bluuebunny/crossref_metadata_2025 huggingface dataset:
Regeneration and the immune system I. In vitro and in vivo activation of lymphocytes by liver regeneration and the role of Kupffer cells in stimulation
Authors: Miyahara Seiko , Yokomuro Kozo , Takahashi Hidemi , Kimura Yoshitami
Published: February 2007

When syngeneic lymphocytes and mitomycin C‐treated regenerating liver cells, prepared from partially hepatectomized mice, are cultured together, the in vitro DNA synthesis is activated (sMLHLR: syngeneic mixed hepatectomized‐liver cell‐lymphocyte culture). The Ia+ Kupffer cells play an important role as stimulators in the response, since the stimulating activity of regenerating liver cells is lost either by the pretreatment with anti‐Ia monoclonal antibody plus complement or by the removing Kupffer cells from them. The lymphocytes are also activated in vivo during liver regeneration following partial hepatectomy. When lymphocytes, prepared from hepatectomized mice, are cultured with regenerating liver cells, the lymphocytes are stimulated to accelerate their DNA synthesis in a manner typical of the secondary immune responses (secondary sMLHLR). In primary sMLHLR, the responder cells are mainly Lyt‐1+, whereas, in secondary sMLHLR, they are mainly Lyt‐2+. The mechanism of changing the Lyt phenotype of major responder cells from Lyt‐1 to Lyt‐2 during sMLHLR is discussed.
Polyclonal activation of the murine immune system by an antibody to IgD III. Ontogeny
Authors: Muul Linda M. , Mond James J. , Finkelman Fred D.
Published: February 2007

It has recently been demonstrated that the injection of adult mice with an affinitypurified goat antibody to mouse IgD (GaMδ) stimulates activation of the humoral immune system that resembles, on a polyclonal level, specific B cell activation by a T cell‐dependent antigen. One to 2 days after adult BALB/c mice are injected with 200 μg of GaMδ, their splenic B lymphocytes undergo a series of T‐independent activation steps that include increases in surface (s) Ia expression, cell size and DNA synthesis. Seven days after GaMδ injection, these cells undergo T‐dependent activation steps, that include further proliferation as well as differentiation into IgG1‐secreting cells. We have now studied the ontogeny of the T‐independent (day 2) and T‐dependent (day 7) activation steps by injecting 100–200 μg of GaMδ into 3‐day‐ to 10‐week‐old BALB/c mice. GaMδ failed to induce increases in B cell sIa expression or size 2 days after injection of mice 2 weeks old or younger and failed to stimulate increased DNA synthesis 2 days after injection of 4‐week‐old mice. In contrast, increases in spleen cell sIa expression, size and DNA synthesis were seen 7days after injection of 6‐ to 8‐day‐old mice. Furthermore, increases in the numbers of spleen cells with large amounts of intracytoplasmic IgG1 were seen at the same time, although these increases were much less than were seen in GaMδ‐treated adult mice. Thus, the ability of GaMδ to induce T help and to act in concert with such help to stimulate B cell proliferation and differentiation precedes in ontogeny the ability of GaMδ to directly induce B cell proliferation and early differentiative events. In addition, the early activating events that we have studied are not required for T‐dependent B cell proliferation and antibody production to occur, althougt they appear to contribute to the magnitude of clonal expansion and antibody production.
RNA synthesis in activated macrophages I. Poly(I) · poly(C)‐induced triggering of cytolytic activity is associated with decrease in RNA synthesis
Authors: Varesio Luigi , Issaq Haleem J. , Taramelli Donatella
Published: February 2007

The effects of polyinosinic, polycytidylic acid [poly(I) · poly(C)] on the activation and RNA metabolism in murine peritoneal macrophages (MΦ) elicited by proteose‐peptone (pMΦ) was investigated. Poly(I) · poly(C) triggered the cytolytic activity of pMΦ and augmented their glucose oxidation. In contrast, a profound depression of [3H]uridine incorporation into RNA was observed in poly(I) · poly(C)‐activated pMΦ. The degree of depression of RNA labeling paralleled the dose of poly(I) · poly(C) used to activate the pMΦ and the expression of tumoricidal activity. This decrease in [3H]uridine incorporation into MΦ RNA could not be accounted for by decreased permeability of the activated MΦ to [3H]uridine, or by instability of the labeled RNA. Moreover, analysis of the specific activity of the intracellular uridine triphosphate (UTP) pool and studies on the labeling of MΦ RNA with [32P] orthophosphate indicated that the decreased RNA labeling was not due to changes in the specific activity of UTP. We concluded that poly(I) · poly(C)‐activated pMΦ exhibit a depressed rate of RNA synthesis. We suggest that the rate of RNA synthesis may be investigated as a potential new indicator for MΦ activation.
The kinetics of in vivo sensitization of rat peritoneal and lung mast cells: temporal dissociation from circulating levels of IgE
Authors: Coleman John W. , Layton Guy T. , Stanworth Denis R.
Published: February 2007

Following i.p. injection of ovalbumin (OVA) plus Bordetella pertussis vaccine into Hooded Lister rats, the time‐course of sensitization of peritoneal and lung mast cells (MC) did not parallel kinetic changes in the levels of circulating OVA‐specific and total IgE. OVA‐induced secretion of 5‐hydroxytryptamine from isolated peritoneal and lung MC and the presence of OVA‐specific IgE in serum were first demonstrated at day 14 post‐immunization. However, subsequent to day 14, the responsiveness of both types of MC to OVA declined, while circulating levels of OVA‐specific IgE continued to rise. Peritoneal MC, but not lung MC, showed increased responsiveness to challenge with anti‐IgE on day 7 post‐immunization, whereas circulating levels of total IgE were not elevated until day 14, thus demonstrating that nonantigen‐specific IgE was acquired by peritoneal MC before it entered the circulation. Lung MC generally showed decreased reactivity to both OVA and anti‐IgE, compared with peritoneal MC; no significant correlations were demonstrated between the responses of MC from these two tissue sites.
Induction of suppressive cells within the T4 subset by a human T‐T hybridoma‐produced factor
Authors: Grillot‐Courvalin Catherine , Labaume Sylvaine , Brouet Jean‐Claude
Published: February 2007

This study examines the target cell and the mode of action of a factor (TsF), produced by a human T‐T hybridoma, which suppresses pokeweed mitogen‐induced B cell differentiation as previously reported (J. Immunol. 1982. 129: 1008.). The suppressive effect still exists when peripheral blood leukocytes are depleted of T8+ cells or when TsF is added to a mixture of purified T4+ and B cells. This suppressive effect disappears when the T4 cells are irradiated. The suppression, however, is restored when a large number of T8 cells are added to the irradiated T4 cells. Lastly, a 24 h preincubation of T4 but not T8 cells, with TsF is able to generate suppressive effectors. These results suggest that the target cell for TsF is a radioresistant T4 cell which, upon activation, is able to induce radiosensitive T4 or T8 cells to suppress B cell differentiation.
Anti‐viral immune response of allogeneic irradiation bone marrow chimeras: cytotoxic T cell responsiveness depends upon H‐2 combination and infectious agent
Authors: Zinkernagel Rolf M. , Sado Toshi , Althage Alana , Kamisaku Hitoko
Published: February 2007

Allogeneic irradiation bone marrow chimeras C57BL/10 (H‐2b) →B10.BR (H‐2k) and B10.BR (H‐2k)→C57BL/10 (H‐2b) were raised under specific pathogen‐free (SPF) conditions; they survived very well and were healthy under SPF for 3–4 months and subsequently, under conventional housing conditions, for 1 to 8 months. Their immune response against third‐party alloantigens was comparable with that of controls. Anti‐vaccinia virus responses were very low when compared with syngeneic control chimeras or unmanipulated control animals; if anti‐vaccinia virus cytotoxic T cell reactivity was measurable, it was specific for the bone marrow donor, rather than the recipient thymic H‐2 type. In contrast, the anti‐LCMV (lymphocytic‐choriomeningitis virus) response was excellent and comparable to that in controls for B10.BR (H‐2k)→C57BL/10 (H‐2b) chimeras, but was completely absent for C57BL/10 (H‐2b)→B10.BR (H‐2k) chimeras. LCMV‐specific cytotoxic effector T cells from B10.BR→C57BL/10 chimeras were restricted entirely to recipient H‐2b. In contrast to the asymmetric cytotoxic T cell response, both types of chimeras developed good primary footpad swelling reactions after local infection, which arose somewhat slower with LCMV than in control of chimeras. The capacity to control infection by Listeria monocytogenes was excellent for all controls and B10.BR→C57BL/10 chimeras but apparently absent in C57BL/10→B10.BR chimeras.

Differentiation of T cell restriction specificity for thymic H‐2 is apparently most efficient, but it remains unclear whether the observed asymmetry reflects exaggerated immune response regulation in H‐2‐incompatible stem cell‐thymus chimeras or differential cross‐reactivities between restricting transplantation antigens.
Quantitative representation of all T cells committed to develop into cytotoxic effector cells and/or interleukin 2 activity‐producing helper cells within murine T lymphocyte subsets
Authors: Pfizenmaier Klaus , Scheurich Peter , Däubener Walter , Krönke Martin , Röllinghoff Martin , Wagner Hermann
Published: February 2007

A limiting dilution culture system based on stimulation with concanavalin A (Con A) has been used to study the quantitative distribution of helper and of cytotoxic precursor cells in Lyt‐2‐defined subpopulations of murine T cells. Virtually all of the selected Lyt‐2+ and Lyt‐2− T cells grow and expand to large clonal colonies within an 8–9‐day culture period. Our data show that upon stimulation with Con A, 90% of the Lyt‐2− T cells were capable to produce interleukin 2 (IL2) activity. In addition, IL2 activity is produced by 8–10% of Lyt‐2+ T cells. However, at the clonal level, the average of the IL 2 activity produced by Lyt‐2+ T cells is about 8‐fold less as compared to Lyt‐2− T cells. Precursors of cytotoxic T cells were almost exclusively found in the Lyt‐2+ population, of which about 70% displayed lytic activity in a lectin‐dependent cytolysis test. For the vast majority of clones analyzed the capacity to produce IL 2 activity and the capacity to express lytic activity, was found to be mutually exclusive. A minority of clones (< 3%) was found to simultaneously produce IL2 activity and to express cytotoxicity. These latter cells are therefore considered as bifunctional T cells.
The cytolytic T lymphocyte response to the murine cytomegalovirus. II. Detection of virus replication stage‐specific antigens by separate populations of in vivo active cytolytic T lymphocyte precursors
Authors: Reddehase Matthias J. , Keil Günther M. , Koszinowski Ulrich H.
Published: February 2007

During the acute cytolytic T lymphocyte (CTL) response of mice to infection with the murine cytomegalovirus two independent populations of activated interleukin‐receptive CTL precursors can be demonstrated. One population is specific for cell membrane‐incorporated viral structural antigens, whereas the second population detects an antigen, whose appearance is correlated with the synthesis of viral immediate early proteins. Since this new type of antigen is only defined by lymphocyte recognition, it is referred to as the lymphocyte‐detected immediate early antigen (LYDIEA). Expression of immediate early antigen precedes the production of viral progeny and, therefore, it is possible that LYDIEA‐specific CTL could serve as indicator cells for the very first activities of the viral genome, even during nonproductive infection.
Ontogeny of surface markers on functionally distinct T cell subsets in the chicken
Authors: Traill Karine N. , Böck Günther , Boyd Richard L. , Ratheiser Klaus , Wick Georg
Published: February 2007

Three subsets of chicken peripheral T cells (T1, T2 and T3) have been identified in peripheral blood of adult chickens on the basis of fluorescence intensity after staining with certain xenogeneic anti‐thymus cell sera (from turkeys and rabbits). They differentiate between 3–10 weeks of age in parallel with development of responsiveness to the mitogens concanavalin A (Con A), phytohemagglutinin (PHA) and pokeweed mitogen (PWM). Functional tests on the T subsets, sorted with a fluorescence‐activated cell sorter, have shown that T2,3 cells respond to Con A, PHA and PWM and are capable of eliciting a graft‐vs.‐host reaction (GvHR). In contrast, although T1 cells respond to Con A, they respond poorly to PHA and not at all to PWM or in GvHR. There was some indication of cooperation between T1 and T2,3 cells for the PHA response. Parallels between these chicken subsets and helper and suppressor/cytotoxic subsets in mammalian systems are discussed.
Regulatory effects of isologous anti‐idiotypic antibodies on the formation of different immunoglobulin classes in the immune response to phosphorylcholine in BALB/c mice
Authors: Blaser Kurt , Heusser Christoph H.
Published: February 2007

BALB/c mice were immunized with purified phosphorylcholine (PC)‐specific myeloma protein of the TEPC‐15 tumor, which bears the major idiotype of the anti‐PC response (T15 Id) in this mouse strain. Four months after establishing the anti‐idiotypic response, animals were immunized with PC‐keyhole limpet hemocyanin under conditions which normally lead to IgE antibody formation. The expression of anti‐PC antibodies of each immunoglobulin class was compared to a group of matched control mice not immunized with T15. In BALB/c mice producing anti‐idiotypic antibodies the formation of anti‐PC IgM, IgG1, IgG2, IgG3 and IgE was suppressed to various extents and for different lengths of time. An exception to this observation was the formation of anti‐PC IgA antibodies, in that no significant difference was measured between the two groups of mice. BALB/c mice immunized in the same way with the PC‐specific myeloma proteins of the MOPC‐167, MOPC‐511 and MOPC‐603 tumors produced normal levels of anti‐PC IgE and IgG antibodies. These results suggest that T15 idiotype‐positive antibodies are probably formed within all classes of specific antibodies expressed during an immune response to PC. The possibility of extending the phenomena of anti‐idiotype‐induced suppression to the level of various classes of specific antibodies, including IgE, might be of interest in the treatment of some allergic diseases.
Absence of cell surface fixation of a monoclonal antibody detectable by conventional immunoassays does not exclude expression of and interaction with the corresponding antigenic determinant
Authors: Layet Corine , Bouteiller Philippe P. Le , Olive Daniel , Mishal Zohair , Caillol Danielle , Kourilsky François M. , Jordan Bertrand R. , Lemonnier François A.
Published: February 2007

No specific binding of anti‐HLA class I B.10.6 monoclonal antibody (mAb) could be demonstrated by cell surface radioimmunoassay and cytofluorographic studies at the surface of murine transformed L cells expressing HLA‐A3 or Cw3 molecules. However, specific interaction of this antibody with these molecules at the surface of these transformed cells was indirectly established, since it inhibited specifically the binding to the same HLA class I molecules of other anti‐HLA class I mAb. Therefore, the absence of detectable binding of mAb, in conventional immunoassays, does not exclude expression by these cells of the corresponding antigenic determinant.
The regulation of macrophage activity in congenitally athymic mice
Authors: Sharp A. Kay , Colston M. Joseph
Published: February 2007

It has been suggested that non‐T cell‐mediated cellular immune mechanisms might be elevated in nude mice, which could contribute to their partial resistance to intracellular infectious agents and to the development of spontaneous tumors. In this study we have examined macrophages (MΦ) from athymic, euthymic and athymic T cell‐reconstituted mice in terms of their phagocytic capacity, microbicidal and tumoricidal activity, and the production of hydrogen peroxidase and superoxide anions. These studies have demonstrated the presence of activated MΦ in nude mice and suggest that this activation is associated with the absence of T cell‐mediated suppression. We have also compared MΦ activation levels in germ‐free nude mice which have a defined intestinal bacterial flora. We have found MΦ activity to be significantly elevated in microbiologically defined nude mice when compared to germ‐free nude mice, indicating that a resistant gut flora is capable of nonspecifically stimulating nude mouse MΦ.
Mitogenic and polyclonal B cell activation activities of synthetic lipid A analogues
Authors: Kumazawa Yoshio , Matsuura Motohiro , Nakatsuru‐Watanabe Yoko , Fukumoto Mitsuru , Nishimura Chiaki , Homma J. Yuzuru , Inage Masaru , Kusumoto Shoichi , Shiba Tetsuo
Published: February 2007

The activation of murine B cells and macrophages by synthetic lipid A analogues, solubilized with triethylamine and complexed with bovine serum albumin, were investigated in vitro. The analogues used are nonphosphorylated, C‐1 or C‐4′ monophos‐phorylated and C‐1,4′ diphosphorylated derivatives of β‐1,6‐linked D‐glucosamine disaccharide possessing both ester‐ and amide‐bound fatty acid substituents. They were divided into 4 groups, A, B, C and D in terms of the fatty acid substitution. Ester‐ and amine‐bound fatty acids of the analogues are both tetradecanoic acids (C14) in group A, C14 and (R)‐3‐hydroxytetradecanoic acids (C14‐OH) in group B, both C14‐OH in group C and C14‐OH and (R)‐3‐tetradecanoyloxytetradecanoic acids in group D.

Mitogenic activity was exhibited in spleen cells from C3H/HeN mice by the C‐1 monophosphorylated analogues in groups A and B, and by the C‐4′ monophosphorylated analogues in groups B, C and D, but not in cells from C3H/HeJ mice. Nonphosphorylated analogues in groups A, B and C, and C‐4′ monophosphorylated analogue in group A showed negative mitogenic activity. Only the nonphosphorylated analogue in group D exhibited mitogenic activity in spleen cells from C3H/HeJ mice as well as those from C3H/HeN mice. None of the other analogues exhibited the activity in C3H/HeJ spleen cells.

Polyclonal B cell activation activity was exhibited by the C‐1 monophosphorylated analogues in groups A and B, and by the C‐4′ monophosphorylated analogues in groups C and D. Nonphosphorylated analogues in all groups and C‐4′ monophosphorylated analogues in groups A and B showed negative PBA activity.

None of the analogues tested could induce any cytostatic macrophages from thioglycollate‐elicited peritoneal macrophages.
Assignment of human natural killer (NK)‐like cells to the T cell lineage. Single allospecific T cell clones lyse specific or NK‐sensitive target cells via distinct recognition structures
Authors: Moretta Alessandro , Pantaleo Giuseppe , Mingari Maria Cristina , Melioli Giovanni , Moretta Lorenzo , Cerottini Jean‐Charles
Published: February 2007

The aim of the present study was to define the cell lineage of mixed lymphocyte culture (MLC)‐induced natural killer (NK) effector cells. Human MLC cells were plated under limiting microculture conditions in the presence of irradiated spleen cells and interleukin 2‐containing supernatant. After 18 days, microcultures were scored for proliferation and for cytolytic activity against specific lymphoblasts and NK‐sensitive K562 target cells. About 1 in 7 and 1 in 5 proliferating microcultures had specific or NK‐like cytolytic activity, respectively. Moreover, several microcultures exhibited dual (specific and NK‐like) cytolytic activity, even when they had been established at relatively low numbers of responding cells/well (0.5–0.25) to ensure a high probability of monoclonality. Direct evidence for the existence of cytolytic effector cells with dual activity was achieved by using clones derived from single MLC T cells by micromanipulation. Out of 26 cytolytic clones so derived, 16 exhibited specific cytolytic activity, whereas 22 lysed K562 target cells. More interestingly, 12 of these 26 clones were active against both types of target cells. Only one of these clones was able to lyse autologous or unrelated target cells. In contrast, all such clones lysed the NK‐sensitive cell lines G11, MOLT‐4, Raji, Daudi, Chang and T‐24. Addition of saturating amounts of B9‐4 monoclonal antibody in the lytic assays resulted in the inhibition of the specific cytolysis, but not the NK‐like activity of clones with dual cytolytic activity. It thus appears that (a) alloreactive cytotoxic T lymphocytes can mediate both specific and NK‐like cytolysis and (b) two independent recognition structures are involved in this dual activity.
Functional significance of the regulation of macrophage Ia expression
Authors: Beller David I.
Published: February 2007

The relationship of la expression and antigen‐presenting function by macrophages has been evaluated. When macrophages are maintained in standard culture media, both Ia antigens and accessory cell function are lost. The reacquisition of these properties follows exposure to an Ia‐inducing lymphokine, for which cDNA‐derived interferon‐γ may substitute. The induction of function is related quantitatively to the level of Ia expression. Moreover, both properties reflect newly expressed Ia determinants, since treatment with anti‐I‐A plus complement at the beginning of culture diminishes neither the subsequent level of Ia expression nor function. Treatment with anti‐Mac‐1 plus complement, however, reduces function commensurate with the effectiveness of macrophage depletion. Finally, we find that fixation of macrophages after exposure to antigen does not inhibit antigen presentation, indicating that metabolic activity, while required for antigen processing, is not necessary for presentation.
Characterization of a major human antibody clonotype (1A) by monoclonal antibodies to combining siteassociated idiotopes
Authors: Zenke Gerhard , Eichmann Klaus , Emmrich Frank
Published: February 2007

To identify major human antibody clonotypes with specificity to N‐acetyl‐D‐glucosamine (GlcNAc), affinity‐purified antibody preparations of different human individuals were analyzed by isoelectric focusing (IEF). A major clonotype (1A) was identified representing 7% of all anti‐GlcNAc antibodies of one donor (no. 371). Anti‐GlcNAc antibodies of donor 371 were used as immunogen to prepare monoclonal anti‐idiotopic antibodies (mAb). Two anti‐idiotopic mAb were specific for the major clonotype 1A, as shown by blotting of the anti‐GlcNAc antibodies after IEF in thin‐layer agarose to nitrocellulose filters and staining with either iodinated antigen or iodinated anti‐idiotopic mAb. The two anti‐idiotopic mAb 13F15 and 10F59 were further analyzed with respect to the antigenic determinant which they recognize. Both were directed to combining site‐related determinants as shown by inhibition studies. Although 13F15 has a 500‐fold higher binding capacity (relative affinity) equal amounts do not inhibit more than 50% of 10F59 binding, suggesting that the two mAb detect two closely related, but not identical, idiotopes in the antigen combining site of clone 1A. Although clonotype 1A is a unique antibody exclusively found in IEF of donor 371, the idiotope 1A.1, defined by mAb 13F15, is a recurrent determinant detectable on different anti‐GlcNAc spectrotypes in various human sera.
T‐T cell interactions during in vitro cytotoxic T cell responses. VI. The role of T cell‐derived colony‐stimulating factor in helper T cell activation
Authors: Krönke Martin , Scheurich Peter , Pfizenmaier Klaus , Heeg Klaus , Wagner Hermann
Published: February 2007

A limiting dilution culture system has been used to analyze the role of T cell‐derived colony stimulating factor (CSF) during the activation of IL2‐producing helper T lymphocytes (HTL). EL4 thymoma‐derived supernatant (EL4‐SN) increased about 4‐fold the frequency of HTL precursors responding to metabolically active allogeneic stimulator cells. Upon biochemical separation this biological activity within the EL4‐SN segregated from interleukin 2 (IL2) and gamma interferon but was associated with or identical to CSF. Further, a comparable rise in HTL precursor frequencies was observed when semipurified interleukin 1 (IL1) derived from the P388 D1 cell line was added to limiting dilution cultures. In contrast to IL1, semipurified CSF failed to facilitate the activation of HTL precursors when heat‐treated allogeneic spleen cells were used as stimulator cells. Because EL4‐derived CSF was found to induce IL1 production by macrophages we conclude that T cell‐derived CSF amplifies, via stimulation of antigen‐presenting cells the number of inducible HTL precursors. Therefore, the CSF/IL 1‐dependent increase in HTL precursor frequencies reported here may reflect the differential activation threshold of HTL precursors, most of which will not be activated by antigen per se but only in presence of additional cytokines.
Heterogeneous and monoclonal helper T cells induce similar anti‐(4‐hydroxy‐3‐nitrophenyl)acetyl (NP) antibody populations in the primary adoptive response I. Isotype distribution
Authors: Tesch Hans , Smith Frances I. , Müller‐Hermes Werner J. P. , Rajewsky Klaus
Published: February 2007

In the response to NP‐lipopolysaccharide or NP‐Ficoll predominantly anti‐NP antibodies of the IgM class are produced in mice with lower amounts of IgG3 and IgG2b but little or no IgG1 and IgG2a. In contrast, in the primary T‐dependent response to NP‐keyhole limpet hemocyanin (KLH) or NP‐chicken gamma globulin high amounts of all IgG isotypes are induced. To investigate whether isotype‐specific T cells are responsible for these differences we carried out cell transfer experiments using carrier‐specific T cell lines. Two such lines were established and one of the two could be cloned. Upon activation by antigen the T cell lines induced unprimed syngeneic splenic B cells to proliferate and differentiate into antibody‐secreting cells in vitro in an antigen‐nonspecific way. Antigen‐specific activation of unprimed B cells in a cell transfer system in vivo showed that high concentrations of hapten‐specific antibodies of all IgG isotypes are induced through both carrier‐specific T helper lines. The isotypic pattern of these antibodies is similar to that produced via heterogeneous splenic T cells in the cell transfer system, or in normal animals on immunization with the same antigen. These results suggest that isotype‐specific T cells are not required for the production of IgG isotypes in a primary anti‐NP response and thus not responsible for the differences seen in isotypic patterns between T‐dependent and T‐independent responses.
Antibody‐free target cells stimulate chemiluminescence in polymorphonuclear leukocytes: an artifact due to mycoplasma contamination
Authors: Peterhans Ernst , Bertoni Giuseppe , Köppel Peter , Wyler Robert , Keller Robert
Published: February 2007

The generation of toxic oxygen species represents a significant mechanism in the killing of microorganisms and antibody‐coated cells by phagocytic cells. We have investigated the possibility that antibody‐free target cells stimulate reactive oxygen generation in polymorphonuclear leukocytes (PMNL) using the measurement of luminol‐dependent chemiluminescence (CL). It was found that the induction of CL consistently correlated with a mycoplasma contamination of the target cells. Free mycoplasma organisms of two species found frequently as contaminants in cell culture also stimulated CL. Upon artificial infection with mycoplasma, cultured cells acquired the capacity to evoke CL generation in PMNL. Our experiments strongly suggest that the induction of reactive oxygen generation by antibody‐free target cells is an artifact due to mycoplasma contamination of the target cell cultures.
Suppression of the cytotoxic T cell response to minor alloantigens in vivo Linked recognition by suppressor T cells
Authors: Gascoigne Nicholas R. J. , Crispe I. Nicholas
Published: February 2007

This report describes the activity of transferable suppressor T cells (Ts) generated in vivo in response to minor alloantigens. These Ts cells are antigen specific in both primary and secondary in vivo cytotoxic T lymphocyte responses to minor alloantigens and are the result of a host response rather than of a graft‐vs.‐host reaction. The Ts cells are produced soon after immunization and their activity is transient. They act via “linked recognition”, since they can suppress the cytotoxic T lymphocyte response to noncross‐reactive minor antigens, but only if these are presented on the same antigenic cell. A model for dominant low responsiveness in (high × low responder)F1 animals is proposed, whereby Ts cells, activated via the low responder allele, work by linked recognition to suppress helper cells activated via the high responder allele.
The surface expression of a tumor‐associated antigen on human kappa myeloma cells
Authors: Boux Heather A. , Raison Robert L. , Walker Karen Z. , Musgrove Elizabeth , Basten Antony
Published: February 2007

The monoclonal antibody K‐1‐21 defines an antigen, KMA (kappa myeloma antigen) on the surface of human χ myeloma cells. K‐1‐21 also recognizes human χ light chains in free form but not when covalently bonded to heavy chains. To examine the relationship between KMA and this determinant on free χ chains, the surface expression of KMA was examined on the IgG,χ myeloma line LICR LON/HMy2 (HMy2). No patching or capping was observed in the presence of K‐1‐21 alone, but KMA could be capped if the cells were incubated with K‐1‐21 followed by fluorescein isothiocyanate‐conjugated sheep F(ab′)2 anti‐mouse immunoglobulin. Capping was not affected by the inhibitors calcium ionophore or dibucaine. When IgG molecules were removed from the cell surface by capping with anti‐IgG antiserum both KMA and free χ light chains could still be detected with K‐1‐21 and a polyvalent anti‐χ antiserum, respectively. By contrast, after removal of all surface χ chains with the polyvalent anti‐χ serum, no staining was observed with K‐1‐21 indicating that KMA may be an epitope on free χ chains inserted in the membrane of χ myeloma cells but absent from normal cells. KMA cell surface expression varied with the stage of the cell cycle. Flow cytometric analysis of K‐1‐21‐stained HMy2 cells from either continuous cultures or from elutriated fractions enriched for various cell cycle phases showed that, within the cycling population, cells in G2 + M expressed KMA at a higher frequency and density than did cells in G1.
Analysis of antigen uptake and presentation by Epstein‐Barr virus‐transformed human lymphoblastoid B cells
Authors: Chu Edward , Umetsu Dale , Lareau Marianne , Schneeberger Evelyn , Geha Raif S.
Published: February 2007

Epstein‐Barr virus‐transformed human B cells (EBV‐B cells), but not resting B cells or B cells activated by T cell‐derived factors, have been shown to support the proliferation of tetanus toxoid (TT)‐specific autologous T cell clones in response to TT antigen. The accessory cell function of EBV‐B cells was compared to that of monocytes with regard to antigen uptake and processing.

After an 18‐h incubation period with 125I‐labeled TT, the amount of radioactivity associated with the cells (∼ 50 ng/107 cells) and the percentage of cells containing radiolabeled material (∼ 50%) were equivalent for EBV‐B cells and monocytes. Like with monocytes, EBV‐B cells pulsed with TT for 18 h or more were equivalent in their capacity to induce T cell proliferation to EBV‐B cells to which soluble TT was added for the duration of the culture period.

The requirements for antigen uptake and presentation to T cells were similar for both EBV‐B cells and monocytes. Both processes were energy dependent, inhibited by cold (4 °C), 2‐deoxyglucose, and azide, and both required no de novo protein synthesis as they were not affected by pretreatment of the cells with the irreversible protein inhibitor pactamycin. Trypsin treatment of antigen‐pulsed EBV‐B cells and monocytes followed by fixation for 1 min in 0.03% paraformaldehyde completely abolished the capacity of both cell types to induce T cell proliferation.

In both EBV‐B cells and monocytes, antigen presentation, but not antigen uptake, was inhibited by the addition of the lysosomotropic agent chloroquine during the antigen‐pulse period suggesting that the mechanisms of antigen processing are similar for both cell types. Vacuoles positive for acid phosphatase with an electron microscopic structure similar to that of lysosomes were found in EBV‐B cells but not in resting B cells or B cells activated by T cell‐derived factors.

The present observations indicate that EBV‐B cells take up antigen and process it in a fashion similar to monocytes. The presence of lysosomes appears to correlate with the capacity of B cells to present antigen.
HLA‐DC antigens can serve as recognition elements for human cytotoxic T lymphocytes
Authors: Spits Hergen , Borst Jannie , Giphart Marius , Coligan John , Terhorst Cox , de Vries Jan E.
Published: February 2007

The specificity of four cytotoxic T lymphocyte (CTL) clones which recognize class II major histocompatibility complex (MHC) antigens was analyzed. All clones recognized antigens associated with the serologically defined HLA‐DRw6 specificity. The activity of two of these clones, JR‐2‐2 and JR‐2‐10, could be inhibited by a monoclonal antibody Q 5/13 specific for a monomorphic determinant present on HLA‐DR. In contrast, the activity of the two other CTL clones, JR‐2‐19 and JR‐2‐26, was not blocked by Q 5/13, but by a new monoclonal reagent, SPV‐L3. This latter monoclonal antibody precipitated a two‐chain structure of 28 kDa and 33 kDa and reacts with a monomorphic determinant. The molecular weight of the polypeptides precipitated with SPV‐L3 was slightly less than those precipitated with a HLA‐DR‐specific monoclonal reagent. In addition two‐dimensional gel electrophoresis showed that the antigen precipitated by SPV‐L3 differed in charge from those precipitated with the anti‐HLA‐DR antibody. These results indicate that SPV‐L3 recognizes a class II MHC product different from HLA‐DR. This observation was confirmed by partial amino acid sequence analysis of the two chains which revealed that the molecule precipitated by SPV‐L3 is homologous to HLA‐DC/DS molecules. Therefore this report provides the first evidence that human cytotoxic T cells can recognize HLA‐DC/DS antigens.
Independent inhibition of IL 2 synthesis and cell proliferation by anti‐Ia antibodies in mixed lymphocyte responses to Mls
Authors: Macphail Stuart , Stutman Osias
Published: February 2007

Whole anti‐I region antisera and monoclonal anti‐Ia antibodies are capable of totally inhibiting the response of murine spleen cells to the non‐H‐2 Mls alloantigens. This inhibition was shown to be highly specific for the appropriate Ia and operative at the level of the stimulator cell. Both cell proliferation and interleukin (IL 2) production were inhibited; however, the inhibition of cell proliferation was not due only to the inhibition of IL 2 production and the inhibition of IL 2 production was not solely due to the inhibition of IL1 production. We propose that both the IL 2‐producing cell and the proliferating cell need to “see” an intact Ia+ cell and perhaps Ia antigens in association with Mls determinants to respond.
Surface immunoglobulin phenotype of complement receptor‐positive and complement receptor‐negative B cells of normal and xid immunodeficient mice
Authors: Lindsten Tullia , Finkelman Fred D. , Thompson Craig B. , Andersson Birger , Scher Irwin
Published: February 2007

Single‐ and dual‐parameter fluorescence‐activated cell sorter analyses of complement receptor‐positive and ‐negative (CR+ and CR−) splenic B cells of normal and xid immunodeficient mice were performed to determine the frequencies of surface (s) IgM+ and sIgD+ cells, as well as the quantities of sIgM and sIgD on these cells. Single parameter analysis revealed identical frequencies of sIgM+ and sIgD+ cells in the CR+ B cell population of normal mice. In contrast, the percentage of sIgM+ cells exceeded that of sIgD+ cells in the CR− normal and CR+ xid B cell population to a similar extent. CR− xid B cells showed an even greater discrepancy between sIgM+ and sIgD+ cells.

Dual‐parameter analysis of CR+ normal B cells showed the typical sIg phenotype of mature B cells, i.e. an intermediate density of sIgM and a high density of sIgD. The CR+ xid B cell population displayed an increase in cells with high sIgM and no‐to‐low sIgD and a decrease in cells with intermediate sIgM and high sIgD when compared to CR+ normal cells. CR− B cells from normal and xid immunodeficient mice were similar in that both consisted largely of a cell population with low sIgM and no‐to‐low sIgD. However, the CR− normal B cell population also contained cells with intermediate sIgM and high sIgD, which were lacking in the CR− xid cell population. Similar results were obtained when CR+ and CR− B cells from normal and xid mice were purified from a small resting B cell population, isolated by elutriation. This indicates that the sIg phenotype of CR− cells is not due to the presence of large cells which have lost sIgD during activation.
Polyclonal B cell responses in the presence of defined filler cells: complementary effects of lipopolysaccharide and anti‐immunoglobulin antibodies
Authors: Zubler Rudolf H.
Published: February 2007

The signal requirement for polyclonal B cell responses in the presence of T helper (Th) cells, lipopolysaccharide (LPS), anti‐Ig antibodies coupled to Sepharose beads (anti‐Ig) and/or T cell supernatants (SN) was studied in a murine system using (a) low numbers of B cells/culture in order to reduce the effects of contaminating Th cells and (b) defined sources of irradiated filler cells in the form of EL4 thymoma cells or cloned male H‐Y antigen‐specific Th cells. The results demonstrate that for optimal proliferation as well as (protein A) plaque‐forming cell (PFC) generation B cells required at least two activation signals in addition to factor(s) present in Th or EL4 SN, i.e. either a specific (or concanavalin A‐dependent nonspecific) Th signal and anti‐Ig or (in cultures with EL4 filler cells) LPS and anti‐Ig. While confirming several previous studies, including our own, which showed a requirement for two activation signals in conjunction with T cell factors in antigen‐specific B cell responses, the present system differs from previous polyclonal systems by showing nonoverlapping effects of LPS or a specific Th signal on the one hand and anti‐Ig on the other in the induction of growth factor responsiveness of B cells. In addition, limiting dilution analysis showed that in cultures with EL4 filler cells, LPS, anti‐Ig and EL4 SN 1/8 surface Ig‐positive cells generated > 10 PFC with a mean clone size of 70 PFC and indicated that only the B cells were limiting. This system using defined thymoma filler cells should be useful for assaying factors potentially replacing the LPS or anti‐Ig signals.
Chicken thymocyte‐specific antigen identified by monoclonal antibodies: ontogeny, tissue distribution and biochemical characterization
Authors: Chen Chen‐lo H. , Chanh Tran C. , Cooper Max D.
Published: February 2007

Two monoclonal antibodies, CT‐1 (IgG1,χ) and CT‐1a (IgG3,χ), were prepared against chicken thymocytes. The antigen identified by these antibodies was found to be a glycoprotein with a major polypeptide component having an apparent molecular weight of 63000 and a minor polypeptide component of approximately 103000. Immunoprecipitation and blocking experiments revealed that the two antibodies react with different antigenic determinants of the molecule. Ontogenic studies employing immunofluorescence failed to reveal the antigenic determinants on cells from the embryo or embryonic yolk sac on days 3 and 6 of incubation. The number of embryonic thymocytes bearing the molecules detected by these antibodies increased from 6% on day 12 to 54% on day 13; the frequency of thymocytes expressing this glycoprotein reached adult levels (> 90%) by the 15th day of embryonic age. In contrast, the CT‐1 and CT‐1a antibodies reacted with only 2‐5% of blood and splenic cells and less than 0.05% of cells from bursa or bone marrow of young adult chickens. In the quail CT‐1 reacted with cortical, but not medullary, thymocytes, while the CT‐1a antibody was unreactive with quail thymocytes. The surface glycoproteins detected by these discriminating monoclonal antibodies may provide an important discriminating marker for thymic lymphocytes in the chicken and the quail.
The alloantigenic organization of RT1Aa, a class I major histocompatibility complex molecule of the rat
Authors: Diamond Austin G. , Larkins Audrey P. , Wright Bruce , Ellis Susan T. , Butcher Geoffrey W. , Howard Jonathan C.
Published: February 2007

Over 300 monoclonal IgG alloantibodies have been prepared against RT1Aa, the class I major histocompatibility complex molecule of the DA rat. In this study a combination of techniques is exploited to show that all these antibodies can be allocated to 9 antigenic sites which form a continuous antigenic surface, that is, no site is completely isolated from the rest. The results suggest that techniques for the identification of antigenic sites using competitive inhibition of monoclonal antibody binding are generally valid, in the sense that competition between antibodies appears most commonly to represent competition between combining sites for a structural feature of the antigenic surface. From the distribution of antibodies between sites, it is clear that the RT1Aa molecule has three immunogenic areas against which nearly all the antibodies studied were directed. Of these areas one is both antigenically complex, consisting of four closely spaced sites, and remarkably immunodominant. Antibodies directed at sites between the major areas are extremely rare.
Immunodominant protein epitopes I. Induction of suppression to hen egg white lysozyme is obliterated by removal of the first three N‐terminal amino acids
Authors: Wicker Linda S. , Katz Michael , Sercarz Eli E. , Miller Alexander
Published: February 2007

The lack of response to hen egg white lysozyme (HEL) by C57BL (H‐2b) mice has been demonstrated previously to be related to the induction of suppressor T (Ts) cells which recognize the amino terminal region of HEL. In this report, the nature of the protein determinant required for Ts cell induction is more precisely detailed using des‐1,2,3‐HEL (AP‐HEL) prepared with an aminopeptidase purified from Aeromonas proteolytica. Remarkably, the removal of just these three amino acids obliterates the ability of HEL to induce Ts cells specific for HEL. Additionally, in contrast to HEL, AP‐HEL is able to prime for an in vitro T cell proliferative response to either AP‐HEL or HEL. Thus, removal of a very limited region of a protein antigen can drastically alter its immunogenic properties.
Pre‐B cells in bone marrow: peanut agglutinin binding and separation of cytoplasmic μ chain‐bearing cell populations in normal, post‐irradiation and polycythemic mice using fluorescence‐activated cell sorting
Authors: Osmond Dennis G.
Published: February 2007

Mouse bone marrow cells exposed to fluorescein‐conjugated peanut agglutinin (PNA) showed subsets of highly labeled cells when analyzed in a fluorescence‐activated cell sorter. After separating three cell fractions of large and small PNA‐binding cells and PNA‐nonbinding cells, respectively, the B lymphocyte precursor (pre‐B) cells, having cytoplasmic μ chains (cμ) without surface μ chains (sμ), were recovered solely in the PNA‐binding fractions. Only a minority of sμ+ small lymphocytes having the lowest densities of sμ bound PNA. Small and large cμ+sμ− pre‐B cell populations were separated in high degrees of purity in the PNA‐binding fractions, especially when obtained from bone marrow undergoing lymphoid regeneration after sublethal X‐irradiation and during stimulation of lymphocyte production in post‐polycythemic erythroid suppression. Characteristic shifts in the size distribution profile of PNA‐binding cells reflected changes in the maturation stage of the pre‐B cells. The results demonstrate that surface membrane components with strong PNA‐binding capacities characterize cμ+sμ− pre‐B cells in the bone marrow during both normal and perturbed primary B lymphocyte genesis. The PNA‐binding sites become undetectable soon after the first expression of sμ. This property permits the isolation from the bone marrow of high concentrations of subsets of large and small cμ+sμ− cells in a viable state suitable for use in further functional studies.
Poly(Glu60, Ala30, Tyr10) (GAT)‐specific T cells do not express B cell public idiotopes but can be primed by monoclonal anti‐idiotypic antibodies
Authors: Bismuth Georges , Sommé Gérard , Roth Claude , Gougeon Marie‐Lise , Thèze Jacques
Published: February 2007

Eight monoclonal anti‐idiotypic antibodies directed against public idiotopes have been further characterized: (a) they bind to public idiotopes with a high affinity; (b) they recognize all anti‐poly(Glu60, Ala30, Tyr10) (GAT) antibodies as measured by inhibition of the anti‐GAT plaque‐forming cell response. This has been verified in three strains of mice. These reagents were not able to detect idiotope expression on eight GAT‐specific helper T cell lines and clones. This result was obtained by two techniques: (a) idiotope expression at the T cell surface was measured by indirect immunofluorescence using a cell sorter with surface antigens H‐2D, Thy‐1.2, Lyt‐1 and L3T4 as positive controls; (b) after immunoadsorption of [35S]methionine‐labeled cellular extracts from two lines, no unique molecule was retained by the HP‐idp22 monoclonal anti‐idiotypic antibody coupled to Sepharose. Despite these negative results, this antibody was found to prime lymph node cells in vivo, which were able to proliferate specifically in response to GAT. Two T cell lines derived from this lymphocyte population do not express any of the idiotopes tested. These results suggest that monoclonal anti‐idiotypic antibodies may be influencing T lymphocyte activity indirectly.
Monoclonal anti‐cardiolipin antibodies bind to DNA
Authors: Rauch Joyce , Tannenbaum Hyman , Stollar B. David , Schwartz Robert S.
Published: February 2007

BALB/c mice immunized with the phospholipid, cardiolipin, produced anti‐cardiolipin and anti‐DNA antibodies. Seven hybridomas derived from spleen cells of the cardiolipin‐immunized mice produced cardiolipin‐binding monoclonal antibodies that also bound to the polynucleotides DNA, poly(dT), and poly(I). The seven cardiolipin‐induced monoclonal antibodies shared idiotypic determinants with a high frequency idiotypic marker of spontaneously expressed anti‐DNA autoantibodies of lupus‐prone MRL‐lpr/lpr mice. The monoclonal antibodies presumably bound to phosphodiester phosphate groups that occur in both polynucleotides and phospholipids. The results imply that production of anti‐DNA autoantibodies does not require immunization by DNA.