bluuebunny/crossref_metadata_2025 huggingface dataset:
Purification of functional, determinant-specific, idiotype-bearing murine T cells
Authors: Lewis GK , Goodman JW
Published: June 2004

Strain A/J mice immunized with azobenzenearsonate (ABA)-mouse IgG conjugates develop suppression for anti-trinitrophenyl(TNP) responses to doubly conjugated (ABA,TNP) proteins. This suppression is specific for the ABA epitope and is mediated by T cells in cell transfer experiments. ABA-binding T cells from suppressed animals were purified by a two-stage procedure in which B cells were removed from spleen cell populations by adherence to plastic surfaces coated with anti-mouse Ig antibody, followed by binding the nonadherent population (more 95 percent Thy-1-positive) to surfaces coated with ABA-protein conjugates. Approximately 90 percent of the cells recovered by temperature-dependent elution from the ABA plates (similar to 2 percent of the spleen cells) bound antigen immediately afterward, and up to 50 percent of the cells bound anti-cross-reactive idiotype antibody. On the other hand, the nonadherent T-cell population was completely negative in the antigen- binding and idiotype assays. Another distinguishing feature of the two T-cell populations was that 78 percent of the adherent cells, but only 2 percent of the nonadherent cells, were Ia positive, although the specific I-region marker(s) expressed on the cells was not identified. The biological function of the antigen-binding T cells was investigated using a standard cell transfer protocol. Suppressor cells were enriched in the adherent population by a factor of at least 25, establishing that functional, epitope-specific, idiotype-bearing T cells can be significantly purified by this procedure.

Note Added in Proof. We have recently isolated two types of ABA-binding molecules biosynthetically labeled with (35)S-methionine from NP-40 lysates of purified antigen-specific T cells. The molecules were purified by adsorption onto an ABA-Sepharose immunoadsorbent followed by elution with 9 M urea. Autoradiograms of SDS-PAGE of the eluates revealed components with tool wt of approximately 60,000 and 33,000 dahons. These molecules were not present in eluates from a bovine IgG-Sepharose control immunoadsorbent and thus represent specific ABA-binding products synthesized by T cells.
In vitro sensitization of thymocytes. Role of H-21 I region determinants and cell-free mixed leukocyte culture supernates in generation of cytotoxic responses.
Authors: Sopori M , Bernstein A , Bach F H
Published: June 2004

Stimulation of thymocytes in vitro by spleen cells differing for the entire H-2 complex leads to a significant proliferative response without a significant cell-mediated lympholysis (CML) response. Addition of soluble cell-free supernates (SF), (taken from a 7-day mixed leukocyte culture) enables these cultures to develop CML response. For optimal CML response, the SF has to be added within 48 h of onset of cultures. Although with spleen cells as responding cells, SF could quantitatively replace I-region different stimulating cells for generation of CML responses, with thymocytes as responding cells, stimulation with I-region cells appeared obligatory for the generation of CML responses. The implications of these findings are discussed.
Immunological studies of aging. IV. The contribution of thymic involution to the immune deficiencies of aging mice and reversal with thymopoietin32-36.
Authors: Weksler M C , Innes J D , Goldstein G
Published: June 2004

Aged mice preferentially lose the capacity to make IgG and high affinity PFC after immunization with the T-dependent antigen DNP-BGG. We have found that thymectomy accelerates the appearances of these immune deficiencies associated with aging. When splenocytes from old mice are transferred to young lethally irradiated, syngeneic mice and the recipients immunized 7 wk later, the number of IgG and high affinity PFC was increased compared to the response of old splenocytes transferred to young thymectomized mice. These immune deficiencies of aged mice were also reversed when old mice were treated with thymopoietin in vivo or splenocytes from old mice were incubated with thymopoietin before adoptive transfer to young irradiated, thymectomized syngeneic mice. The T-cell independent response to DNP-Ficoll was less impaired than the T-cell dependent response to DNP-BGG in old animals. These data suggest that a decline in thymic function that occurs during aging may contribute to the immunological deficiencies of old animals.
On a regulatory gene controlling the expression of the murine lambda1 light chain.
Authors: Geckeler W , Faversham J , Cohn M
Published: June 2004

We describe here two alleles, an allele of the lambda1 locus present in the SJL strain (rlambda1lo) and an allele of the lambda1 locus present in the BALB/c strain (rlambda1 +), of a regulatory gene locus which specifically influences the expression of the mouse lambda1 light chain structural gene. The rlambda1 regulatory gene is not linked to either the major histocompatibility complex or to the heavy-chain allogroup but appears to be linked to the lambda1 structural gene locus. In the homozygous state, the present of the rlambda1lo allele results in a 50-fold reduction in the number of lambda1 antigen-sensitive, bone-marrow derived lymphocytes (ASCs) compared to the presence of the rlambda1 + allele. However, those few lambda1ASCs present in rlambda1lo homozygotes can be induced normally to produce lambda1 light chains indistinguishable from those found in rlambda1 + homozygotes. The reduction in lambda1ASC's due to the rlambda1lo allele results both in a reduction in the amount of lambda1 Ig in the serum and also in a large variation in the magnitude of the lambda1 antibody response to alpha(1,3) dextran by individual animals. This variation permits the estimate that, on the average, 50 B cells of anti-alpha(1,3) specificity must be present per animal to permit a measurable response. Surprisingly, the expression of a gene locus regulating lambda1 light chain expression (rlambda1 locus) shows a clear gene dosage effect with rlambda1lo/rlambda1 + heterozygotes having 1/2 the number of lambda1ASCs and 1/2 the amount of serum lambda1 Ig as rlambda1 +/rlambda1 + homozygotes. This fact permits an analysis of the relationship between germ-line v-genes and their individual expression in serum Ig. The rlambda1 locus controls specifically a DNA-level event which occurs in stem cells as they become committed to lambda1 light chain expression. We postulate that the rlambda1 locus represents one of the DNA level recognition sites involved in the translocation event which places the vlambda1 and clambda1 structural genes in a transcriptional unit.
Structural and functional differences between the H-2 controlled Ss and Slp proteins.
Authors: Ferreira A , Nussenzweig V , Gigli I
Published: June 2004

Based on functional and structural data, it is concluded that the Ss protein in the mouse expresses the activity of the fourth component of complement. Removal of the Ss, but not of Slp, antigen correlates with a high degree of significance (P less than 0.001) with decrease of C4 hemolytic activity. In phenotypically Slp negative mice the plasma/serum levels of Ss correlate with the C4 activity (P less than 0.001). Structurally, Ss is a 209,000-mol wt protein, consisting of three covalently linked polypeptide chains (alpha,beta,gamma). Treatment of Ss with C1 cleaves a 7,000-8,000-mol wt fragment from the alpha-chain. Slp is also a three chain covalently linked protein of 209,000 daltons, however its three chains differ in size from those of the Ss protein. Slp does not express hemolytic activity and its alpha-chain is not cleaved by C1.
Nonspecific resistance to bacterial infections. Enhancement by ubiquinone-8.
Authors: Block L H , Georgopoulos A , Mayer P , Drews J
Published: June 2004

A lipid fraction from Escherichia coli was extracted with apolar solvents and was found to protect mice from a number of experimental bacterial infections. The benzoquinone, ubiquinone-8, was isolated from this extract by high pressure liquid chromatography and identified as such by nuclear magnetic resonance and mass spectrometry. At a dose of 25 mg/kg this substance was found to provide complete protection against otherwise lethal infections with gram-negative and gram-positive bacteria in mice. Treatment was most effective when given intravenously 24 h before infection. In comparative studies, ubiquinone-8 had a clearly higher activity than ubiquinones-4, Q6, and Q10. A highly significant increase in the clearance rate of bacteria from the blood by the spleen and the liver of treated animals, correlated well with the protective effect of ubiquinone-8. The compound stimulated the ability of mouse macrophages to incorporate sheep erythrocytes and significantly increased the number of antibody-producing cells in spleens of mice.
Maturation of bone marrow lymphocytes. II. Development of Fc and complement receptors and surface immunoglobulin studied by rosetting and radioautography.
Authors: Yang W C , Miller S C , Osmond D G
Published: June 2004

Radioautographic DNA labeling and rosetting techniques were combined to study the development of surface IgM, Fc, and complement receptors (FcR, CR) on small lymphocyte populations in mouse bone marrow. [3H]thymidine was either infused continuously to label newly formed cells for periods up to 4 days, or injected daily, 21--35 days before use, to label a sample of long-lived cells. Bone marrow cells were incubated with sensitized sheep erythrocytes to detect surface IgM, FcR, and CR, respectively, and examined radioautographically after cytocentrifugation. During [3H]thymidine infusion, marrow small lymphocytes lacking surface markers were the first to show [3H]thymidine labeling. Most of these cells became labeled by 4 days (IgM--ve, 89%; FcR--ve, 92%; Cr--ve, 88%). Labeling of small lymphocytes bearing surface IgM, FcR, and Cr began after an initial lag and increased to high values by 4 days (IgM + ve, 73%; FcR + ve, 82%; CR + ve, 83%). Labeled IgM + ve small lymphocytes formed progressively larger rosettes as cell age increased. Some proliferating large lymphoid cells formed rosettes for IgM, FcR, and CR. Labeled long-lived small lymphocytes expressed surface IgM, FcR, and CR, the incidence of each receptor being uniformly high (38--43%) and the rosettes tending to be larger than those formed by newly formed lymphocytes. In double-surface marker studies, FcR and CR rosettes were formed by some IgM--ve small lymphocytes as well as IgM + ve cells in the marrow. After transfusion of marrow cells from donor mice infused with [3H]thymidine for 24 h, many labeled newly formed lymphocytes homed into the splenic red pulp of unlabeled syngeneic recipients. Subsequently, these cells showed a rapid increase in the incidence of rosettes for surface IgM, FcR, and CR, together with a progressive enlargement of each type of rosette. Although all the labeled small lymphocytes recovered from the spleen developed both surface IgM and FcR by 3 days, only approximately one-half developed CR. The results demonstrate that most of the small lymphocytes bearing FcR, CR, and surface IgM in mouse bone marrow are newly formed indigenous cells. Each receptor becomes detectable by rosetting soon after the small lymphocytes are first produced. The newly formed, marrow-derived small lymphocytes are able to continue their development of surface IgM, FcR, and CR after migrating into the spleen, consistent with a maturation of primary B lymphocytes. In addition, the data indicate the genesis in mouse marrow of a non-B lineage of lymphocytes (notably, IgM--ve FcR + ve cells.). A minority of small lymphocytes bearing IgM, FcR, and CR in mouse marrow are long-lived cells, presumptive recirculating immigrants, differing in receptor status from the newly formed cells. The results are discussed with regard to the heterogeneity of marrow lymphocytes and proposed models of primary B lymphocyte and null lymphocyte production.
Malaria immunization in Rhesus monkeys. A vaccine effective against both the sexual and asexual stages of Plasmodium knowlesi.
Authors: Gwadz R W , Green I
Published: June 2004

Rhesus monkeys were immunized with a preparation of Plasmodium knowlesi parasites containing principally microgametes with lesser numbers of macrogametes and asexual trophozoites. The antigen mixture was emulsified in Freund's complete adjuvant (FCA) and administered intramuscularly. After one or two inoculations of from 10(5) to 10(7) microgametes in FCA, monkeys showed high levels of circulating anti-gamete antibodies as demonstrated by various in vitro microgamete immobilization or transmission blocking tests. After challenge with P. knowlesi, immunized monkeys developed low level asexual parasitemias and were not infectious to feeding mosquitoes as measured by growth of the parasite on the mosquito gut. Control monkeys developed rapidly rising, usually fatal infections and were highly infectious to mosquitoes. Anti-gamete antibodies appear to neutralize the sexual parasites and prevent mosquito infection within the gut of the recently fed mosquito vector. Suppression of asexual parasitemia in immunized monkeys may be due to the presence of asexual trophozoites in the antigen mixture or to antigens common to both sexual and asexual stages of the parasite. A vaccine effective as a single injection capable of interrupting malaria transmission from man to man whereas reducing the severity of the disease in infected individuals offers a new approach to the control of one of the major diseases affecting man.
The involvement of suppressor T cells in Ir gene regulation of secondary antibody responses of primed (responder X nonresponder)F1 mice to macrophage-bound L-glutamic acid60-L-alanine30-L-tyrosine.
Authors: Germain R N , Benacerraf B
Published: June 2004

(Responder [R] X nonresponder [NR])F1 mice give indistinguishable primary in vitro plaque-forming cell (PFC) responses to either R or NR parental macrophages (Mphi) pulsed with the Ir-gene controlled antigen L-glutamic acid60-L-alanine30-L-tyrosine10 (GAT). However, such (R X NR)F1 mice, if primed to GAT, retained in vitro responsiveness to GAT-R-Mphi, but no longer responded to GAT-NR-Mphi. This suggested (a) a possible Mphi-related locus for Ir gene activity in this model, and (b) the occurrence of active suppression after priming with GAT leading to a selective loss of the usual primary responsiveness of (R X NR)F1 mice to GAT-NR-Mphi. This latter interpretation was tested in the current study. [Responder C57BL/6 (H-2b) X nonresponder DBA/1 (H-2q)]F1 mice were primed with 100 microgram GAT in pertussis adjuvant. 4-8 wk later, spleen cells from such mice were tested alone or mixed with normal unprimed F1 spleen cells for PFC responses to GAT-R-Mphi and GAT-NR-Mphi. The primed cells failed to respond to GAT-NR-Mphi, and moreover, actively suppressed the normal response of unprimed F1 cells to GAT-NR-Mphi. If the primed spleen cell donor had been treated with 5 mg/kg cyclophosphamide 3 days before priming or with 5-10 microliter/day of an antiserum to the I-Jb subregion [B10.A(5R) anti B10.A(3R)] during the first 4 days postpriming (both procedures known to inhibit suppressor T-cell activity), cells from such mice responded in secondary culture to both GAT-R-Mphi and also GAT-NR-MPhi. In addition, such spleen cells no longer were capable of suppressing normal F1 cells in response to GAT-NR-Mphi. Similar data were obtained using [CBA (H-2k) X DBA/1 (H-2q)]F1. Further, it was shown that (a) primary responsiveness to GAT-NR-Mphi was not an artifact of in vitro Mphi pulsing, because in vivo GAT-pulsed Mphi showed the same activity and (b) the secondary restriction for Mphi-antigen presentation was controlled by H-2 linked genes. These data suggest an important role for suppressor T cells in H-2 restricted secondary PFC responses, and also provide additional support for the hypothesis that Ir-gene controlled differences in Mphi antigen presentation are related to both suppressor cell generation and overall responsiveness in the GAT model.
Secondary induction of cytotoxic T lymphocytes with solubilized syngeneic tumor cell plasma membranes.
Authors: Alaba O , Law L W
Published: June 2004

Secondary induction of in vitro cytotoxic T lymphocytes in a syngeneic system has been achieved with plasma membrane, both in the particulate and solubilized forms. Both the induction and the lytic phases were shown to be immunologically specific. The effector cells generated were completely susceptible to treatment with anti-theta antibody and complement, suggesting that they are T lymphocytes.
Idiotypic cross-reactivity between antibodies of different specificities.
Authors: Karol R , Reichlin M , Noble R W
Published: June 2004

Cross-idiotypic specificity has been demonstrated between antibody populations of different specificities using antibodies directed toward human sickle cell hemoglobin (HbS). A site-specific antibody directed toward the beta6-position of HbS, anti-Val, was used to elicit an anti-idiotypic response in rabbits. Using this anti-idiotypic serum, idiotypic cross-reactivity was demonstrated between antibody populations that bind to human adult hemoglobin (HbA). It was demonstrated that in the case of the goat antibodies, these idiotypically cross-reacting antibodies are directed towards the beta6-position of the hemoglobin molecule. However, they differ in their specificity, binding to this site on HbA, whereas anti-Val binds only to HbS. The sheep antibody populations directed toward HbS differ qualitatively from those of the goat. Using rabbit anti-idiotypic serum specific for sheep anti-Val, cross-reactivity could be demonstrated with antibodies directed toward the alpha-chain of the hemoglobin molecule, as well as the beta-chain. There was also a low level of cross-reactivity with antibodies from a goat immunized with HbA.
T-T-cell interactions during the vitro cytotoxic allograft responses. I. Soluble products from activated Lyl+ T cells trigger autonomously antigen-primed Ly23+ T cells to cell proliferation and cytolytic activity.
Authors: Wagner H , Röllinghoff M
Published: June 2004

Secondary murine cytotoxic T lymphocyte responses from alloantigen-primed T cells can be induced in vitro by apparently unrelated regimens, such as addition of either concanavalin A (Con A), conditioned medium from Con A stimulated lymphocyte cultures, conditioned medium from secondary mixed lymphocyte cultures (MLC), or stimulator cells sharing only the I-region with the stimulating cells used for primary sensitization. We now report that upon polyclonal (Con A), or antigen-specific (MLC) stimulation, Lyl+ T cells release a factor, which in turn triggers alloantigen primed Ly23+ T cells to proliferation and cytolytic activity. The secondary cytotoxic T lymphocyte inducing factor (SCIF) is produced within 24 h. For its production, an intact protein metabolism, not DNA metabolism, is required. Once induced, the functional activity of SCIF is nonspecific and not H-2 restricted. SCIF allows exponential growth and long-term propagation of cytolytic Ly23+ T cells with specificity to alloantigens used for primary sensitization. SCIF induced activation of alloantigen primed Ly23+ T cells does not require the presence of alloantigens. The results therefore reveal a process by which Lyl+ T-cell-derived nonspecific factor(s) induce autonomously Ly23+ T-cell-mediated, antigen-specific, cytotoxic T lymphocyte responses.
B-cell precursors specific to sheep erythrocytes. Estimation of frequency in a specific helper assay.
Authors: Schreier M H
Published: June 2004

A sensitive, specific, and reproducible in vitro helper assay is described which is suited to limiting dilution analysis of murine B cells. 1 in about 3,000 syngeneic splenic B cells can be induced to form plaque-forming cells (PFC) to sheep erythrocytes in this system. The induction of PFC is absolutely dependent on antigen and specific helper T cells.
The induction of virus-specific cytotoxic T lymphocytes with solubilized viral and membrane proteins.
Authors: Finberg R , Mescher M , Burakoff S J
Published: June 2004

Reconstituted membranes were prepared from detergent solubilized P815 (H-2d) tumor cell membranes and solubilized Sendai virus protein. These reconstituted membranes stimulated a virus-specific H-2-restricted secondary CTL response. Stimulating activity was dependent upon the presence of both viral and P815 protein in the same membrane and was restricted to the H-2 specificity present in the reconstituted membrane. Liposomes prepared from solubilized Sendai virus proteins and partially purified H-2 alloantigen also had activity for CTL induction. The results demonstrate the feasibility of using detergent solubilized membrane proteins to study antigen recognition by virus-specific, H-2 restricted cytolytic T lymphocytes.
Activation of mouse lymphocytes by anti-immunoglobulin. II. A thymus-independent response by a mature subset of B lymphocytes.
Authors: Sieckmann D G , Scher I , Asofsky R , Mosier D E , Paul W E
Published: June 2004

Mouse spleen cells can be stimulated to proliferate in vitro by purified anti-mu or anti-gamma,kappa antibodies. These responses can be obtained in cell populations bearing membrane immunoglobulin (Ig), purified by the fluorescence activated cell sorter (FACS), but they are not observed in FACS-purified Ig- cell populations. Furthermore, treatment of spleen cell populations with anti-Thy 1.2 and complement does not impair the response, nor does addition of nylon wool-purified T lymphocytes enhance it. These results indicate that B lymphocytes respond to anti-Ig and that their response does not require T cells. On the other hand, cells from athymic nude (nu/nu) mice respond slightly less well to anti-mu than do cells from heterozygous littermate (nu/+) controls; nu/nu cells are almost unresponsive to anti-gamm,kappa and addition of nylon wool-purified T cells from nu/+ controls does not restore the response. This suggests that T lymphocytes or the thymus may control the appearance of cells responsive to anti-gamma,kappa. Responsiveness of normal mice to anti-mu does not appear until 4 wk of age and does not reach maximum levels until 8 wk of age. Acquisition of full responsiveness to anti-gamma,kappa is even more delayed. This, together with the failure of mice with the CBA/N B-cell defect to respond to anti-Ig, suggests that cells stimulated to proliferate by anti-Ig are a mature subset of B cells. Depletion of adherent cells by Sephadex G-10 treatment or by treatment with carbonyl iron and exposure to a magnetic field does not diminish anti-mu or anti-gamma,kappa responses, suggesting that the responsiveness does not require the presence of macrophages. Thus, activation of B-cell proliferation by anti-Ig appears to be a T-cell independent, macrophage-independent process in which membrane Ig plays a direct role in signal generation.
Involvement of H-2L gene products in virus-immune T-cell recognition. Evidence for an H-2L-restricted T-cell response.
Authors: Biddison W E , Hansen T H , Levy R B , Doherty P C
Published: June 2004

The H-2L locus is closely linked to H-2D and codes for antigenic specificities present on a 45,000 mol wt glycoprotein that is distinct from the molecule which bears the D region private specificity. It was found that BALB/c-H-2db mice, which lack detectable cell-surface H-2L gene products, were able to generate influenza- and vaccinia-immune cytotoxic T cells which lyse D region-compatible target cells, although they have been reported to be incapable of making a similar response to ectromelia virus (7). Thus, the lack of H-2L antigenic specificities does not produce a general loss of responsiveness for other viruses even when a highly cross-reactive pox virus (vaccinia) was studied. Antisera-blocking experiments utilizing sera specific for either L or D molecules indicated that BALB/c mice generate influenza virus-immune cytotoxic T-cell subsets which independently recognize H-2L and H-2D gene products in association with viral antigens. These results are the first indication that products of the H-2L locus can operate analogously to H-2K/D gene products in virus-immune T-cell recognition.
Assembly of the cytolytic alternative pathway of complement from 11 isolated plasma proteins.
Authors: Schreiber R D , Müller-Eberhard H J
Published: June 2004

The known cytolytic function of the alternative pathway in serum was quantitatively reproduced by combining 11 isolated plasma proteins at their respective serum concentrations. These proteins are: C3, Factor B, Factor D, C3b inactivator, beta1H, native properdin, C5, C6, C7, C8, and C9. In absence of activators of the alternative pathway, this mixture was stable at 37 degrees C as evidenced by lack of consumption of Factor B, C3, and C5. Upon addition of either rabbit erythrocytes or neuraminidase-treated sheep erythrocytes, cell lysis ensued and the extent of lysis was dependent on dose of the component mixture. The dose-response curves obtained with the isolated component mixture and with C4-depleted serum were virtually indistinguishable. Nonactivator erythrocytes (untreated sheep erythrocytes) were not lysed by the component mixture. Deletion of properdin resulted only in a twofold diminution of the hemolytic activity of the component mixture. No immunoglobulin requirement was apparent. These results indicate that the cytolytic systems studied are internally sufficient and capable of coupling the initiation and amplification sequence with the cytolytic membrane attack sequence.
Extracellular cytolysis by activated macrophages and granulocytes. II. Hydrogen peroxide as a mediator of cytotoxicity.
Authors: Nathan C F , Silverstein S C , Brukner L H , Cohn Z A
Published: June 2004

When deprived of oxygen, Bacille Calmette-Guérin (BCG)-activated macrophages no longer lysed P388 lymphoma cells. Both H2O2 release and cytotoxicity by BCG-activated macrophages and by granulocytes triggered with phorbol myristate acetate (PMA) were markedly inhibited when the glucose concentration in the medium was reduced to 0.03 mM or less, or if glucose were replaced with galactose. Catalase abolished PMA-triggered cytotoxicity by both types of effector cells, whereas superoxide dismutase had no effect. Ferricytochrome C reduced the cytotoxicity of BCG-activated macrophages, an effect which was largely reversed by superoxide dismutase. 10 drugs, thought to quench singlet oxygen and/or scavenge hydroxyl radical, did not affect cytotoxicity in this system. Neither azide nor cyanide reduced cytolysis, but there was marked inhibition by lactoperoxidase and iodide. This suggested that cytotoxicity was not dependent upon myeloperoxidase, and that lactoperoxidase may have diverted H2O2 from the oxidation of target cells to oxidation of substances in serum. Mouse erythrocytes, although sensitive targets, interfered with the cytolysis of lymphoma cells, probably by competition for H2O2. Starch particles with covalently bound glucose oxidase resembled macrophages in their spatial relation to the target cells and in the flux of H2O2 they generated from their surface, but were not expected to produce any other potentially toxic products. Such particles lysed lymphoma cells, and the lysis was prevented by catalase. Neither arginase nor thymidine appeared to be involved in cytolysis by BCG-activated macrophages under the conditions used. These findings demonstrated that release of H2O2 was both necessary and sufficient for cytolysis by BCG-activated macrophages and by granulocytes when pharmacologically triggered.
Vaccinia-specific cytotoxic T-cell responses in the context of H-2 antigens not encountered in thymus may reflect aberrant recognition of a virus-H-2 complex.
Authors: Doherty P C , Bennink J C
Published: June 2004

BALB/c (H-2Kd-Dd) spleen and lymph node populations were specifically depleted of alloreactive potential by filtration through H-2 different, irradiated recipients. These negatively selected T cells were then stimulated with vaccinia virus in mice expressing the foreign H-2 determinants encountered previously in the filter environment. Strong virus-immune cytotoxic T-cell responses were seen in the context of H-2Kk and H-2Ks, but not 2H-2Kb. The T cells generated were not cross-reactive for the H-2Kk and H-2Kd alleles, and responsiveness was independent of concurrent presence of effector populations operating at H-2D. These findings are consisent with the idea that recognition is mediated via a complex receptor, part of which is specific for virus and part for self H-2. The capacity to interact with allogeneic, virus-infected cells may then reflect aberrant recognition of a virus-H-2-antigen complex by this single, large binding site. For instance, the T cell which would normally recognize H-2Kd-virus x, or H-2Dd-minor histocompatibility antigen Z, may now show specificity for H-2Kk-vaccinia virus. Implications for both the selective role of the thymus and for mechanisms of tolerance are discussed.
Ultrastructure of mononuclear phagocytes developing in liquid bone marrow cultures. A study on peroxidatic activity
Authors: Van Der Meer JWM , Beelen RHJ , Fluitsma DM , Van Furth R
Published: June 2004

Monoblasts, promonocytes, and macrophages in in vitro cultures of murine bone marrow were studied ultrastructurally, with special attention to peroxidatic activity. Monoblasts show peroxidatic activity in the rough endoplasmic reticulum and nuclear envelope as well as in the granules. The presence of peroxidatic activity in the Golgi apparatus could not be determined. Promonocytes have peroxidase-positive rough endoplasmic reticulum, Golgi apparatus, nuclear envelope, and granules, as previously reported.

During culture, cells are formed with peroxidatic activity similar to that of monocytes or exudate macrophages (positive granules; negative Golgi apparatus, RER, and nuclear envelope); we call these cells early macrophages. In addition, transitional macrophages with both positive granules and positive RER, nuclear envelope, negative Golgi apparatus (as in exudate- resident macrophages in vivo), and mature macrophages with peroxidatic activity only in the RER and nuclear envelope (as in resident macrophages in vivo) were found. A considerable number of cells without detectable peroxidatic activity were also encountered.

Our finding that macrophages with the peroxidatic pattern of monocytes (early macrophages), exudate-resident macrophages (transitional macrophages), and resident macrophages (mature macrophages), develop in vitro from proliferating precursor cells deriving from the bone marrow, demonstrates once again that resident macrophages in tissues originate from precursor cells in the bone marrow. Therefore, this conclusion can no longer be challenged on the basis of a cytochemical difference between monocytes and exudate macrophages on the one hand and resident macrophages on the other.
Disproportion in T-cell subpopulations in immunodeficient mutant hr/hr mice.
Authors: Reske-Kunz A B , Scheid M P , Boyse E A
Published: June 2004

Mice of the HRS strain, which carry the mutant gene hr, were examined for abnormalities in representation of the three T-cell sets Ly1, Ly23, and Ly123 in the spleen. The salient feature of hr/hr mice, which are immunologically deficient, in comparison with +/hr segregants, was a gross disproportion in numbers of cells belonging to the Ly1 and Ly123 sets, at the age of 3--3.5 mo. At this age, Ly123 cells of hr/hr spleen outnumbered Ly1 cells by 2:1, whereas in +/hr spleens Ly123 cells were outnumbered by approximately 1:2. Cells from pooled lymph nodes of hr/hr mice did not show a correspondingly gross disporprotion of Ly1 and Ly123 cells. Total counts of splenic T cells, and of B cells, were not significantly different in hr/hr and +/hr mice.
Bactericidal activity of a superoxide anion-generating system. A model for the polymorphonuclear leukocyte.
Authors: Rosen H , Klebanoff S J
Published: June 2004

The acetaldehyde-xanthine oxidase system in the presence and absence of myeloperoxidase (MPO) and chloride has been employed as a model of the oxygen-dependent antimicrobial systems of the PMN. The unsupplemented xanthine oxidase system was bactericidal at relatively high acetaldehyde concentrations. The bactericidal activity was inhibited by superoxide dismutase (SOD), catalase, the hydroxyl radical (OH.) scavengers, mannitol and benzoate, the singlet oxygen (1O2) quenchers, azide, histidine, and 1,4-diazabicyclo[2,2,2]octane (DABCO) and by the purines, xanthine, hypoxanthine, and uric acid. The latter effect may account for the relatively weak bactericidal activity of the xanthine oxidase system when purines are employed as substrate. A white, carotenoid-negative mutant strain of Sarcina lutea was more susceptible to the acetaldehyde-xanthine oxidase system than was the yellow, carotenoid-positive parent strain. Carotenoid pigments are potent 1O2 quenchers. The xanthine oxidase system catalyzes the conversion of 2,5-diphenylfuran to cis-dibenzoylethylene, a reaction which can occur by a 1O2 mechanism. This conversion is inhibited by SOD, catalase, azide, histidine, DABCO, xanthine, hypoxanthine, and uric acid but is only slightly inhibited by mannitol and benzoate. The addition of MPO and chloride to the acetaldehyde-xanthine oxidase system greatly increases bactericidal activity; the minimal effective acetaldehyde concentration is decreased 100-fold and the rate and extent of bacterial killing is increased. The bactericidal activity of the MPO-supplemented system is inhibited by catalase, benzoate, azide, DABCO, and histidine but not by SOD or mannitol. Thus, the acetaldehyde-xanthine oxidase system which like phagocytosing PMNs generates superoxide (O.2-) and hydrogen peroxide, is bactericidal both in the presence and absence of MPO and chloride. The MPO-supplemented system is considerably more potent; however, when MPO is absent, bactericidal activity is observed which may be mediated by the interaction of H2O2 and O.2- to form OH. and 1O2.
RNA and protein synthesis in human peripheral blood polymorphonuclear leukocytes.
Authors: Granelli-Piperno A , Vassalli J D , Reich E
Published: June 2004

Polymorphonuclear leukocytes purified from human peripheral blood synthesized RNA and proteins when placed in cell culture. Autoradiography of the cultured cells revealed that a majority of mature PMNs were engaged in macromolecule synthesis, and an analysis of newly synthesized proteins by SDS-polyacrylamide gel electrophoresis showed that many different polypeptide chains were synthesized by these cells. The rate of [3H]uridine incorporation and the pattern of newly synthesized proteins were modulated by Con A and glucocorticoids. These results suggest that in spite of their short lifetime and a large performed enzymatic apparatus, mature PMNs retain a substantial capacity for RNA and protein synthesis; and, further, that modulation of macromolecule synthesis forms part of the mechanism by which PMNs respond to inflammatory and anti-flammatory stimuli.
Differential cyclophosphamide sensitivity of precursor cells in allogeneic and H-2 restricted cytotoxic responses.
Authors: Hurme M
Published: June 2004

Spleen cells from cyclophosphamide-treated mice responded in vitro to allogeneic stimulator cells but not to TNP-coupled syngeneic spleen cells. Similarly, cells from female mice, primed in vivo with syngeneic male cells, could not respond in vitro to male spleen cell stimulation if the mice were pretreated with cyclophosphamide. These results suggest that the precursor cells for H-2-restricted cytotoxic responses belong to a different T-cell subpopulation than the precursor cells for allogeneic responses.
Isolation of type-specific polysaccharide antigen from group B type Ib streptococci.
Authors: Tai J Y , Gotschlich E C , Lancefield R C
Published: June 2004

Group B streptococcus type Ib (strain H36B) was subjected to digestion with extracellular muralytic enzymes prepared from Streptomyces albus. Type Ib-specific polysaccharide antigen was isolated from the lysate by alcohol precipitation and Sepharose 6B chromatography. The purified type Ib antigen has a Kd value of 0.31 on a Sepharose 4B column and contains four sugars, galactose, glucose, N-acetyl glucosamine, and sialic acid in a molar ratio of 2.05:0.86:1.00:0.90. Acid treatment (pH 2.0) of this polysaccharide results in partial degradation of the antigen (Kd = 0.41 on Sepharose 4B) with the loss of 93% of the sialic acid. The molar ratio of the remaining sugars in the polysaccharide remains identical to that in the native one. This suggests that the sialic acid is at the terminal position in the molecule. Both intact and acid-treated antigen cross-react with some type Ia and type Ic antisera as a result of the common Iabc determinant, but not with type II and type III antisera. Absorption studies indicate that Ib-specific determinant and Iabc determinant are on the same molecule and that sialic acid is not the cross-reactive determinant.
Role of a nonimmunoglobulin cell surface determinant in the activation of B lymphocytes by thymus-independent antigens.
Authors: Subbarao B , Mosier D E , Ahmed A , Mond J J , Scher I , Paul W E
Published: June 2004

Lyb 5 is a B-cell alloantigen which is expressed on 50-60% of B cells. It was defined originally on the basis of cytotoxicity. We have described a new reactivity within the anti-Lyb 5 serum on the basis of selective inhibition of antibody responses in vitro by this antiserum in the absence of complement. This inhibitory activity of anti-Lyb 5.1 serum appears to be due to recognition of antigenic determinants different from the prototype antigens detected in the cytotoxicity assay. Anti-Lyb 5 serum incorporated into spleen cell cultures selectively inhibits antibody responses to a class of thymus-independent antigens (TI-2) previously characterized by their failure to elicit antibody formation in immature mice or in the defective CBA/N strain. Responses to optimal concentrations of TI-1 antigens, which can induce antibody synthesis in these mice, are unaffected by the addition of anti-Lyb 5.1 serum. The B-cell alloantigen defined by this functional assay is designated tentatively Lyb 7 and it is shown to be distinct from cell surface immunoglobulins. Lyb 7 appears to have a role in the activation of B lymphocytes by the TI-2 class of thymus-independent antigens.
Form variation in Escherichia coli K1: determined by O-acetylation of the capsular polysaccharide.
Authors: Orskov F , Orskov I , Sutton A , Schneerson R , Lin W , Egan W , Hoff G E , Robbins J B
Published: June 2004

The chemical basis for the alternating antigenic change called form variation noted for the Escherichia coli K1-capsular polysaccharide has been shown by 13C nuclear magnetic resonance to be a result of random O-acetylation of C7 and C9 carbons of the alpha-2-8-linked sialic acid homopolymer. A serologic method (antiserum agar) was developed to identify and isolate the form variants. The O-acetyl positive and O-acetyl negative K1 polysaccharides had unique biochemical and immunologic properties. The O-acetyl-positive variants resisted neuraminidase hydrolysis in contrast to the susceptibility of the O-acetyl negative variant to this enzyme. In addition, O-acetylation altered the antigenicity of the O-acetyl polysaccharides. When injected as whole organisms, O-acetyl positive organisms produced anti-K1 -antibodies in rabbits specific for this polysaccharide variant. O-acetyl negative organisms were comparatively less immunogenic; however, antibodies induced by these organisms reacted with both K1 polysaccharide variants. Burros, injected with either variant, produced antibodies reactive with both K1 polysaccharides.
Cell-surface antigens associated with recombinant mink cell focus-inducing murine leukemia viruses.
Authors: Cloyd M W , Hartley J W , Rowe W P
Published: June 2004

Distinct type-specific antigens were detected on cells infected with cloned mink cell focus-inducing (MCF) murine leukemia viruses by means of cell surface immunofluorescence absorption assays with rabbit antisera raised against naturally-occurring AKR MCF viruses. The MCF type-specific antibodies were present in high titer and not absorbable by cells infected with ecotropic, xenotropic, or wild mouse amphotropic murine leukemia viruses, or combinations of ecotropic and xenotropic viruses. Three MCF subtype-specific reactions were identified. One subspecificity (operationally designated MCFA-1) defined antigenic determinant(s) distributed among MCF viruses in general. Another (MCFA-2) specified determinant(s) induced by all naturally occurring MCF isolates not of Friend or Moloney origin. A third subspecificity (MCFA-3) was induced by some MCF isolates, and not by others; the presence of this antigen did not correlate with the source of any presently known biological property of the viruses. In addition, type-specific antigenic determinants of ecotropic and xenotropic murine leukemia viruses were expressed on MCF virus-infected cells. The serological profile of MCF viruses thus supports the contention that they are env gene recombinants between ecotropic and xenotropic murine leukemia viruses. However, new, distinct MCF-specific determinants are also generated, and these could be useful markers in studying MCF viruses.
The tumor dormant state. Quantitation of L5178Y cells and host immune responses during the establishment and course of dormancy in syngeneic DBA/2 mice.
Authors: Weinhold K J , Goldstein L T , Wheelock E F
Published: June 2004

Subcutaneous implantation of DBA/2-derived L5178Y cells into DBA/2 mice followed 10 d later by nodule excision protected 100% of mice from the rapid outgrowth of an intraperitoneal challenge of L5178Y cells given 7 d postexcision. Challenged mice remained clinically normal for 48--250 d before onset of an ultimately fatal tumor outgrowth. The numbers of L5178Y cells in the peritoneal cavity increased logarithmically for 4 d after challenge and then declined to low but detectable levels which persisted throughout the clinically normal period. Cells active in 18-h in vitro cytolytic assays against 51Cr-labeled L5178Y target cells were found in the peritoneal cavity. The effector cells were determined to be Thy1.2 positive. Their activity was tumor specific and reached peak levels 4 d after tumor challenge and then gradually declined to undectable levels during the following 70 d. Tumor emergence occurred most frequently during the period when CMC activity was no longer demonstrable in the remaining clinically normal mice. A transient peak of low level cytophilic antitumor antibody was detected about 30 d after tumor cell challenge. The temporal associations between the numbers of tumor cells and the levels of cell-mediated lysis against L5178Y cells indicate the importance of the cell-mediated cytolysis response in limiting initial tumor outgrowth and suggest its role as one of the factors responsible for long-term tumor suppression during tumor dormancy.
The tumor dormant state. Comparison of L5178Y cells used to establish dormancy with those that emerge after its termination.
Authors: Weinhold K J , Miller D A , Wheelock E F
Published: June 2004

The tumor dormant state established in L5178Y immunized and challenged mice is characterized by a prolonged period of clinical normalcy followed by rapid tumor outgrowth. The tumor cells which emerged after termination of the tumor dormant state had abnormal marker chromosomes identical to those in the L5178Y cells used in the original challenge inoculum, indicating that the emergent tumor cells were progeny of the challenge inoculum. Original and emergent L5178Y cells had equivalent in vivo growth rates, when inoculated into normal DBA/2 mice. The emergent L5178Y cells were less susceptible than original cells to in vitro lysis by tumor dormant PC. Original and emergent L5178Y cells expressed common tumor-associated target antigens for cytolytic effector cells. Both modulation and masking of these target antigens were ruled out as mechanisms for decreased susceptibility to cell-mediated cytolysis. Immunofluorescence revealed heterogeneity in tumor-associated antigen expression within both original and emergent cell populations, with a decreased intensity of staining in the emergent population. Both populations were equally susceptible to lysis by alloimmune cells, alloantiserum, and anti-Thy 1.2 serum, but emergent cells were less susceptible to lysis by serum directed against L5178Y TAA. Quantitative absorption revealed that the emergent L5178Y cells expressed eightfold less serologically detectable TAA than the original cells. These findings indicate that the host immune response developing during establishment of the tumor dormant state selects a stable tumor cell subpopulation which expresses decreased amounts of surface tumor-associated target antigens.
The role of H-2-linked genes in helper T-cell function. VI. Expression of Ir genes by helper T cells.
Authors: Marrack P , Kappler J W
Published: June 2004

We examined the expression of (TG)-A--L specific Ir genes in helper T cells using T cells from low responder leads to (B10, high responder x low responder) F1 chimeric mice. In this paper, the low responder strain studied was B10.M, H-2f. B10.M T cells from these chimeric animals do not help anti-TNP-(TG)-A--L responses, even though they have matured in a high responder thymus and been primed and challenged with antigen on high responder Mphi and B cells. These findings indicate that in the H-2f haplotype an Ir-gene controlling anti-(TG)-A--L activity is expressed in helper T cells. The findings are in contrast to those we have obtained and previously reported with T cells of another low responder haplotype, H-2a. Taken together with our previous findings that (TG)-A--L specific Ir genes are expressed by B cells and Mphi of both the H-2a and H-2f haplotypes, the results indicate two sites of action for Ir genes, and suggest two different gene products acting at different stages of the response, both of which are defective in H-2f cells, and only one of which is defective in H-2a cells.
Cellular requirements for lipopolysaccharide adjuvanticity. A role for both T lymphocytes and macrophages for in vitro responses to particulate antigens.
Authors: McGhee J R , Farrar J J , Michalek S M , Mergenhagen S E , Rosenstreich D L
Published: June 2004

By employing primary cultures of purified spleen cells from lipopolysaccharide (LPS) responder (C3H/HeN or C57BL/10Sn) or nonresponder (C3H/HeJ or C57BL/10ScN) mice incubated with particulate antigen and LPS prepared by phenol-water extraction (Ph), we have presented evidence that both T cells and macrophages (MO) are required for LPS-induced adjuvanticity. First, MO derived from C3H/HeN spleen cells, when mixed with responder, C3H/HeN lymphocytes and Ph-LPS, elicited enhanced antibody responses to sheep erythrocytes (SRC) antigen, whereas lymphocytes from the nonresponder, C3H/HeJ mouse strain did not evoke this response. Similarly, purified T cells from C3H/HeN spleens, when cultured with responder, nu/nu spleen cells, and Ph-LPS yielded enhanced anti-TNP PFC responses; whereas, C3H/HeJ T cells did not potentiate immune responses when mixed with optimal concentrations of Ph-LPS. LPS prepared by butanol-water extraction elicited significant adjuvant effects with all cell combinations. Finally, purified responder T cells and MO enabled either responder or nonresponder B cells to elicit LPS potentiation. These data indicate that T cells and MO are controlling LPS-induced augmentation of B-cell responses.