bluuebunny/crossref_metadata_2025 huggingface dataset:
Proliferative Response to Anti-IgM Antibodies of Various B Lymphocyte Subpopulations Isolated by Cell Sorting
Authors: Sidman Charles L. , Unanue Emil R.
Published: December 2022

We have utilized several methods to isolate populations of murine splenic B lymphocytes bearing different classes of surface Ig molecules. Antibody and complement-mediated cytotoxicity was used to remove IgD-bearing B cells, and fluorescein-conjugated anti-IgM and polyspecific anti-Ig-F(ab′)2 antibody fragments were used with a fluorescence-activated cell sorter to isolate cells bearing IgM or all Ig classes. First, the technique of isolating cells in the sorter was evaluated. It was found that among cells sorted as negative for Ig there was always a certain percentage where IgM reappeared on the membrane after ovenight culture. Second, evidence is presented that surface IgM is present on and controls the proliferation of the majority of B lymphocytes in an adult mouse spleen. This includes cells bearing predominantly IgM and cells bearing predominantly IgD. Finally, a theory of B cell stimulation is presented, based on distinctive roles for surface IgM and IgD molecules, as well as a balance between their signals to an individual cell.
Syngeneic GvH Induced in Popliteal Lymph Nodes by Spleen Cells of Old C57BL/6 Mice
Authors: Gozes Yeoshua , Umiel Tehila , Meshorer Asher , Trainin Nathan
Published: December 2022

The frequency of autoimmune processes seems to increase with age. We have studied here whether lymphoid cells of aged mice have the potential to express autoreactivity by the use of the in vivo graft-vs-host (GvH) assay. It was found that spleen cells from old (104 weeks) C57BL mice caused significant enlargement of the popliteal lymph node upon injection into the footpads of syngeneic young or old recipients. Histologically this enlargement presented characteristics of a GvH reaction. This effect, which was not abolished by irradiation of the hosts, was totally cancelled by in vitro irradiation or by anti-θ treatment of the donor cells. These results indicate that T cells from aged mice have the potential to manifest autoimmune reactivity.
Increased Spontaneous Polyclonal Activation of B Lymphocytes in Mice with Spontaneous Autoimmune Disease
Authors: Izui Shozo , McConahey Patricia J. , Dixon Frank J.
Published: December 2022

Early in life, mice of four kinds [NZB, (NZB × NZW)F1, MRL/1, and male BXSB] with autoimmune disease spontaneously produced far more (>3 S.D.) anti-hapten antibody-forming cells in spleens and greater concentrations of anti-hapten antibodies in sera than immunologically normal strains of mice (AKR, BALB/c, C3H, C57BL/6, DBA/1J, DBA/2J, LB/J, 129, NZW, and female BXSB). This increased nonspecific antibody production by the abnormal animals' B cells correlated well with the spontaneous development of anti-single-stranded DNA antibodies, but not with serum levels of the viral envelope glycoprotein, gp70. These results suggest that the spontaneous formation of autoantibodies in mice whose immunologic disorder is manifested by a lupus-like disease may result from polyclonal activation of B cells by endogenous or exogenous B cell activators.
Interaction of Cytotoxic T Lymphocytes with Target Cells
Authors: Whisnant Carol C. , Singer Kay H. , Amos D. Bernard
Published: December 2022

H-2 antigens from EL4 (H-2b) and RL♂ 1 (H-2d) leukemias were solubilized with deoxycholate and partially purified based on their adherence to a Lens culinaris hemagglutinin column. In double reciprocal experiments we have shown that these preparations specifically inhibit conjugate formation between target cells and alloimmune peritoneal lymphocytes, a preparation rich in cytotoxic T lymphocytes.
Mechanisms of Action of “Lymphocyte-Activating Factor” (LAF)
Authors: Katz Stephen P. , Kierszenbaum Felipe , Waksman Byron H.
Published: December 2022

LAF (lymphocyte-activating factor), a nondialyzable, nonmitogenic substance obtained from rat macrophage-conditioned medium, potentiates the response of LNC to the T cell mitogen PHA and substitutes for macrophages in the DNA synthetic response of mitogen-stimulated, macrophage-depleted (Mφ-) LNC. The active factor has a m.w. between 50,000 and 100,000 as determined by membrane filtration. LAF acts between 12 and 24 hr after mitogen addition, apparently stimulating lymphocytes in G1 to enter S phase of the cell cycle. It induces an increase in the number of stimulated LNC synthesizing DNA without affecting the amount of DNA synthesized, as demonstrated by autoradiographic analysis. Its action is not due to generally enhanced cell survival, to prolongation of the S phase, to a mercaptoethanol effect, or to presence of free cold nucleotides in the LAF preparation. The LAF effect on mitogen-stimulated LNC or Mφ-LNC is associated with a rise in cGMP, starting at 12 to 16 hours, and reaching as much as 10 times normal levels by 24 hr. Addition of exogenous DBcGMP (10-7 M), agents that activate guanylate cyclase such as carbachol (10-8 M), or imidazole, a cGMP phosphodiesterase inhibitor (10-9 M), to PHA-stimulated Mφ-LNC mimics the LAF effect, maximally if the reagents are added 12 hr after mitogen. Inclusion of EGTA, an effective chelator of Ca++ (plus excess Mg++) between 12 and 26 hr of culture ablates both the LAF effect and the concomitant rise in cGMP. The evidence supports the conclusion that LAF acts by raising cGMP in G1 cells.
Studies on Lymphocyte-Mediated Cytolysis
Authors: Ozato Keiko , Henney Christopher S.
Published: December 2022

Lymphocyte and mastocytoma (P815) cell populations incubated with DNP-phosphatidylethanolamine in the form of liposomes displayed cell surface hapten as evidenced by: i) 125I-anti-DNP antibody binding; ii) C-mediated lysis, and iii) K-cell mediated lysis. Under optimal conditions, P815 cells incubated with DNP-liposomes showed a higher DNP density than was obtained by direct surface haptenation with dinitrobenzene sulfonic acid (DNBS). Moreover, the DNP transferred via cell-liposome interactions persisted on the cell surface at least as long as that introduced by DNBS.

Despite ready accessibility to antibody, the cell surface hapten introduced by cell-liposome interaction was not recognized by hapten-specific cytotoxic T cells. Thus, cells bearing liposome-derived DNP- (or TNP-) could not serve as targets for H-2 restricted hapten-specific effector cells; did not competitively inhibit the lysis of DNBS- (or TNBS-) modified cells; and, furthermore, did not stimulate hapten-specific cytotoxic cells in either primary or secondary responses. Interestingly, membrane preparations obtained from TNBS-treated cells did not “trigger” primary or secondary hapten-specific cytotoxic responses, although the same preparations readily stimulated “primed” allogeneic cells.

In contrast to the results obtained with cells bearing lipid-associated hapten, the addition of haptenated (DNP- or TNP-) bovine serum albumin (BSA) to lymphocyte cultures readily stimulated H-2 restricted, hapten-specific, cytotoxic T cell production. Cells incubated with DNP- (or TNP-) BSA for even brief (60 to 90 min) periods became susceptible to attack by syngeneic hapten-specific effector cells and specifically inhibited the lysis of syngeneic cells modified with the appropriate nitrobenzene sulfonic acid.
Role of the Murine Major Histocompatibility Complex in Macrophage-Mediated Cytolysis
Authors: Somers Scott D. , Zwilling Bruce S.
Published: December 2022

Peritoneal cells from congeneic resistant mice infected with BCG displayed differential cytotoxicity toward tumor cells, destroying more allogeneic tumor cells than syngeneic tumor cells. This observation was made regardless of the tumor cells used or the effector cell source. The responsible effector cell remained in a doubly adherent population, was sensitive to carrageenan and silica, insensitive to anti-thymocyte sera, and is probably a macrophage. Activated macrophages were capable of reacting against tumor cells as well as histoincompatible embryonic cells. These observations may indicate that macrophages are capable of discriminating cell surface components linked to the major histocompatibility complex.
Specific Antibodies to Hemoglobin A1 (anti-Glu) and Hemoglobin S (anti-Val) in the Guinea Pig: Immunologic and Structural Correlations
Authors: Otieno Samuel , Bloor Anita , Karol Robin , Reichlin Morris , Noble Robert W.
Published: December 2022

Like goats and sheep, guinea pigs can produce, in response to human sickle cell hemoglobin (β6 Glu → Val), an antibody population (anti-Val) that will bind sickle cell hemoglobin but not normal hemoglobin HbA. Unlike goats and sheep, guinea pigs can produce in response to human hemoglobin A1 an antibody fraction, anti-Glu, that will not react with human sickle cell hemoglobin. These anti-Glu antibodies have been isolated by affinity chromatography and their specificity confirmed by fluorescence-quenching titrations. The sequence of the first 10 amino acids of the β-chain of guinea pig hemoglobin has been determined. This sequence differs from those of both hemoglobin HbA and sickle cell hemoglobin by two residues, those at positions 5 and 6. This explains the similarity of the immunogenicity of this site on the two human hemoglobins when administered to guinea pigs. Both goats and sheep are identical to hemoglobin A1 at the β-6 position.
Immune Response to Alloantigens in Vitro, Amplification of the Development of Cytotoxic T Lymphocytes by Lipopolysaccharide and Polyadenylic:Polyuridylic Acid
Authors: Narayanan P. R. , Kloehn D. B. , Sundharadas G.
Published: December 2022

We have examined the effects of lipopolysaccharide (LPS) and the double stranded RNA, polyadenylic:polyuridylic acid (poly A:U) on the generation of cytotoxic T lymphocytes in a mixed leukocyte culture (MLC) reaction of allogeneic mouse spleen cells. Both LPS and poly A:U augment the generation of cytotoxic cells in responder-stimulator combinations, which differ for the entire H-2 region or only for the H-2K or H-2D subregion. Also, in the presence of optimal concentrations of LPS and poly A:U the dose of antigen required for optimal sensitization and generation of cytotoxic cells is reduced significantly. The cytotoxic cells generated are specific for the sensitizing antigen and they resemble the cytotoxic T lymphocytes in a normal MLC in that they show a log-linear relationship between effector:target ratio and percent cytotoxicity. Furthermore, both LPS and poly A:U appear to bring about their effect by acting on the responding cells as the enhanced generation of cytotoxic cells is brought about if the responding cells are preincubated with these agents and washed before adding the stimulating cells. The effect of poly A:U depends on the presence of adherent cells whereas the effect of LPS is independent of the presence of these cells suggesting that the two agents exert their effect at different steps in the development of the cytotoxic cells. These results suggest that elucidation of the mode of action of LPS and poly A:U in this system might shed light on the mechanisms underlying development of cytotoxic T lymphocytes in vitro.
Mouse T Cell-Mediated Cytolysis Specifically Triggered by Cytophilic Xenogeneic Serum Determinants: A Caveat for the Interpretation of Experiments Done under “Syngeneic” Conditions
Authors: Golstein Pierre , Luciani Marie-Françoise , Wagner Hermann , Röllinghoff Martin
Published: December 2022

We have demonstrated by using a combination of in vivo priming and in vitro incubation that the injection into mice of cultured cells can lead to the development of cytolytic T cells as a function of the serum used for growing the injected cells. Moreover, mere injection of xenogeneic serum followed by in vitro incubation of the primed cells in serum-containing medium led to the development of cytolytic T cells. The induction and the cytolysis phases of this system had distinct characteristics. For the induction of the development of cytolytic cells, cytophilic xenogeneic serum determinants were necessary and were recognized in a specific way. For cytolysis, T cells were required, which recognized unidentified antigens (but apparently not xenogeneic serum determinants) at the surface of syngeneic or allogeneic target cells. In addition, different strains of mice had different patterns of reactivity in this system.

These results are discussed from two points of view. First, they lead us to issue a serious caveat for the interpretation of experiments done in “syngeneic” conditions, when the sensitizing cells have been in contact with xenogeneic serum. Second, we underline the interest of this system for further studies on the induction of T cell-mediated cytolysis.
B Lymphocyte Subpopulations Separated by Velocity Sedimentation
Authors: Teale Judy M. , Howard M. C. , Falzon E. , Nossal G. J. V.
Published: December 2022

Spleen cells from unprimed adult, 7-day-old germ-free, and 3-day-old neonatal mice were separated by 1 × G sedimentation velocity, and the resulting fractions were tested in an in vitro splenic focus assay for NIP-specific precursor frequencies, kinetics of the response, and the class of antibody produced by the NIP-specific clones. The results of this study indicate that the splenic focus assay, contrary to other functional B cell assays, is able to detect the responses of small, medium, and large lymphocytes from both adult and neonatal B cell sources. Evidence is presented for a time-dependent appearance of hapten-specific clones in some of the B cell fractions. Most striking is the very large cell subset sedimenting at an s value >5.5 in the adult and >6.1 in the neonate which shows a progressive increase in the frequency of hapten-specific precursors. This would suggest the presence of a less mature B cell. An analysis of the class of antibody produced by the various sizes of B lymphocytes from both adults and neonates indicated that the majority of B cells in the smallest and largest cell fractions tested produce IgM only, whereas the majority of B cells in the medium-sized lymphocyte fraction produced IgM or IgM plus IgG classes.

In addition, the cycling status of adult and neonatal lymphocytes in the various size distributions was examined. It was found that the smallest lymphocyte fraction contained no detectable lymphocytes in rapid cell cycle. The largest lymphocyte fraction, on the other hand, was enriched for cycling cells over unfractionated controls, and in the neonatal fraction the majority of lymphocytes were in rapid cell cycle. The splenic focus assay is presumably capable of detecting a wide range of B cell subsets as defined by the criteria above including less mature B cells and is therefore a very useful cloning assay for studying B cell development.
B Lymphocyte Subpopulations Separated by Velocity Sedimentation
Authors: Teale Judy M. , Howard M. C. , Nossal G. J. V.
Published: December 2022

Neonatal and adult splenic B lymphocyte subpopulations, separated by velocity sedimentation, were tested in an in vitro splenic focus assay for their susceptibility to tolerance induction with hapten-protein conjugates. At least two tolerizable B cell subsets have been defined in the neonatal spleen; one comprising the slowly sedimenting, small lymphocytes and the other comprising the very rapidly sedimenting, large lymphocytes. In addition, a rapidly sedimenting large B cell subset in the adult spleen was found to be highly susceptible to tolerance induction. It was suggested that the inability to detect this susceptibility in unfractionated adult spleen was due to the low proportion of these very large lymphocytes in the total spleen cell population. The tolerizable B cell subsets presently defined probably represent the least mature B lymphocytes detectable in the splenic focus assay.
EAC Rosette-Forming Lymphocytes in Commercial Preparations of Sheep Erythrocytes
Authors: O'Neill Peggy A. , Woodson Dora L. , Mackler Bruce F.
Published: December 2022

In vitro enumeration of human complement (C3) receptor-bearing lymphocytes is routinely performed with sheep erythrocytes (E) sensitized with 19S anti-sheep erythrocyte antibody (A) and fresh mouse complement (C), termed the EAC reagent (1). Sheep whole blood diluted with an equal volume of Alsever's solution is washed with saline and used to prepare the EAC reagent. It is assumed that the residual sheep leukocytes in the solution are rapidly lost upon storage after the blood is collected. However, here we report finding considerable numbers of residual sheep lymphocytes in sheep blood in Alsever's solution up to 30 days after storage at 4°C. In addition, one finds that about 75% of these residual sheep lymphocytes form autorosettes with the EAC reagent.

Preparations of whole sheep blood diluted with an equal volume of Alsever's solution were obtained from a commercial source, and samples were assessed for the presence and viability of lymphocytes starting with the day the blood was drawn and continuing for 4 weeks.
Immune-Type Interferon-Induced Transfer of Viral Resistance
Authors: Blalock J. E. , Georgiades J. , Johnson H. M.
Published: December 2022

Mouse immune-type interferon (type II), a lymphokine, caused the transfer of viral resistance from mouse L cells to human WISH cells. The interferon was incapable of protecting WISH cells in the absence of L cells. The transfer of viral resistance occurred with interferon preparations of various specific activities, and was in proportion to the interferon concentration in te preparations. The transferred resistance had the characteristics of an interferon-induced antiviral state in that it was blocked by actinomycin D, effective against different types of viruses, and resulted from an action on the cell rather than on the virus. Mouse immune-type interferon was more efficient than virus-type (type I) at eliciting the transfer of protection. The transfer phenomenon may represent a mechanism for amplification of the interferon system as a host defense against viral infection. Further, it serves as a model for studying the mechanism of lymphokine-induced transfer of information between cells.
Physiology of Mixed Leukocyte Reaction Suppressor Factor
Authors: Belmont John W. , Rich Robert R. , Rich Susan Solliday
Published: December 2022

The secretory physiology of the T cell-produced lymphokine, mixed leukocyte rection suppressor factor (MLR-TsF), was characterized with respect to its kinetics of secretion and its sensitivity to a variety of metabolic blocking agents. It was found that spleen cells from alloantigen-immunized mice released active MLR-TsF after freeze-thaw lysis. Upon restimulation with the same priming alloantigen, MLR-TsF was secreted into culture supernatants, and the rate of secretion was determined to be nearly constant. Although colchicine and vinblastine, which bind to microtubules, are known inhibitors of lectin-induced proliferation, it was demonstrated that these drugs had no effect on the secretion of MLR-TsF. However, cytochalasin B, an inhibitor which also binds to some cytoskeletal and membrane-associated proteins, did inhibit the production of MLR-TsF. The above findings indicated that the activation-secretion mechanism of MLR-TsF was much like that described for lymphotoxin and macrophage migration inhibition factor. The dissociation between DNA synthesis and lymphokine secretion was also demonstrated in the MLR-TsF system. DNA synthesis plays no role in the in vitro production of suppressor factor, as determined by resistance to treatment with mitomycin C and γ-irradiation. However, new protein synthesis is required as indicated by the potent inhibitory effects of cycloheximide. Experiments utilizing timed addition and removal of cycloheximide defined a broad period of drug sensitivity, starting from the beginning of culture and lasting for 12 to 16 hr. In addition, experiments measuring the effect of cycloheximide on the MLR-TsF content of cell lysates demonstrated that the cell-associated lymphokine activity is lost when protein synthesis is interrupted. These experiments support the conclusion that MLR-TsF is synthesized de novo in culture. In addition, the secretory process itself may require protein synthesis.
Inhibition of T Cell-Mediated Cytolysis by 2-Deoxy-D-Glucose (2-DG): Differential Effect of 2-DG on Effector Cells Isolated Early or Late After Alloantigenic Stimulation in Vitro
Authors: MacDonald H. Robson , Cerottini Jean-Charles
Published: December 2022

The effect of the hexose analogue 2-deoxy-D-glucose (2-DG) on the functional activity of various populations of cytolytic T lymphocytes (CTL) has been compared. Under aerobic conditions, CTL harvested at the peak of the response (day 4) in primary or secondary mixed leukocyte cultures (MLC) were much more readily inhibited by 2-DG than CTL obtained from MLC at later times (day 11 to 18) or from the peritoneal cavity of alloimmune mice. Quantitatively, 0.4 mM 2-DG was sufficient to inhibit cytolysis by 50% in day 4 CTL populations, whereas 25 mM had little or no effect on day 11 to 18 CTL. Evidence was obtained that inhibition of cytolysis by 2-DG under these conditions was accompanied by a parallel inhibition of effector:target cell binding. In contradistinction to these findings, the cytolytic activity of both day 4 and day 11 MLC cells was readily inhibited by 2-DG under conditions where cell respiration was blocked by sodium azide. Furthermore, uptake of radiolabeled 2-DG was observed under aerobic conditions in both day 4 and day 11 MLC cells. These results strongly suggest that inhibition of cytolysis by 2-DG under aerobic conditions is mediated via a direct effect on CTL which is independent of the consequences of energy depletion. An indirect method by which CTL may be inhibited by 2-DG is suggested.
Release of E-Rosette Augmenting Factor (E-RAF) After Stimulation of Human Leukocytes with Mitogens or Antigens
Authors: Agbata Anthony I. , Kirkpatrick Charles H.
Published: December 2022

Stimulation of human peripheral blood leukocytes (HPBL) with mitogens such as phytohemagglutinin or concanavalin A increases the total number of lymphocytes that form rosettes with sheep erythrocytes. A similar effect is seen when HPBL from skin test-positive, but not skin test-negative, donors are stimulated by a specific antigen. It was also found that the culture fluids from mitogen-stimulated lymphocytes contained a substance that significantly increased the percentage of E-rosette forming cells (85 to 95%) over that of control culture fluids (40%). A similar phenomenon was also observed with supernatant fluids derived from antigenic stimulation of cells from skin test-positive donors, but not from skin test-negative subjects.

The factor that produces this effect has been designated E-rosette augmenting factor (E-RAF). It is nondialyzable; it appears in the supernatants of antigen or mitogen-stimulated cells within 12 hr; and its production is not blocked by mitomycin C. It is produced by cells that do not adhere to glass wool columns and by mitogen-stimulated thymocytes. Sephadex G-100 chromatography showed that mitogen-induced E-RAF eluted from the column in advance of antigen-induced E-RAF. E-RAF has many properties that are characteristic of lymphokines.
Human Transfer Factor: Structural Properties Suggested by HPRP Chromatography and Enzymatic Sensitivities
Authors: Burger Denis R. , Vandenbark Arthur A. , Dunnick Wesley , Kraybill William , Daves G. Doyle , Vetto R. Mark
Published: December 2022

Leukocyte extracts containing human transfer factor (TF) were fractionated by exclusion chromatography, and the active fraction (Sephadex G25, Fraction IIIa) was subjected to high pressure, reverse phase (HPRP) chromatography and enzymatic degradation. TF activity was assessed by the systemic transfer of dermal skin test reactivity from KLH-immunized donors to naive recipients. Preparative HPRP chromatography resolved Fraction IIIa into multiple chromophoric regions, two of which demonstrated transfer of KLH reactivity. Alkaline phosphatase treatment of Fraction IIIa converted the major ultraviolet-absorbing component, 5′-inosine monophosphate, to inosine and resulted in TF activity being restricted to one region. This HPRP region (R1A) contained less than 1% of the UV254 active material in Fraction IIIa but greater than 90% of the reactivity.

The sensitivity of TF to pronase, proteinase K, phosphodiesterase I, and phosphodiesterase II was evaluated by inhibition of systemic transfer of KLH reactivity. Pronase and proteinase K destroyed systemic transfer activity and the pronase destruction could be inhibited with traysylol. Phosphodiesterase I, a 3′ exonuclease, destroyed activity, whereas phosphodiesterase II, a 5′ exonuclease, did not. The data are consistent with a phosphodiester-containing polypeptide in the structure of human TF for KLH reactivity.
The Immunochemistry of Antibodies Sharing Concanavalin A's Anti-Mannosyl Binding Specificity
Authors: Reichert Cheryl M. , Goldstein Irwin J.
Published: December 2022

Procedures were developed for the synthesis of the disaccharide hapten, p-isothiocyanatophenyl 2-O-α-D-mannopyranosyl-α-D-mannopyranoside, and for its conjugation to hemocyanin. The synthetic carbohydrate: protein antigen was then emulsified in complete Freund's adjuvant and injected into the footpads of New Zealand White rabbits. A population of the resulting anti-conjugate antibodies displayed some binding properties analogous to concanavalin A, the carbohydrate-binding protein of the jack bean. The antisera weakly precipitated mannans from Saccharomyces rouxii, S. cerevisiae, and an α-(1 → 3)-mannopyranosyl transferase-deficient mutant from Kluyveromyces lactis Y58a. These polysaccharides, possessing side chains containing terminal α-(1 → 2)-mannobiosyl residues, produce strong precipitation reactions with concanavalin A. In addition, various saccharides were tested for their ability to inhibit the interaction of anti-conjugate antisera with α-(1 → 2)-mannobiosyl-containing polymers. p-Nitrophenyl 2-O-α-D-mannopyranosyl-α-D-mannopyranoside showed a strong complementarity for the binding sites of both the anti-conjugate antisera and concanavalin A. However, the antibody failed to bind a concanavalin A-reactive mouse fibrosarcoma or to stimulate mitogenesis of human peripheral lymphocytes.
Further Characterization of Thymic Suppressor Cells and a Factor That Suppress the Generation of Cells Cytotoxic for a Syngeneic Tumor in DBA/2 Mice
Authors: Levy Julia G. , Maier Thomas , Kilburn Douglas G.
Published: December 2022

Antigen-specific suppressor cells and suppressive extracts obtained from the thymuses of DBA/2 mice bearing small syngeneic P815 mastocytomas were compared for their immunogenetic properties and requirements. The assay for specific suppression involved the ability of either cells or extracts to inhibit the primary in vitro cytotoxic response of normal DBA/2 splenocytes to mitomycin-treated P815 cells. It was shown that pretreatment of suppressor cell populations with anti-Iad antiserum plus rabbit complement removed the suppressive activity. Similarly, absorption of the suppressive factor with anti-Iad antiserum removed the suppressive properties of the material. It was found that the suppressor cells, generated in DBA/2 tumor bearers, were capable of specifically suppressing the anti-P815 response of B6D2 F1 radiation chimeras possessing lymphoid cells of the H-2b or H-2t2 haplotype equally as well as they could suppress the response of H-2d-bearing effector cells. This indicates that the suppressor cells are not H-2 restricted with respect to K or D markers on the responder cells in this system.
Influence of Different Routes of Anti-Tumor Immunization: Alternative Induction of Tumor Immunity and Tumor Enhancement
Authors: Bauer Heinz , Hayami Masanori , Stehfen-Gervinus Johannes C.
Published: December 2022

Chickens and quails were immunized in parallel either i.v. or intramuscularly (i.m.) with lectin column-purified antigens from chick embryo cells that were transformed in vitro by avian sarcoma virus (ASV). After five to six injections, immunity of the animals was tested by challenge with ASV into the wing webs. Whereas tumor growth was inhibited after i.v. immunization with respect to incidence rate and time of tumor appearance, tumor growth was enhanced after i.m. injection. Animals that were injected with normal cell antigens served as controls. Spleen cells from only those animals that were immunized i.v. exerted a cytotoxic effect in vitro against ASV-transformed cells, whereas spleen cells from i.m. injected animals, in contrast, suppressed such cytotoxicity. The search for serum blocking or arming factors suggested that sera from i.m. injected animals block cellular cytotoxicity whereas sera from i.v. immunized animals render normal spleen cells cytotoxic (arming effect). The use of viruses from different subgroups and of antigens from gp85-lacking ASV-transformed cells indicates that immune effects were obtained against tumor cell surface antigens that differ from the antigen that is involved in virus neutralization (s-gp85).
Structural Comparison of Murine Ia Antigens Determined by the I-A and I-E Subregions
Authors: Cullen Susan E. , Kindle Cathy S. , Littman Dan R.
Published: December 2022

After nonionic detergent solubilization, the Ia molecules determined by the I-A subregion are distributed between a covalently linked “58K” structure composed of one α chain (35,000 daltons) and one β chain (25,000 daltons) separable by treatment with SDS and mercaptoethanol, and a form in which α and β chains are separable by treatment with SDS alone (α, β form). In contrast, the Ia molecules determined by the I-E subregion have α and β chains that are not covalently linked, and can be entirely separated by SDS-PAGE under rigorous (100°C) dissociating conditions without reducing agent. Milder dissociating conditions (37°C) permitted observation of a noncovalently linked 58K form for I-E molecules. Therefore, in their native state, both I-A and I-E molecules probably are composed of associated α and β chains.

Iodoacetamide treatment of intact cells before detergent solubilization decreased the fraction of I-A molecules in the covalently linked 58K form. This finding probably means that the covalent linkage is an artifact of solubilization, but because it is restricted to I-A molecules, there still must be an underlying structural difference between I-A and I-E molecules. Selective destruction by iodoacetamide of the antigenic activity of the 58K form of I-A molecules could also account for the loss in 58K form, but only if there is an iodoacetamidesensitive antigenic and/or structural difference between I-A molecules in the 58K and α, β form. We compared these two forms by isoelectric focusing, but found no clear differences between them.
The Fine Specificity of Regulatory T Cells
Authors: Adorini Luciano , Miller Alexander , Sercarz Eli E.
Published: December 2022

We have demonstrated the induction of suppressor T cells of highly restricted specificity in a nonresponder strain. HEL (hen egg-white lysozyme) fails to elicit an antibody response in C57BL/10 (B10) mice whereas the closely related REL (ring-necked pheasant egg-white lysozyme) is strongly immunogenic. However, an anti-HEL primary antibody response can be obtained in B10 mice in vivo or in vitro by coupling HEL to red blood cells (RBC). Preimmunization with HEL-CFA induced reduction of the anti-HEL PFC response to a subsequent challenge with HEL-RBC. Suppression was demonstrated by in vitro mixing experiments and was mediated with remarkable efficiency by I-J positive suppressor T cells. Suppression was specific for the anti-HEL response and did not affect the anti-RBC response. Both IgM and IgG anti-HEL in vivo antibody responses showed 70 to 80% suppression. Relatively high doses of antigen were required to generate efficient suppression. Optimal suppression was reached within 4 weeks after HEL priming and remained maximal at least 9 weeks after priming. Fine specificity in the expression of suppressive activity was displayed by HEL-specific suppressor cells that were not able to suppress an anti-REL response. Furthermore, REL priming did not suppress an in vitro anti-HEL response to HEL-RBC. The close similarity in amino acid sequence and in B cell cross-reactivity between HEL and REL, contrasted with the lack of REL recognition by HEL-specific suppressor T cells, indicates that a limited region on the nonimmunogenic HEL, absent on REL, can account for the induction of suppression.
Secretory Component: Interactions with Intracellular and Surface Immunoglobulins of Human Lymphoid Cells
Authors: Crago Sylvia S. , Mestecky Jiri
Published: December 2022

Unstimulated and PWM-stimulated lymphocytes from normal human peripheral blood, cord blood, peripheral blood of patients with panhypogammaglobulinemia and selective IgA deficiency, as well as human lymphoblastoid cell lines were examined for their ability to bind secretory component (SC) on the surface and in the cytoplasm. SC binding was not detected on the cell surface at any stage of differentiation in these cells. However, binding of SC was detected in the cytoplasm of 2.3% of normal peripheral blood lymphocytes cultured in the presence of PWM for 6 to 7 days, and in two IgA-producing lymphoblastoid cell lines. The capability of lymphoid cells to bind SC was not concurrent with J chain production. Although IgA was detected in the cytoplasm of PWM-stimulated lymphocytes from IgA-deficient patients, these cells did not bind SC. The failure to detect surface receptors indicates that SC is not a probable factor determining the homing of IgA precursor cells into exocrine tissues.
Cyclical Changes in Susceptibility of A Myeloma Tumor (LPC-1) to Immune Destruction
Authors: Hale Arthur H. , Celis Esteban , Rusell John H. , Eisen Herman N.
Published: December 2022

Cells of the LPC-1 myeloma tumor (of BALB/c origin) were tested periodically for susceptibility to lysis by anti-H-2 cytotoxic T lymphocytes (CTL) and by complement-dependent anti-H-2 antibodies (Ab), and for ability to adsorb these Ab, as the cells grew in culture and in ascites form in nude (BALB/c·nu) and in normal BALB/c mice. Cells harvested from 2 to 8 days after inoculation into the peritoneal cavity had the “early” phenotype: they were extensively lysed by the CTL and Ab and effectively adsorbed the Ab. Cells harvested on day 8 and afterward had the “late” phenotype: they were not lysed by CTL, required more Ab for lysis, and had about 8-fold less ability (per cell) to adsorb the Ab. All of these activities were completely reversed 2 to 4 days after transferring late cells into fresh BALB/c hosts. LPC-1 cells growing in nude BALB/c mice also changed from the early, CTL-susceptible to the late CTL-resistant phenotype. With repeated cell counts and measurements of 125I-iododeoxiuridine incorporation to estimate growth, it was found that exponentially growing cells had the early phenotype and that cells in stationary phase had the late phenotype. When placed in culture, the early cells retained their phenotype indefinitely (up to 40 days). However, the late cells switched phenotype: initially resistant to CTL and Ab, after 1 to 4 days in culture they became fully susceptible to CTL and highly reactive with the Ab, and these features persisted in culture thereafter. This transition occurred even when the late, resistant cells were prevented from dividing in culture by prior irradiation (10,000 R) or by trinitrophenylation with 2,4,6-trinitrobenzenesulfonic acid. The findings in both nude mice and in culture indicate that the reversible, early-late phenotypic changes are due to changes in individual cells, rather than to immunoselection of preexisting variant cells.
Three Suppressor Systems in Human Blood That Modulate Lymphoproliferation
Authors: Rice Lawrence , Laughter Arline H. , Twomey Jeremiah J.
Published: December 2022

Three suppressor systems of T lymphocyte proliferation found in normal blood were characterized. The adherent cell suppressor system (ACSS) is effected by a steroid and radioresistant monocyte that survives well in culture. The prostaglandin-related suppressor system (PgSS) is effected by a similar cell but is distinct from the ACSS in terms of magnitude of suppression, effective monocyte concentration, and carrageenan or indomethacin sensitivity. The induced suppressor system (ISS) is effected by a T lymphocyte that is partially radiosensitive and loses activity after 24 hr in culture. Although the ISS is unaffected by age, ACSS and PgSS activity is, overall, higher among elderly than among young adult subjects. Activation of these three suppressor systems does not require cell replication. They are not restricted by histocompatibility barriers.
Thermal Injury-Associated Immunosuppression: Occurrence and in vitro Blocking Effect of Post Recovery Serum
Authors: Ninnemann John L. , Fisher Jack C. , Wachtel Thomas L.
Published: December 2022

Sera from 38 of 72 burn patients have been found to be significantly suppressive to the PHA-induced blastogenesis of normal human lymphocytes in culture. In many of these patients, we have observed that suppression levels decline with recovery. In a study of eight of these patients, we have found that the addition of post recovery serum to cultures of normal lymphocytes blocked the suppressive effect of autologous serum obtained earlier. Blocking appears to be achieved through the formation of antibodies since: a) IgG levels are greatly elevated in serum samples having blocking activity, b) the time of appearance of blocking substances in the serum is compatible with the generation of antibody, and c) blocking activity is contained in the protein-A isolated IgG fraction of such post recovery serum.
Search for Suppression of T Cells Specific for the Second Nonhost H-2 Haplotype in F1 → P Irradiation Bone Marrow Chimeras
Authors: Zinkernagel Rolf M. , Althage Alana
Published: December 2022

Irradiation bone marrow chimeras were made by lethally irradiating parental (P) mice and reconstituting them with T cell-depleted bone marrow cells from F1 hybrid offsprings. These F1 → P chimeric lymphocytes were then analyzed for suppressor mechanisms that could cause the differentiation of effector T cells whose responsiveness was restricted in specificity to the host parent's H-2 haplotype. No evidence was detected for acute or chronic suppression of the second parent's restriction specificity in the following tests of cytotoxic T cell activity against virus-infected targets: 1) Mixing normal F1 spleen cells with various amounts of chimeric spleen and lymph node cells during the cytotoxicity test or, during sensitization in F1 mice tested 5 days after irradiation and infection, revealed no suppression of the activity for the second parental type target cells. 2) Chimeric F1 → P1 lymphocytes did not suppress alloreactivity directed against the second parental H-2 type. 3) Presence or absence of T cells in the transferred, reconstituting F1 bone marrow cells did not change the hosts' influence on the selection of restriction specificity. 4) F1 → P chimeric lymphocytes transferred to 500 R irradiated normal F1 mice did not suppress reactivity for the second parental type targets. 5) F1 → P chimeric lymphocytes or bone marrow cells transferred to lethally irradiated F1 could still differentiate to express the second parental restriction specificity; similarly, when F1 → P lymphocytes were mixed with normal but anti-ϑ plus complement-treated F1 bone marrow cells and transferred to lethally irradiated F1, both restriction specificities were generated.

Therefore, F1 → P irradiation bone marrow chimeras as described and tested here do not contain mechanisms that suppress the restriction specificity of the second parent, which is not a host to the maturing F1 T cells, at least at the effector stage or during differentiation.
Immunoglobulin Class Responsible for Gonococcal Bactericidal Activity of Normal Human Sera
Authors: Schoolnik Gary K. , Ochs Hans D. , Buchanan Thomas M.
Published: December 2022

“Natural” gonococcal bactericidal antibody is present in the serum of most normal adults and may prevent the systemic dissemination of serum sensitive strains of Neisseria gonorrhoeae. The bactericidal activity of normal human serum for gonococcal strain F62 resides entirely in the IgM serum fraction and is destroyed by treatment with 2-mercaptoethanol. Its absence from cord, but not paired maternal sera, provides additional evidence that this activity is mediated by IgM. Serum bactericidal activity is low and comparable to cord serum for the first 4 weeks of life, rising progressively thereafter to 25, 50, and 100% of the adult geometric mean titer at ages 6 months, 1 year, and 2 years, respectively. The sera of eight patients with x-linked agammaglobulinemia were deficient in gonococcal bactericidal antibody. In contrast, sera from a single patient with x-linked immunodeficiency with increased IgM and nine patients with ataxia telangiectasia possessed normal bactericidal activity for strain F62. The development of gonococcal bactericidal antibody may be delayed in the Wiskott-Aldrich syndrome, since bactericidal activity was low for age in two patients studied at age 2 years, but normal in a third child studied at age 12. The magnitude and nature of the antibody response to bacteriophage antigen and the IgM serum concentration were correlated with the serum bactericidal activity of 22 patients with variable immunodeficiencies. The characterization of the natural occurrence of gonococcal bactericidal antibody should serve as a useful baseline in studies of the immune response to gonococcal infection or immunoprophylaxis.
Immunologic Vasculitis in Beige Mice with Deficiency of Leukocytic Neutral Protease
Authors: Johnson Kent J. , Varani James , Oliver Janet , Ward Peter A.
Published: December 2022

Immune complex vasculitis has been induced in normal mice and in mice with features of the Chediak-Higashi syndrome (“beige” mice). The accumulation of neutrophils in peritoneal exudates after the injection of C5a is not quantitatively depressed in beige as compared with normal mice. Immune complex-induced vasculitis in these two strains of mice is not quantitatively different, as assessed by vascular damage (vasopermeability changes and histologic criteria). Measurements of leukocyte enzymes confirm the findings of Vassalli et al. that leukocytes of beige mice lack neutral protease activity. The data suggest that the neutral protease of murine leukocytes does not account for the vascular damage of immune complex vasculitis.
Detection and Characterization of Human Leukemia-Associated Antigens on Leukemic Cell Lines and Thymocytes
Authors: Seon Ben K. , Yoshizaki Kazuyuki , Pressman David
Published: December 2022

Common human thymus-leukemia-associated (HTL) antigens were identified for seven leukemic T cell lines and several thymocyte preparations derived from different donors by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The leukemic T cell lines tested were MOLT-4, CCRF-CEM, RPMI 8402, CCRF-HSB-2, JM, HPB-ALL, and HPB-MLT. The leukemic T cells and normal thymocytes along with normal peripheral blood lymphocytes (PBL) and other control cells were labeled with 125I by lactoperoxidase-catalyzed iodination and then lysed with Renex 30, a nonionic detergent, in the presence of protease inhibitors Trasylol and phenylmethanesulfonyl fluoride. The supernatant of the cell lysate was subjected to various treatments to minimize the occurrence of nonspecific precipitation of radio-activity in the subsequent analyses. The treated supernatant was subjected to immunoprecipitation by adding the γG fraction of rabbit antiserum rendered specific to HTL antigens (Abs-RαMOLT) followed by the subsequent addition of a slight excess of goat antiserum to the Fc fragment of rabbit IgG. The resulting immunoprecipitate was reduced with dithiothreitol and analyzed by SDS-PAGE. Two different types of human thymocytes were identified and designated types I and II. Type II thymocytes showed three HTL antigenic components with approximate sizes of 150,000, 120,000, and 63,000 daltons, whereas type I thymocytes showed only the 63,000 dalton component. The seven leukemic T cell lines studied were similar to type II thymocytes in the SDS-PAGE pattern of HTL antigens rather than to type I thymocytes, although there were some differences in the pattern among the different cell lines. The HTL antigens were not detected on PBL, on normal B cell lines, on the leukemia cells of a patient with chronic lymphocytic leukemia, and on the leukemia cells of a patient with acute myelogenous leukemia. Control experiments were carried out with normal rabbit serum or with Abs-RαMOLT absorbed with thymocytes in place of Abs-RαMOLT.
Production and Characterization of DR Xenoantisera: Use for Detection of Serum DR Antigens
Authors: Wilson B. S. , Indiveri F. , Pellegrino M. A. , Ferrone S.
Published: December 2022

Immunoadsorbents consisting of rabbit anti-DR antibodies bound to Staphylococcus aureus were used to bind DR antigens from NP40 cellular extracts of cultured B lymphoid cells. When injected into rabbits, these DR antibody adsorbents elicited the production of antibodies that were shown by both serologic and immunochemical techniques to react specifically with the DR antigen molecule. A rosette inhibition assay with B lymphoid cells and anti-DR xenoantisera was used to detect DR antigens in human serum. DR serum antigens could be found in an enriched preparation of serum lipoproteins obtained by centrifugation in KBr. In addition, sera from some patients with neoplastic diseases contained higher levels of DR serum antigens than those found in normal individuals.
Murine Plasma Cells Secreting More Than One Class of Immunoglobulin
Authors: Mckeever Paul E. , Neiders Mirdza E. , Nero Gregory B. , Asofsky Richard
Published: December 2022

The antigenic and molecular characteristics of the two immunoglobulins secreted by a single cell line of plasmacytoma SAMM 368 were analyzed by polyacrylamide gel electrophoresis of biosynthesized proteins. Adapted to continuous in vitro cultivation, this BALB/c plasmacytoma secretes at least 98% of its heavy chains as components of fully assembled and isotypically uniform IgG2b and IgA molecules. The IgA is secreted as monomers, dimers, and multimers with chemical properties typical of BALB/c myeloma IgA including disulfide bonded J chain and noncovalently bonded light chains. The noncovalently bonded light chains are monomers rather than dimers. Free light chains are also secreted. The ability to segregate heavy chains is attributed either to chemical, enzymatic, or compartmental regulatory factors operating within these plasma cells.
Shared N-Terminal Sequences in Monoclonal IgMκ and IgGκ Proteins from a Patient with a Complex Multiple Paraprotein Disorder
Authors: Hopper John E. , Noyes Claudia , Hsu Robert , Heinrikson Robert , Gallagher William
Published: December 2022

The N-terminal sequence analyses were performed on the heavy (H) and light (L) chains of the idiotypically identical IgMκ and IgGκ paraproteins isolated from the serum of patient, Cam. The N-terminal 39 residues of the κ chains of the IgM and IgG were identical and belonged to the human VκIII subgroup. This sequenced stretch included the first L chain hypervariable region. The N-terminal 27 residues of the variable regions (VH) of the respective µ and γ heavy chains were also identical and belonged to the human VHIII subgroup. These identical VH sequences were unique with lysine residues at positions 13 and 19. This dual lysine substitution has not been seen in 37 other human VHIII sequences reported in the literature.

This N-terminal sequence homology in the V-regions of Cam IgMκ and IgGκ paraproteins and the shared idiotypy expressed by Cam IgM, IgG, and IgA proteins strongly suggest the existence of complete structural homology in the variable regions of the H and L chains of these Ig molecules of three separate Ig classes. At the cellular and genetic level, these results point toward a common clonal origin for the idiotypically related Ig molecules and suggest that identical V-region (VH and VL) genes were utilized by the Cam lymphoid clone in the biosynthesis of the respective IgM, IgG, and IgA proteins.