expression of PKC ε and ζ were determined using high concentrations of PMA over 24‐ and 48‐h time periods in these cells. PKC ε expression was not altered by incubation of K‐4 cells with up to 200 ng/ml PMA over a 24‐ or 48‐h period. PKC ε was down‐regulated in a concentration‐dependent manner by PMA after both 24‐ and 48‐h of activation. Expression of PKC ε was not completely depressed, however, even at the highest concentration of the phorbol ester after 48‐h incubation with PMA. The presence of PKC ε, ζ, and δ was confirmed by immunohistochemistry with distinct patterns of expression observed. PMA‐induced PKC activation for a 1‐h period resulted in a translocation of PKC δ from resting cytoplasmic/nuclear staining to a cytoplasmic aggregate. Following 1‐h activation through the T cell receptor‐associated complex CD3, PKC δ translocated from a peri‐nuclear/cytoplasmic compartment to a putative cytoskeletal location in K‐4 cells. This translocation was time dependent and redistributed to a cytoplasmic aggregate prior to the cytoskeleton. Similarily, following 1‐h activation through the T cell receptor, PKC ζ redistributed directly to what is possibly a cytoskeletal cell compartment. The cytoplasmic distribution of PKC ζ was unaltered following activation with PMA over a 1‐h time period. There was no apparent redistribution of PKC ε cytoplasmic staining pattern following a 1‐h direct or indirect activation. These results underline the differences in individual PKC isoform distribution, and responses to different stimuli, thereby providing additional evidence for the use of discrete PKC isoform signaling pathways in T cells. Furthermore, this data underlines the differences in PMA‐mediated PKC activation and activation through the T cell receptor.
Actions of the chemotactic cytokines MCP‐1, MCP‐2, MCP‐3, RANTES, MIP‐1α and MIP‐1β on human monocytes
Authors: Uguccioni Mariagrazia , D'Apuzzo Massimo , Loetscher Marcel , Dewald Beatrice , Baggiolini Marco
Published: March 2007

The activities of six synthetic CC chemokines, MCP‐1, MCP‐2, MCP‐3, RANTES, MIP‐1α and MIP‐1β on human blood monocytes were studied. All CC chemokines elicited a bimodal migration response in vitro. Highest numbers of migrating cells were obtained with the monocyte chemotactic proteins (MCP) and RANTES, somewhat lower numbers with MIP‐1α, and only weak migration with MIP‐1β. The most potent attractants were MCP‐1 and MIP‐1α which reached maximum efficacy at 0.1 to 1 nM. All CC chemokines also induced the release of N‐acetyl‐β‐D‐glucosaminidase from cytochalasin B‐pretreated monocytes. The MCP were most effective (MCP‐1 > MCP‐3 > MCP‐2), RANTES and MIP‐1α showed moderate (1/3 of MCP‐1 activity), and MIP‐1β only minimal activity. Cytosolic free Ca2+ changes and exocytosis were used to monitor receptor desensitization. Marked cross‐desensitization was observed among MCP‐1, MCP‐2 and MCP‐3 on the one hand, and RANTES, MIP‐1α and MIP‐1β on the other, indicating receptor sharing within these two subgroups of CC chemokines. The responses to RANTES, MIP‐1α and MIP‐1β were also moderately to markedly desensitized by pretreatment with MCP‐1, MCP‐2 or MCP‐3, while the responses to the MCP were virtually unaffected by pretreatment with RANTES, MIP‐1α and MIP‐1β. These results suggest that the MCP also interact with receptors recognized by RANTES, MIP‐1α and MIP‐1β, but not vice versa. Binding studies were performed with radiolabeled MCP‐1 or MIP‐1α. All MCP competed readily for labeled MCP‐1 yielding a concentration‐dependent sigmoidal displacement curve. Displacement with RANTES, MIP‐1α and MIP‐1β was observed at higher concentrations, but was not complete. Radiolabeled MIP‐1α was displaced efficiently by MIP‐1α or MIP‐1β, but only partially by RANTES. Of the MCP, only MC‐3 completely displaced MIP‐1α, while only partial displacement was observed with MCP‐1 and MCP‐2.
The clonal composition of myelin basic protein‐reactive encephalitogenic T cell populations is influenced both by the structure of relevant antigens and the nature of antigen‐presenting cells
Authors: Sun Deming , Hu Xian‐Zhen , Coleclough Christopher
Published: March 2007

Studies of experimental autoimmune encephalomyelitis (EAE) in rodents have revealed that encephalitogenic T cell lines reactive with myelin basic protein (BP) are frequently dominated by clones expressing a restricted T cell receptor repertoire. Using the rat EAE model, we have begun to examine the basis for clonal dominance within BP‐reactive T cell lines. We find that variations introduced into the standard protocol of periodic antigen stimulation produce marked shifts in the representation of different clones within encephalitogenic T cell populations. For example, altering the source of antigen‐presenting cells (APC), while holding antigen (BP) constant, and substituting BP from guinea pig (GPBP) for that of the rat antigen (RBP) with constant APC, both cause shifts in the composition of the dominant clones within BP‐reactive T cell lines. Our results suggest that: (i) adherence to an invariant protocol of antigen challenge may lead to an underestimation of the diversity of BP‐reactive encephalitogenic T cell populations; and (ii) the minor structural differences between GPBP and RBP not only cause the weak immunogenicity of RBP but also result in the alteration of different T cell subsets. These observations indicate that apparent restrictions upon the repertoire of autoimmune T cells should be interpreted with caution when such cells are elicited by immunization with foreign antigens.
Distinct splicing of CD45 mRNA in activated rat γδ cytotoxic T lymphocytes
Authors: Hansson Johan , Dohlsten Mikael , Sjögren Hans‐Olov , Hedlund Gunnar
Published: March 2007

Previous studies have demonstrated tumor‐ and allo‐specific cytotoxic γδ T lymphocytes in rats. In this report we define the surface phenotype of these T cell receptor (TCR)γδ+ T cells and demonstrate distinct CD45 mRNA splicing in activated γδ cytotoxic T lymphocytes (CTL). γδ T lymphocytes in the blood and the peritoneal cavity were TCRαβ−‐CD3+CD8α+CD45RC+ but expressed variable levels of LFA‐1 molecules. Normal peritoneal γδ T lymphocytes, peritoneal γδ T cells from rats injected with the bacterial superantigen staphylococcal enterotoxin A (SEA) as well as γδ T lymphocytes in peripheral blood were all LFA‐1low, Peritoneal γδ T cells from tumor‐, and allo‐sensitized rats were either LFA‐1low or LFA‐1high and specific cytotoxicity was highly enriched in the LFA‐1high subset. No cytolytic activity against SEA‐presenting cells was recorded in γδ T cells from SEA‐injected rats. Different isoforms of CD45 in T cells are generated by alternative mRNA splicing of exons 4, 5, 6 (or A, B and C, respectively) and the recently described alternate exon 7. CD45 splicing in sorted γδ T cells was evaluated utilizing reverse transcription polymerase chain reaction. Normal peritoneal γδ T cells expressed exon(578), exon(678), exon(78) and the extensively spliced exon(8) variant. Peritonealγδ T cells from rats sensitized with irradiated syngeneic tumor cells, allogeneic cells or bacterial superantigen SEA as well as γδ T lymphocytes in peripheral blood contained the full‐length exon(45678), as well as the exon(5678), exon(578), exon(678) and exon(78) splicing products. Notably, the exon(8) variant was also seen in peritoneal γδ T cells of SEA‐sensitized rats. Sorted tumor‐specific LFA‐1high γγ CTL expressed exon(45678), exon(5678), exon(578), exon(678) and exon(78) CD45 splicing products whereas the non‐cytolytic LFA‐1low γδ T cell subset also contained exon(8) variant. In summary, it is concluded that antigen‐specific TCRγδ+ CTL express high levels of LFA‐1 and that the splicing machinery in these cytolytic cells favors expression of the exon(45678) and exon(5678) CD45 splicing products whereas the exon(8) variant is lost. TCRαβ+ CTL express high levels of LFA‐1 but are devoid of the full‐length exon(45678) splicing product. The different CD45 splicing patterns found in αβ CTL and γδ CTL indicate different molecular requirements in respect to CD45 during activation and differentiation of these T lymphocyte subsets.
Epstein‐Barr virus isolates with the major HLA B35.01‐restricted cytotoxic T lymphocyte epitope are prevalent in a highly B35.01‐positive African population
Authors: Lee Stefan P. , Morgan Suzanne , Skinner Julia , Thomas Wendy A. , Jones Sarah Rowland , Sutton Julian , Khanna Rajiv , Whittle Hilton C. , Rickinson Alan B.
Published: March 2007

An influence of cytotoxic T lymphocyte (CTL) response over Epstein‐Barr virus (EBV) evolution was first suggested by the finding that virus isolates from highly HLA‐A11‐positive Oriental populations were specifically mutated in two immunodominant A11‐restricted CTL epitopes. Here we turn to a second HLA allele, B35.01 and show that B35.01‐restricted CTL responses in Caucasian donors reproducibly map to a single peptide epitope, YPLHEQHGM, representing residues 458–466 of the type 1 EBV nuclear antigen 3 A protein (B95.8 strain). In this case, however, most EBV isolates from a highly B35.01‐positive population (in The Gambia) either retained the CTL epitope sequence or carried a mutation (P → S at position 2) which conserved antigenicity; changes leading to reduced antigenicity (Y → N at position 1) were found in only a minority of cases. Furthermore, CTL recognizing the YPLHEQHGM epitope could be reactivated from the blood of some B35.01‐positive Gambian donors by in vitro stimulation with the synthetic peptide, indicating that epitope‐specific immunity does exist in this population. Possible differences between the A11‐based and B35.01‐based studies are discussed.
Human eosinophils express a receptor for secretory component. Role in secretory IgA‐dependent activation
Authors: Lamkhioued Bouchaib , Gounni Abdelillah Soussi , Gruart Valérie , Pierce Annick , Capron André , Capron Monique
Published: March 2007

The existence of a functional receptor for secretory component (SC) on the eosinophil membrane might explain the preferential degranulation induced by secretory IgA (sIgA) when compared to serum IgA. Indeed, flow cytometry analysis revealed that purified human SC could bind to a subpopulation (4–59%) of blood eosinophils purified from 19 patients with eosinophilia. Binding of radiolabeled human SC could be competitively inhibited using unlabeled SC or secretory IgA but not with serum IgA or IgG. Immunoprecipitation and immunosorbent chromatography using human SC revealed the presence of a major component at 15 kDa in eosinophil extracts as well as in culture supernatants but not in neutrophils. The 15‐kDa protein eluted from the human SC immunosorbent was able to bind to SC or to sIgA but not to serum IgA. Eosinophils preincubated with human SC or sIgA released eosinophil cationic protein (ECP) and eosinophil peroxidase (EPO) after addition of anti‐SC or anti‐IgA monoclonal antibody as respective cross‐linking reagents. These results indicated that binding of free or complexed SC to human eosinophils could induce eosinophil degranulation. Furthermore, the dose‐dependent inhibition by SC of mediator release induced by sIgA but not by serum IgA, suggested that the receptor for SC could be involved in the preferential degranulation mediated by sIgA. These results indicate a novel pathway of eosinophil activation and its potential involvement in mucosal immunity, particularly in inflammatory diseases associated with infiltration of eosinophils and the enhanced production of sIgA.
Involvement of the interleukin‐2 receptor γ subunit in interleukin‐4‐dependent activation of mouse hematopoietic cells and splenic B cells
Authors: Watanabe Sumiko , Kondo Motonari , Takatsu Kiyoshi , Sugamura Kazuo , Arai Ken‐Ichi
Published: March 2007

Interleukin‐4 (IL‐4) has various activities on B cells and on hematopoietic cells. We previously reported that TUGm2, a monoclonal antibody to the γ subunit of the IL‐2 receptor (IL‐2Rγ), inhibited IL‐4‐dependent proliferation of CTLL2, a cytotoxic T cell line. We proposed that IL‐2Rγ is required for the functional IL‐4 receptor (IL‐4R) in T cells. In the present work, we further examined whether or not IL‐2Rγ is involved in IL‐4R function in mouse myeloid cell lines and splenic B cells. TUGm2 suppressed the IL‐4‐induced proliferation of BA/F3 or IC2 cells, as well as of purified splenic B cells. TUGm2 partially suppressed proliferation of B cells induced by the combination of IL‐4 and anti‐immunoglobulin M (IgM) antibody. In contrast, TUGm2 had no effect on proliferation of B cells induced by anti‐IgM antibody alone or lipopolysaccharide (LPS). TUGm2 also inhibited IgE production induced by IL‐4 of LPS‐stimulated B cells. The induction of major histocompatibility complex class II molecules or CD23 by IL‐4 was virtually unaffected by TUGm2 antibody. These results indicate that IL‐2Rγ is differentially involved in various IL‐4‐dependent reactions.
T helper subset involvement in two high antibody responder lines of mice (Biozzi mice): HI (susceptible) and HII (resistant) to collagen‐induced arthritis
Authors: De Franco Marcelo , Gille‐Perramant Marie‐Françoise , Mevel Jean‐Claude , Couderc Jacques
Published: March 2007

CD4+ T helper cells play a critical role in the chronicity of collagen‐induced arthritis (CIA) in mice. The present results focus on the involvement of Th1 and Th2 subsets at the initial stage of the experimental disease in two lines of mice selected for high antibody production: HI that is susceptible, and HII that is resistant to CIA. Both lines are known to be H‐2q, display an identical full set of V‐β genes, and mount similar antibody responses to both heterologous and autologous CII. The kinetic analysis of local T cell and anti‐bovine CII antibody responses was followed by Elispot assays, the production of interferon‐γ (IFN‐γ) and IgG2a being considered indicative of a Th1 profile, and interleukin‐5 (IL‐5) as well as IgG1‐IgE, of a Th2 profile. The number of IL‐5 Elispots is constantly higher in susceptible than in resistant mice. The IFN‐γ production is rather low in HI compared to HII, and besides, preferential help is observed for the Th2‐dependent IgG1‐IgE isotype‐producing B cells in HI, while a switch‐over toward IgG2a anti‐CII isotype is found in HII. These results suggest that a Th1 preeminence at the onset of the anti‐CII response is decisive in the resistance to CIA.
Reciprocal expression of co‐stimulatory molecules, B7‐1 and B7‐2, on murine T cells following activation
Authors: Das Mercy R. Prabhu , Zamvil Scott S. , Borriello Frank , Weiner Howard L. , Sharpe Arlene H. , Kuchroo Vijay K.
Published: March 2007

The co‐stimulatory B7 molecules (B7‐1 and B7‐2) are expressed on professional antigen‐presenting cells in mice. In this study, we demonstrate that B7‐1 (CD80) and B7‐2 (CD86) are also expressed on murine T cells in the absence of major histocompatibility complex class II molecules. The temporal expression of these two molecules on T cells varies with the state of activation where resting T cells express B7‐2 but show little or no expression of B7‐1. Following activation, B7‐2 expression is down‐regulated and there is a concomitant increase in the expression of B7‐1 on the cell surface which peaks at about 72 h. Thus these two co‐stimulatory molecules are reciprocally expressed on the T cell surface. This pattern of expression of B7‐1 and B7‐2 on T cells suggests that these two molecules may have different roles in the generation and regulation of immune responses.
Spleen cells from antigen‐minimized mice are superior to spleen cells from germ‐free and conventional mice in the stimulation of primary in vitro proliferative responses to nominal antigens
Authors: Hooper D. Craig , Molowitz Eric H. , Bos Nico A. , Ploplis Victoria A. , Cebra John J.
Published: March 2007

T lymphocytes from mice reared under conditions of differential exposure to food, environmental and microbial antigens were compared for phenotypic shifts that may be associated with prior exposure to antigens as well as functional variations in the ability to respond to antigens de novo. While the intra‐epithelial CD8 T cell compartment was found to differ significantly in the type of T cell receptor predominantly expressed, CD4 T cells from various lymphoid organs of conventionally reared specific pathogen‐free (CL‐SPF) mice showed only subtle phenotypic differences from cells obtained from antigen‐minimized germ‐free (AF) and germ‐free (GF) mice. Cells derived from mice exposed to a reduced antigen load exhibited primary in vitro proliferative responses to antigens such as dinitrophenyl‐keyhole limpet hemocyanin which were significantly enhanced when compared with similar responses of cells from conventional mice. In cell mixing experiments, differences in the reactivity of T cells from the spleens of AF, GF and CL‐SPF mice were dependent on the source of the spleen cells employed as antigen‐presenting cells (APC). Experiments in which the T cell population was held constant revealed that, as APC, spleen cells from AF mice were most often superior to spleen cells from GF mice which were in turn considerably better than a similar population from SPF mice. We conclude that the enhanced primary reactivity of spleen cells from AF mice to nominal antigen in vitro is likely to be the result of a difference in the function and/or regulatory activities of the cell population employed as APC in this investigation.
Insertion of a short human immunodeficiency virus (HIV)‐2 gp36 sequence into an HIV‐1 p24 recombinant protein results in a polypeptide with potent and TCRBV‐restricted T cell triggering activity
Authors: Imberti Luisa , Sottini Alessandra , Roldan Eugenia Quirós , Albertini Alberto , Mattioli Sonia , Prati Enrica , Primi Daniele
Published: March 2007

In the present work we investigate whether artificial alterations of the structure of an inactive retrovirus‐encoded protein could transform it in a superantigen. As a model system we used a recombinant human immunodeficiency virus (HIV)‐1 p24 protein and two of its variants in which a short peptide corresponding to sequences of gp41 of HIV‐1 (HIV‐1 p24*) or gp36 of HIV‐2 (HIV‐1‐2 p24*) has been inserted nearby the carboxy‐terminal end of HIV‐1 p24. As expected both HIV‐1 p24 and HIV‐1 p24* were inactive, while HIV‐1‐2 p24* was a potent inducer of human, but not murine, T cell proliferation. The possibility that the observed activity was due to contaminants was ruled out since the proliferative response could be specifically inhibited by a monoclonal anti‐p24 antibody and by a peptide encompassing the area of HIV‐1 p24/HIV‐2 gp36 junction. Furthermore, the data exclude the possibility that the gp36 insertion is per se responsible for the observed proliferative activity.

The analysis of the functional, phenotypic and molecular properties of the responding cells demonstrated that the response was class II dependent and that the activated cells were predominantly CD4+CD8− expressing a strongly biased repertoire of TCRBV segments. Collectively, these data strongly suggest that the HIV‐1‐2 p24* fusion protein shares common functional properties typical of superantigen molecules.

Thus, our demonstration that a viral protein can be transformed into a superantigen simply by the insertion of a short peptide at the carboxy‐terminal end has important implications for understanding the mode of action of retrovirus‐encoded superantigens.
Evidence for intact costimulation via CD28 and CD27 molecules in hyporesponsive T cells from human immunodeficiency virus‐infected individuals
Authors: Meyaard Linde , Kuiper Heleen , Otto Sigrid A. , Wolthers Katja C. , van Lier René A. W. , Miedema Frank
Published: March 2007

In the activation of T cells, the primary signal is antigen‐specific and given through T cell receptor (TcR)/CD3 ligation. Furthermore, costimulatory molecules such as CD28 and CD27, provide an essential signal for activation through interaction with their ligands, present on the membrane of antigen‐presenting cells. During asymptomatic human immunodeficiency virus (HIV)‐1 infection, T cell function is progressively lost. Here, we investigated whether in the presence of impaired responses of T cells from HIV‐infected individuals to signal one, costimulation through CD28 and CD27 after interaction with their natural ligands CD80 and CD70 is intact. T cell proliferative responses to signal one in combination with CD80 or CD70 were decreased in a large fraction of asymptomatically HIV‐infected individuals. This was due to impaired responses of signal one but not to impaired responses to costimulation, since CD80 or CD70 did enhance signal one‐mediated proliferative responses to a normal extent. Moreover, in individuals with proliferative responses to signal one that were decreased to 50% of normal T cell responses, costimulation even was increased compared to controls.

Our results demonstrate that in HIV‐infected individuals the response to costimulation is relatively preserved compared to responses to the first signal and point to the defect in T cells in HIV infection being primarily in the CD3/TcR‐mediated pathway.