bluuebunny/crossref_metadata_2025 huggingface dataset:
Identification of intrathymic T progenitor cells by expression of Thy‐1, IL 2 receptor and CD3
Authors: Nakano Naoko , Hardy Richard R. , Kishimoto Tadamitsu
Published: March 2007

Immature (L3T4−/Lyt‐2− “double‐negative”) thymocytes were separated into several functionally distinct fractions based on their expression of IL 2 receptors, Thy‐1 and CD3. The majority (60–70%) of double‐negative thymocytes in young adult mice lack detectable IL 2 receptor expression, have high levels of Thy‐1 and rapidly “progress” to a L3T4+ or L3T4+/Lyt‐2+ stage when cultured for 20 h in simple medium. In contrast, the IL 2 receptor‐positive fraction retains the double‐negative phenotype for as long as it survives in culture and addition of IL 2 has little or no effect on such cells. IL 2 does generate strong proliferation from a fraction of cells expressing low levels of Thy‐1, but not detectable IL 2 receptors. Such culture generates an unusual population of double‐negative cells that expresses the pan‐B cell molecule B220 and which kill both the NK target cell line YAC‐1 and the NK‐resistant line EL4. This Thy‐1‐low fraction includes all of the double‐negative thymocytes capable of T cell reconstitution. Thy‐1‐low fraction could be further separated into two populations with regards to CD3 expression. CD3− but not CD3+ population could reconstitute mature T cells, indicating that Thy‐1‐low, IL 2R− and CD3− cells are the most enriched population of intrathymic T cell progenitors.
Inhibition by anti‐HLA class II monoclonal antibodies of monocyte‐dependent T cell proliferation induced by monoclonal antibody OKT3
Authors: Ruggiero Giuseppina , Manzo Ciro , Fontana Silvia , Scala Giuseppe , Pirozzi Giuseppe , Ferrone Soldano , Zappacosta Serafino
Published: March 2007

The inhibitory effect of monoclonal antibodies (mAb) to monomorphic (locusrestricted and locus‐shared) and polymorphic determinants of HLA class II antigens on the monocyte‐dependent proliferation of T cells stimulated with mAb OKT3 has been studied. The effect appears to be specific, dose dependent, is not mediated by the Fc portion of mAb and reflects their interaction with the corresponding determinants. The anti‐HLA class II mAb do not have to be present in the culture throughout the incubation period, but are essential in early phases of mAb OKT3 T cell activation. Both monocytes and T cells are the targets of the inhibition exerted by the anti‐HLA class II mAb. Their inhibitory effect involves several steps in the sequence of events which leads to T cell proliferation, including interleukin (IL) 1 and 2 secretion, and IL 2 receptor expression.
T cell receptor γ chain variable gene rearrangements in acute lymphoblastic leukemias of T and B lineage
Authors: Boehm Thomas L. J. , Werle Andreas , Ganser Arnold , Kornhuber Bernhard , Drahovsky Dusan
Published: March 2007

The status of immunoglobulin and T cell receptor genes in acute lymphoblastic leukemia (ALL) of T and B lineage has been studied. Our data indicate that illegitimate gene rearrangements at immunoglobulin heavy chain (in T cell ALL), and T cell receptor β chain (in pre‐B ALL) genes are only rarely found (2 out of 30 patients). In contrast, T cell receptor γ chain gene rearrangements, characteristically found in T‐ALL, are also present in 7 of 18 patients with pre‐B ALL. Several features distinguish these illegitimate T cell receptor γ chain gene rearrangements from those in normal and leukemic T cells. Vγ genes located far upstream of the Jγ/Cγ complexes (Vγ2, Vγ3, Vγ4, Vγ5) appear to be preferentially used in normal adult peripheral blood T cells. In contrast, Vγ genes located immediately 5′ to the Jγ/Cγ complexes (Vγ8, Vγ9, Vγ10, Vγ11) predominate in Vγ‐Jγ recombinations observed in T‐ALL and pre‐B ALL. Whereas the Jγ2 region is primarily used in T cell receptor γ gene rearrangements observed in T‐ALL, those in pre‐B ALL are confined mostly to the Jγ1 region. These data suggest a limited accessibility of the T cell receptor γ chain gene locus for recombination processes in early stages of B cell differentiation.
A novel serine proteinase (HuTSP) isolated from a cloned human CD8+ cytolytic T cell line is expressed and secreted by activated CD4+ and CD8+ lymphocytes
Authors: Fruth Uli , Sinigaglia Francesco , Schlesier Michael , Kilgus Jochen , Kramer Michael D. , Simon Markus M.
Published: March 2007

A novel proteinase, termed human T cell‐associated serine proteinase (HuTSP), has been highly purified from a human CD8+ T lymphocyte clone. By using a panel of chromogenic model peptide substrates the enzyme was found to specifically recognize L‐arginine and to cleave Tos‐Gly‐Pro‐Arg‐nitroanilide with high efficiency at a pH optimum of 10.5–11. Exposure to class‐specific proteinase inhibitors revealed that HuTSP is a serine endopeptidase. The enzyme has a mol. mass of ∼ 50 kDa (non‐reduced) and of ∼ 25–30 kDa (reduced) when analyzed by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis suggesting HuTSP to be a disulfide‐linked dimer. The enzyme is shown to be inducible by lectin in both CD4+ and CD8+ lymphocytes. Moreover, HuTSP was detected in a number of independent CD4+ and CD8+ T cell clones and was found to be released from effector cells upon ligand binding to the CD3‐T cell receptor complex.
Rearrangement and organization of T cell receptor gamma chain genes in human leukemic T cell lines
Authors: Kimura Nobuhiro , Du Ran‐Pan , Mak Tak W.
Published: March 2007

Seventeen T cell leukemic lines, which are arrested at different stages of differentiation, were analyzed for γ‐chain rearrangement and expression using the constant region gene and representatives of the different families of Vγ chain genes. With the exception of the T cell line P30/OKUBO, which is arrested at the earliest stage (stage I), all the leukemic lines have rearranged their γ‐chain genes and transcripts can be detected in all but MOLT‐4 and P30/OKUBO cell lines. Although Vγ1 genes were rearranged and expressed in leukemic lines arrested at stages II, III and IV, Vγ2 and Vγ3 were only rearranged and expressed in cells at stage III (0/5 in stage II, 5/9 in stage III and 0/1 in stage 4). Thus, there may be a sequential usage of Vγ gene during human T cell ontogeny (although it is possible that this may just be a coincidence), similar to that which has been shown during murine ontogeny. Based on the rearrangement data, the genomic order of 5′ Vγ1‐Vγ2‐Vγ3‐J‐C 3′ is proposed.
A genetically determined lack of oral tolerance to ovalbumin is due to failure of the immune system to respond to intestinally derived tolerogen
Authors: Mowat Allan McI. , Lamont Alan G. , Bruce Maureen G.
Published: March 2007

In this study we have examined whether differences between mouse strains in the induction of tolerance after feeding ovalbumin (OVA) are due to differences in intestinal processing of OVA or are determined by the systemic immune system. Compared with major histocompatibility complex (MHC)‐congenic BALB/c mice, BALB/B mice develop much less tolerance of systemic delayed‐type hypersensitivity (DTH) and humoral immunity after feeding OVA and this defect is also expressed partially in (BALB/B × BALB/c)F1 animals. Serum taken from either BALB/c or BALB/B mice fed OVA 1 h before produced significant suppression of systemic DTH responses in BALB/c, but not in BALB/B mice. Although OVA‐fed BALB/B serum was slightly less tolerogenic than BALB/c serum, we conclude that the defective induction of oral tolerance in BALB/B mice is due primarily to a MHC‐influenced defect with the immune system. These findings support the idea that clinical food‐sensitive enteropathy reflects an immune response gene‐controlled defect in tolerance to dietary proteins.
Two types of gamma T cell receptors expressed by T cell acute lymphoblastic leukemias
Authors: van Dongen Jacques J. M. , Wolvers‐Tettero Ingrid L. M. , Seidman J. G. , Ang Siew‐Lan , van de Griend René J. , De Vries Evert F. R. , Borst Jannie
Published: March 2007

CD3+ cells, isolated from peripheral blood of two patients with T cell acute lymphoblastic leukemia (T‐ALL), did not react with the monoclonal antibody WT31, which is thought to recognize a framework determinant on the conventional T cell receptor (TcR), consisting of disulfide‐linked α and β chains. The T‐ALL cells of neither patient synthesized TcRα mRNA; the cells of patient DD contained only truncated (D‐J) TcRβ mRNA, while the cells of patient HZ contained truncated as well as mature (V‐D‐J) TcRβ mRNA. The leukemic cells of both patients made TcRγ mRNA. At the cell surface, the T‐ALL cells of patient DD expressed a CD3‐associated disulfide‐linked dimer, which contained the TcRγ protein. On the leukemic cells of patient HZ the TcRγ protein was present as a 41‐44‐kDa CD3‐associated subunit in a noncovalently linked form. The TcRγ genes in the T‐ALL cells of patient DD were rearranged exclusively to the Cγ1 locus, while in the T‐ALL cells of patient HZ both Cγ1 alleles were deleted and rearrangement to the Cγ2 locus had occurred. The Cγ1 gene segment, just like the TcRα and TcRβ gene segments, contains a cysteine codon in its second exon. This cysteine residue is involved in the formation of the interchain disulfide bond. The human Cγ2 gene segment, however, does not contain a cysteine codon in its second exon. The absence of the cysteine residue in Cγ2 encoded TcRγ chains explains the lack of an interchain disulfide bond in the TcR on the T‐ALL cells of patient HZ. The TcR gene configuration, as well as the expression of TcR mRNA and TcR protein as observed in these two leukemias, is consistent with a model for T cell differentiation in which the TcRγ gene rearranges first to the Cγ1 locus prior to or coinciding with D‐J joining of the TcRβ gene, followed by rearrangement to the Cγ2 locus and V‐D‐J joining of the TcRβ gene.
Response of human B cells to different anti‐immunoglobulin isotypes: Absence of a correlation between early activation events and cell proliferation
Authors: Roifman Chaim M. , Mills Gordon B. , Stewart David , Cheung Roy K. , Grinstein Sergio , Gelfand Erwin W.
Published: March 2007

Cross‐linking of surface immunoglobulin (sIg) by antibodies against IgM, IgG and IgD activates B cells and in some circumstances can induce cell proliferation. We studied the potential link between anti‐Ig‐induced changes in the cytosolic free Ca2+ concentration ([Ca2+]i), inositol phosphate production and the ability to induce cell proliferation in the presence or absence of the phorbol ester 12‐O‐tetradecanoylphorbol 13‐acetete (TPA). Anti‐IgM, but not anti‐IgD or anti‐IgG, induced cell proliferation in the presence but not the absence of TPA. Each of the antibodies induced a rapid increase in [Ca2+]i which appeared to be due to release of Ca2+ from internal stores. This was followed by a sustained increase in [Ca2+]i, apparently due to Ca2+ uptake from the extracellular medium. Anti‐IgD induced the greatest increase in [Ca2+]i, anti‐IgM induced intermediate changes and anti‐IgG the lowest change. Since inositol 1,3,5‐trisphosphate (IP3) can release Ca2+ from internal stores, we tested the ability of each anti‐Ig isotype to increase concentrations of IP3. In contrast to the change in [Ca2+]i and proliferation, anti‐IgG induced the most significant increase in IP3 concentrations. Taken together these data indicate that changes in [Ca2+]i, inositol phosphate production and anti‐Ig‐induced human B cell proliferation are not directly linked. They also demonstrate that changes in [Ca2+]i, inositol phosphate production and activation of protein kinase C are not sufficient to induce proliferation of human B cells. It appears that anti‐IgM induces an additional Ca2+‐independent, inositol phosphate‐independent and protein kinase C‐independent activation signal which can collaborate with TPA to induce B cell proliferation. The molecular events involved in this signal remain to be identified.
Recombinant human interleukin 5 is an eosinophil differentiation factor but has no activity in standard human B cell growth factor assays
Authors: Clutterbuck Elaine , Shields John G. , Gordon John , Smith Susan H. , Boyd Andrew , Callard Robin E. , Campbell Hugh D. , Young Ian G. , Sanderson Colin J.
Published: March 2007

Following the observation that mouse interleukin 5 (IL 5) is active as a B cell growth factor (BCGF) as well as an eosinophil differentiation factor, this work was carried out to test recombinant human IL 5 for BCGF activity. A highly active, partially purified batch of recombinant human IL 5 was prepared and tested for BCGF activity in four laboratories. This batch gave a 50% endpoint of 1:77450 in the human eosinophil differentiation assay, 1:983 in the mouse eosinophil differentiation assay and 1:42 in the mouse BCL1 assay, thus demonstrating that, like mouse IL 5, human IL 5 has cross‐species activity. By comparison with the assays in the mouse this batch would be expected to have 50% maximal human BCGF activity of about 1:4000. In each assay a known positive factor was used as a positive control, and there was no inhibitory activity in the preparation. However, despite the activity towards the mouse B cell lymphoma, the results showed no detectable activity in a panel of assays used to identify human BCGF and B cell differentiation factors. These assays included (a) proliferation assays with tonsillar or splenic B cells in the presence of the co‐stimulators anti‐μ or phorbol myristate acetate; (b) a restimulation assay in which tonsillar B cells are first activated with either Staphylococcus aureus Cowan 1 or a mixture of phorbol dibutyrate and ionomycin, or splenic B cells are first activated with anti‐μ; (c) production of immunoglobulin by B cells in a restimulation assay with Staphylococcus aureus Cowan 1; (d) production of immunoglobulin by the Epstein‐Barr virus‐transformed B lymphoblastoid CESS cell line; (e) the ability to stimulate proliferation of chronic lymphocytic leukemia (B‐CLL) cells freshly explanted from three different patients; (f) the ability to stimulate the B lymphoma (L4) cell line and the mature B cell (HBF1) line, and (g) the ability to replace T cells in specific antibody responses.

It therefore seems unlikely that recombinant human IL 5 is either a growth or a differentiation factor for human B cells, and raises the interesting question of the biological significance of the BCGF activity of this factor in the mouse.
Idiotypic characteristics of λ1‐bearing anti‐α(1→3)dextran response in wild mice: high frequency of responder strains and co‐expression of IdI 558 and IdI 104 on the same molecule
Authors: Juy Dominique , Drapier Anne‐Marie , Guenet Jean‐Louis , Cazenave Pierre‐André
Published: March 2007

In laboratory mouse strains, only mice with the IghCa phenotype are able to mount an immune response against the α(1→3) glycosidic linkage of the B1355 dextran. The response is highly restricted with respect to utilization of only Vγ1 and a few related members of the VH558 family, and characterized by the production of cross‐reacting (IdX), and individual (IdI) 104 and 558 idiotopes. We report in this study data showing a large degree of polymorphism in the anti‐α(1→3)Dex response among 35 wild‐derived mouse strains belonging to 4 different species. In contrast to the anti‐α(1→3)Dex response of common laboratory strains, neither this response nor expression of the IdX, and IdI 104 and 558 idiotopes are linked to the IghCa haplotype. We also demonstrate co‐expression on the same molecule of IdI 104 and IdI 558 idiotopes in wild mouse‐derived hybridomas. These data challenge the idea of the exclusive expression at the molecular level of IdI 104 or IdI 558.
Induction of sporozoite‐specific memory cells in mice immunized with a recombinant Plasmodium vivax circumsporozoite protein
Authors: Nardin Elizabeth H. , Barr Philip J. , Gibson Helen L. , Collins William E. , Nussenzweig Ruth S. , Nussenzweig Victor
Published: March 2007

A recombinant Plasmodium vivax circumsporozoite (CS) protein (rPvCS‐1) has been investigated as a possible malaria sporozoite vaccine candidate. Experiments were carried out to determine whether sporozoite‐specific memory cells develop in Swiss Webster mice immunized with rPvCS‐1. Challenge of rPvCS‐1‐immunized mice with P. vivax sporozoites resulted in a 100‐fold increase in the mean serum anti‐sporozoite antibody titer. The presence of parasite‐specific T helper cells was demonstrated using an in vitro assay. Anti‐CS antibodies were detected in the culture supernatants of spleen cells of rPvCS‐1‐immunized mice following in vitro challenge with P. vivax sporozoite extract. Immune spleen cells depleted of T cells did not produce antibodies when challenged with sporozoite extract in vitro. In conclusion, immunization of mice with the rPvCS‐1 protein induced memory T cells which recognized native CS antigen and functioned as T helper cells in the production of anti‐sporozoite antibodies both in vivo and in vitro.
Evidence that murine hematopoietic cell subset marker J11d is attached to a glycosyl‐phosphatidylinositol membrane anchor
Authors: Pierres Michel , Naquet Philippe , Barbet Jacques , Marchetto Sylvie , Marics Irène , Devaux Christian , Barad Marc , Hyman Robert , Rougon Geneviève
Published: March 2007

Glycosyl‐phosphatidylinositol (G‐PI) has been shown to serve as membrane anchor for cell surface molecules such as Thy‐1, Ly‐6‐controlled ThB and Qa antigens. Here, we present several lines of evidence indicating that the hematopoietic cell lineage (i.e. thymocytes, B cell subset and red blood cells) marker defined by the rat monoclonal antibody J11d is also a G‐PI‐linked structure. First, surface expression of the J11d‐defined molecules, and that of the related antigen B2A2, was found to be specifically reduced by treatment of thymocytes and B lymphoma or hybridoma cells with excess of Staphylococcus aureus PI‐specific phospholipase C; this enzyme also solubilizes a 35–40‐kDa material from erythrocyte microsomal membranes corresponding to the predominant J11d‐reactive red cell surface molecules. Second, Thy‐1− mutants of the BW5147, T1M1, S1A or S49 murine T lymphoma cells of the complementary classes A, B, C and E (i.e. shown to be defective in the enzymatic machinery that posttranslationally modify Thy‐1 molecules) also lack J11d, or express it at a very low level. Although directed at a G‐PI‐linked structure, the J11d monoclonal antibody, unlike other reagents to Thy‐1 or Ly‐6‐controlled antigens, failed to induce thymocyte proliferation even in the presence of phorbol myristate acetate and cross‐linker monoclonal antibody.
Human T cells from autoimmune and normal individuals can produce tumor necrosis factor
Authors: Turner Martin , Londei Marco , Feldmann Marc
Published: March 2007

T cell clones derived from patients with autoimmune diseases were found to be capable of producing tumor necrosis factor (TNF). This was demonstrated by stimulating the clones, in the absence of accessory cells, with antibodies against the Ti/T3 complex and with recombinant interleukin 2 (IL 2). Analysis of RNA extracted from these clones showed that TNF mRNA was more abundant than lymphotoxin (LT) mRNA. We also found that TNF protein in the supernatants of these clones was generally more abundant than LT as assessed by using the murine L929 cell assay. TNF production was not limited to T cells from autoimmune individuals, since the T cell tumor HUT 78 and T cells purified from the peripheral blood of healthy individuals also made TNF. Unlike the T cell clones, HUT 78 produced greater amounts of LT mRNA than TNF mRNA. Induction of TNF mRNA in T cells from healthy individuals displayed a two‐signal requirement (phorbol myristate 13‐acetate and phytohemagglutinin or OKT3 and phorbol myristate 13‐acetate), similar to that described for the induction of the T cell lymphokines IL2 and interferon‐gamma (IFN‐γ). Additionally we found that IL 2 alone was sufficient to induce TNF in these cells when they had been precultured with phytohemagglutinin for 7 days to express IL 2 receptors. The cloned T cells we have characterized also produce IFN‐γ which was detected in the supernatants of the clones using a radioimmunoassay. The evidence suggests that T cells can produce TNF and have the potential to deliver by themselves the dual and synergistic signals of TNF/LT and IFN‐γ to target cells, a process which may be of importance in the pathogenesis of human autoimmunity.
CD3+4−8−WT31− (T cell receptor γ+) cells and other unusual phenotypes are frequently detected among spontaneously interleukin 2‐responsive T lymphocytes present in the joint fluid in juvenile rheumatoid arthritis. A clonal analysis
Authors: De Maria Andrea , Malnati Mauro , Moretta Alessandro , Pende Daniela , Bottino Cristina , Casorati Giulia , Cottafava Franco , Melioli Giovanni , Mingari Maria Cristina , Migone Nicola , Romagnani Sergio , Moretta Lorenzo
Published: March 2007

T lymphocytes (E rosetting cells) isolated from the joint fluid of four patients with juvenile rheumatoid arthritis (JRA) were first analyzed for surface antigen expression. Approximately 15% of cells were CD25+ (interleukin, IL, 2 receptor positive), in addition, a remarkable proportion of cells expressed the CD2+3− phenotype. CD3+ cells outnumbered the sum of CD4+ and CD8+ cells as well as the cells reactive with the WT31 monoclonal antibody (which recognizes a framework determinant of the α/β T cell receptor). Purified T cells were cloned under culture conditions (1% phytohemagglutinin, PHA plus IL 2) which allow clonal expansion of most peripheral blood T lymphocytes. Under these conditions proliferating cells ranged from 25 to 65%; clones (derived from microcultures containing 0.5 or 0.25 cells/well) were tested for cytolytic activity against P815 cells (in the presence of PHA) or against the natural killer (NK)‐sensitive K562 target cells. Fifty‐four percent and 73% of clones obtained from the two patients with the polyarticular form of the disease displayed cytolytic activity in the lectin‐dependent assay. Cytolytic clones were 22 and 29% in the two patients with single joint involvement. About half of all cytolytic clones displayed NK‐like activity. Surface antigen analysis revealed that, in addition to conventional CD3+4+8− and CD3+4−8+, a noticeable fraction of clones (50/202) displayed unusual surface phenotypes. In particular, 33/50 coexpressed CD4 and CD8 antigens; 7/50 were CD2+3−4−8− and displayed NK‐like activity; 10/50 expressed CD3 but lacked both CD4 and CD8 antigen and did not react with the WT31 monoclonal antibody. In order to allow selective growth of IL 2‐responsive cells, T lymphocytes were also cloned directly in IL 2. As much as 57% of all clones thus obtained (48/84) displayed cytolytic activity. Moreover, about half expressed unusual surface phenotypes including CD2+3−4−8−, CD3+4+8+ and CD3+4−8−WT31−. Given the accumulation at the site of the joint involvement of unusual T cells, most of which displayed cytolytic activity and were likely to represent cells activated in vivo (IL 2 responsive), one may speculate that these cells may be involved in the injury process.
gp140, the EBV/C3d receptor (CR2) of human B lymphocytes, is involved in cell‐free phosphorylation of p120, a nuclear ribonucleoprotein
Authors: Delcayre Alain X. , Fiandino Anny , Barel Monique , Frade Raymond
Published: March 2007

gp140, the EBV/C3d receptor (EBV/C3dR; CR2), is a membrane site involved in human B cell regulation. Cross‐linking of this receptor on the cell surface by its specific ligands led to the enhancement of B cell proliferation in synergy with T cell factors. In vitro activation of human peripheral B lymphocytes by cross‐linking membrane immunoglobulins with anti‐μ antibody induced EBV/C3dR phosphorylation. These studies were pursued by analyzing cell‐free phosphorylation of EBV/C3dR isolated from Raji cell fractions, and immobilized on OKB7, a monoclonal anti‐EBV/C3dR antibody. Three EBV/C3dR‐related antigens which could be cell‐free phosphorylated were detected: gp140, the EBV/C3dR, p130 and p120. gp140, the mature form of EBV/C3dR, was isolated from plasma membrane and from purified nuclei. p130 was identified as an intracellular intermediate of EBV/C3dR glycosylation, localized in low‐density microsomes. Phosphoamino acid analysis of EBV/C3dR allowed the detection of phosphotyrosine and phosphoserine residues. These data suggest that EBV/C3dR could carry an autophosphorylation activity and could be associated to serine kinases. Using polyclonal anti‐p120 antibody and anti‐120 kDa nuclear ribocucleoprotein monoclonal antibody (mAb), p120 was identified as a nuclear ribonucleoprotein antigenically not related to EBV/C3dR. Detection of p120 on EBV/C3dR, immobilized on OKB7, was due to interactions between both antigens, instead of anti‐EBV/C3dR mAb cross‐reactivity with p120. Cell‐free phosphorylation of p120 was under the control of EBV/C3dR. However, it is not yet established whether other nuclear or membrane components were involved in the control of p120 cell‐free phosphorylation by EBV/C3dR. From the data presented herein, we propose that phosphorylation of a 120‐kDa nuclear ribonucleoprotein by EBV/C3dR‐associated kinases could represent a crucial step in in vivo regulation of human B cell activation.
Recombinant tumor necrosis factor can induce interleukin 2 receptor expression and cytolytic activity in a rat × mouse T cell hybrid
Authors: Plaetinck Geert , Declercq Wim , Tavernier Jan , Nabholz Markus , Fiers Walter
Published: March 2007

Tumor necrosis factor (TNF), an endotoxin‐induced macrophage monokine, is known for its cytotoxic and cytostatic effect on some tumor cell lines. Here we show that highly purified recombinant TNF, in combination with interleukin 2 (IL 2), can induce IL 2 receptor expression and cytolytic activity in a rat × mouse T cell hybrid (PC60). Previously, it was shown that IL 1 had a similar effect on PC60 cells. The ability of TNF to co‐induce IL 2 receptor expression suggests that it may play a role in the activation of certain lymphoid effector cells. This observation augments the growing list of biological activities attributed to TNF.
CD4+ helper T cells are required for resistance to a highly metastatic murine tumor
Authors: Schild Hans‐Jörg , Kyewski Bruno , von Hoegen Paul , Schirrmacher Volker
Published: March 2007

A role of CD4+ T helper cells in induction of tumor transplant rejection leading to complete regression of a highly metastatic DBA/2 mouse lymphoma was analyzed. Using an anti‐CD4 monoclonal antibody (GK1.5) which eliminates T helper cells in vivo and in vitro, we found that CD4+ cells are required for tumor resistance in syngeneic DBA/2 mice or allogeneic but major histocompatibility complex‐identical B10.D2 mice. In contrast, in allogeneic C57BL/6 mice tumor rejection was independent of CD4+ cells. An analogous requirement for immune CD4+ cells for in vitro induction of CD8+ tumor‐specific cytotoxic T cells was found in these respective strains. The requirement for immune CD4+ cells in vitro could be replaced by recombinant interleukin 2. These results demonstrate a role of CD4+ regulatory T cells and T‐T cell cooperation in the induction of anti‐tumor immunity and tumor rejection, and point to possible therapeutic interventions in the afferent phase of anti‐tumor immune responses.
Human monoclonal immunoglobulins that bind the human acetylcholine receptor
Authors: Eng Hubert , Lefvert Ann Kari , Mellstedt Håkan , Österborg Anders
Published: March 2007

Myeloma immunoglobulins, once thought to be without any immunological function, are now known to be reactive with many antigens, including self components. We have screened 149 monoclonal immunoglobulin samples and found 14 (9%) to react with the human acetylcholine receptor (AChR). Such anti‐AChR antibodies are often associated with the autoimmune disease myasthenia gravis (MG). The anti‐AChR binding of the myeloma components was restricted to the F(ab′)2 fragment and the affinities were similar to anti‐AChR antibodies isolated from MG patients. Despite the presence of anti‐AChR antibodies none of the patients exhibited any symptoms of MG.
Expression of Fcγ receptors on splenic T cells of mice infected with Plasmodium chabaudi adami
Authors: Langhorne Jean , Titus Julie A.
Published: March 2007

In previous studies, it was shown that mice infected with Plasmodium chabaudi adami have a deficiency in their production of IgG1 immunoglobulin, suggesting isotype‐specific immunoregulation. In order to examine this phenomenon in further detail the expression of Fcγ receptors (FcγR) on T cells obtained from mice infected with P. chabaudi was studied by flow cytometry. There was an increase in the number of splenic T cells which expressed FcγR during infection. At the peak of the acute stage of infection (10–15 days) up to 40% of T cells were positive for FcγR expression. These FcγR were present on about 40% of both Lyt‐2+and L3T4+ T cells. The isotype preference of these receptors on control Thy‐l+ T cells is IgG1 IgG2b > IgG2a as determined by an inhibition assay and fluorescence‐activated cell sorter (FACS) analysis. However, 2 to 3 weeks after infection this pattern was altered such that IgG2b and IgG2a represented the major isotypes binding to the FcγR of the L3T4+ T cell. At this stage of infection FcγR on L3T4+ cells fail to bind IgG1. In the Lyt‐2+ T cells IgG1 and IgG2b remained the best inhibitors. These data suggest that there may be changes in FcγR expression on T cells during infection reflected particularly in a decreased ability of IgG1 to bind to the FcγR of L3T4+ cells.
A new antigen identified by the monoclonal antibody UCHB1 delivers a costimulatory signal to a subset of human B cells
Authors: Armitage Richard J. , Rowe Deborah J. , Beverley Peter C. L.
Published: March 2007

A novel anti‐B cell monoclonal antibody UCHB1 is described which detects an antigen present on a selected subpopulation of both normal and leukemic B cells. Approximately 20% of normal tonsil and peripheral blood (PB) B cells are UCHB 1+ and the majority of B cells from all cases of prolymphocytic leukemia (PLL) tested, together with a low, variable percentage of PB B cells from non‐Hodgkin's lymphoma patients with centroblastic centrocytic leukemia also express this antigen.

Chronic lymphocytic leukemia and hairy cell leukemia B cells, pre‐B acute lymphoblastic leukemia and Epstein‐Barr virus transformed cell lines all lack UCHB1 positivity as do all non‐B lineage leukocytes and cell lines so far tested.

In tonsil sections UCHB 1 staining is almost completely confined to surface IgM+ mantle zone lymphocytes and follicular dendritic cells, but not B cells, within the germinal centers. UCHB 1 can induce a rise in the level of intracellular free calcium ([Ca2+]i) in PLL B cells and low concentrations of monoclonal antibody can result in the entry of these cells into cell cycle in the absence of additional factors. The proliferative effect of UCHB 1 is greatly enhanced in the presence of Staphylococcus aureus Cowan and, to a lesser extent, phorbol ester. A similar costimulatory effect is exerted on a proportion of normal tonsil B cells although here, despite inducing a rise in [Ca2+]i, UCHB1 alone does not cause cells to proliferate. UCHB 1 may well prove to be a useful antibody both to distinguish between different B cell leukemias and to study the phenotype and function of leukemic and normal B cell subpopulations.
Establishment of mouse cell lines which constitutively secrete large quantities of interleukin 2, 3, 4 or 5, using modified cDNA expression vectors
Authors: Karasuyama Hajime , Melchers Fritz
Published: March 2007

Mouse cell lines of different lineages have been established which constitutively secrete large quantities of recombinant mouse interleukins (mIL2, mIL3, mIL4 or mIL5). An existing bovine papilloma virus‐based expression vector, pBV‐lMTHA, was modified to allow transformed X63Ag8‐653 myeloma cells, NIH 3T3 fibroblasts and C127 mammary tumor cells to stably carry multiple copies of the vector, to express the inserted cDNA encoding a single interleukin constitutively, and to secrete the interleukin in high quantities.

Cell lines transformed with mIL2 cDNA stably carried 30–100 copies of the plasmid per cell and constitutively secreted biologically active mIL2 in quantities similar to those produced by murine EL4 thymoma cells or rat spleen cells stimulated with mitogens. Deletion of the 3′ untranslated region containing AT‐rich sequences from the mIL2 cDNA resulted in a 100‐fold increase in the constitutive production and secretion of mIL2 by the transformants. Addition of a heavy metal further increased the production 2 to 6‐fold. Cells transformed with 3′‐deleted mIL3 cDNA constitutively secreted 300–1000 times higher activities of mIL3 than the myelomonocytic leukemia line WEHI 3. mIL 4 produced by the similar transformants induced [3H]thymidine uptake of a T cell line, a mast cell line and B leukemia cells, and enhanced the production of IgG1 by B cells. IL 4 liters were 150 times higher than those produced by the concanavalin A‐stimulated T cell line 2.19. mIL 5 was secreted by similar transformants at 10‐fold higher titers than those produced by concanavalin A‐stimulated 2.19 T cells, as judged by the proliferation and maturation of B cell leukemia BCL1. The expression vectors should be useful in establishing eukaryotic cell lines producing proteins from full length cDNA clones at higher rates. The established cell lines secreting IL2, 3, 4 or 5 at high rate should be useful sources for these interleukins in the investigation of their function in the immune system.
Induction of lymphocyte locomotor capacity by a substratum‐dependent factor from mononuclear cells
Authors: Sundqvist Karl‐GÖSta , Otteskog Per
Published: March 2007

Media conditioned by human mononuclear cells contain a factor which renders freshly purified T lymphocytes motile and which is distinct from interleukin 1 and interleukin 2. The motility induced by this factor is morphologically indistinguishable from spontaneous lymphocyte motility. The factor is most effective when allowed to attach to the substratum. The effect of the factor on lymphocyte motility is detectable within 30 min but increases over a period of several hours. The relatively rapid action taken together with the ability to interact with both lymphocytes and the substratum point to the possibility that the factor rendering lymphocytes motile is a ligand which mediates cell‐substrate adhesion. Attempts to characterize this putative ligand using gel filtration chromatography yielded activity peaks at 500 to 600 kDa.
Human recombinant interleukin 4 induces normal B cells to produce soluble CD23/IgE‐binding factor analogous to that spontaneously released by lymphoblastoid B cell lines
Authors: Bonnefoy Jean Yves , Defrance Thierry , Peronne Catherine , Menetrier Christine , Rousset Françoise , Pène Jerome , Vries Jan E. De , Banchereau Jacques
Published: March 2007

Surface‐labeled Epstein‐Barr virus (EBV)‐transformed lymphoblastoid RPMI 8866 cells release in their supernatant a radiolabeled 25‐kDa polypeptide which reacts with the Fee RL/CD23‐specific monoclonal antibody (mAb) 25 and which binds to IgE but not IgG (IgEBF/sCD23). IgE BF/sCD23 had an isoelectric point of 4.5–5.0. The reactivity of mAb 25 with IgE BF/sCD23 allowed us to set up a radioimmunoassay for detection of IgE BF/sCD23 in cell culture supernatants. Supernatants from Fee RL/ CD23+ cell lines were found to contain IgEBF/sCD23. Addition of human recombinant interleukin 4 (IL4) to normal human B cells cultures induced the production of IgEBF/sCD23. Activation of B cells with anti‐IgM antibody coupled to beads enhanced the IL4‐induced production of IgEBF/sCD23 when compared to nonactivated B cells. This correlates with the finding that anti‐IgM antibody‐activated B cells cultured with IL4 express more FceRL/CD23 than B cells cultured with IL4 alone. The biochemical characteristics of radiolabeled IgE BF/sCD23 immunoprecipitated by mAb 25 from the supernatants of normal B cells cultured with IL 4 were identical to those of the IgEBF/sCD23 isolated from EBV‐transformed cell line supernatants. Addition of interferon‐γ to B cells cultured with IL 4 strongly decreased the level of IgE BF/sCD23 in culture supernatants correlating with the observed decrease of FceRL/CD23 on B cell surface. These data demonstrate that normal human B cells cultured in the presence of IL 4 produce an IgE‐binding factor (sCD23) biochemically and antigenically equivalent to that spontaneously produced by EBV‐transformed lymphoblastoid cell lines.
Anti‐CD3 antibodies induce T helper function for human B cell differentiation in vitro by an interleukin 2‐independent pathway
Authors: Sauerwein Robert W. , Van Meer Walter G. J. Der , Oostveen Johan W. Van , Miedema Frank , Aarden Lucien A.
Published: March 2007

Previously we have shown that interleukin 2 (IL 2) is an essential mediator in T cell‐dependent B cell differentiation induced by pokeweed mitogen. Here we show that activation with monoclonal anti‐CD3 antibodies of peripheral blood T cells led to the induction of helper activity for IgM secretion by human B cells from a prolymphatic leukemia. With the use of monoclonal antibodies against the IL2 receptor and CD3+ CD4+ CD8− chronic lymphatic leukemia T cells with a strongly reduced capacity to produce IL2, it was demonstrated that the anti‐CD3‐driven Ig secretion was obtained by an IL 2‐independent pathway. The T cell help in this system is mediated by soluble factors.
The induction of organ‐specific antibodies during the graft‐vs. ‐host reaction
Authors: Kupers Rudolf C. , Suiter Tobias , Gleichmann Ernst , Rose Noel R.
Published: March 2007

During the parent (P) into F1 hybrid graft‐vs.‐host reaction (GVHR), nuclear, leukocyte and erythrocyte autoantibodies are commonly seen. The specificity of these autoantibodies is reminiscent of those found in systemic lupus erythematosus (SLE) patients and SLE‐prone mice. Organ‐specific antibodies, however, including thyroglobulin (Tg) antibodies do not arise spontaneously. There have been conflicting reports about the ability of exogenous Tg to induce an anti‐Tg response during the GVHR. We have re‐examined this question in greater detail.

Using the murine P → F1 GVHR system, the results of this work demonstrate that mouse thyroglobulin (MTg)‐specific antibodies can be induced during a GVHR. However, mice must both be undergoing a GVHR, and have received exogenous MTg. The highest autoantibody response occurs if mice are injected with mouse thyroid extract or purified MTg at the time of P → F1 cell transfer. The anti‐MTg response is MTg dose dependent. The ability to induce anti‐MTg antibody was not major histocompatibility complex restricted, for both the DBA/2→B6D2F1 (low responder H‐2 haplotypes to MTg), and AKR or DBA/2→ AKD2FI (high/low responder → high responder haplotype) GVHR gave similar responses. The anti‐MTg titers peaked between days 7–10 and declined thereafter. In contrast, antibodies to dsDNA were not present at this early time, but developed after several weeks. We conclude that organ‐specific autoantibodies can be induced during a GVHR if the appropriate antigen(s) are presented near the time of GVHR induction.
Signals involved in T cell activation. T cell proliferation induced through the synergistic action of anti‐CD28 and anti‐CD2 monoclonal antibodies
Authors: Lier René A. W. Van , Brouwer Miranda , Aarden Lueien A.
Published: March 2007

Monoclonal antibodies (mAb) directed against the T cell differentiation antigen CD28 (Tp44) induce proliferation of resting T lymphocytes in the presence of phorbol esters. Moreover, it has been reported that such antibodies augment and sustain T cell proliferation induced by soluble antigens, phytohemagglutinin and anti‐CD3 mAb. Recently, we have shown that in monocyte‐depleted T cell suspensions, anti‐CD28 mAb 9.3 and Kolt‐2 were able to circumvent the requirement for interleukin 2 (IL2) in T cell proliferation induced by soluble anti‐CD3 antibodies. Apart from the synergy of anti‐CD28 antibodies with phorbol myristate acetate and anti‐CD3 antibodies, we found that anti‐CD28 mAb were able to induce T cell mitogenesis in combination with an E rosette‐blocking anti‐CD2 antibody. In this report, we show that antibodies directed against different epitopes on the CD2 antigen can synergize with anti‐CD28 mAb. Furthermore, we demonstrate that proliferation induced through the synergistic action of anti‐CD28 mAb with anti‐CD2 antibodies can be induced in the absence of accessory cells and is accompanied by the production of IL 2 and the excpression of IL2 receptors. We were unable to induce detectable Ca2+ mobilization through the simultaneous binding of anti‐CD28 and anti‐CD2 mAb. Taken together, these data show that IL2‐dependent proliferation can be induced through the simultaneous binding of anti‐CD28 and anti‐CD2 antibodies, possibly through phosphatidyl inositol‐independent pathways. The observations may provide further insight into the activation mechanisms of human T cells.
Nonrandom TRGγ variable gene rearrangement in normal human T cells and T cell leukemias
Authors: Migone Nicola , Casorati Giulla , Celle Paola Francia Di , Lusso Paolo , Foa Robin , Lefranc Marie‐Paule
Published: March 2007

To estimate the extent of the TRGγ variable (V) gene repertoire used in human T cell ontogeny, we have analyzed the variety of Vγ, gene rearrangements in a large series of T and non‐T acute and chronic leukemias. A limited heterogeneity of rearranged fragments was observed: only 13 types of differently rearranged fragments, four of which occurred only once, were found among 80 rearranged chromosomes. Furthermore, in the leukemic population as a whole, the frequency distribution of the most common types of rearranged Vγ, gene‐containing fragments appeared to be nonrandom (p < 0.01). Of interest is the clear preference for functional vs. nonfunctional V, genes (nonfunctional genes being those which carry frameshifts or nonsense mutations but which presumably can still rearrange due to their conserved signal sequences). We discuss the possibilities that this preference may result either from selection of the TRGγ product at some stage during T cell development or, alternatively, from an intrinsic, antigen‐independent polarity in Vγ, gene activation.
Down‐regulation of the T cell receptor by a mitogenic anti‐Thy‐1 antibody
Authors: Tomonari Kyuhei
Published: March 2007

Antigen‐specific unresponsiveness lasting at least 2 weeks can be induced in a T cell clone by 24‐h pretreatment with mitogenic anti‐T cell receptor antibodies. In this report the relationship is explored between the antigen‐specific unresponsiveness and activation pathways triggered via the T cell receptor and Thy‐1: the latter pathway is dependent on the former. A mitogenic anti‐Thy‐1 antibody (KT16) made the T cell clone unresponsive to specific antigen and to an anti‐T cell receptor antibody coupled to Sepharose. The unresponsiveness lasted for at least 7 days. However, cells made unresponsive to specific antigen in these ways (the T cell receptor and Thy‐1) could be activated by both interleukin 2 and KT16. KT16 down‐modulated the T cell receptor immediately after the pretreatment, but not on day 7 after the pretreatment. These facts indicate that the state of the unresponsiveness was caused by blocking transduc‐tion of an activation signal triggered by the T cell receptor to an activation pathway shared by the T cell receptor and Thy‐1.
The T11 (CD2) cDNA encodes a transmembrane protein which expresses T111, T112 and T113 epitopes but which does not independently mediate calcium influx: analysis by gene transfer in a baculovirus system
Authors: Alcover Andres , Chang Hsiu‐Ching , Sayre Peter H. , Hussey Rebecca E. , Reinherz Ellis L.
Published: March 2007

To determine whether a full‐length human T11 (CD2) cDNA encodes a protein which expresses all three T11 epitopes (T 111, T112, T113) and independently triggers activation in a cell other than a T lymphocyte, a baculovirus expression system was employed. Here we show that a recombinant T11 cDNA‐containing baculovirus can induce high‐level expression of T111, T112 and T113 epitopes on the surface of gut epithelial SF9 cells. However, in this environment the T11 protein cannot be triggered to transduce a signal resulting in an elevation of the cytosolic free Ca2+ as is known to occur in T lymphocytes. These results support the notion that T11 functions in a coordinate fashion with other intracellular lymphoid components.
Recombinant human granulocyte‐macrophage colony‐stimulating factor induces secretion of autoinhibitory monokines by U‐937 cells
Authors: Lindemann Albrecht , Riedel Detlev , Oster Wolfang , Mertelsmann Roland , Herrmann Friedhelm
Published: March 2007

Colony‐stimulating factors are required for survival proliferation, differentiation and functional activation of granulocytes, macrophages and their precursor cells. In the present report, however, we demonstrate antiproliferative activity of recombinant human (rh) granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) on mono‐blast cell line U‐937 and provide evidence for the involvement of tumor necrosis factor alpha TNF‐α and interleukin 1 beta (IL 1β) in its growth inhibitory action. GM‐CSF (but not granulocyte CSF, G‐CSF or macrophage CSF, M‐CSF) suppressed DNA synthesis and self renewal of U‐937 cells. Similarly, medium conditioned by U‐937 cells in response to GM‐CSF (GM‐CSF U‐937‐CM) was able to reduce clonogenicity and [3H] thymidine uptake by U‐937 cells. Since neutralization of GM‐CSF present in GM‐CSF U‐937‐CM by monoclonal antibody to GM‐CSF did not abrogate the autoinhibitory activity present in GM‐CSF U‐937‐CM, we considered the possibility that other soluble molecules are released by U‐937 cells upon GM‐CSF stimulation. Neutralization by antibodies to IL1β and TNF‐α suggested that both monokines could be the antiproliferative principle operating in GM‐CSF U‐937‐CM. Moreover, employing IL 1β‐specific enzyme‐linked immunosorbent assay, TNF‐α specific radioimmunoassay, Northern analysis using a cloned TNF‐α‐specific cDNA and an oligonucleotide probe for IL 1β, we demonstrate GM‐CSF‐inducible IL 1β and TNF‐α gene expression by U‐937 cells at the mRNA and protein level. Although M‐CSF expression was induced under similar conditions, M‐CSF failed to inhibit growth of U‐937 cells.
The molecular interactions with helper T cells which limit antigen‐specific B cell differentiation
Authors: Julius Michael H. , Rammensee Hans‐Georg , Ratcliffe Michael J. H. , Lamers Marinus C. , Langhorne Jean , Köhler Georges
Published: March 2007

Helper T (Th) cell‐dependent activation requirements for 2,4,6‐trinitrophenyl (TNP)‐specific resting B cells obtained from mice transgenic for Sp‐6 μ,χ genes were analyzed. Carrier‐specific T cell help required linked recognition of TNP carrier and was functionally restricted by the B cell major histocompatibility complex. However, histoincompatible T cell‐B cell conjugates formed by bridging surface immunoglobulin and Th cell receptor for antigen (TcR) through TNP‐conjugated anti‐TcR antibodies resulted in the efficient differentiation of TNP‐specific B cells. Thus, Th cell‐dependent cognate recognition of B cells is not obligatory. Specific conjugate formation could be obviated by using unconjugated fragments of anti‐TcR antibodies. If dimeric, these fragments supported the Th cell‐dependent differentiation of co‐cultured histoincompatible resting B cells. Unconjugated monomeric fragments were ineffective, demonstrating the necessity for TcR cross‐linking. Resting B cells from Sp‐6' mice rendered TNP‐conjugated monomeric fragments of anti‐TcR antibodies effectively multivalent, thereby satisfying conditions for the activation of co‐cultured Th cells. The results demonstrate that Th cells do not transduce activation signals through TcR recognition of B cell membrane‐associated ligand which limit the induction of B cell differentiation. Cross‐linking of TcR on Th cells is required, sufficient and can be induced through interaction with the antigen‐specific B cell surface.