bluuebunny/crossref_metadata_2025 huggingface dataset:
Antigen-inducible, H-2-restricted, interleukin-2-producing T cell hybridomas. Lack of independent antigen and H-2 recognition
Authors: Kappler JW , Skidmore B , White J , Marrack P
Published: June 2004

We developed a method for production of antigen-specific, H-2-restricted T cell hybrids. The tumor cell partner in the fusions was itself a T cell hybrid, FS6-14.13.AG2 (or its derivatives), which could be induced to produce the growth factor, interleukin-2 (IL-2), in response to a challenge with concanavalin A, but had no known antigen specificity. The normal T cell partner in the fusions was a population of lymph node T cell blasts that had been highly enriched in antigen-specific, H-2-restricted T cells by in vivo immunization, followed by in vitro challenge with antigen and clonal expansion in IL-2-containing medium. These fusions produced hybrids that grew constitutively in culture. A sizable proportion of the hybrids demonstrated the ability to produce IL-2 in response to a challenge with specific antigen presented by irradiated spleen cells of the appropriate H-2 type.

Four cloned antigen/H-2-specific hybrid lines were produced. AO-40.10 responded to chicken ovalbumin (OVA) when presented by I-A(k)-bearing cells. DC1.18.3 responded to the apo form of beef cytochrome c when presented with I-A(d). AODK-10.4 responded to keyhole limpet hemocyanin (KLH) presented with I-A (d). AODK-1.16 also responded to KLH presented by a product of the I region of H-2(d), but the data were consistent with either a product of the I-J-I-E(d) region or a combinatorial molecule with elements from both I-A(d) and I-E(d)/I-C(d). Coincidentally, AO-40.10 was shown to have an unexpected alloreactivity with a product of H-2(b) mapping to the K-I-A region. These hybrids should prove invaluable as sources of monoclonal material for the study of the receptor(s) on T cells with H-2-restricted antigen specificities.

We also generated T cell hybrids with two antigen/H-2 specificities by fusing an azaguanine-resistant clone of AO-40.10 to normal T cells with a different antigen/H-2 specificity. Many of the hybrids retained reactivity to OVA plus H-2(a) and to the second antigen/H-2 combination. None reacted to either OVA plus the second H-2 type or to the second antigen plus H-2(a).

One of these hybrids was successfully cloned to produce the line AOFK- 11.11.1. It retained the ability to recognize OVA plus I-A(k) inherited from one parent, and KLH plus IA(f) inherited from the other. It did not recognize OVA plus IA(f) or KLH plus I-A(k).

These results have some bearing on models describing the nature of T cell receptors for antigen recognized in association with H-2 products. They do not support models in which antigen and H-2 are recognized separately by two independent T cell receptors.
Priming of T helper cells by antigen-activated B cells. B cell-primed Lyt-1+ helper cells are restricted to cooperate with B cells expressing the IgvH phenotype of the priming B cells.
Authors: L'Age-Stehr J
Published: June 2004

Activated B cells isolated shortly after primary immunization of BALB/c donor mice with sheep erythrocytes (SRBC), were transferred to normal syngeneic recipients or to low-dose cyclophosphamide-pretreated syngeneic recipients. In pretreated recipients, the transfer of activated B cells, but not of T cells or macrophages, resulted in an augmented production of indirect plaque-forming cells in the primary immune response to SRBC but not to horse erythrocytes. It was shown in double-transfer experiments that T helper cells (Lyt-1+) had been stimulated by the transfer of antigen-activated B cells. Criss-cross double-transfer experiments using the mouse strains CB20 and BAB14 (congenic to BALB/c at the loci coding for the immunoglobulin heavy chain) indicate that those T helper cells are primed after recognition of B cell products that are encoded for by genes linked to the loci coding for the variable region of the immunoglobulin heavy chain (IgVH). The thus-primed Ig-dependent T helper cells (THIg) are adaptively restricted to cooperate with B cells that display IgVH-linked gene products similar to those that originally stimulated the THIg. These findings suggest that in the course of an immune response to T cell-dependent antigens, help for the production of specific IgG can be provided by THIg that have been primed and/or clonally expanded after recognition of IgVH-linked gene products by (e.g., complementary) T cell receptors.
Antigen-specific t lymphocyte clones. II. Purifcation and biological characterization of an antigen-specific suppressive protein synthesized by cloned T cells
Authors: Fresno M , McVay-Boudreau L , Nabel G , Cantor H
Published: June 2004

We have generated an antigen-specific T suppressor clone that synthesizes 70,000-mol wt peptides that have antigen-specific-binding activity. Although these data also indicated that antigen-binding peptides completely inhibited the in vitro primary response to a complex antigen, suppression might reflect the combined biologic activities of many different 70-mol wt polypeptides or polypeptides associated with the 70,000-mol wt material by noncovalent interactions. The protein responsible for antigen-specific suppression was therefore purified to virtual homogeneity after sequential separation of internally labeled supernate peptides on Sephacryl S-200 and DEAE-cellulose columns followed by isoeleetrofocusing. The resulting protein is greater than 95 percent homogeneous according to sodium dodeeyl sulfate-polyacrylamide electrophoresis and represents two peptides having two very close but distinguishable isoelectric point values of approximately 5.0.

The purified molecules are retained by columns coated with lentil lectin or antigen but not by columns coated with antisera specific for immunoglobulins, the I region of the major histocompatibility complex or Ly-1 or Ly-2 antigens. Less than 50 pg of the purified glycoprotein specifically and completely suppresses production of anti-sheep erythrocyte plaque-forming cell by mixtures of 10(6) Ly-1 cells and B cells and this is a result of inactivation of Ly-l-mediated helper function. Specific inactivation of T (Th) cells by the 70,000-mol wt molecule is rapid, specific, and requires the presence of antigen. The mechanism of specific suppression of Th function may depend upon two functionally distinct regions of the 70,000-mol wt molecule: one that binds antigen and a second that mediates suppression.
Stimulation of primary in vitro IgE antibody responses in culture of human peripheral mononuclear cells.
Authors: Nonaka M , Zuraw B L , O'Hair C H , Katz D H
Published: June 2004

Experiments are presented herein that demonstrate the capacity to stimulate human peripheral mononuclear cells to synthesize and secrete significant quantities of IgE molecules in vitro by exposure to appropriate concentrations of 2,4-dinitrophenyl (DNP)-protein conjugates, pokeweed mitogen (PWM), or a combination of DNP-proteins and PWM. Cultures stimulated in this fashion synthesize increased quantities of both total IgE and DNP-specific IgE antibody molecules. This in vitro human IgE antibody system should provide a useful tool for further exploration of regulatory control of IgE responses in both normal humans and those manifesting various forms of IgE-mediated allergic disorders.
Bone marrow origin of Ia-positive cells in the medulla rat thymus.
Authors: Barclay A N , Mayrhofer G
Published: June 2004

Irradiated rats were reconstituted with bone marrow from F1 hybrids. Ia antigen of donor-bone marrow origin was detected by an immunoperoxidase technique on cryostat sections and found predominantly in the medulla of rat thymus 2 wk after reconstitution. These Ia-bearing cells increased in number with time after reconstitution, but the Ia on the cortical epithelial cells remained of host origin. The nature of the bone marrow-derived cells and their implication for major histocompatibility complex restriction are discussed.
Interaction of Leishmania with a macrophage cell line. Correlation between intracellular killing and the generation of oxygen intermediates.
Authors: Murray H W
Published: June 2004

The promastigote form of Leishmania donovani and Leishmania tropica, the etiologic agents of visceral and cutaneous leishmaniasis, respectively, readily parasitize unstimulated J774G8 macrophage-like cells, whereas 80-95% of the same promastigotes are killed within resident macrophages from normal BALB/c mice. This striking difference in intracellular anti-leishmanial activity correlated closely with the capacity to generate toxic oxygen intermediates. Thus, after triggering with phorbol myristate acetate or phagocytosis of zymosan or promastigotes, 90% of the J774G8 cells failed to reduce nitroblue tetrazolium, and released 5-10-fold less O2- and H2O2 than BALB/c macrophages. Exposure to concanavalin A-stimulated lymphokine, however, effectively enhanced the oxidative response of J774G8 cells, and, similarly, induced intracellular anti-leishmanial activity. Inhibiting macrophage H2O2 production consistently decreased the killing of Leishmania by lymphokine-treated J774G8 cells. These observations illustrate the usefulness of examining homogeneous macrophage cell lines that are deficient in a particular effector function, and also serve reemphasize the important role of oxygen intermediates in the microbicidal response of mononuclear phagocytes to intracellular parasites.
Studies on the antigenic determinants in the self-association of IgG rheumatoid factor
Authors: Nardella FA , Teller DC , Mannik M
Published: June 2004

The number, location, and other characteristics of the antigenic determinants for self-association of IgG-rheumatoid factors (IgG-RF) were examined using the IgG-RF isolated from the plasma of one patient as a model system. Affinity chromatography was employed for isolation of the IgG-RF. Sedimentation equilibrium ultracentrifugation was used to study the various interactions.

The antigenic valence of IgG-RF Fc, normal human Fc, and rabbit Fc fragments was two for the interaction with Fab fragments from IgG-RF, as might be expected from the molecular symmetry of IgG. The antigenic valence of intact normal IgG, however, was only one, indicating that when one of the available antigenic determinants interacted with the Fab fragment of IgG-RF, the other determinant becomes sterically inaccessible. Reduction and alkylation, known to increase the flexibility of the hinge region, did not alter the antigenic valence of IgG for Fab fragments of IgG-RF. The antigenic valence of IgG-RF in self-association could not be experimentally determined but must be two to permit the observed concentration-dependent further polymer formation of IgG-RF dimers.

Unique antigenic determinants on the Fc fragments of IgG-RF were sought and not found, thus reaffirming the formation of two antigen-antibody bonds as the basis for dimerization of IgG-RF molecules.

The pFc' and Fc' fragments, representing Cγ3 domains of IgG, failed to show significant interaction with Fab fragments of IgG-RF, indicating that the antigenic determinants were not expressed by the Cγ3 regions but are located either on Cγ2 region or require intact Cγ2 and Cγ3 regions for expression. These conclusions were corroborated by the antigenic valence of one for the Fc(i) fragment, a new papain-generated intermediate fragment of Fc, composed of two intact Cγ3 domains and one intact Cγ2 domain.

Normal IgG, because of its valence of one for interaction with IgG-RF, would effectively terminate further polymerization of IgG-RF dimers. This may well in part explain the finding of smaller IgG-RF complexes in the serum than in synovial fluid of patients with rheumatoid arthritis.
Inhibition of T cell activation in vivo with mixtures of monoclonal antibodies specific for I-A and I-A/E molecules.
Authors: Sprent J , Lerner E A , Bruce J , Symington F W
Published: June 2004

(CBA x B6)F1 (Iak x Iab) T cells were activated to sheep erythrocytes in irradiated F1 mice in the presence of various monoclonal anti-Ia reagents and then tested for their capacity to collaborate with B cells from B10.BR (I-Ak, I-Ek) (kk), B10.A(4R) (kb), and B10 (bb) mice. Anti-I-Ak antibodies blocked the generation of help for B10.A(4R) B cells, but not B10.BR or B10 B cells. An anti-I-Ab antibody blocked help for B10 B cells, but not for B10.BR or B10.A(4R) B cells. An antibody (Y-17) specific for I-Ak/Ek and I-Ab/Ek molecules, but not for I-Ak or I-Ab molecules, failed to impair the generation of help for B10.BR, B10.A (4R), or B10 B cells. In marked contrast to injecting each antibody separately, a mixture of anti-I-Ak and anti-I-Ak,b/Ek (Y-17) antibodies virtually abolished the generation of help for B10.BR B cells. A mixture of anti-I-Ak and anti-I-Ab antibodies effectively blocked help for (4R x B10)F1 B cells, i.e., cells expressing hybrid I-A molecules. These two antibodies only marginally impaired help for (CBA x B6)F1 B cells. To block help for (CBA x B6)F1 B cells required selection in the presence of a cocktail of anti-I-Ak, anti-I-Ab, and anti-I-Ak,b/Ek antibodies. The implications of these findings are discussed.
Immune response to stage-specific surface antigens of the parasitic nematode Trichinella spiralis.
Authors: Philipp M , Taylor P M , Parkhouse R M , Ogilvie B M
Published: June 2004

Rats were infected with the nematode Trichinella spiralis and the primary serum antibody response to antigenic surface proteins of infective larvae, intestinal worms, and newborn larvae was studies. 1 wk after infection, the sera contained antibodies to surface antigens of both infective larvae and intestinal worms. These early sera, however, failed to react with newborn larvae surface antigens. In addition, adsorption of sera with living intestinal worms or infective larvae removed antibodies to surface antigens of the homologous stage only. Finally, the time-course of appearance of antibodies that mediate eosinophil adherence to the surface of each stage of the parasite. We concluded that in a primary infection in rats, the surface proteins of T. spiralis used in this study are antigenically stage specific. Furthermore, they could be targets for the stage-specific, antibody-dependent eosinophil-mediated destruction of this parasite, known to occur in vitro.
Surface properties of bacillus Calmette-Guérin-activated mouse macrophages. Reduced expression of mannose-specific endocytosis, Fc receptors, and antigen F4/80 accompanies induction of Ia.
Authors: Ezekowitz R A , Austyn J , Stahl P D , Gordon S
Published: June 2004

Infection of the mouse peritoneal cavity by bacillus Calmette-Guérin (BCG) markedly alters the surface properties of the macrophages induced, compared with cells obtained from uninfected control animals or after injection of thioglycollate broth. Quantitative binding assays with radiolabeled ligands or antibodies showed that BCG-activated peritoneal macrophages (BCG-PM) expressed one-fourth or less receptor activity for mannose-terminal glycoconjugates as well as reduced levels of Fc receptors and of antigen F4/80 compared with nonactivated macrophages. Endocytosis mediated by mannose-specific receptors was reduced in parallel. In contrast, surface Ia antigen was increased threefold in the same adherent cell population. Radioautographic analysis confirmed that greater than 80% of adherent cells still expressed low levels of the macrophage-specific mannosyl receptor and antigen F4/80, and that I antigens had been induced on 64% of macrophages rather than on other cells. Control experiments established that only the BCG-PM macrophages released H2O2 after stimulation with phorbol myristate acetate, whereas both BCG-PM and thioglycollate-induced macrophages produced superoxide anion and plasminogen activator. The BCG-PM were viable, secreted normal levels of lysozyme, and displayed a stable phenotype after cultivation for 60 h. Inhibitors of oxygen products, prostaglandins, and proteases did not alter reduced endocytosis by BCG-PM. These studies indicated that expression of macrophage surface markers is reversed by BCG-activation, and that their known enhanced ability to lyse target cells extracellularly is associated with decreased endocytosis via specific receptors. Whether these changes are a result of an altered cell population or of modulation of selective surface properties is not known.
Biological properties of an influenza A virus-specific killer T cell clone. Inhibition of virus replication in vivo and induction of delayed-type hypersensitivity reactions.
Authors: Lin Y L , Askonas B A
Published: June 2004

We tested two biological properties of a continuously growing mouse cytotoxic T cell line, L4, which is specific for influenza A virus and has been cloned and recloned many times. We previously reported that L4 cells are H-2 restricted and cross-reactive for all type A influenza viruses, whereas they do not recognize type B influenza viruses. They bear Thy-1 and Lyt-2 markers. In the present study, we show that L4 cytotoxic T cells protect mice against a lethal influenza infection on transfer to syngeneic recipients, and reduce virus titers in the lungs of mice challenged with a heterologous type A influenza virus. This provides further support for the active role of cytotoxic T cells in limiting virus replication in influenza infection. We could also demonstrate that the cloned cytotoxic T cells induce a delayed-type hypersensitivity skin reaction in the footpads of mice challenged with live or inactivated influenza virus. This reaction can be observed at 24 h, but has declined by 48 h. A clone of cells derived from L4 that has lost its cytotoxic potential and its ability to recognize infected cells did not induce a delayed-type hypersensitivity reaction in the presence of virus. Thus, cytotoxic T cells actively killing influenza virus-infected cells are able to induce a delayed-type hypersensitivity skin reaction to homologous and heterologous type A influenza viruses.
Heterogeneity of natural killer cells in the mouse.
Authors: Lust J A , Kumar V , Burton R C , Bartlett S P , Bennett M
Published: June 2004

Mice were treated with the bone-seeking isotope, 89Sr, cyclophosphamide, and short-term lethal irradiation in vivo, and murine spleen cells are treated with anti-Nk-1.2 plus complement (C) in vitro. Fresh spleen cell suspensions from the above groups and from beige and neonatal mice were subsequently tested for natural killer (NK) cell activity against a panel of lymphoid and nonlymphoid tumor cell target. NK cell reactivities against YAC-1, MPC-11, and Cl.18 tumors were markedly and consistently reduced in (a) mice treated with 89Sr, (b) spleen cells treated with anti-Nk-1.2 plus C, and (c) C57BL/6 bg/bg mice. In contrast, NK activities against FLD-3 and WEHI-164.1 tumors were usually normal in mice treated with 89Sr, in beige mutant mice, and in spleen cells after treatment with anti-Nk-1.2 antibody and C. It appears, therefore, that two major groups of NK cells exist in fresh mouse spleen cells suspensions. NK-A cells are marrow dependent, Nk antigen positive, and deficient in beige mice; these lyse YAC-1, MPC-11, and Cl.18 tumors. NK-B cells, which are responsible for the lysis of WEHI-164.1 and FLD-3, are Nk antigen negative, marrow independent, and unaffected by the bg/bg mutation. Other features of NK-B cells, suggest that these NK cells, although they share the characteristics mentioned above, differ among themselves especially with respect to age of maturation and susceptibility to cyclophosphamide and total body irradiation. The NK-B group may therefore induce subsets that remain to be defined.
Physiology of IgD. I. Compensatory phenomena in B lymphocyte activation in mice treated with anti-IgD antibodies.
Authors: Jacobson E B , Baine Y , Chen Y W , Flotte T , O'Neil M J , Pernis B , Siskind G W , Thorbecke G J , Tonda P
Published: June 2004

The role of delta-positive cells in the immune response was studied by comparing the effects of treatment with allotype-specific IgD hybridoma antibody on homozygous BALB/c or SJL/J and heterozygous (BALB x SJL)F1 mice. Homozygous mice, injected from birth with the relevant anti-delta antibody, made primary or secondary immune responses to intravenously injected trinitrophenyl (TNP)-Brucella abortus, TNP-Ficoll, and TNP-keyhole limpet hemocyanin, which did not differ significantly from those of control mice, despite the fact that IgD+ cells were depleted and Ig+ cells were markedly reduced in the spleens of treated mice. Responses in nodes draining a local injection of TNP-Brucella abortus were, however, significantly suppressed. Heterozygous mice, injected from birth with either anti-Ig-5a or anti-Ig-5b, showed a marked reduction in the number cells producing IgG antibody of linked allotype specificity in the secondary response to intravenously injected sheep erythrocytes. A corresponding decrease in the amount of serum IgG2a of that allotype specificity was also noted. However, in agreement with the results obtained in homozygotes, heterozygotes injected simultaneously with anti-IgD directed against each of the allotypes made normal, if not enhanced, plaque-forming cell responses of both allotype specificities. Similarly, serum IgG2a levels were normal in all but one mouse treated in this fashion. These results indicate that IgD+ cells are not essential for an immune response in vivo. Although the delta-positive cell is used preferentially under normal conditions, it appears that an alternative mechanism exists by which, in the absence of these cells, the animal is able to make a normal immune response.
Natural antibodies to the human T cell lymphoma virus in patients with cutaneous T cell lymphomas.
Authors: Posner L E , Robert-Guroff M , Kalyanaraman V S , Poiesz B J , Ruscetti F W , Fossieck B , Bunn P A , Minna J D , Gallo R C
Published: June 2004

Sera from patients with cutaneous T cell lymphoma and leukemia were screened for the presence of natural antibody to the human T cell lymphoma (leukemia) virus, HTLVCR, using a solid-phase radioimmunoassay. Sera from two patients, including patient CR, from whose cultured T lymphoblastic cell line (HUT102), the retrovirus HTLVCR was isolated, reacted specifically with proteins of HTLVCR. Serum from patient CR also reacted specifically with proteins of HTLVMB, an independent but highly related retroviral isolate from a patient with Sezary T cell leukemia. The specificity for HTLVCR proteins was demonstrated by solid-phase immunocompetition assays and competition radioimmunoprecipitation assays. Analysis of radioimmunoprecipitates indicated that the natural antibodies were directed against HTLVCR core proteins with molecular weights of 24,000 and 19,000 (p24 and p19). Whereas the serum reactivities for HTLVCR proteins were shown to be highly specific, additional reactivities seen against proteins of animal retroviruses including GaLV, SSV, FeLV, and BaEV were clearly shown not to be viral specific but rather were due to reactivity with cellular antigens contaminating the viral preparations or with related antigens present in fetal calf serum. These results demonstrating natural antibodies to HTLVCR provide the first evidence for a specific antibody response to a retrovirus in humans.
Demonstration and characterization of Ia-positive dendritic cells in the interstitial connective tissues of rat heart and other tissues, but not brain.
Authors: Hart D N , Fabre J W
Published: June 2004

In this study, we have used a mouse monoclonal antibody to rat Ia (RT1-B or class II) antigens to demonstrate, by immunofluorescence on frozen sections, intensely Ia-positive dendritic cells in the interstitial connective tissues of every tissue we have examined (heart, liver, thyroid, pancreas, skin, kidney, ureter, and bladder) with the striking exception of brain. The characteristics of the interstitial dendritic cell found in heart were studied in detail, and this cell was shown to be negative for acid phosphatase, beta-glucuronidase, and ATPase activity, and certainly some and probably all of the cells were negative for nonspecific esterase activity. Experiments with colloidal carbon suggested that the cell was either poorly or not at all phagocytic. The cells were negative for surface immunoglobulin and the W3/13 antigen, but positive for the leukocyte common antigen and the SD (Class I) antigens of the major histocompatibility complex. The cell was shown to be of bone marrow origin, and either the cell itself, or more probably its precursor, was shown to be sensitive to irradiation and to cyclophosphamide. All strains tested--including the nude rat--had large numbers of interstitial dendritic cells. The widespread distribution, except in brain, of this cell, which resembles in every respect the dendritic cell described by Steinman et al. (4) in the spleen and lymph nodes of the mouse, is of interest, and the implications in this finding are discussed.
Purification of murine T cell growth factor. A lymphocyte mitogen with helper activity.
Authors: Granelli-Piperno A , Vassalli J D , Reich E
Published: June 2004

Mouse T cell growth factor was purified from the serum-free conditioned medium of lectin-stimulated spleen cells. A 3,000-fold purification was achieved with a final yield of 12%. The purified protein, with an apparent Mr of 23,000 by sodium dodecyl sulfate polyacrylamide gel electrophoresis, was active at concentrations of 4 x 10(-11) M, both in the T cell growth factor and T cell replacing factor assays. In addition, purified T cell growth factor alone was mitogenic for spleen cells from both nude and normal mice.
Specificity of the helper T cell for the cytolytic T lymphocyte response to influenza viruses
Authors: Reiss CS , Burakoff SJ
Published: June 2004

We have described a model system in which helper T cells are required to mount a primary antiviral cytolytic T lymphocyte response. The radioresistant helper cell can be found in the spleens of mice that have been immunized subcutaneously with influenza viruses 6-8 d previously. These helper cells appear to be type specific but cross-reactive among the subtypes of influenza A viruses. The phenotypes of the interacting cell populations were determined.
Antibody-specific immunoregulation and the immunodeficiency of aging.
Authors: Klinman N R
Published: June 2004

Experiments were carried out to assess the role of naturally acquired antibody-specific immunoregulation in the immunodeficiency of aged individuals. It was found that greater than 50% of the primary dinitrophenyl-specific BALB/c B cells did not respond in carrier-primed 2-yr-old BALB/c adoptive hosts as compared with similarly primed younger recipients. Similar suppression was observed in carrier-primed younger BALB/c mice that had received 4 x 10(7) spleen cells from 2-yr-old BALB/c mice, as opposed to those that had received 4 x 10(7) spleen cells from younger mice. This diminution in responsiveness was noted only for syngeneic BALB/c B cells because B cells of strains differing from BALB/c in the heavy chain allotype-idiotype locus were not suppressed. These findings indicate that old, but not young, mice had developed the capacity to suppress primary B cells bearing receptors expressing much of the syngeneic antibody repertoire. This suppression may play an important causative role in the relatively poor humoral immune responsiveness of aged individuals.
Escape of metastasizing clonal tumor cell variants from tumor-specific cytolytic T lymphocytes.
Authors: Bosslet K , Schirrmacher V
Published: June 2004

A metastasizing variant of a chemically induced lymphoma from a DBA/2 mouse is shown to carry a distinct tumor-associated transplantation antigen (TATA), which can be recognized by syngeneic secondary anti-tumor cytolytic T lymphocytes (CTL). During metastasis of twice-cloned cell lines of this tumor, variants develop that are specifically immunoresistant to lysis by anti-tumor CTL. The variants are detected in the spleen of normal syngeneic mice. They remain stable over long-term passage in tissue culture. The high frequency with which these immunoresistant metastatic variants develop was found to explain the relative ineffectiveness of specific immunization against this metastatic tumor. Compared with organ-selective metastatic variants, the immunoresistant tumor variants seem to arise with a much higher frequency. The change in TATA expression described here differs from antibody-induced antigenic modulation in that it is more stable and genetically transmitted.
Origin and fate of IgE-bearing lymphocytes. I. Peyer's patches as differentiation site of cells. Simultaneously bearing IgA and IgE.
Authors: Durkin H G , Bazin H , Waksman B H
Published: June 2004

Peyer's patches (PP) from adult conventionally raised (C) and germ-free (GF) rats of the same age differed strikingly in the distributions of cells bearing membrane-bound immunoglobulin of the different isotypes. High concentrations of cells with membrane-bound IgE (approximately 20% of total cells determined by indirect immunofluorescence) were found in PP of rats, whereas IgE+ cells were absent from PP of C rats. The numbers of IgA+ cells were almost threefold higher in PP of GF rats when compared with C rats. In contrast, PP of GF rats had greatly reduced numbers of IgM-bearing cells (4%) when compared with C rats (23%); in some experiments virtually no IgM-bearing cells were detected. The levels of IgA- and IgE-bearing cells in spleen of GF rats were increased (to 11% and 7%, respectively). Of the IgE-bearing cells in PP and spleen of GF rats, approximately one-half were simultaneously positive for IgA. When these PP cells were treated with pronase to remove membrane bound immunoglobulins and maintained in culture, both IgE and IgA reappeared within 12 h. The proportion of doubly labeled cells was similar to that of the untreated population. No IgE+ cells were detected in bone marrow of C of GF rats at any time, in agreement with the findings of Ishizaka et al., although up to 20% of bone marrow cells bore other immunoglobulin isotypes, suggesting that IgE-bearing cells arise in the PP either de novo or by switching from precursors carrying IgM or IgA.
On the heterogeneity of murine natural killer cells.
Authors: Minato N , Reid L , Bloom B R
Published: June 2004

The heterogeneity of cells capable exerting spontaneous cytotoxicity in vitro was explored using antisera to several genetically determined surface markers on mouse lymphocytes. Four phenotypes of cells derived either from fresh or cultured murine lymphoid tissue were found to exert natural killer (NK) activity in vitro. One affector cell subset, termed NKI cells, had the serological phenotype of Thy-1-, Lyt-2-, Qa5+, and lysed measles virus persistently infected target cells (HeLa-Ms) but not P815 mastocytoma cells. It corresponds with the NK cells described in most systems in which lymphoma targets are commonly used. A second subset, with the same target cell specificity, termed NKT is a thymus-independent cell with the phenotype Thy-1+, Lyt-2-, Qa-5+, Ly-5+. A third subset of NK cells, termed T killer (TK) cells deriving from cultures of conventional but not nude mouse spleens, mediated spontaneous cytotoxicity of P815 mastocytoma cells, but not of virus-infected targets. It has a phenotype of Thy-1+, Lyt-2+, Qa-5-, Ly-5+, apparently identical with that of conventional, antigen-specific cytotoxic T lymphocytes. The fourth phenotype of NK cells, termed NKM, derived primarily from cultures of bone marrow, is cytotoxic for HeLa-measles but not P815, and expresses only Ly-5+ among the various markers tested. Beige mice possess normal TK and NKM activities, but had normal NKI, NKT as well as NKM activity. All NK cell subsets express the Ly-5 surface marker. The existence of four phenotypically distinct NK effector cells was strengthened by studies on selective regulation of their activity by two different biological factors. Interferon (IFN) augmented NK activity of primarily one of the subsets examined, the NKI cell; the activity of IFN on NKT cells could not be directly tested, but IFN was without positive effect on TK or NKM cells. In contrast, partially purified IFN-free interleuken 2 (IL-2) augmented the activities of both the TK and NKT subsets, but not of NKI or NKM cell. IL-2 was active in augmenting NK activity in spleen cells obtained from both conventional and nu/nu mice, but was without effect on spleens of nu/nu mice depleted of Thy-1+ cells. These and other data suggest that IL-2 acts primarily, if not exclusively, on THy-1+ cells. These results strengthen the view that natural cytotoxicity in vitro can be mediated by several distinct cell populations under different genetic and regulatory control and indicate the importance of defining and delineating the cell lineages of each and the role of the independent subsets in resistance to virus infections and tumors in vivo.
Formation of the initial C3 convertase of the alternative complement pathway. Acquisition of C3b-like activities by spontaneous hydrolysis of the putative thioester in native C3.
Authors: Pangburn M K , Schreiber R D , Müller-Eberhard H J
Published: June 2004

Activation of the alternative pathway of complement commences with the formation of an initial fluid-phase C3 convertase. Treatment of C3 with the nucleophilic reagent methylamine has previously been shown to result in the cleavage of an intramolecular thioester bond and to induce C3b-like properties, including the ability to form a fluid-phase C3 convertase. This report examines the hypothesis that spontaneous hydrolysis of the thioester generates a derivative of C3 that is responsible for the formation of the initial C3 convertase of the alternative pathway. The rate of spontaneous decay of C3 hemolytic activity in buffer was found to be between 0.2 and 0.4%/h. In the presence of other alternative pathway proteins, the rate of inactivation was 1%/h. The rate of spontaneous inactivation was greatly accelerated by low concentrations of chaotrophic agents such as KSCN or guanidine. Liberation of a sulfhydryl group, not present in native C3, correlated with loss of hemolytic activity, indicating that exposure to chaotropic agents resulted in thioester hydrolysis. Unlike native C3, C3 bearing a single reactive sulfhydryl group was capable of generating fluid-phase C3 convertase with Factors B, D, and P and was cleaved by Factor I (C3b inactivator) in the presence of Factor H (beta 1H). The fragmentation patterns indicated that the C3a domain was covalently associated with the functionally C3b-like C3. Organomercurial agarose was employed for the rapid removal of sulfhydryl-bearing, hemolytically inactive forms of C3 and C3b from native hemolytically active C3.
H-2-restricted helper factor secreted by clone hybridoma cells.
Authors: Lonai P , Puri J , Bitton S , Ben-Neriah Y , Givol D , Hämmerling G J
Published: June 2004

Biological and serological characteristics of a helper factor secreted by cloned hybridoma cells was described. The factor is carrier specific and contains determinants shared with immunoglobulin VH bu does not react with V kappa- or V lambda-specific antibodies. Presence of four H-2I-controlled antigenic specificities, Ia.ml, Ia.m2, Ia.17, and Ia.m7, was detected. Hence, it is possible that both A beta and E alpha loci may be involved in its control. Helper effect could be obtained only toward B cell sources that shared the H-2K and I-A antigens with the hybridoma cells. Similarly, the factor was absorbed only by spleen cells syngeneic in I-A. Previous studies have demonstrated that this clone binds antigen in an H-2-restricted manner. It follows that H-2-restricted helper cells produce H-2-restricted helper factors. Hence, they support the view that specific T cell factors may represent secreted T cell receptors.
Monoclonal antibody directed against interleukin 2. I. Inhibition of T lymphocyte mitogenesis and the in vitro differentiation of alloreactive cytolytic T cells.
Authors: Gillis S , Gillis A E , Henney C S
Published: June 2004

Our recent studies have detailed the generation of B cell hybridomas whose IgG product significantly inhibits interleukin 2 (IL-2)-dependent T cell replication. Given the capacity of such hybridoma antibody to interfere with the activity of mouse, rat, and human IL-2, we asked whether anti-IL-2 IgG would mediate similar inhibitory effects on other in vitro immune responses. In this communication, we report that addition of purified anti-IL-2 monoclonal antibody to either mitogen- or alloantigen-stimulated spleen cells exerted markedly deleterious effects on both resultant T cell proliferation and the generation of cytolytic effector cells. These results provide serological evidence in support of the integral role that IL-2 plays in controlling antigen/mitogen-induced T cell proliferation and serves further to define the ability of monoclonal antibody against IL-2 to function as an immunosuppressive agent.
Mast cell mediator release as a function of cyclic AMP-dependent protein kinase activation.
Authors: Winslow C M , Lewis R A , Austen K F
Published: June 2004

Stereo-specific perturbation of the IgE-receptor (shown in previous studies) produces a monophasic rise in cyclic AMP that peaks at 15 s and a depletion of cyclic AMP-dependent protein kinase that plateaus at 30-60 s. The previously observed linear relationship between the attenuation in the monophasic rise in cyclic AMP and the quantity of mediator release in the presence of incremental concentrations of the adenosine analogue 2',5',-dideoxyadenosine, DDA, which is known to inhibit adenylate cyclase, indicated a direct relationship between receptor perturbation, transmembrane activation of adenylate cyclase, and granule secretion. The role of cyclic AMP as a second messenger in this sequence is now apparent from the linear relationship between net percent mediator release and net percent activation of cyclic AMP-dependent protein kinase isoenzyme when IgE-dependent activation of adenylate cyclase is suppressed by incremental quantities of DDA. There was a comparable percent activation of both types I and II mast cell cyclic AMP-dependent protein kinase isoenzymes with anti-IgE-induced activation and secretion, and there was a parallel suppression of the activation of both isoenzymes in the presence of DDA. Although these studies firmly link the activation of cytoplasmic cyclic AMP-dependent protein kinase to the IgE receptor-initiated transmembrane activation of adenylate cyclase. they do not discriminate among the functions of the two isoenzymes.
Role of prostaglandin E in the biphasic fever response to endotoxin
Authors: Skarnes RC , Brown SK , Hull SS , McCracken JA
Published: June 2004

Biphasic fevers were induced in sheep with intravascular infusions or injections of 4-10 μg (80-200 ng/kg) of endotoxin, whereas monophasic fevers were obtained with doses of 1-2/μg (20-40 ng/kg). A marked increase in arterial blood pressure invariably accompanied the onset of fever; the latency of responses to the higher and lower doses of endotoxins averaged 26 min and 42 min, respectively. Prostaglandin (PG) assays of plasma from the carotid artery and jugular vein during fever episodes revealed a surge of PGE and PGF coincident with the pressor response and the first phase of fever, but PG were not detected in plasma samples taken throughout the second phase of fever. PG measurements of arterial and venous plasma collected at a distal site (hind limb) showed a similar surge of PGE and PGF in association with the early fever response, indicating that intravascular PG synthesis and release represents a generalized systemic response to circulating endotoxin. Carotid arterial infusions of PGE(2) produced immediate monophasic fevers and pressor responses, whereas PGD(2) infusions produced an immediate pressor effect but no fever. Infusions of PGF(2α) or prostacyclin, however, evoked neither fever nor pressor effects. Intracarotid infusions of leukocyte pyrogen (LP) caused monophasic fevers with latent periods of 15-20 min but pressor responses were not seen and neither PGE nor PGF were detected in plasma samples from the carotid artery or jugular vein before or during fever. Indomethacin, a potent inhibitor of arachidonic acid metabolism, blocked fever responses to endotoxin and to LP. These findings implicate PGE as the mediator of the early phase of endotoxin fever and imply a role for another pyrogenic metabolite ofarachidonic acid in the mediation of the second phase of fever, i.e., the phase associated with circulating LP. It is possible that both pyrogenic metabolites are generated within the vascular compartment, reaching thermoregulatory centers of the brain by transfer across the blood-brain interface.
Biosynthesis of Pb44, the protective antigen of sporozoites of Plasmodium berghei.
Authors: Yoshida N , Potocnjak P , Nussenzweig V , Nussenzweig R S
Published: June 2004

In a previous paper (2) we identified a protective antigen (Pb44) of the surface membrane of sporozoites of Plasmodium berghei by means of a monoclonal antibody. Immunoprecipitation of extracts of mature salivary gland sporozoites, metabolically labeled with L[35S]methionine using the same monoclonal antibody, revealed three specific polypeptides: *Pb44, *Pb52, and *Pb54. Metabolically labeled *Pb44 is probably identical to the protective antigen previously identified by surface labeling. Both proteins have the same molecular weights and isoelectric points under denaturing conditions, and they share an epitope. Moreover, *Pb44 also seems to be located on the cell membrane. The results of pulse-chase experiments strongly suggest that *Pb52 is the precursor of *Pb44. The relationship between *Pb54 and the protective antigen is unknown. The three polypeptides seem to be strictly associated with only one of the developmental stage of the parasite. They were not detected in blood forms and were found in minute amounts in sporozoites from the midgut of mosquitoes. In contrast, in mature salivary gland sporozoites they constitute main products of protein synthesis.
Functional characterization of synthetic leukotriene B and its stereochemical isomers.
Authors: Lewis R A , Goetzl E J , Drazen J M , Soter N A , Austen K F , Corey E J
Published: June 2004

Leukotriene B (LTB), a potent lipid chemotactic factor for neutrophils, is 5S,12R-dihydroxy-6,14-cis,8,10-trans-eicosatetraenoic acid (Fig 1), based upon direct comparison of natural LTB with synthetic 5S,12R-dihydroxy-6,8,10,14-eicosatetraenoic acid (5,12-di-HETE) stereoisomers in three biological assays. Of the six synthetic stereoisomers evaluated, only the 5S,12R,6,14-cis,8,10-trans compound had chemotactic potency for human neutrophils in vitro that was comparable to that of natural LTB, with a concentration of 3 X 10(9-9) M eliciting a one-half maximum response. In contrast, the racemic mixture of 5R,12R- and 5S,12S-6,10-trans,8,14-cis, the racemic mixture of 5S,12R- and 5R,12S-6,10-trans,8,14-cis, the 5S,12R-6,8-trans,10,14-cis, the 5S,12R-6,8,10-trans,14-cis, and the 5S,12S-6,8,10-trans,14-cis stereoisomers required concentrations of 3 X 10(-7) to 1 X 10(-6) M to elicit comparable responses. Only natural LTB and its synthetic counterpart elicited a local neutrophil infiltration when injected into the skin of the rhesus monkey at 10 ng and 100 ng per site. Natural and synthetic LTB at a concentration of 3 X 10(-8) M each provoked an EC25 contractile response of guinea pig pulmonary parenchymal strips in vitro, whereas the other four tested stereoisomers of 5,12-di-HETE were inactive at this concentration. Structure-function analyses suggest that the neutrophil chemotactic activity depends critically upon the C-1 to C-12 domain, including the stereochemistry of the 6-,8-,and 10-olefinic bonds and the presence of both hydroxyl groups.
T cell clones reactive with sperm whale myoglobin. Isolation of clones with specificity for individual determinants on myoglobin.
Authors: Infante A J , Atassi M Z , Fathman C G
Published: June 2004

We have been able to isolate clones of sperm whale muscle myoglobin (Mb)-reactive T cells from (C57BL/6 x A/J)F1 [(B6A)F1] mice. Four types of clones were isolated, distinguished by their patterns of recognition of Mb cyanogen bromide (CNBr) fragments and antigen presenting cell (APC) requirements. Individual T cell clones proliferated in response to one of three CNBr fragments of Mb. Dose-response curves of all clones were identical for native Mb and the appropriate fragment. T cell clones reactive to fragment 1-55 did not proliferate in response to peptide 15-22 (a peptide that binds to serum antibody directed against 1-55). These data support previous findings suggesting differences between antigen recognition by T and B cells, i.e., T cells may not recognize antigen in its native conformation and/or T and B cells may recognize distinct epitopes on the same antigen. Using T cell clones to analyze genetic control of responsiveness to Mb, we found that certain (B6A)F1 T cells recognize Mb presented by low responder strain APC. Thus, genetically determined low responsiveness in this case is probably not due to failure of APC function. We also found that responsiveness to certain Mb epitopes mapped to the I-A subregion whereas others mapped, via gene complementation, to the I-A and I-E subregions. We found no examples of responsiveness mapping to the I-C subregion and suggest an alternative explanation for previous reports mapping genetic control of responsiveness to certain Mb determinants to I-C.
Alteration of clonal profile. III. T15 ontogenetic advantages are not sufficient for establishing idiotypic dominance in adoptive transfer.
Authors: Quan Z S , Quintáns J
Published: June 2004

We have examined the ontogeny of BALB/c plaque-forming cell (PFC) responses to phosphorylcholine (PC) from fetal and neonatal liver by using the (CBA/N x BALB/c)F1 transplantation model. In this system, thymus-dependent (PC-keyhole limpet hemocyanin) and thymus-independent class 1 (PC-Brucella abortus, PC-lipopolysaccharide) PC antigens stimulate B cell subpopulations, which functionally emerge early after transfer. Responsiveness to a thymus-independent class 2 antigen, C-polysaccharide extract of a Streptococcus pneumoniae mutant, is acquired later. The response to PC antigens tested initially exhibited T15 dominance. Non-T15 clones, which are not expressed to a great degree in normal BALB/c mice, are inherently slow in their rate of maturation; in adoptive transfer, however, they eventually comprise much of the transplanted anti-PC PFC response. Obviously, the advantages the T15 subset has in ontogeny do not result in idiotypic dominance once the immature cells are removed from the intact BALB/c environment. We discuss possible regulatory mechanisms involved in the alteration of the T15+:T15- ratio.
Anti-phosphocholine hybridoma antibodies. II. Functional analysis of binding sites within three antibody families.
Authors: Andres C M , Maddalena A , Hudak S , Young N M , Claflin J L
Published: June 2004

The present investigation extends our immunochemical characterization of binding site heterogeneity among a large series of monoclonal anti-phosphocholine (PC) antibodies. Hybridoma proteins (HP) from eight genetically distinct strains are included in this study, yet no strain specific characteristics are observed. These HP, as previously shown (5), are divided into three well-defined families based on public or family-specific Id and L chain isotypes characteristic of three PC-binding myeloma proteins: T15, M603, and M511. All antibodies exhibited some degree of inter- or intra-family heterogeneity, or both. Some of this intra-family diversity was reflected by differential reactivity for PC when attached to three different carriers. In spite of this, the specificity profiles for hapten analogues of PC, as measured by hapten inhibition of binding, were the same for all members of the T15 family. Altering the carrier had no effect, thus suggesting that the binding site pocket for PC is essentially preserved, whereas that for carrier is variable. Similar conclusions were reached for most of the M603 HP, although the binding site is different from the T15 HP. The M511 HP stand in sharp contrast to the HP in the other two families because their binding sites exhibit extensive variability. The independence in reactivity for PC and PC plus carrier offers a rational explanation for idiotypic and/or structural heterogeneity within a family. More importantly it suggests interesting strategies for diversification within one group of antibodies.