bluuebunny/crossref_metadata_2025 huggingface dataset:
Transfection of human CD59 complementary DNA into rat cells confers resistance to human complement
Authors: Walsh Louise A. , Tone Masahide , Waldmann Herman
Published: March 2007

We have examined the role of the human CD59 antigen in inhibiting complement‐mediated lysis by transfer and expression of a CD59 cDNA in rat cells. A cDNA encoding CD59 was subcloned into the expression vector pSFSVneo and stably transfected into the rat T cell line NB2‐6TG. Indirect immunofluorescence staining using the anti‐CD59 monoclonal antibody YTH53.1 demonstrated the presence of human CD59 antigen on transfected cells and its attachment to the cell surface by a rat glycolipid anchor. Transfected cells were found to contain a single 3.3‐kb species of CD59 mRNA by Northern blot hybridization. Immunoblotting revealed that this encoded a protein band of the same size as that observed in human erythrocytes. To determine the biological effect of expression of human CD59 in rat cells, an assay was devised which measured the relative lysis of transfected cells compared to untransfected cells in the presence of human complement and a lytic monoclonal antibody. It was observed that CD59‐transfected rat cells are less susceptible to lysis by human complement and that this effect was blocked by a F(ab')2 fragment of YTH53.1. These experiments provide a direct demonstration that CD59 can function as an homologous complement restriction factor for nucleated cells.
Molecular mechanisms underlying lymphocyte recirculation II. Differential regulation of LFA‐1 in the interaction between lymphocytes and high endothelial cells
Authors: Tamatani Takuya , Kotani Masaharu , Tanaka Toshiyuki , Miyasaka Massayuki
Published: March 2007

Although it has been suggested that LFA‐1 is one of the important molecules mediating interaction between lymphocytes and high endothelial (HE) cells, the implication was based on the observation that lymphocyte binding to high endothelial venules in frozen lymph node sections was partially inhibited by anti‐LFA‐1 monoclonal antibody at 4°. However, it has previously been unequivocally demonstrated that LFA‐1 molecule is unable to function at this low temperature.

To assess the actual involvement of LFA‐1 in lymphocyte‐HE cell interaction at body temperature, we examined effects of newly developed anti‐rat LFA‐1 and anti‐rat ICAM‐1 monoclonal antibody on binding of resting or activated lymphocytes to a rat HE cell line at 37°. We found that (a) LFA‐1/ICAM‐1‐independent pathway was predominant in the interaction between resting lymphocytes and HE cells, indicating that LFA‐1 functions little, if any, in the interaction and (b) lymphocyte triggering through CD3 significantly increased binding inducing a LFA‐1/ICAM‐1‐dependent pathway, whereas phorbol 12‐myristate 13‐acetate stimulation also enhanced lymphocyte binding but inducing a LFA‐1‐dependent/ICAM‐1‐independent pathway. In both cases no apparent alteration in LFA‐1 expression was observed, suggesting that the increase in lymphocyte adhesion was not due to a quantitative but a qualitative change induced in LFA‐1 molecule upon lymphocyte stimulation. These findings suggest that LFA‐1 is normally inert but can be “switched” upon lymphocyte stimulation to participate in lymphocyte‐HE cell interaction, and that the LFA‐1's ligand specificity can be differentially regulated at the lymphocyte‐HE venule interface.
Regulation of LFA‐1‐mediated T cell adhesion by CD4
Authors: Mazerolles Fabienne , Hauss Pascale , Barbat Christiane , Figdor Carl. G. , Fischer Alain
Published: March 2007

Heterotypic adhesion of T lymphocytes to monocytes. B lymphocytes, or other target cells is mainly mediated by LFA‐1 and CD2 molecules. Low‐affinity binding of resting T cells can be transiently up‐regulated by cross‐linking of CD3. We have previously found that binding of specific ligands to CD4 can down‐regulate adhesion of resting T cells to B cells. We now show that the enhanced adhesiveness of CD4+ T cells induced by CD3 cross‐linking using plastic‐bound anti‐CD3 antibody can also be inhibited by several CD4 ligands, i.e. anti‐CD4 antibodies, the gp160 env protein of human immunodeficiency virus, as well as by putative CD4 ligands, i.e. synthetic peptides analogous to the gp160‐binding site to CD4 (positions 418–434 and 449‐464) and a 12‐mer synthetic peptide (DR‐12) analogous to positions 35–46 of HLA class II β subunit and including the highly conserved Arg‐Phe‐Asp‐Ser (RFDS) sequence.

After CD3 cross‐linking, maximal binding of T cells to HLA class II‐positive and ‐negative B cells was similar, although binding to HLA class II‐negative B cells was more prolonged. T cells that were passively induced to up‐regulate adhesion by binding of a CD11a‐specific antibody, NKIL16, known to enhance LFA‐1‐dependent adhesiveness, were less sensitive to the inhibitory effect of the DR‐12 peptide, whereas the inhibitory effects of gp160 were preserved. The kinetics of adhesion of NKIL16‐pretreated T cells was not influenced by HLA class II expression at the B cell surface. Together, these results strongly suggest that CD4‐HLA class II interaction may down‐regulate low‐affinity adhesion of resting T cells and, to some extent, high‐affinity adhesion of T cells actively induced by CD3 cross‐linking but not passively induced by an anti‐CD11a antibody.
Interleukio 2 stimulates serine phosphorylation of CD45 in CTLL‐2.4 cells
Authors: Valentine Mary A. , Widmer Michael B. , Ledbetter Jeffrey A. , Pinault Fran , Voice Robert , Clark Edward A. , Gallis Byron , Brautigan David L.
Published: March 2007

Ligation of interleukin 2 (IL2) is known to regulate both protein tyrosine and serine/threonine phosphorylation. A family of leukocyte transmembrane proteins whose cytoplasmic domain exhibits intrinsic protein tyrosine phosphatase activity is collectively called CD45 and is identified by a set of common cell surface epitopes. Although CD45 is known to be a phosphoprotein, it is not known how phosphorylation specifically regulates its function. We therefore identified a cell line, the IL 4‐dependent line CTLL‐2.4, in which CD45 could be phosphorylated in response to addition of IL 2. These cells are a variant of an IL 2‐dependent murine cell line which were selected for long‐term growth on IL 4 but which retain the ability to proliferate on exposure to IL 2. Incubation of CTLL‐2.4 in low serum concentrations followed by stimulation with IL 2 caused a three‐ to fivefold increase in the phosphorylation of CD45 in a time‐ and concentration‐dependent manner. CD45 in non‐stimulated cells contained one major tryptic phosphopeptide, whereas, after exposure of the cells to IL 2, two new phosphopeptides were present in CD45. The pattern of IL 2‐induced phosphorylation was different from that found following addition of phorbol 12‐myristate 13‐acetate (PMA) to the cells. Although IL 2 induced rapid and potent tyrosine phosphorylation in CTLL‐2.4 cells, all of the basal and cytokine‐activated phosphorylation of CD45 occurred on serine residues. The IL 2‐stimulated phosphorylation caused no change in the amount of cell surface CD45 and no alteration of its catalytic activity using an artificial tyrosine phosphorylated substrate‐RCM‐lysozyme. We speculate that the increase in phosphorylation of CD45 may modify its association with potential substrates. The differences in the phosphorylation patterns induced by IL 2 and PMA further suggest that more than one kinase can use CD45 as substrate and that IL2 activates a protein serine/threonine kinase different from protein kinase C.
Targeting behavior of rat monoclonal IgG antibodies in vivo: role of antibody isotype, specificity and the target cell antigen density
Authors: Yousaf Nasim , Howard Jonathan C. , Williams Bryan D.
Published: March 2007

The studies described in this report were designed to investigate factors that could influence the behavior of erythrocytes following their interaction with monoclonal antibodies (mAb) in a fully homologous experimental opsonization system in vivo. The clearance profiles and tissue distribution of target erythrocytes were examined in both normal and decomplemented rats preinjected with rat IgG2a or IgG2b mAb directed against the same or different sites on RT1Aa, the classical class I major histocompatibility complex antigen of the DA rat. Complement played a major role in augmenting the clearance and promoting hepatic sequestration of target erythrocytes in rats preinjected with IgG2a mAb directed against the S site. In contrast, an intact complement system was not an essential requirement for erythrocyte clearance when S site‐specific IgG2b mAb were used. With each antibody tested, (DA × PVG)F1 cells, expressing about half as much antigen, were removed significantly slower than DA erythrocytes, this finding being more pronounced when the animals had been preinjected with mAb of the IgG2a isotype. A comparison of the tissue distribution of DA and (DA × PVG)F1 erythrocytes indicated that hepatic uptake was greater for target cells expressing higher antigen density. A considerable degree of heterogeneity was observed in the in vivo behavior of the target erythrocytes with three groups of IgG2b mAb that recognized different sites on the class I molecule. The S site‐specific IgG2b mAb were much more efficient in the hepatic Fc receptor‐mediated clearance system than were the P site‐directed mAb of the same subclass. Our results suggest that antibody specificity may also be a contributory factor, in addition to antibody isotype and target cell antigen density, in determining the fate of target cells in vivo.
Correlation between the occurrence of lupus nephritis, anti‐erythrocyte autoantibodies and Vx haplotype in NZB × 129/J and NZB × SM/J recombinant inbred murine strains
Authors: Bailey Naila C. , Bona Alexandra , Dikman Steven , Bonilla Francisco , Alt Frederick W. , Bona Constantin
Published: March 2007

The NZB mouse is genetically predisposed to the development of autoimmune disease that resembles the human autoimmune systemic lupus erythematosus and autoimmune hemolytic anemia, with increased titers of anti‐DNA, and Coombs' autoantibodies. The various autoimmune traits are controlled separately by a limited number of genes. Genetic studies have shown that several immune loci are involved in autoimmunity: T cell abnormalities, H‐2 complex and immunoglobulin genes have been implicated. In this report, we present evidence for a significant correlation of NZB Vχ1 haplotype defined by restriction fragment length polymorphism analysis with anti‐erythrocyte autoantibodies in NZB × 129/J and NZB × SM/J recombinant inbred lines.
CD45R CD4 T cell subset‐reconstituted nude rats: subset‐dependent survival of recipients and bi‐directional isoform switching
Authors: Sparshott Sheila M. , Bell Eric B. , Sarawar Sally R.
Published: March 2007

High and low molecular weight variants (CD45R) of the leukocyte common antigen (CD45) divide CD4 T helper cells into subpopulations which display distinct characteristics. In vitro and in vivo evidence suggested that the presence of the high molecular weight splice variants CD45RA or CD45RB distinguished naive CD4 T cells from memory T cells which underwent an irreversible switch to the low molecular weight isoform as a consequence of antigen encounter. In the rat monoclonal antibody MRC OX22 identifies an epitope on the CD45RB molecule. We investigated this proposed differentiation pathway by reconstituting athymic nude rats with highly purified OX22+ or OX22− CD4 T lymphocyte subsets obtained from the thoracic duct (TDL) of euthymic, congenic, allotype‐marked donors. Injection of CD4+CD45RB− (45R−) cells ensured long‐term survival of nude recipients; recipients of CD4+CD45R+ (45R+) cells died within 2–3 months of injection. Early after transfer (3–4 weeks) the progeny of both 45R+ and 45R− TDL were uniformly 45R−. With time (by 2 months) progeny of both parental types expressed the high molecular weight CD45RB isoform. Nude recipients of 45R− TDL always generated progeny a proportion of which bore the 45R+ phenotype; 3 months to 2 years post‐injection, 30%–50% of the donor‐derived CD4 T cells were 45R+. 45R− progeny isolated from primary recipients of either 45R− or 45R+ cells transferred into secondary nude recipients induced skin allograft rejection with equal effectiveness and also generated 45R+ offspring. The results indicated that CD4 T cell subsets switched between CD45R isoforms and that the change between high and low molecular weight expression was bi‐directional. The splice variants apparently are not lineage or maturation markers, but rather identify CD4 T cells that exist transiently in different functional states.
Differential reactivity of Vβ9 T cells to minor lymphocyte stimulating antigen in vitro and in vivo
Authors: Utsunomiya Yasunori , Kosaka Hiroshi , Kanagawa Osami
Published: March 2007

The monoclonal antibody (mAb) MR10‐2 is specific for mouse T cells expressing the Vβ9 chain of T cell antigen receptor. This mAb reacts with 2% of peripheral T cells in a number of different strains of mice, but in mice expressing the minor lymphocyte stimulating (Mls)‐1a antigen, Vβ9+ T cells recognized by MR10‐2 are deleted. However, we failed to demonstrate the Mls‐1a antigen‐specific response of naive, non‐activated Vβ9 T cells in vitro. On the other hand, T cells activated initially with anti‐Vβ9 (MR10‐2) mAb in vitro are capable of responding to the Mls‐1a antigen. These results demonstrate that Mls‐1a reactivity of Vβ9 T cells depends on the activation state of the T cells, suggesting a differential activation requirement between naive, non‐activated T cells and previously activated T cells.
Functional importance of CD4+ and CD8+ cells in cytotoxic lymphocytes activity and associated gene expression. Impact on the age‐related decline in lytic activity
Authors: Bloom Eda T.
Published: March 2007

Previously we found that the age‐related decline in cytotoxic lymphocyte (CTL) activity in a murine allogeneic system is associated with declining expression of the perforin and two serine esterase genes by senescent CTL. To identify the cell subsets responsible for the age‐related decrement in the generation of CTL, splenic T cells, subsets, and mixtures of subsets from aging and young BALB/c female mice were analyzed for their allo‐responsiveness to C57BL/6 spleen cells. Depletion studies revealed that CD8+ cells expressed both the lytic activity and the CTL‐associated genes, although both CD8+ and CD4+ cells were required for generation of optimal lytic and proliferative responses. Mixture experiments demonstrated that the reduced lytic activity generated by CD8+ cells from the spleens of old mice is a consequence of alterations in both CD8+ and CD4+ cells. The results of experiments in which CD4+ cells from young and old mice were mixed revealed that the alteration in CD4+ cells is consistent with a loss of function. These findings show that (a) the expression of the perforin and serine esterase genes in primary CTL is associated with CD8+ T cells in old and young mice, and exhibits an age‐related decrement consistent with that for lytic activity, and (b) the senescent decline in CTL activity is a consequence of aging decreasing activity in both the CD4+ and CD8+ subsets.
Independent regulation of 55‐kDa and 75‐kDa tumor necrosis factor receptors during activation of human peripheral blood B lymphocytes
Authors: Erikstein Bjørn K. , Smeland Erlend B. , Blomhoff Heidi Kiil , Funderud Steinar , Prydz Kristian , Lesslauer Werner , Espevik Terje
Published: March 2007

We have studied the expression of two different tumor necrosis factor receptors (TNFR; 55 kDa and 75 kDa) on resting and activated human peripheral blood B lymphocytes using specific monoclonal antibodies (mAb). Flow cytometric analysis revealed that most resting B cells expressed small amounts of the 75‐kDa TNFR, and that the 75‐kDa TNFR was markedly up‐regulated upon stimulation with anti‐μ or Staphylococcus aureus Cowan strain I (SAC). In contrast, the expression of the 55‐kDa TNFR was low on resting as well as on activated cells. B cell activation was accompanied by an increased binding of biotinylated TNF‐α, and this binding could be blocked by preincubation by utr‐1 (anti‐75‐kDa TNRF), but not the htr (anti‐55‐kDa TNFR) antibodies. Notably, a number of cytokines tested (interleukin 1 to 8, interferon‐γ, TNF‐α and ‐β) did not influence the expression of either the 75‐kDa or the 55‐kDa TNFR when given to resting B cells. Moreover, phorbol 12‐myristate 13‐acetate led to an early, marked down‐regulation of the 75‐kDa TNFR expression, followed by a later modest increase after > 24 h. In contrast to other cell systems where htr mAb have been found either to mimic or to inhibit TNF action, htr mAb had insignificant effects in assays for restimulation of preactivated B cells. However, utr‐1 markedly inhibited the TNF‐β but only partly inhibited the TNF‐α‐induced proliferation. Taken together, our data suggest that changes in 75‐kDa protein expression is responsible for the increased TNFR expression on activated vs. resting peripheral blood B cells and that this protein also may play an important functional role.
Characterization of the induction of persistence of major histocompatibility complex class II by hybrids of macrophages from Bacillus Calmette Guerin‐resistant mice
Authors: Faris Mary , Zwilling Bruce S.
Published: March 2007

Peritoneal macrophages (MΦ) from mice that are resistant to infection by Mycobacterium bovis (strain BCG) (Bcgr) can be induced to express major histocompatibility complex (MHC) class II glycoproteins (I‐A) continuously upon treatment with 100 units of recombinant interferon‐γ (rIFN‐γ). In contrast, MΦ from mice that are susceptible to BCG (Bcgs) express I‐A transiently. Persistent expression of I‐A does not require the continued synthesis of the glycoprotein. Thus, treatment with cycloheximide (CHX) reduces I‐A expression by MΦ that express I‐A transiently but does not affect the expression of I‐A that is persistently expressed. It was not possible, in these studies, to characterize the induction of persistence independent of MHC class II expression because of the 24–48 h required for MHC class II synthesis and cycling to the cell surface. During this time, persistence was also induced. To characterize persistence independent of MHC class II induction we have produced MΦ‐MΦ somatic cell hybrids that express I‐A constitutively by fusing cells from a Bcgs MΦ cell line with MΦ from Bcgr mice. Treatment of some of the hybrids with CHX reduced MHC class II expression. The MΦ hybrids required treatment with high doses of rIFN‐γ to induce CHX‐resistant I‐A expression. The induction of the persistence of I‐A, following the addition of rIFN‐γ, required a short burst of protein synthesis as well as the presence of rIFN‐γ for at least 3 h. The addition of actinomycin D simultaneously with rIFN‐γ did not prevent the induction of the persistence of I‐A expression by one of the MΦ hybrids (F6.4). In contrast, the induction of persistence of I‐A expression required a longer period of induction than was observed for hybrid F6.4, which was attributed to the requirement for new RNA and protein synthesis by the A1.8 hybridoma.
A major fraction of human intraepithelial lymphocytes simultaneously expresses the γ/δ T cell receptor, the CD8 accessory molecule and preferentially uses the Vδ1 gene segment
Authors: Deusch Kai , Lüling Friedrich , Reich Kristian , Classen Meinhard , Wagner Hermann , Pfeffer Klaus
Published: March 2007

The frequency of T cell receptor (TcR) type and the variable gene segment expression in human intraepithelial lymphocytes (IEL) from the large intestinal mucosa were studied by flow cytometry and immunohistochemistry, and compared to those in peripheral blood lymphocytes (PBL) — or lamina propria lymphocytes (LPL). Employing anti‐γ/δ TcR and anti‐α/β TcR monoclonal antibodies (mAb), flow cytometric analysis revealed that a large fraction of IEL (37%) are γ/δ T cells, whereas within LPL and PBL γ/δ T cells comprise a minor population (4.6% and 3.8% respectively). At these sites the number of γ/δ T cells labeled with anti‐CD8 mAb were 58.3% (IEL), 43.3% (LPL) and 24.4% (PBL). In situ staining of serial sections of large intestine confirmed these results. Hence, these data suggest a selective accumulation of CD8+ γ/δ T cells in the human epithelium of the large intestine. Furthermore, analysis of γ/δ TcR bearing IEL+ disclosed a marked preponderance of cells using the Vδ1 gene segment, whereas γ/δ TcR+ PBL preferentially express Vδ2. Strikingly, the majority of these Vδ1+ IEL bear the CDS molecule on their surface. These results are taken as evidence for a selective localization of Vδ1+ CD8+ γ/δ T cells in the epithelium of the large instestine.
Molecular characterization of human T cell receptor α chains including a Vδ1‐encoded variable segment
Authors: Miossec Christine , Caignard Anne , Ferradini Laurent , Roman‐Roman Sergio , Faure Florence , Michalaki Hélène , Triebel Frédéric , Hercend Thierry
Published: March 2007

Previously we have shown that a small fraction of human peripheral T cells expresses a surface receptor recognized both by the BMA031 mAb, specific for a TcR α/β framework epitope, and by the A13 mAb, putatively specific for an epitope encoded by the Vδ1 gene segment. An interleukin 2‐dependent polyclonal cell line (termed T2) was derived from such A13+BMA031+ circulating lymphocytes. The molecular characterization of the TcR chains expressed by T2 cells demonstrated indeed that the Vδ1 gene (one of the two major Vδ genes) was transcribed with the Cα gene segment. In the T2 polyclonal cell line, distinct Vδ1/Cα transcripts were all found to include the same Jα segment suggesting the existence of “hybrid” TcR α/δ chains encoded by unique Vδ1/Jα rearrangements. The present study was designed to characterize further the Vδ1/Jα rearranged genes expressed in A13+BMA031+ cells. Three additional cell lines were generated from peripheral blood of distinct adult healthy donors. Using the anchored polymerase chain reaction, it was found that 17 different Jα segments were used in the 20 Vδ1JαCα transcripts which have been studied. Together, these data indicate that Vδ1 is a “mixed” (i.e. α/δ) TcR V segment which can join with most (if not all) J segments in the α/δ locus. In addition, it can be definitely concluded that the A13 mAb recognizes a Vδ1‐encoded antigenic determinant and not a Vδ1J epitope (i.e. it can be defined and used as an anti‐Vδ1 mAb, as opposed to reagents such as for example δ‐TCS‐1).
A human monoclonal IgG reactive with a private idiotope of a monoclonal IgM with autoantibody activity against myelin‐associated glycoprotein
Authors: Aissa‐Fennira Fatma Ben , Benammar‐Elgaaied Amel , Bouguerra Annie , Schmitt Christian , Brouet Jean‐Claude , Dellagi Koussay
Published: March 2007

One hundred and thirteen sera from patients with monoclonal IgG were tested for reactivity against a panel of 13 human monoclonal IgM having various autoantibody activities: 6 to myelin‐associated glycoprotein (MAG), 2 to vimentin intermediate filament protein and 5 to red blood cell antigens [cold agglutinins with specificity directed to I antigen (3 cases), i antigen (1 case) or Pr antigen (1 case)]. One IgG was found to react with a monoclonal IgM with anti‐MAG activity. This reactivity was characterized as idiotypic and directed against a private idiotope of the monoclonal IgM. This work provides further evidence for the existence of anti‐idiotypic antibody activity of monoclonal Ig occurring in human B cell neoplasias.
Lymphokine requirements for the development of specific cytotoxic T cells from single precursors
Authors: Rogers Lesley A. , Zlotnik Albert , Lee Frank , Shortman Ken
Published: March 2007

A high cloning efficiency, filler cell‐free culture system was developed for the growth of single murine cytotoxic T lymphocyte precursors (CTLp) and their differentiation into cytotoxic T lymphocytes (CTL). The system used nonspecific stimulation with phorbol ester and calcium ionophore in the presence of recombinant lymphokines. The optimal lymphokine combination was interleukin 2 throughout, together with interferon‐γ during the first 6 days and interleukin 6 during the last 2 days of culture. Under these conditions half of all CD4−CD8+ T cells became CTL clones. The CTL were CD4−CD8+CD3+ TcR α/β+ and were derived from CD4−CD8+ Pgp‐1− precursors.
Elevated production of interferon‐γ and interleukin 4 by mature T cells from autoimmune Ipr mice correlates with Pgp‐1 (CD44) expression
Authors: Budd Ralph C. , Schumacher Jodi H. , Winslow Genine , Mosmann Tim R.
Published: March 2007

The cell surface glycoprotein, Pgp‐1 (CD44), has been shown to be a marker of murine memory T lymphocytes. When activated, Pgp‐1hi memory T cells produce strikingly higher amounts of interferon‐γ (IFN‐γ) than naive Pgp‐1lo T cells, yet both subsets make similar levels of interleukin (IL)2. Whereas Pgp‐1hi cells represent only 20%–25% of peripheral T cells from most strains, this marker is expressed by the vast majority (> 90%) of T cells from autoimmune MRL mice homozygous for the lymphoproliferation (lpr) gene. The massive lymphadenopathy that develops in lpr/lpr mice is composed of both non‐mature (CD4−CD8−) T cells as well as a greatly expanded number (up to 300‐fold) of mature (CD4+CD8−, CD4−CD8+) T cells. Paralleling the expression of high levels of Pgp‐1, we find that compared to normal mouse T cells, the lpr mature T lymphocyte subsets are also very high producers on a per cell basis of IFN‐γ and, for the CD4+ subset, IL4. Increased concentrations of IFN‐γ and IL4 produced by large numbers of lpr Pgp‐1hi mature T cells could contribute to the autoimmune syndrome in MRL lpr/lpr mice through the effects of these cytokines on augmenting MHC class II expression and production of certain classes of antibodies.
Characterization of pancreatic islet cell infiltrates in NOD mice: effect of cell transfer and transgene expression
Authors: O'reilly Lorraine A. , Hutchings Patricia R. , Crocker Paul R. , Simpson Elizabeth , Lund Torben , Kioussis Dimitris , Takei F. , Baird Joyce , Cooke Anne
Published: March 2007

Insulin‐dependent diabetes mellitus can be transferred into young irradiated non‐obese diabetic (NOD) mice by spleen cells from a diabetic NOD donor. T cells (both L3T4+ and Ly‐2+) enter the pancreas 2 weeks following transfer. They are present initially at peri‐islet locations but progressively infiltrate the islet with accompanying β cell destruction. The infiltrate is heterogeneous with respect to Vβ usage. Inflammatory macrophages (Mac‐1+, F4/80+) can be detected at peri‐islet locations at 1 week after transfer and continue to be recruited during the disease process. Their presence at the initiation of disease suggests that their primary function may be autoantigen presentation. Increased expression of major histocompatibility complex (MHC) class I molecules is observed on both endocrine and exocrine tissue in areas of infra‐islet infiltration. MHC class II and ICAM‐1 expression was restricted to the cells constituting the inflammatory infiltrate. Expression of these molecules was not observed on β cells implying that presentation of autoantigen by the β cell itself does not play a role in the β cell destruction in NOD mice.
Neonatal complement depletion results in predominant expression of a myeloma M 104E private idiotope among anti‐dextran antibodies
Authors: Weiler Eberhardt , Lehle Georg
Published: March 2007

Myeloma M 104E (IgM, Λ1) with specificity for the α(1 → 3) glucosidic linkage of dextran B 1355 S (Dex) carries two idiotopes (Id) as defined by isogenic anti‐idiotype mAb. The public Id is not influenced by two amino acid replacements in CDR 2 nor by an alternative D region sequence. It is shared by all or nearly all humoral antibodies in the primary immune response against Dex of mice carrying haplotype Igha. The private Id‐5 appears to depend on the integrity of the VH germ‐line sequence and on the particular M 104E D region sequence. It is present on a highly fluctuating but usually small fraction of primary anti‐Dex antibody. We report here that this situation is changed when mice are treated with Cobra venom anti‐complement factor (CVF) after birth and thereby were deprived of complement for the first two weeks of life. When immunized with Dex as adults the majority of anti‐Dex Ab carried the M 104E Id‐5. Thus, humoral antibody in CVF‐treated animals resembled the Ly‐1+ anti‐Dex precursor B cell population in the peritoneal cavity, while anti‐Dex Ab in animals not treated with CVF more closely corresponded to the Ly‐1− precursor B cell pool in spleen (H.‐P. Lehmann and G. Lehle, Eur. J. Immunol. 1991. 21: 1201).
Direct helper T cell‐induced B cell differentiation involves interaction between T cell antigen CD28 and B cell activation antigen B7
Authors: Damle Nitin K. , Linsley Peter S. , Ledbetter Jeffrey A.
Published: March 2007

Cognate interactions between major histocompatibility complex class II antigen (Ag)‐reactive CD4+ T helper (Th) and Ag‐presenting B cells induce first the activation of B cells and their subsequent differentiation into Ig‐secreting cells (IgSC). The Th cell‐associated homodimeric glycoprotein CD28 has been implicated as an important regulator of Th activation. Recently, B cell‐associated early activation Ag B7 has been identified as a ligand for the CD28 molecule. In this study, we have examined using monoclonal antibodies (mAb) the roles of CD28 and B7 molecules during the Th‐B cell cognate interactions leading to the differentiation of B7+ B cells. Anti‐CD28 mAb 9.3 specifically inhibited proliferative responses of CD4+ T cells to both allogeneic B cells and soluble Ag‐presenting autologous non‐T cells. In addition, anti‐CD28 mAb 9.3 inhibited Th‐induced differentiation of alloantigen‐presenting B cells into ISC. Similar inhibition of both Ag‐induced Th activation and B cell differentiation into ISC was observed using mAb BB1 which recognizes a B cell‐associated molecule B7. In contrast, non‐cognate Th‐independent exogenous interleukin 6‐induced differentiation of B7+ B cells into ISC was not inhibited by mAb to either molecule. These results clearly demonstrate the involvement of CD28 on Th and its ligand B7 on B cells during cognate Th‐B interactions leading to the differentiation of B cells. Furthermore, these results also suggest the development of new mAb‐based therapeutic approaches for exaggerated B cell activation associated with certain autoimmune diseases such as systemic lupus erythematosus.
Diversity in antigen recognition by Mycobacterium tuberculosis‐reactive T cell clones from the synovial fluid of rheumatoid arthritis patients
Authors: Res Pieter C. M. , Orsini Daniela L. M. , van Laar Jacob M. , Janson Anneke A. M. , Abou‐Zeid Christiane , de Vries Rene R. P.
Published: March 2007

In a previous study we have shown that synovial fluid mononuclear cells from many rheumatoid arthritis (RA) patients exhibit an enhanced response to M. tuberculosis antigens as compared to peripheral blood mononuclear cells. The 65‐kDa heat‐shock protein of M. tuberculosis was shown not to play an important role in this response, therefore other mycobacterial proteins must be involved. In this study we have investigated the possibility that synovial fluid T cells from RA patients predominantly recognize a limited number of M. tuberculosis antigens, as a result of a lesion‐specific activation of only those M. tuberculosis‐reactive T cells that have cross‐reacted with joint‐related autoantigens. From the synovial fluid of four RA patients M. tuberculosis‐reactive T cell clones were isolated and analyzed for their phenotype, HLA‐DR restriction and proliferation to immunoblot fractions containing sodium dodecyl sulfate‐polyacrylamide gel‐separated M. tuberculosis proteins of known molecular weight range. The overall M. tuberculosis immunoblot recognition pattern of the clones was strikingly heterogeneous. Within a panel of 15 clones 12 different antigenic specificities could be distinguished. In other words, we did not observe a dominant recognition of a few M. tuberculosis antigens by synovial fluid T cells. This argues against the hypothesis that the elevated synovial T cell reactivity against M. tuberculosis is a reflection of an in vivo expansion of a limited number of different types of M. tuberculosis‐reactive T cells as a result of a cross‐reaction with putative joint autoantigens.
Biological half‐life of normal and truncated human IgG3 in scid mice
Authors: Hassan Mohammed Sawkat , Abedi‐Valugerdi Manucher , Lefranc Gérard , Hammarströom Lennart , Smith C. I. Edvard
Published: March 2007

The fractional catabolic rate of IgG in the body is regulated with precision in order to maintain appropriate serum concentrations. We have investigated the turnover rate of normal human IgG3, a newly identified, naturally occurring, truncated form of IgG3 and a human‐mouse chimeric IgG3 antibody in an immunodeficient mouse strain, C.B‐17 scid (SCID), lacking endogenous Ig. The half‐life of non‐truncated, normal serum IgG3 as well as the chimeric antibody was about 7 days, whereas truncated IgG3 had a shorter half‐life of about 5 days. Due to the inherent immune defect, the SCID mouse could represent a versatile animal model for the determination of turnover rates, inasmuch as an immune response will not be mounted against foreign antigens.
Human antibody production in transgenic mice: expression from 100 kb of the human IgH locus
Authors: Brüggemann Marianne , Spicer Carl , Buluwela Lakjaya , Rosewell Ian , Barton Sheila , Surani M. Azim , Rabbitts Terence H.
Published: March 2007

In order to study the requirements for a human antibody repertoire to be engineered in transgenic animals, we have created mouse strains that express human immunoglobulin genes from 100 kb of cosmid DNA. Undisrupted integration of the human DNA in the mouse germ line, encoding germ‐line V, D, J and the Cμ, constant region, was achieved, and proved sufficient for the production of human IgM in the mouse serum. Co‐integration of one cosmid (containing the VH6 gene and a 36‐kb region of the J‐proximal D cluster) with a second cosmid (containing VH26, a further 38‐kb of the D cluster, JH and Cμ) results in V‐D‐J rearrangements which utilise the V gene segments from both cosmids. DNA rearrangements in the transgenic mice, similar to those seen in human DNA, were found only in spleen but not in thymus. Random hybridomas made from these transgenic mice show heterogeneous rearrangements of the human transgenes. Sequences of V‐D‐J units derived from transgene rearrangements reveal extensive N‐region and apparent D segment diversity. These results show that utilisation of human Ig genomic segments does occur in transgenic mice, paving the way for the derivation of a mouse strain that makes authentic human antibodies from inserted heavy and light chain gene loci.
Evidence for T cell receptor‐HLA class II molecule interaction in the response to superantigenic bacterial toxins
Authors: Fleischer Bernhard , Mittrücker Hans‐Willi
Published: March 2007

The staphylococcal enterotoxins and related microbial T cell mitogens stimulate T cells by cross‐linking variable parts of the T cell receptor (TcR) with MHC class II molecules on accessory or target cells. In this report we describe that a given combination of T cell, accessory cell (AC) and toxin can be non‐stimulatory. However, the same T cell can respond to the same toxin on another AC and the same AC can present the same toxin to another T cell. This indicates that in the complex formed between TcR, toxin and class II molecule an interaction between TcR and class II molecule takes place.
A recombinant extracellular domain of the human interleukin 4 receptor inhibits the biological effects of interleukin 4 on T and B lymphocytes
Authors: Garrone Pierre , Djossou Odile , Galizzi Jean‐Pierre , Banchereau Jacques
Published: March 2007

Human interleukin 4 (IL 4) acts on various hematopoietic cell types through interaction with a specific cell surface receptor (IL 4R), whose cDNA has been cloned. We have produced a cDNA encoding a soluble form of the extracellular domain of the human IL 4R (sIL 4R) and describe here the capacity of sIL 4R to antagonize the in vitro activities of IL 4 on normal B and T lymphocytes. sIL 4R inhibited IL 4‐induced proliferation of both phytohemagglutinin‐preactivated peripheral blood mononuclear cells (PBMC) and anti‐IgM co‐stimulated tonsil B cells with similar efficiency. This inhibitory activity was specific since sIL 4R did not affect IL 2‐dependent proliferation of these cells. sIL 4R also blocked IL 4‐dependent induction of the low‐affinity receptor for IgE on B cells and inhibited IgE production by IL 4‐activated PBMC. Thus, in contrast to the IL 6R extracellular domain which stimulates IL 6 biological activity, the IL 4R extracellular domain is a powerful antagonist of its specific ligand.
Identification of a major human immunodeficiency virus‐1 reverse transcriptase epitope recognized by mouse CD4+ T lymphocytes
Authors: Haas Gaby , David Raymond , Frank Reiner , Gausepohl Heinrich , Devaux Christian , Claverie Jean‐Michel , Pierres Michel
Published: March 2007

Delineation of major T helper cell recognition sites of human immunodeficiency virus (HIV‐1) proteins represents one important step in the design of an efficient acquired immune deficiency syndrome (AIDS) vaccine. Towards this end, we have explored the immunogenicity of HIV‐1BRU proteins in the mouse model. Preliminary experiments revealed that inbred mice primed with whole inactivated HIV‐1 developed strong CD4+ T cell proliferative responses to a variety of recombinant viral proteins including reverse transcriptase (RT). To characterize further the mouse T cell responses to this protein, several Ad‐ or Ed‐restricted T hybridoma cells (THC) were established from BALB/c or DBA/2 mice. These THC were tested for their capacity to recognize a series of 15‐mer synthetic overlapping peptides spanning three segments of HIV‐1 RT that had been preselected on the basis of either α‐helicity, amphipaticity, and/or for containing rare amino acid sequence patterns. Peptides corresponding to a C‐terminal region (residues 528–560) of RT were recognized by several of the THC established from RT‐primed mice. Furthermore, a non‐α‐helical peptide from this region (A3, 528–543) was capable of priming mice with different H‐2 haplotypes for both peptide A3 and native RT CD4+ T cell recognition. In addition to the recently identified RT determinant 203–219 capable of triggering both mouse and human CD8+ CTL, the present results identify a good candidate for an immunodominant RT epitope capable of eliciting RT‐specific T helper cell responses.
Mouse LFA‐3 studied with chimeric soluble CD2 shows preferential expression on lymphoid cells
Authors: Rutschmann Roland , Karjalainen Klaus
Published: March 2007

Here we have used a soluble, polyvalent form of mouse CD2, i.e. CD2‐hCμ, to detect its ligand LFA‐3. Mouse LFA‐3 is preferentially expressed on lymphoid cells unlike human LFA‐3 which shows a wide tissue distribution. Mouse LFA‐3 is one of the early surface molecules expressed on developing thymocytes appearing on the surface of fetal thymocytes on day 13 of gestation. Like human LFA‐3, the mouse homolog could be shown to be a phosphatidyl inositol‐linked membrane protein. Since mouse LFA‐3 is preferentially expressed on lymphoid cells and since CD2 is expressed by both Tand B lymphocytes, we would favor the view that the CD2/LFA‐3 adhesion system in the mouse may play a role in interactions between lymphocytes (T‐T and/or T‐B) rather than in cell interactions between lymphocytes and non‐lymphoid cells in the thymic, bone marrow or spleen microenvironments.
T cell response to myelin basic protein epitopes in multiple sclerosis patients and healthy subjects
Authors: Liblau Roland , Tournier‐Lasserve Elisabeth , Maciazek Jacqueline , Dumas Gérad , Siffert Odile , Hashim George , Bach Marie‐Anne
Published: March 2007

T cell lines and clones specific for human myelin basic protein (BP) were selected from three multiple sclerosis (MS) patients and two healthy subjects and tested for their proliferative responses to a battery of synthetic peptides, 9 to 21 amino acid residues long. The combined amino acid sequence of the peptides spanned the complete sequence of the human BP. The results suggest the development of T cells sensitized to at least four independent regions of the human BP, indicating some diversity of the human T cell repertoire to BP. However, an immunodominant T cell epitope was located in the C‐terminal region, defined by residues 149–162. This epitope was recognized by T cells from three subjects out of five (one MS patient and both healthy controls) in the context of different DR specificities. Another epitope (located in the 57–75 region) which triggered one MS patient's T cell response was also recognized by a mycobacteria‐specific T cell clone cross‐reacting with BP.
CD4+ T cells and B cells are necessary for the transfer of protective immunity to Plasmodium chabaudi chabaudi
Authors: Meding Sally J. , Langhorne Jean
Published: March 2007

It is shown here that B cells, in addition to CD4+ T cells, are necessary for the development of protective immunity to Plasmodlum chabaudi chabaudi (P. chabaudi) in mice. Reconstitution of severe combined immunodeficient (SCID) mice with immune or normal CD4+ T cells protected the majority of mice against an otherwise lethal challenge but the mice were unable to clear their parasitemias. By contrast, transfer of the same T cell populations into athymic nu/nu mice enabled the recipients to control and clear their infections, immune CD4+ T cells being most effective. Furthermore, SCID mice given CD4+ T cells from immune and normal donors simultaneously with immune B cells also could eliminate their infection. Clearance of parasitemia correlated with the presence of malariaspecific antibodies in the serum. The role of B cells and CD4+ T cells in the protective immune response to P. chabaudi is discussed.
Interleukin 2 counteracts the inhibition of cytotoxic T lymphocytes by cholera toxin in vitro and in vivo
Authors: Moscovitch‐Lopatin Miriam , Petrillo Raymond J. , Pankewycz O. G. , Hadro Edward , Bleackley Chris R. , Strom Terry B. , Wieder Kenneth J.
Published: March 2007

Cholera toxin irreversibly activates a 43‐kDa guanosine triphosphate (GTP)‐binding protein by adenosine diphosphate ribosylation, resulting in activation of adenylate cyclase and increased intracellular levels of cyclic adenosine monophosphate (cAMP). Because increases in intracellular cAMP inhibit interleukin 2 (IL 2) expression and cytotoxic T lymphocyte (CTL) generation and function in vitro and in vivo, we hypothesized that IL 2 may counteract the inhibition of CTL by cholera toxin. Activated CTL treated with IL 2 were protected from the inhibitory effects of cholera toxin. IL 2 also counteracted the inhibitory effect of cholera toxin on steady‐state levels of CTL‐specific serine esterase mRNA. Given the putative role of serine esterase for in vitro generated CTL effector activity, these results may account for recovery of CTL activity. Although IL 2 restored CTL function and serine esterase transcription, it did not block cholera toxin‐catalyzed ribosylation of the 43‐kDa GTP‐binding protein, nor did it prevent the accumulation of intracellular levels of cAMP. In vivo, C57BL/6 mice challenged with the allogeneic tumor P815 had suppressed CTL function when cholera toxin was administered. These cholera toxin‐treated mice died of tumor overgrowth, whereas untreated mice rejected the allogeneic tumor. Co‐treatment of alloimmunzed mice with cholera toxin and IL 2 prevented death from tumor overgrowth and restored CTL function; 67% of these mice survived. These data provide evidence that IL 2 acts in CTL through a mechanism independent of cholera toxin‐sensitive GTP‐binding protein in vitro and in vivo, despite elevated intracellular cAMP levels.
T and B cell collaboration: induction of motility in small, resting B cells by interleukin 4
Authors: Clinchy Birgitta , Elenström Carina , Severinson Eva , Möller Göran
Published: March 2007

In this report we investigate if IL 4 can work as a chemoattractant factor by inducing locomotion in B cells. We found that murine recombinant IL 4 (rIL 4) induced motile morphology and migration through polycarbonate micropore filters of murine, splenic B cells at an optimal concentration of 3 ng/ml. Kinetic studies revealed optimal migration at 8–16 h, although a significant response could be detected already after 1 h. Flow cytometric studies confirmed that the migrated cells were indeed B cells. We also compared the activity of small, dense B cells and large, low‐density B cells, based on Percoll gradient separation. We found no difference in IL 4‐induced motility among the two groups. Furthermore, we looked at B cells activated in vitro by preculture in lipopolysaccharide (LPS) or IL 4. Our data indicate that both LPS and IL 4 can increase the general capacity for motility in B cells after preculture for 24 h.

Tand B cell collaboration requires close cell‐cell contacts in order for T cell help to be administered to the B cell. One way of enhancing such cell contacts could be through directional cell migration induced by helper factors (chemotaxis). We suggest that IL 4 can play a role as a chemoattractant factor that enhances cell contacts between T helper cells and B cells.
Allo‐cross‐reactivity of a human neuraminidase‐specific T cell clone dependent on presentation of an endogenous B cell‐specific antigen
Authors: Kuijpers Karel C. , Leeuwen Pammy Treep‐Van , Miedema Frank , Lucas Cornells J.
Published: March 2007

T cell specific for foreign antigen recognize a complex of peptides and self‐major histocompatibility complex (MHC) molecules and can also cross‐react with allo‐MHC molecules. It remains controversial, however, what alloreactive T cells exactly recognize. It has been proposed that alloreactive T cells recognize endogenous peptides presented by allo‐MHC molecules. To test this hypothesis, we examined an influenza virus‐specific T cell clone (6H5), specific for neuraminidase N2 and restricted by HLA‐DR1. In the absence of influenza virus, this clone cross‐reacted with HLA‐DR1Dw1+ but not with HLA‐DR1Dw20+ Epstein‐Barr virus‐transformed lymphoblastoid cells (B‐LCL). Cold target inhibition experiments and the rearrangement pattern of the T cell receptor β chain indicated that 6H5 was a monoclonal T cell population most likely using the same T cell receptor for both responses. To determine whether determinants other than HLA‐DR1Dw1 itself were involved in the allorecognition of 6H5, this clone was tested on several cell types. In the absence of virus, T cell clone 6H5 responded to HLA‐DR1Dw1+ B‐LCL or activated B cells, but, surprisingly, not to other cell types expressed HLA‐DR1Dw1, including monocytes and transfected L cells.

These experiments further support the concept that recognition of allogeneic MHC (in this case HLA‐DR1Dw1) may result from a cross‐reactivity of T cells specific for a complex of foreign antigen and self‐MHC (neuraminidase N2 and HLA‐DR1Dw20). Furthermore, allorecognition of T cell clone 6H5 appears to depend upon the recognition of a complex of allogeneic MHC and a cell‐type specific endogenous peptide presented by activated B cells.