bluuebunny/crossref_metadata_2025 huggingface dataset:
Role of cellular and humoral immunity and helper cell involvement in permissiveness to infection by Schistosoma mansoni
Authors: Mendlovic Fela , Tarrab‐Hazdai Rebeca , Arnon Ruth
Published: March 2007

Cellular and humoral parameters were studied in permissive (mouse) and nonpermissive (rat) animals during the first few weeks following infection by Schistosoma mansoni. Comparison was made between the antigen‐induced proliferation of lymphocytes of the two species. Extract of cercaria, schistosomula and adult worms obtained from infected mice served as the test antigen in 3 and 6 days incubation in vitro thymidine uptake assay. Both spleen and thymus cells of mice and rats differed in their reactivity with the schistosome antigens, but the difference was much more pronounced in the thymocytes. In this case, whereas cells obtained from mice were not induced to proliferate by any of the antigens, thymocytes of the infected rat showed specific increasing proliferation rates as a function of time after infection.

In the rat, a subset of T cell population, the T helper cell, appears to be involved in the immune response, as established by the use of anti‐helper monoclonal antibody W3/25. In the presence of these antibodies the immune reactivity of the rat thymocytes with antigens from different stages of the parasite was markedly reduced. The effect of W3/25 antibodies was observed not only in vitro, in the lymphocyte proliferation assay, but also in vivo, by a significant increase in the extent of infection in the W3/25‐treated rats, as compared to untreated controls.

Differences between the two hosts were also observed in their humoral immune response against the schistosome antigens, as manifested by complement‐dependent cytotoxicity. In these assays mouse sera were more effective than rat sera. When autologous complement was evaluated, mouse complement had a higher cytotoxic effect than rat complement in killing of the parasite. Both sources of complement were less effective than guinea pig complement in causing antibody‐mediated cytotoxicity.
CD4 monoclonal antibody pairs for immunosuppression and tolerance induction
Authors: Qin Shixin , Cobbold Steve , Tighe Helen , Benjamin Richard , Waldmann Herman
Published: March 2007

A pair of rat anti‐mouse CD4 monoclonal antibodies (mAb) have been selected which bind to different epitopes of the molecule. Both the mAb are rat IgG2b and show clear synergistic activity in complement lysis in vitro. When injected together in vivo, they exhibit an improved immunosuppressive effect, compared to each antibody alone, on allogeneic graft rejection, humoral responses and on tolerance induction. Limiting dilution analysis indicates that the in vivo depletion of interleukin 2‐producing cells is improved using both mAb by 2–3‐fold over that obtained with the individual antibodies. As little as 60 ng per mouse of the CD4 antibody pair was sufficient to allow the induction of tolerance to human γ‐globulin, even without elimination of the CD4+ cells. The results suggest that appropriate antibody pairs may be good candidates for effective immunosuppressive serotherapy in man.
Selective up‐regulation by interferon‐γ of surface molecules of the Ly‐6 complex in resting T cells: the Ly‐6A/E and TAP antigens are preferentially enhanced
Authors: Dumont Francis J. , Dijkmans Roger , Palfree Roger G. E. , Boltz Robert D. , Coker Lisa
Published: March 2007

Surface molecules encoded by the murine Ly‐6 locus can transduce triggering signals in T cells and thus may play important roles in T cell function. Previously, we found that Ly‐6 molecules are up‐regulated by interferon (IFN)‐α/β in resting T cells. Here, we examined the possible influence of IFN‐γ on these molecules. Purified T cells from C57BL/6 (Ly‐6.2) and BALB/c (Ly‐6.1) mice were incubated in vitro with recombinant murine IFN‐γ and the expression of Ly‐6 antigens was measured by flow cyto‐fluorometry. It was found that both Ly‐6A/E and T cell‐activating protein (TAP) molecules are markedly enhanced while Ly‐6C is less affected. Under the same conditions, other T cell surface molecules showed no or marginal changes. The effect of IFN‐γ on Ly‐6A/E and TAP expression reached a maximum with as little as 10 U/ml and required only 18–24 h of incubation. Moreover, the enhancement of Ly‐6A expression induced by IFN‐γ was stable for at least 5 days. Analysis of T cell subsets further revealed that IFN‐γ‐induced augmentation of Ly‐6A (C57BL/6 mice) involves both Lyt‐2+ and L3T4+ cells while the increase of Ly‐6E (BALB/c mice) is more pronounced in Lyt‐2+ cells. The functional consequence of these phenotypic alterations was evaluated by studying the mitogenic responses of T cells to antibody‐mediated Ly‐6 cross‐linking in the presence of phorbol myristate acetate. Pretreatment of resting T cells with IFN‐γ dramatically increased the responses to anti‐Ly‐6A and anti‐Ly‐6E monoclonal antibodies. IFN‐γ treatment also boosted the stimulation induced by anti‐TAP monoclonal antibody when this stimulation was performed under suboptimal conditions. Therefore, IFN‐γ selectively up‐regulates the Ly‐6A/E and TAP activation pathways in resting T cells. We speculate that this effect may contribute to the immunoregulatory activities of IFN‐γ.
Human CD3+4−8−WT31−T lymphocyte expressing the putative T cell receptor γ‐gene product. A limiting dilution and clonal analysis
Authors: Moretta Lorenzo , Pende Daniela , Bottino Cristina , Migone Nicola , Ciccone Ermanno , Ferrini Silvano , Mingari Maria Cristina , Moretta Alessandro
Published: March 2007

The small peripheral blood CD3+ T cell population lacking both CD4 and CD8 surface antigens has been analyzed in the present study. Enriched CD3+4−8− populations were obtained by depletion with anti‐CD4 or anti‐CD8 monoclonal antibodies (mAb) and complement. The resulting populations contained >99% CD2+ cells, whereas CD3+ represented approximately 50%. Virtually all of the cells were CD4−8− and did not react with the WT31 mAb, specific for a framework determinant of the α/β T cell receptor (TCR). In order to analyze the molecular nature of CD3‐associated molecules in CD3+WT31− populations, cells were stimulated with 0.5% phytohemagglutinin (PHA) for 24 h and expanded for an additional 7–14 days in interleukin 2 (IL2). The resulting cells were >95% CD3+ and expressed neither CD4/CD8 nor WT31 antigen.

Cell surface iodination followed by cross‐linking and immunoprecipitation with anti‐CD3 mAb showed that CD3‐associated molecules consisted of a major 45‐kDa band and a minor band of 43 kDa. Thus, whereas CD3‐associated molecules isolated from polyclonal CD3+WT31+ populations (expanded in IL 2 under the same culture conditions) appeared as diffuse bands, CD3‐associated molecules isolated from CD3+WT31− populations displayed a homogeneous molecular mass. Northern blot analysis revealed the presence of mRNA for the TCR γ chain whereas the mRNA for the α chain was mostly represented by a truncated (1.2 kb) form. Also small amounts of a nonproductive mRNA for the β chain were detected. Freshly isolated CD3+WT31−‐enriched populations proliferated in response to PHA and concanavalin A, moreover, IL 2 was detected in the culture supernatants after cell stimulation. By applying culture conditions which allow virtually all T cells to undergo clonal expansion, approximately 1/3 CD3+WT31− were clonogenic. In addition, the large majority of proliferating microcultures lysed the K562 cell line and about half the natural killer (NK)‐resistant fresh melanoma target cells. A large number of clones derived from CD3+WT31− enriched populations by limiting dilution has been further analyzed. More than 95% of the clones were CD3+4−8−WT31−; 12/15 clones analyzed in more detail displayed NK activity and 6/15 lysed melanoma cells; in addition, all lysed P815 target cells in the presence of PHA, thus indicating that all the clonogenic CD3+WT31− cells have a cytolytic potential.
Synthesis and expression of MHC class II molecules in the absence of attached invariant chains by recombinant interferon‐γ‐activated bone marrow‐derived macrophages
Authors: Schneider Franz‐Josef , Opel Beate , Ballhausen Wolfgang , Henkes Wolfgang , Steinlein Peter , Reske Konrad
Published: March 2007

Pure populations of in vitro propagated bone marrow‐derived macrophages are constitutively Ia negative. Co‐culturing of these cells with recombinant interferon‐gamma (rIFN‐γ) resulted in the appearance of high amounts of Ia antigens at the cell surface of essentially all cells. The continuous presence of the stimulus was a prerequisite for sustained Ia expression because removal of the stimulus resulted in rapid decline of surface Ia.

Two‐dimensional (2D) gel analysis (1D isoelectric focusing, 2D sodium dodecyl sul‐fate‐polyacrylamide gel electrophoresis) of class II molecules synthesized by rIFN‐γ‐stimulated bone marrow macrophages (BMMΦ) revealed that, in contrast to class II complexes hitherto described, BMMΦ‐derived I‐A and I‐E subregion‐encoded subunits are synthesized without invariant chains. The invariant chain‐deficient α,β heterodimers are expressed at the cell surface in high proportions demonstrating that their correct assembly and transport to the cell surface is accomplished in the absence of invariant chains. The lack of invariant chains appears not to be due to a failure of rIFN‐γ to induce transcription of the γ‐chain gene because rIFN‐γ‐induced, in contrast to uninduced, BMMΦ accumulate high levels of invariant chain‐specific transcripts as evidenced by Northern blot analysis. These findings suggest that translation of γ‐chain‐specific mRNA is blocked in BMMΦ for as yet unknown reasons. Alternatively, newly synthesized γ chains might have escaped their regular intracellular maturation pathway as a result of unidentified modifications mediated by altered posttranslational processing mechanisms.
Histamine‐producing cell‐stimulating activity. A biological activity shared by interleukin 3 and granulocyte‐macrophage colony‐stimulating factor
Authors: Dy Michel , Schneider Elke , Gastinel Louis‐Noel , Auffray Charles , Mermod J. J. , Hamburger J.
Published: March 2007

The histamine‐producing cell‐stimulating factor (HCSF) was first described as a lymphokine which is produced during secondary mixed leukocyte culture and which induces increased histamine synthesis by murine hematopoietic cells. It has been shown that it is different from interleukin 3 (IL3), despite the fact that pure IL3 expresses HCSF activity. Our results provide evidence that this factor (constitutively produced by the P388 D1 cell line) is identical with granulocyte‐macrophage colony‐stimulating factor (GM‐CSF) i.e.: (a) physicochemical properties of HCSF and GM‐CSF, such as molecular weight, isoelectric charge, hydrophobicity and behavior during affinity chromatography, are indistinguishable and both activities coelute during all biochemical purification procedures; (b) increased bone marrow cell histamine synthesis induced by P388 D1‐derived HCSF is inhibited by anti‐GM‐CSF antiserum; (c) the GM‐CSF cDNA probe hybridizes with a poly(A)+RNA from P388 D1 cells while no hybridizing signal was obtained with poly(A)+RNA from WEHI‐3 and from P815 cells. On the other hand, the IL3 cDNA probe hybridizes with a 1.0‐kb poly‐(A)+RNA from WEHI‐3 but not with those from P388 D1 and P815. Moreover, well known sources of GM‐CSF, such as lung conditioned medium and semi‐purified GM‐CSF from phytohemagglutinin‐induced supernatant of the murine T lymphoma LBRM‐33‐5 A4 (preparation devoid of IL 3), as well as recombinant murine GM‐CSF, induce increased histamine synthesis by hematopoietic cells. All these results demonstrate that, in our culture conditions, the P388 D1 cell line spontaneously produces GM‐CSF which is responsible for the P388 D1‐induced HCS activity. Consequently, the latter is a property shared by the two distinct hematopoietic growth factors acting on the less committed cells, i.e. IL 3 and GM‐CSF, whereas M‐CSF or G‐CSF are unable to induce histamine production. Interestingly, IL 4 which is known to support established mast cell line proliferation cannot induce HCS activity. In addition, none of the other cytokines tested, such as IL1, IL2, interferons or tumor necrosis factor can express HCS activity. This expression seems to be a specific property of IL3 and GM‐CSF.
Differential regulation of T cell receptor gamma genes in immature thymocyte populations
Authors: Kinnon Christine , McGuire Kathleen L. , Rothenberg Ellen V.
Published: March 2007

Immature thymocytes that lack both Lyt‐2 (CD8) and L3T4 (CD4) expression can respond rapidly to stimulation with phorbol ester and calcium ionophore by expressing some gene products characteristic of mature, activated T cells. Here we studied the effect of such short‐term stimulation on the number of copies per cell of RNA for components of the T cell receptor complex. Although, upon stimulation, mRNAs for T cell receptor β chain accumulated to higher levels, the cells did not rapidly increase their expression of α‐chain transcripts from rearranged or germ‐line genes. Transcripts from the Cγ1 (Cγ13.4) and Cγ2 (Cγ10.5) genes were differentially regulated. The rarer Cγ1 transcripts were strongly induced, while the initially abundant Cγ2 transcripts showed a modest decrease in transcripts per cell within 24 h. Thus, the ratio of these two transcripts could be shifted dramatically prior to any significant change in the cellular composition of the population. These results suggest regulatory processes that may contribute to the observed expression of γ products in vitro or in normal development.
A monoclonal antibody (HML‐1) defining a novel membrane molecule present on human intestinal lymphocytes
Authors: Cerf‐Bensussan Nadine , Jarry Anne , Brousse Nicole , Lisowska‐Grospierre Barbara , Guy‐Grand Delphine , Griscelli Claude
Published: March 2007

A monoclonal antibody, HML‐1, was produced by fusion of NSI myeloma cells with spleen cells of a mouse immunized with isolated human intestinal intraepithelial lymphocytes (IEL). Immunofluorescence studies of isolated cells, as well as immunoperoxidase staining of tissue sections, indicated that HML‐1 labeled all the various subsets of human intestinal IEL, approximately 40% of lamina propria T cells, 30% mesenteric lymphoblasts and some lymphocytes in other mucosae, particularly IEL. Conversely, it revealed only rare cells in all other lymphoid compartments. Analysis by polyacrylamide gel gradient electrophoresis showed that HML‐1 precipitated two major noncovalently bound components of approximate mol. masses of 105 and 150 kDa from human IEL. HML‐1 thus defines a novel human membrane antigen present on a subpopulation of lymphocytes preferentially associated with epithelia, and particularly with the intestinal epithelium. The characteristics of this human antigen are very similar to those of an antigen we had previously described in the rat. The possible functional role of this novel class of lymphocyte membrane antigens as well as the nature of the mechanism that triggers their expression remain to be elucidated.
Differential requirements for the processing and presentation of soluble and particulate bacterial antigens by macrophages
Authors: Ziegler H. Kirk , Orlin Christine A. , Cluff Christopher W.
Published: March 2007

The requirements for antigen processing and presentation by macrophages using various forms of antigens derived from Listeria monocytogenes have been studied. Antigen presentation was monitored by T cell‐macrophage binding and interleukin production using T cells from Listeria monocytogenes‐infected mice and specific T cell hybridomas. Antigen processing requirements were defined by three criteria: (a) inhibition by lysosomotropic agents, NH4Cl and chloroquine; (b) kinetic relationships between antigen uptake and antigen presentation; and (c) antigen presentation by macrophages pre‐fixed with glutaraldehyde. In comparing heat‐killed Listeria monocytogenes (HKLM) with soluble listerial proteins (SLP), the presentation of SLP was less sensitive to lysosomotropic agents, showed faster antigen processing kinetics than with HKLM and could occur using pre‐fixed macrophages. Transitions between particulate and soluble forms had dramatic influences on processing requirements. Antigens associated with HKLM could be converted to soluble forms which did not require processing by preculture with macrophages and also by physical (e.g. sonication) and chemical (sodium dodecyl sulfate) treatments in the presence of protease inhibitors. Conversely, antigen processing was required when SLP were converted to a particulate form by covalent binding to latex beads. Analysis of SLP by molecular sieve chromatography and preparative SDS‐polyacrylamide gel electrophoresis revealed that high molecular weight proteins (> 60 kDa) could be presented by prefixed macrophages without prior processing. We conclude that the transition from a particulate to soluble antigenic form can be a significant antigen processing event.
T cell activation by anti‐CD3 antibodies: function of Fc receptors on B cell blasts, but not resting B cells, and CD18 on the responding T cells
Authors: Austyn Jonathan M. , Smith Kenneth G. C. , Morris Peter J.
Published: March 2007

Mouse anti‐human CD3 (T3) antibodies can induce T cell proliferation in the presence of Fc receptor (FcR)‐bearing accessory cells. Depending on whether the particular antibody can interact with the FcR, it can be mitogenic or otherwise. Previously, some of us (Smith, K. G. C. et al., Eur. J. Immunol. 1986. 16: 478) examined human T cell responses to the murine anti‐CD3 antibody switch variants UCHT1 (IgG1) and UCHT1B (IgG2b). Using a novel xenogeneic system with mouse macrophages (MΦ) and an anti‐FcR antibody, 2.4G2, we obtained direct evidence for accessory function of FcR in these responses. However, mouse B cells which also possess FcR were not accessory cells.

Here we show that resting B cells do not inhibit anti‐CD3 responses in the presence of other accessory cells, and they do not synergize with them. They appear to be inert in these responses but this is not simply because of their radiosensitivity. In contrast, B cell blasts proved to be potent stimulators of responses with UCHT1, UCHT1B and OKT3 (IgG2a). All three responses were inhibited by 2.4G2, whereas we have shown previously that the OKT3 response with MΦ was not, in keeping with the known specificities of B cell and MΦ FcR. These findings are discussed in relation to the molecular cloning of FcR, and we consider the possibility that distinct FcR could be expressed on resting and activated B cells.

A report that anti‐CD18 (LFA‐1) antibodies blocked the UCHT1 response with human monocytes raised the possibility that this molecule might also be involved in accessory function. However, we show that this inhibition is in fact at the level of the T cell, since anti‐human, but not anti‐mouse CD 18 antibodies, inhibited proliferative responses and clustering with both human and mouse accessory cells. Our results demonstrate that the principal contribution of accessory cells to anti‐CD3 responses may be the provision of an FcR, and that CD18 is most probably required at the level of the T cell.
Regulation of the antibody response to sheep erythrocytes by monoclonal IgG antibodies
Authors: Quintana Ilana Zalcberg , Silveira Angela V. , Möller Güran
Published: March 2007

Monoclonal antibodies directed against sheep erythrocytes of the isotypes IgG1,IgG2b and IgG2a were used to analyze the specificity of antibody‐induced suppression of the immune response. It was first shown that all monoclonals reacted against different antigenic determinants and they all suppressed the immune response to sheep erythrocytes when given shortly after the antigen to more than 50% as compared to 90–96% inhibition obtained with a polyclonal antiserum. Increasing the doses of monoclonals did not increase suppression. However, two different monoclonals administered together caused an additive, but not a synergistic inhibitory effect. No enhancement of the immune response was observed with any of the Ig classes tested. These findings show that four different antigenic determinants on sheep erythrocytes induced the synthesis of corresponding antibodies, with little or no signs of a dominant determinant. Passively administered monoclonal antibodies, even at supraoptimal doses, never suppressed the immune response to the same extent as a polyclonal antiserum, suggesting that each monoclonal only suppressed the synthesis of the corresponding antibody and did not affect antibody synthesis to other determinants.
Endothelial cell lysis induced by lymphokine‐activated human peripheral blood mononuclear cells
Authors: Miltenburg André M. M. , Meijer‐Paape Marion E. , Daha Mohamed R. , Paul Leendert C.
Published: March 2007

In vitro exposure of human peripheral blood mononuclear cells (PBMC) to interleukin 2 results in the generation of lymphokine‐activated killer (LAK) cells. Such LAK cells exhibit cytotoxicity against a spectrum of tumor target cell lines whereas they apparently do not affect normal tissues. In this report we show that PBMC that have been activated with T cell growth factor lyse trypsinized human umbilical cord venous endothelial cells as well as endothelial cell monolayers in a dose‐dependent manner. Microscopic analysis showed that during the 4‐h incubation period cell clumps containing detached endothelial cells and LAK cells were formed. When these clumps were evaluated with trypan blue the endothelial cells stained positive whereas LAK cells excluded the dye. No lysis occurred when fresh PBMC were added to target endothelial cells. The endothelial cell kill could not be blocked with an anti‐LFA‐1 antibody nor with intact OKT3 or F(ab′)2 fragments of WT32. We conclude that lymphokine‐activated PBMC exhibit cell‐mediated endothelial cell detachment and lysis.
Identification of a novel 45‐kDa cell surface molecule involved in activation of the human Jurkat T cell line
Authors: Carrel Stefan , Isler Patrick , Salvi Suzanne , Giuffrè Laura , Pantaleo Giuseppe , Mach Jean‐Pierre , Cerottini Jean‐Charles
Published: March 2007

This report describes a surface molecule, Tp45, which appears to be involved in interleukin 2 production and Ca2+ mobilization by Jurkat cells. The Tp45 molecule was identified by a monoclonal antibody, MX13, on the surface of either T3/TCR+ or T3/TCR− human T cell lines. Biochemical data showed that mAb MX13 precipitated a single polypeptide chain of 45 kDa both under reduced and nonreduced conditions from lysates of 125I‐surface‐labeled cells. Sequential immunodepletion experiments using lysates of 125I‐labeled T3/TCR+ cells showed that Tp45 was distinct from the α chain of the TCR complex. However, incubation of such cells with either anti‐T3 or anti‐TCR monoclonal antibody induced complete modulation of both the T3/TCR complex and Tp45. Conversely, complete modulation of both Tp45 and the T3/TCR complex was observed after incubation with anti‐Tp45 antibody. Functional studies showed that anti‐Tp45 antibody induced high levels of interleukin 2 production in Jurkat cells. In addition, incubation of these cells with the antibody resulted in Ca2+ mobilization from internal stores.

Anti‐Tp45 antibody reacted with 3–19% peripheral blood (E‐rosette‐positive) T cells in individual donors. The magnitude of the proliferative response elicited by anti‐Tp45 antibody for peripheral blood T cells was lower than that induced by an anti‐T3 antibody. This observation is compatible with the idea that only a subpopulation of T cells is reactive with anti‐Tp45. Multicolor flow cytometry analysis showed that the Tp45+ cells belong preferentially to the T8 subset.
Production of hybridoma growth factor by human monocytes
Authors: Aarden Lucien A. , De Groot Els R. , Schaap Otto L. , Lansdorp Peter M.
Published: March 2007

Human mononuclear leukocytes produce a growth factor (HGF) for hybridoma and plasmacytoma cells. HGF has recently been proven to be identical to IFN‐β2, 26‐kDa protein and BSF‐2. HGF can be quantitated in a proliferation assay with the HGF‐dependent hybridoma cell line B13.29. By selection of an extremely sensitive variant of this cell line, we were able to measure HGF production of single cells. Limiting dilution analysis of the producing cells in combination with size, density and adherence characteristics showed that HGF is produced by monocytes and not by lymphocytes. There was no need for the monocytes to be stimulated but the cells did require the presence of serum. This serum requirement could be met by purified bovine serum albumin, but not by other proteins like ovalbumin or human γ‐globulin. HGF production in vitro by monocytes starts after 2 h of incubation and is completed within 24 h.
Experimental Trypanosoma brucei infections selectively suppress both interleukin 2 production and interleukin 2 receptor expression
Authors: Sileghem Maarten , Hamers Ramond , De Baetselier Patrick
Published: March 2007

Experimental infections with Trypanosoma brucei AnTat 1.1. E not only account for a suppression of interleukin 2 (IL2) production, but also induce an impairment of IL2 receptor expression. Indeed, lymph node cells derived from infected mice failed to express IL2 receptors following mitogenic stimulation as compared to normal controls. This impairment was not attributed to a modulation of the number of T cells and was not caused by the presence of living parasites. Furthermore, the basal level of IL2 receptor expression could not be re‐established through the exogenous supply of recombinant IL2. This impairment of receptor expression was found to be mediated by suppressive cells that affect the relative number of receptor‐positive cells as well as the mean receptor density. On the other hand, the mitogen‐induced secretion of other T cell‐derived lymphokines was not inhibited during infection, indicating that the severe suppression of IL2 regulation was not attributed to a total paralysis of the T cell responsiveness.
Immuno‐electron microscopic characterization of large granular lymphocytes (natural killer cells) from rat liver
Authors: Bouwens Luc , Wisse Eddie
Published: March 2007

The immunocytochemical characteristics of large granular lymphocytes (LGL), isolated from the liver sinusoids of euthymic and athymic (nude) rats, were investigated in electron microscopy by the immunoperoxidase technique. The LGL were found positive for MRCOX‐8 (natural killer cells and cytotoxic/suppressor T cells) and negative for MRCOX‐19 (pan‐T marker) in both rat strains. The LGL were heterogeneous in the expression of the natural killer cell marker asialo‐GM1 which was found on 56% of the LGL from euthymic and on 71% of the LGL from athymic rats. LGL were easily distinguished from the other cells in the preparations, “conventional” lymphocytes and monocytes, thanks to their highly characteristic ultrastruc‐tural features, in particular by the presence of specific electron‐dense cytoplasmic granules and rod‐cored vesicles. These features have been described formerly for the so‐called “pit cells” and are more reliable than the classical LGL characteristics at the light microscopic level, i.e. the presence of azurophilic granules. Our results give further support for the existence of an important population of natural killer cells in the liver sinusoids.
Involvement of specific Lyt‐2+ T cells in the immunological control of experimentally induced murine cutaneous leishmaniasis
Authors: Titus Richard G. , Milon Geneviève , Marchal Gilles , Vassalli Pierre , Cerottini Jean‐Charles , Louis Jacques A.
Published: March 2007

The involvement of Lyt‐2+ T cells in the immunological control of cutaneous leishmaniasis was assessed by studying the course of the disease in mice in which the number of these cells was reduced by treatment with anti‐Lyt‐2 monoclonal antibody (mAb). Administration of anti‐Lyt‐2 mAb exacerbated Leishmania major‐induced cutaneous lesions in both genetically resistant and susceptible mice. This effect correlated with a drastic reduction in the number of specific Lyt‐2+ T cells generated during infection. These results, together with the observation that resistant mice possess higher numbers of specific Lyt‐2+ T cells in lymph nodes draining their lesions than susceptible mice, 3 weeks after infection, indicate that Lyt‐2+ T cells also play a role in the immunological control of Leishmania major‐induced lesions.
Interaction between the CD45 antigen and phytohemagglutinin. Inhibitory effect on the lectininduced T cell proliferation by anti‐CD45 monoclonal antibody
Authors: Bernabeu Carmelo , Carrera Ana C. , De Landázuri Manuel O. , Sánchez‐Madrid Francisco
Published: March 2007

Immunoprecipitation studies from 125I‐labeled T cells, previously activated with the lectin phytohemagglutinin (PHA) from Phaseolus vulgaris, showed a preferential association between the CD45 glycoprotein family containing members of 220, 205, 190 and 180 kDa, and a protein of 33 kDa. This 33‐kDa protein, with an identical molecular mass as PHA, was not associated with other highly expressed molecules such as class I histocompatibility antigens (HLA‐A,B,C), transferrin receptor, CDw44 and CD11a. Further immunoprecipitation analysis from peripheral blood lymphocytes incubated with 125I‐labeled PHA confirmed the identification of the CD45‐associated 33‐kDa protein as the lectin PHA. These results prompted us to analyze the possible involvement of the CD45 molecules in the T cell activation process induced by PHA. Thus, two monoclonal antibodies specific for CD45 antigens were able to inhibit the T cell proliferation induced by the lectin. This inhibitory ability was exerted by affecting both the interleukin 2 production and the interleukin 2 receptor expression.
Human complement factor H: expression of an additional truncated gene product of 43 kDa in human liver
Authors: Schwaeble Wilhelm , Zwirner Jörg , Schulz Thomas F. , Linke Reinhold P. , Dierich Manfred P. , Weiss Elisabeth H.
Published: March 2007

The human complement factor H is an important factor in the control of the alternative pathway and also induces the stimulation of B cells and macrophages in vitro. Using a human factor H cDNA clone as probe, two factor H‐specific transcripts of 4.4 and 1.8 kb were detected in four human livers. Both mRNAs were found independently of the expression of an acute phase marker SAA, in these livers, indicating that their presence is not linked to an acute phase state. The shorter transcript was cloned in two cDNA plasmids H‐19 and H‐20, lacking only seven amino acids of the N‐terminus of the mature factor H protein. The deduced protein sequence showed that this protein is identical to the N‐terminal portion of the large classical factor H of 150 kDa mol. mass. Parallel to the finding that the N‐terminal sequence of factor H is expressed by two distinct mRNA species, evidence is presented that the C‐terminal sequence is also contained on two different transcripts, the common 4.4‐kb mRNA and an additional 1.0‐kb mRNA. A novel, short form of human complement factor H of 43 kDa was detected in human sera which represents the translation product of the 1.8‐kb factor H‐specific mRNA detected in human liver. Five distinct epitopes detected with 6 monoclonal antibodies are present on both the 150‐kDa factor H protein and the truncated 43‐kDa molecule. We conclude that additional H‐specific mRNAs are found in human liver and that at least one of them is translated yielding a truncated form of factor H.
A murine macrophage cell line, immortalized by v‐raf and v‐myc oncogenes, exhibits normal macrophage functions
Authors: Blasi Elisabetta , Radzioch Danuta , Durum Scott K. , Varesio Luigi
Published: March 2007

In vitro immortalized cell lines with the morphology and phenotype of mature macrophages (MΦ) have been generated by infecting freshly isolated bone marrow cells from C3H/HeJ mice with a recombinant retrovirus carrying v‐raf and v‐myc oncogenes. All of the clones obtained had MΦ‐like phenotypes, and one such clone, GG2EE, has been compared to normal MΦ to ascertain the effects of immortalization on the expression of the biological functions of the lines. GG2EE cells expressed cytotoxic activity against L5178Y, P815 or RL 1 target cells in response to stimulation with interferon‐γ (IFN‐γ) and heat‐killed Listeria monocytogenes; in contrast, they failed to kill YAC‐1 target cells. GG2EE cells did not constitutively express I‐A or I‐E antigens; nevertheless, I region‐coded antigens could be induced by IFN‐γ treatment. GG2EE cells produced interleukin 1 upon stimulation with a T cell‐derived lymphokine; they were weakly phagocytic, yet became highly phagocytic following IFN‐γ treatment. Since c‐fos mRNA is augmented in peritoneal exudate MΦ by protein kinase C activators but not by IFN‐γ, we evaluated the effects of calcium ionophore, phorbol myristate acetate, L‐α‐1‐oleoyl‐2‐acetoyl‐sn‐3 glycerol (OAG) and IFN‐γ on the levels of c‐fos mRNA in GG2EE cells. We found that calcium ionophore, PMA and OAG stimulation enhanced the expression of c‐fos mRNA, but IFN‐γ treatment did not. The kinetics of c‐fos induction in GG2EE cells were also comparable to those observed in peritoneal exudate MΦ. Overall, the GG2EE cell line has the same biological properties as normal tissue MΦ. Because it is capable of both constitutive and inducible MΦ‐like functions, this cell line provides a valuable tool for studying the molecular mechanisms controlling induction and/or expression of biological activities in MΦ. It is striking that a cell line immortalized in vitro by two oncogenes, v‐raf and v‐myc, behaves, according to the criteria mentioned above, like a normal MΦ population.
Phorbol diesters increase the phosphorylation of the leukocyte common antigen CD45 in human T cells
Authors: Autero Matti , Gahmberg Carl G.
Published: March 2007

We have studied the phosphorylation of the human leukocyte surface glycoprotein CD45, formerly known as leukocyte common antigen or T200, following activation of peripheral T cells with tumor‐promoting phorbol diesters. The cells were incubated with [32P]orthophosphate, treated with phorbol diesters, and the CD45 was immunoprecipitated using the monoclonal antibody T29/33. CD45 was weakly phosphorylated in nonactivated T cells, but a treatment of the cells with 10, 60 or 200 nM phorbol 12,13‐dibutyrate (PDBu) markedly increased the phosphorylation of the glycoprotein. This effect of PDBu was relatively selective for CD45 since the activation of the cells did not affect the phosphorylation of all membrane phosphoglycoproteins. The effect of another tumor promoter, phorbol 12‐myristate 13‐acetate (PMA), was comparable to that of PDBu, but a nontumor‐promoting phorbol derivative, 4α‐phorbol 12,13‐didecanoate, did not change the level of CD45 phosphorylation. The phosphorylation of CD45 was found to clearly increase after a treatment of T cells with 60 nM PDBu for only 30 s, and the phosphorylation reached its maximum in about 5 min.
In vivo effect of murine recombinant interleukin 3 on early hemopoietic progenitors
Authors: Kindler Vincent , Thorens Bernard , Vassalli Pierre
Published: March 2007

Normal and irradiated mice were perfused with recombinant interleukin 3 (rIL 3) and the number of early hemopoietic progenitors (CFU‐S) was quantified in different organs. Normal mice perfused with rIL 3 for 3 or 7 days showed a dramatic increase in the number of CFU‐S in the spleen, liver and blood, while the bone marrow CFU‐S number was slightly decreased. The total number of CFU‐S per animal was only slightly increased; the major effect of rIL 3 perfusion was thus on the distribution of CFU‐S in the mice. Lethally irradiated mice injected with 5 × 104 syngeneic bone marrow cells and perfused with rIL 3 were killed after 10 days. A significant increase in both the number and size of the splenic colonies was observed. The CFU‐S content of these colonies, determined in a second set of irradiated mice, was increased by more than 25‐fold, indicating that in conditions of hemopoietic depletion, rIL 3 vastly increased the self‐renewal capacity of early progenitors.
Synergistic interaction between interleukin 4 and anti‐Bp50 (CDw40) revealed in a novel B cell restimulation assay
Authors: Gordon John , Millsum Michelle J. , Guy Graeme R. , Ledbetter Jeffrey A.
Published: March 2007

Highly purified resting B lymphocytes stimulated for 3 days to high‐rate DNA synthesis by a synergistic combination of phorbol dibutyrate and ionomycin soon returned to quiescence once those signals had been removed. The maintenance of DNA synthesis in such cultures was found to provide a sensitive assay for revealing the factors that interact with cycling B cells. Whereas several activities ‐ namely, interleukin 4, anti‐Bp50 and a low molecular weight B cell growth factor ‐ were, by themselves, capable of prolonging DNA synthesis over a further day or so, no single factor was capable of sustaining the replication cycle out to day 6 of culture. By contrast, certain combinations of activities displayed significant synergy in the restimulation assay. The most striking observed was that between interleukin 4 and anti‐Bp50 where, by day 6, their combined effect on maintaining DNA synthesis in 3‐day stimulated cells was the same as having kept phorbol dibutyrate and ionomycin in the culture system. The implications of these findings are discussed.
Mott cells: a model to study immunoglobulin secretion
Authors: Alanen Anna , Pira Ulla , Colman Alan , Franklin Richard M.
Published: March 2007

Mott cells are plasma cells defective in immunoglobulin (Ig) secretion. They display this defect by accumulating Ig in the rough endoplasmic reticulum, detectable by Ig+ intracellular inclusion. We have previously produced hybridoma cell lines (Alanen, A. et al., Eur. J. Immunol. 1985. 15: 235) in which this phenotype is preserved, and shown the inability of these cells to secrete the Ig. In order to study this defect further, we fused these hybridoma cells with a x‐secreting hybridoma cell line, Sp1, and, using double selection with hypoxanthine, aminopterin, thymidine and ouabain, obtained hybrid cell lines expressing various combinations of the three Ig chains involved (Mott γ1, Mott χ and Sp1 χ chains). We studied the presence of Ig+ inclusions in these cells as well as Ig secretion by metabolic labeling and immunoprecipitation. All inclusion‐positive clones expressed both Mott heavy and Mott light chains with or without the Sp1 light chain, whereas none of the inclusion‐negative clones produced both Mott‐derived Ig chains. In all of the clones, even those with inclusions, the Ig secretion was at least partially rescued by the fusion. This occurred also in an inclusion‐positive clone which maintained the original Ig status of the Mott without Sp1 χ chain, indicating a complementation by the cell fusion of some cellular factor involved in Ig secretion. Furthermore, we injected Xenopus oocytes with mRNA isolated from three different original Mott cell hybridomas and could show secretion of the Ig, which is not secreted from the original Mott cells, from the oocytes.
T cell activation by cross‐linking anti‐CD3 antibodies with second anti‐T cell antibodies: dual antibody cross‐linking mimics physical monocyte interaction
Authors: Walker Christoph , Bettens Florence , Pichler Werner J.
Published: March 2007

The anti‐CD3 antibody BMA030 (IgG2a isotype) induces T cell activation and proliferation if an interaction with monocytes is provided. In contrast to other anti‐CD3 antibodies, it is unable to induce interleukin (IL)2 responsiveness through cross‐linking by plastic‐bound goat anti mouse Ig antibodies (panning). Cross‐linking BMA030 with a second anti‐T cell antibody is, however, able to induce IL 2 responsiveness in monocyte‐depleted T cell cultures. In this report we show that a large number of different antibodies are suitable for this dual antibody stimulation, and that the extent of proliferation corresponds to the percentage of T cells expressing the respective T cell antigen. Proliferation induced by low concentrations (0.1‐1 ng/ml) of other anti‐CD3 antibodies requires also cross‐linking with second anti‐T cell antibodies. The proliferative response of monocyte‐depleted T cells to two cross‐linked anti‐T cell antibodies plus added IL 2 is of the same magnitude as the one induced by anti‐CD3 antibodies plus monocytes. On the other hand, if monocytes are present, soluble anti‐CD2, ‐CD4, ‐CD8, ‐LFA‐1 antibodies (IgG1 or F(ab′)2 fragments) can inhibit OKT3 or BMA030‐induced T cell activation. Anti‐CD6 antibodies do not interfere with this monocyte‐dependent T cell stimulation. We conclude that dual antibody stimulation mimics the physical contact of T cells with monocyte membranes, where the T cell receptor CD3 complex is cross‐linked with neighboring structures (mainly so‐called adhesion molecules) through the interaction with respective counter‐structures on monocyte membranes. Dual antibody cross‐linking bypasses this interaction and can be used to stimulate IL2 responsiveness of antibodydefined T cells.
Prolonged growth of activated B lymphocytes requires interaction with helper T cells
Authors: Rajasekar Ramanujam , Andersson Jan , Leanderson Tomas
Published: March 2007

Resting B lymphocytes acquire upon activation responsiveness to soluble factors that support their growth and differentiation. We have studied the growth requirements of anti‐μ pre‐activated B lymphocytes in vitro. We found that activated B lymphocytes did respond to soluble factors from helper T cells by entering the cell cycle but cell‐cell contact with specific helper T cells was required for prolonged growth of activated B lymphocytes. Thus, activated B lymphocytes must receive activation signals from helper T cells in order to stay in the proliferative phase of the cell cycle.
Activation of specific T cell lines by the antigens avidin and myelin basic protein in the absence of antigen‐presenting cells
Authors: Altmann Daniel M. , Lider Ofer , Douek Daniel C. , Cohen Irun R.
Published: March 2007

We have investigated the specific activation by soluble antigen of rat or mouse longterm T helper cell lines using antigen‐presenting cell (APC)‐free culture conditions. Some T cell lines specific for avidin or myelin basic protein responded to native antigen in the absence of added APC. Responses in the absence of APC were substantial and specific although, as would be expected, lower than in the presence of APC. Proliferation could not be inhibited by culture with anti‐Ia antibodies and the ability of lines to respond to antigen in the absence of APC did not correlate with the endogeneous surface Ia expression of the lines. Furthermore, irradiated T cells were unable to act as presenting cells for line cells of the same or a different specificity. This suggests that the T cells did not present antigen to each other, and demonstrates, along with other data shown, that activation cannot be attributed to undetected APC remaining in the cultures. Anti‐avidin T cell lines differed markedly in their ability to respond to avidin in the absence of added APC.S2, an anti‐avidin line of H‐2s genotype consistently responded well to avidin seen in the absence of added APC; K2, an H‐2k anti‐avidin line, responded moderately and B3, and H‐2b anti‐avidin line, although the most prolific responder in the presence of APC, never responded to antigen in their absence. Z1a, a Lewis rat‐derived T cell line specific for myelin basic protein, proliferated well in response to the antigen in the absence of added APC. The present findings demonstrate that some T cells can recognize and respond to native antigens without the mediation of specialized APC.
Four epitopes on the rat 55‐kDa subunit of the interleukin 2 receptor as defined by newly developed mouse anti‐rat interleukin 2 receptor monoclonal antibodies
Authors: Mouzaki Athanasia , Diamantstein Tibor
Published: March 2007

Four new mouse monoclonal antibodies (mAb), ART‐38, ART‐35, ART‐75 and ART‐94, directed against the rat interleukin 2 receptor (IL 2R) have been developed. As shown by immunoprecipitation studies they all recognize specifically the 55‐kDa subunit of the rat IL 2R. These mAb were compared to three previously characterized mouse mAb directed against the 55‐kDa molecule of the rat IL 2R, namely the ART‐18, ART‐65 and OX‐39 mAb. Out of all seven mAb, only ART‐18 and OX‐39 were found to inhibit the IL 2 binding to activated T cells, while IL 2R inhibited the binding of ART‐18 alone. ART‐18 was the only mAb found to inhibit the IL 2‐dependent proliferation of cells carrying the IL 2R. Scatchard plot analyses showed gross differences in the numbers of mAb‐binding sites ranging between 30000 (ART‐65) and 165000 (ART‐75) as well as in their affinities which ranged between 2.5 × 10−9 M (ART‐38) and 8.3 × 10−10 M (OX‐39). Cross inhibition studies revealed that the mAb recognize four different epitopes on the 55‐kDa rat IL 2R subunit.
Immunomodulation of murine visceral leishmaniasis by administration of monoclonal anti‐Ia antibodies: differential effects of anti‐I‐A vs. anti‐I‐E antibodies
Authors: Blackwell Jenefer M. , Roberts Morven B.
Published: March 2007

On a B10 genetic background noncure and cure phenotypes for murine visceral leishmaniasis are controlled by H‐2. In this report results are presented which show the effects of administering specific anti‐I‐A and anti‐I‐E monoclonal antibodies to B10.D2/n (H‐2d) noncure mice prior to and during 85 days of infection with Leishmania donovani LV9. The effects of the two anti‐Ia antibodies were precisely equivalent in diminishing circulating anti‐leishmanial IgG levels throughout infection, possibly as a direct effect of the anti‐Ia antibodies in reducing the splenic B cell population. In terms of resolution of liver and spleen parasite loads, which is known to be dependent upon induction of a cell‐mediated immune response, dramatically different results were obtained with the two anti‐Ia antibodies. Anti‐I‐A treatment resulted in prolonged exacerbation of disease in liver and spleen. Anti‐I‐E treatment was associated with enhanced clearance of liver and spleen parasite loads beyond 30 days of infection. The results are consistent with the hypothesis that blocking major histocompatibility complex‐restricted antigen presentation by one class II molecule allows T cell responses controlled by the other to predominate. Hence, in H‐2d mice, I‐E controls suppressor activity while I‐A is associated with helper activity for cell‐mediated control of infection. The results offer some prospect for the development of haplotype‐ and class II molecule‐specific immunotherapeutic regimens in the host which might prevent the undesirable expansion of T cell populations which exacerbate disease without compromizing development of a curative cell‐mediated immune response.
Anti‐Thy‐1‐induced proliferation of immature thymocytes expressing the CD3‐associated γ/δ heterodimer
Authors: MacDonald H. Robson , Miescher Guido C. , Howe Rawleigh C.
Published: March 2007

A series of rat monoclonal antibodies (mAb) directed against the Thy‐1 molecule have been evaluated for their ability to stimulate immature (CD4−8−) murine thymocytes. CD4−8− thymocytes could be induced to proliferate by several anti‐Thy‐1 mAb provided that exogenous inteleukin 1 or interleukin 2 was provided. Apparently nonstimulatory anti‐Thy‐1 mAb could be rendered stimulatory if cross‐linked by a rabbit anti‐rat immunoglobulin reagent. Anti‐Thy‐1‐stimulated CD4−8− thymoctes were found to uniformly express CD3 but did not stain positively with F23.1, a mAb reactive with α/β T cell receptors utilizing the Vβ8 gene family. Immunoprecipitation analysis of such stimulated CD4−8− thymocytes indicated that they expressed a CD3‐associated heterodimer of 45 kDa/35 kDa most likely corresponding to the product of the γ and (putative) δ T cell receptor genes. Taken together, these data demonstrate that anti‐Thy‐1 mAb selectively activate a subset of CD4−8− thymocytes committed to the γ/δ T cell receptor lineage.