bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of Carrageenan on Immune Responses
Authors: Ishizaka Shigeaki , Otani Shuzo , Morisawa Seiji
Published: December 2022

Although C57BL/6 mice showed little response to low dose antigenic stimulation with dinitrophenylated bovine γ-globulin (DNP-BGG), this immune response was enhanced markedly by carrageenan treatment. They also showed a good response to dinitrophenylated ovalbumin (DNP-OA), but this response was much reduced by carrageenan treatment. These results were reversed in C3H/HeN mice. The responder status of these animals seems to depend mainly on macrophages rather than on thymus-derived cell (T cell) or bone marrow-derived cell (B cell) function since similar results were obtained in in vitro experiments with spleen cells from which macrophages had been removed. However, these conclusions do not exclude the possibility that suppressor T cells may have some effect on the low responsiveness of C57BL/6 mice to DNP-BGG.
Genetic Control of the Immune Response to Mammalian Myoglobins in Mice
Authors: Berzofsky Jay A.
Published: December 2022

The antibody response to the major chromatographic component IV of sperm whale myoglobin (MbIV) has been found to be under H-2-linked Ir gene control in several series of congenic resistant strains of mice on the C57BL/10, A, and DBA/2 backgrounds by using an assay which measures molar concentration of antibody independent of affinity. The H-2d and H-2s mice were high responders, whereas the H-2k, H-2b, and H-2q mice were low responders. High responsiveness was dominant in the (H-2d × H-2b)F1 cross. The genetic control was mapped to within the I (or S, G) regions by the low response of A.TL mice. The presence of two (or more) genes within this region controlling the response was indicated by the intermediate response of the B10.A, A/J, and B10.A(5R) strains, indicating at least one gene to the right of H-2K but left of I-C, tentatively called Ir-Mb-1, and at least one other to the right of I-E but left of H-2D (presumably in I-C), tentatively called Ir-Mb-2. The former of these was further mapped to the I-A subregion by the high responsiveness of the D2.GD mice (which are d in K and I-A and b elsewhere), virtually identical to that of the DBA/2 (H-2d) mice with which these are congenic. This last observation suggests that either Ir-Mb-1 can confer full responsiveness by itself, or that coupled complementation is active. In contrast, Ir-Mb-2 alone confers only intermediate responsiveness. Whether these genes control the response to different determinants on the myoglobin molecule remains the subject of further study.

In addition, for a given H-2 haplotype, the response of A background mice was about 5-fold higher than that of B10 background mice. Thus, non-H-2-linked genes also control the level of response. All of the above levels of control, both H-2 and non-H-2 linked, appeared to operate both after a single immunization in complete Freund's and after two subsequent booster immunizations.
Cytotoxic Activity of Lymphocytes
Authors: Yano Keisuke , Lucas Zoltan J.
Published: December 2022

To detect the cellular origins of α-lymphotoxin (α-LT), we cultured various subpopulations of human blood lymphocytes separated by erythrocyte-rosetting techniques with various mitogens. T cell-enriched subpopulations responded to PHA by increased 3H-thymidine uptake into DNA and large amounts of α-LT production. SPL and Con A-Sepharose stimulated DNA synthesis in T cell-enriched cultures if the macrophage content was greater than 1.5%; however, α-LT production was not induced by these two mitogens even when reconstituted with 10% macrophages. B and/or null cell-enriched populations severely depleted of T cells (less than 0.7%) did not respond to PHA, SPL, or Con A-Sepharose. However, reconstitution to 5 or more percent in E-RFC allowed all three mitogens to stimulate DNA synthesis and α-LT production. The LT made by all cell populations 5 and 7 days after stimulation were equally neutralized by a heterologous antiserum to α-LT. These results show that human T and B and/or null cells, when appropriately stimulated, can produce α-LT.
An Antigen-Specific I Region Gene Product That Augments the Antibody Response
Authors: Tokuhisa Takeshi , Taniguchi Masaru , Okumura Ko , Tada Tomio
Published: December 2022

The extracts of thymocytes and spleen cells of mice that had been primed with a carrier antigen (keyhole limpet hemocyanin, KLH) were found to augment the in vitro secondary antibody response of spleen cells against a hapten (2,4-dinitrophenyl, DNP) coupled to KLH, when they were added to the culture 1 to 3 days after the start of cultivation. The factor that augments the antibody response (TaF) had similar physicochemical properties to those of the previously reported antigen-specific suppressive T cell factor (TsF) and was removed by absorption with the immunoadsorbent composed of only relevant antigen (KLH) but not of anti-immunoglobulin antibodies. TaF and TsF were, however, distinguishable by differential absorption with various alloantisera directed at different subregions of the H-2 histocompatibility complex. The activity of TaF was removed by antisera reactive to the products of the I-A subregion, being distinct from TsF which was absorbed by those specific for the I-J subregion. Evidence was presented that the I-region determinants on TaF are distinct from those on Ia antigens that are primarily detectable on B cells, suggesting that TaF is determined by a different locus (Ia-6 ?) from that for B cell Ia antigen (Ia-1). Furthermore, it was shown that TaF is a product of Lyt-1+, 2-, 3- T cells, a fact which clearly distinguishes the cellular origin of TaF from that of TsF which is Lyt-1-, 2+, 3+.

TaF can enhance the antibody response only if the helper T cell having the same carrier-specificity was present in the culture, indicating that TaF is not a helper T cell-replacing factor. The presence of both suppressive and augmenting T cell factors determined by different I subregions in the extract of primed lymphoid cells suggests that the balance of these T cell factors determines the magnitude of the antibody response.
Quantitative Studies on the Precursors of Cytotoxic Lymphocytes
Authors: Teh H.-S. , Phillips R. A. , Miller R. G.
Published: December 2022

Murine cytotoxic lymphocyte precursor cells (CLP) appear to be specificity restricted, yet 2 to 4% of the total pool appears to be activated in a response against a single foreign H-2 alloantigen. The simplest model that provides enough CLP for all possible reactions is that H-2 antigens have cross-reacting determinants. Experimentally, we observed that activation of RNC-nu/+ (H-2k) LN cells with H-2b spleen cells activated 1200 to 1800 CLP per 106 LN cells capable of lysing H-2b target cells. Of these, 3 to 4% could also lyse H-2d target cells. In addition, a small number of clones (approximately 25/106 LN cells) were activated which could lyse H-2d but not H-2b target cells. However, the total number of such apparent nonspecifically activated clones is small since greater than 80% of all the CLP activated on culturing with H-2b alloantigens will lyse H-2b targets. The results are discussed in terms of specificity restriction and possible subpopulations of CLP.
Characterization of the Target Cell Receptor for IgE
Authors: Conrad Daniel H. , Froese Arnold
Published: December 2022

The receptor for IgE from surface-labeled and Nonidet P-40- (NP-40) solubilized rat basophilic leukemia (RBL) cells was isolated with the aid of IgE coupled to Sepharose, by using KSCN as the eluting agent. The IgE-rebinding capacity of this eluate was only 12 to 19% suggesting that some alteration of the receptor had taken place. Upon sucrose density gradient centrifugation and gel filtration the receptor exhibited considerable size heterogeneity, most likely as a consequence of detergent binding. Detergent binding could also be implied from isopycnic centrifugation which yielded a density for receptor which was below that of free IgE. When these experiments were repeated in the presence of SDS, the size heterogeneity disappeared and the density was found to be 1.28 g/ml which is in the normal range for proteins. Analysis of the receptor eluate by SDS polyacrylamide gel electrophoresis on 5% gels indicated a single component with a slightly slower mobility than that of receptor coprecipitated with IgE. When the same receptor eluate was analyzed on 10% gels, two components, with apparent m.w. of 45,000 daltons and 55,000 daltons were seen. Coprecipitated receptor again exhibited only a single peak with an identical relative mobility to that of the 45,000-dalton component in KSCN eluate. The binding of both components to IgE-Sepharose could be inhibited by free IgE; identical concentrations of a purified fraction of rat IgG had no effect. Although complete inhibition of binding was seen only with the lower m.w. component, the inhibition data still suggest some specificity in the interaction of the 55,000 dalton component with IgE.
Fragments Ba and Bb Derived from Guinea Pig Factor B of the Properdin System: Purification, Characterization, and Biologic Activities
Authors: Hamuro Junji , Hadding Ulrich , Bitter-Suermann Dieter
Published: December 2022

Functionally active guinea pig factor B was purified by a combination of chromatographic steps including Sephadex G-25, QAE A25, QAE A50, CM C50, and Sepharose 4B coupled with purified cobra venom factor. Purified factor B had a m.w. of 106,000 daltons and a single subunit structure. It was heat labile. After cleavage of native B with cobra venom factor coupled to Sepharose 4B in the presence of D̄, the resulting two fragments, the larger one (Bb) and the smaller one (Ba), were further purified. The m.w. of Bb and Ba was determined as 64,000 and 53,000 daltons, respectively, by SDS-PAGE. Neither of the fragments evoked a contraction of guinea pig ileum or histamine release from rat mast cells. Only the smaller fragment Ba (at a concentration of 120 nM) stimulated guinea pig peritoneal polymorphonuclear leukocytes to respond with increased movement. This activity as well as the antigenicity of Ba were heat stable, but were sensitive to trypsin digestion, whereas the antigenicity of Bb was heat labile.
Stimulation of an Enhanced in Vitro Immune Response by a Synthetic Adjuvant, Muramyl Dipeptide
Authors: Specter Steven , Cimprich Roberta , Friedman Herman , Chedid Louis
Published: December 2022

Various subcellular bacterial fractions are known to enhance immune responses and serve as potent adjuvants. Muramyl dipeptide (MDP), a synthetic adjuvant mimicking a component of mycobacterial cell walls, enhances humoral immunity to soluble antigens and can increase macrophage cytotoxicity toward mastocytoma cells in vitro. In the present study MDP was found to enhance the hemolytic antibody plaque response of normal mouse spleen cells in vitro to SRBC at a level equal to or greater than that induced by Escherichia coli lipopolysaccharide. Furthermore, MDP was found to enhance the antibody response to SRBC nonspecifically in unimmunized spleen cell cultures, suggesting that similar to LPS the synthetic dipeptide may induce a generalized clonal expansion of committed lymphocytes and thus serve as a “polyclonal activator.” MDP also enhanced the immune responsiveness of normal splenocytes to suboptimum concentrations of SRBC, indicating that this material may be useful in enhancing immunity in situations where there would normally be a poor immune response.
Antigenicity of the Major Polypeptides of Hepatitis B Surface Antigen (HBsAg)
Authors: Shih J. Wai-Kuo , Tan Peter L. , Gerin John L.
Published: December 2022

The major polypeptides (P-1, P-2, and P-6) of HBsAg were isolated from purified preparations of 22-nm HBsAg particles, iodinated, and analyzed by double-antibody radioimmunoprecipitation assays for the presence of hepatitis B virus (HBV)-specific antigens. Each polypeptide fraction contained both group (a) and subtype (d) specific determinants in common by virtue of their immunoreaction with antiserum to native HBsAg and antisera to the other structural polypeptides. The antigenic and structural similarities of the HBsAg polypeptides establish that they are not each unique gene products of the HBV genome.
Acceleration of Human Lymphocyte Activation after Preliminary Culture
Authors: McCombs Candace C. , Michalski Joseph P. , Talal Norman
Published: December 2022

Human peripheral blood mononuclear cells which are precultured in tissue culture medium containing 5% fetal calf serum (FCS) have markedly augmented reactivity to a suboptimal concentration of phytohemagglutinin (PHA). The augmentation of response is due in part to an acceleration of lymphocyte activation. Precultured cells also respond in an accelerated manner to allogeneic cells. The augmentation is apparent after 24 hr of preliminary culture, and reaches a maximum at 4 to 7 days. Kinetic studies of the culturally augmented PHA response indicate that an increased number of reactive lymphocytes rather than an increased rate of proliferation accounts for the effect. Adding fresh cells to the precultured cells does not suppress their augmented response. when human serum is substituted for FCS in the preliminary culture, the augmentation of response does not occur. Serum-mixing experiments indicate a suppressive effect of human serum. These studies may be relevant for understanding immunologic mechanisms related to transplantation and autoimmunity.
Differential Immunosuppressive Effects of Ascites Fluid from Mastocytoma-Bearing Mice on Primary vs Secondary Immunocyte Responses
Authors: Kamo Isao , Friedman Herman
Published: December 2022

The effects of immunosuppressive ascites fluids from mastocytoma-bearing mice on the primary vs secondary immune response to sheep red blood cells (SRBC) was examined. Injection of mice with ascites fluid from tumor-bearing mice markedly depressed the primary immune response of normal syngeneic mice challenged with SRBC. However, there was a preferential depression of the 19S IgM antibody response as compared with the 7S IgG response. Injection of ascites fluid shortly before secondary immunization of mice with SRBC also resulted in depressed IgM PFC responses but only a slight to moderate depression of IgG PFC. Treatment of mice with the ascites fluid before primary immunization had little if any effect on the secondary IgG PFC response, although the IgM response was moderately depressed. These results indicate that the immunosuppressive factor(s) present in the ascites fluid of mastocytomabearing mice has a differential effect on distinct classes of immunocytes. Those immunocytes or their precursors involved in formation of low efficiency 7S IgG antibody are more resistant to immunodepression. Such differences appear due to different sensitivities of cells involved in the immune response system.
BCG-Induced Suppressor Cells
Authors: Klimpel Gary R. , Henney Christopher S.
Published: December 2022

Spleen cells obtained from mice 5 to 40 days after infection with viable BCG organisms (BCG-spleens) were found to be unresponsive in vitro to both mitogenic and alloantigenic stimuli. Moreover, suppressor cells could be demonstrated in the spleens from these infected animals. When spleen cells from BCG-infected mice were added to either syngeneic or allogeneic normal spleen cells, the mixtures neither proliferated nor developed cytotoxic activity when cultured with alloantigen or with concanavalin A (Con A). The development of unresponsiveness post-infection paralleled the onset of suppressive activity. Spleen cells obtained from mice given heat-killed BCG were neither suppressive nor unresponsive.

The suppressive activity of BCG-spleen cells was associated with an adherent, phagocytic cell that lacked membrane-associated Thy-1 antigen. Removal of this cell by passage through nylon wool columns resulted in a cell population that was no longer capable of suppression and that responded normally to alloantigen and to Con A.

It would thus appear that BCG infection results in the development of a “suppressor” macrophage-like cell population within the spleen. The role of this cell type in regulation of the immune response in BCG-infected animals is as yet undefined.
Subpopulations of Splenic T Cells Regulating an Anti-Hapten Antibody Response
Authors: Muirhead Donna Y. , Cudkowicz Gustavo
Published: December 2022

There was a pronounced quantitative difference between the helper activities of B6C3F1 splenic T cells sensitized with unmodified vs modified antigens of SRBC. Modified SRBC induced the greater helper activity which was measured by the magnitude of an anti-TNP response (IgM and IgG) elicited in vivo by virgin B lymphocytes. Antigen modification was produced by conjugating SRBC with the hapten or simply by incubating SRBC in cacodylate buffer. There were restrictions with respect to both erythrocyte species and mouse strains for this differential priming to occur.

The relatively poor performance of SRBC-primed T lymphocytes was apparently not due to suppressor T cells, but rather to activation of only one of two identified T cell subpopulations required for full helper activity. Unmodified SRBC activated a subpopulation of “helper” cells characterized as sensitive to elimination by ATS and long-lived after ATx, but failed to activate in B6C3F1 mice a second subpopulation of “amplifier” cells resistant to elimination by ATS and short-lived after ATx. In contrast, modified SRBC activated both helper and amplifier cells. Under appropriate conditions these subsets of T cells were strongly synergistic in promoting antihapten antibody formation especially of the IgG class. The involvement of two distinct types of T lymphocytes in the positive regulation of antibody responses raises interesting and novel questions concerning the sequence of events in the triggering of B cells and the subsequent development of the response.
In Vitro Mitogenic Stimulation of Murine Spleen Cells by Herpes Simplex Virus
Authors: Kirchner Holger , Darai Gholamreza , Hirt Hans Martin , Keyssner Katharina , Munk Klaus
Published: December 2022

Spleen cells of B6 mice not previously immunized were induced to DNA synthesis by supernatants from HSV-infected tissue culture. The stimulatory principle could be passed through a 45-µm filter and sedimented at 100,000 × G. It was abolished by UV light, heating at 56°C, and by an anti-HSV serum. The possibility that the observed stimulation was caused by LPS was therefore excluded, and there was also no indication of mycoplasma contamination. Partial purification of spleen cells from macrophages resulted in an increased stimulation by HSV. From experiments with nylon columns, anti-θ antibody, and nude mice it was concluded that HSV acted as a B cell mitogen. Strains of both HSV types 1 and 2 were stimulatory for B6 spleen cells. Of nine freshly isolated HSV strains with identical passage history (twice in HEF) four were strongly stimulatory, three showed a moderate stimulation, and two did not stimulate. Spleen cells from A/J and DBA/2 mice were stimulated to the same extent by HSV (WAL) as spleen cells from B6 mice. No viral replication was demonstrable in B6 spleen cell cultures stimulated for DNA synthesis by HSV. Thus our study demonstrates induction of cellular DNA synthesis in B lymphocytes by HSV which is abolished by inactivation of the virus.
Rfv-1 and Rfv-2, Two H-2-Associated Genes That Influence Recovery from Friend Leukemia Virus-Induced Splenomegaly
Authors: Chesebro Bruce , Wehrly Kathy
Published: December 2022

Two genes, Rfv-1 and Rfv-2, that influence recovery from Friend virus leukemia have been identified within or adjacent to the mouse major histocompatibility gene complex (H-2). Rfv-1 was determined to be in the H-2G or H-2D regions by testing of mice with recombinations both to the left and right of these regions. Rfv-2 was located either in the H-2K or H-2I regions or in the Tla region. The Rfv-2 effect was not seen in the high recovery H-2D(Rfv-1)b/b mice. This could be an indication that this gene is weaker than Rfv-1 in its influence on leukemia. The influence of H-2 on recovery from leukemia appeared to operate in a gene-dose fashion (H-2b/b>H-2a/b>H-2a/a).
Effect of Liposomal Model Membrane Composition on Immunogenicity
Authors: Dancey George F. , Yasuda Tatsuji , Kinsky Stephen C.
Published: December 2022

We have examined the effect of composition on the immunogenicity in mice of liposomal model membranes sensitized with dinitrophenyl-ε-aminocaproyl-phosphatidylethanolamine (DNP-Cap-PE) derivatives. Neither cholesterol content nor incorporation of exogenous charged amphiphile (dicetylphosphate, stearylamine) exerted a significant influence on the in vivo anti-DNP response as measured by the appearance of direct plaque-forming cells in the spleen. Similarly, the nature of the fatty acids (saturated vs unsaturated) present in DNP-Cap-PE had no effect. In contrast, the nonpolar region of the basic phospholipids comprising the liposomal bilayers played an important role as revealed by a comparative study of model membranes prepared with beef sphingomyelin (SM), egg phosphatidylcholine (PC), and synthetic distearoyl-, dimyristoyl-, dilauroyl-, and dioleoyl-phosphatidylcholines (DSPC, DMPC, DLPC, DOPC). Thus, liposomes with a large content of phospholipids possessing a high transition temperature (e.g., beef SM, DSPC) were more immunogenic than those containing phospholipids of low transition temperature (e.g., egg PC, DOPC). This correlation held for both unsonicated and sonicated liposomes. These findings may have a bearing on the phenomenon of membrane-localized antigen expression.
Immunologic Responses to Candida Albicans
Authors: Pearsall Nancy N. , Adams Bonnie L. , Bunni Raad
Published: December 2022

Passive transfer of immune serum gave a significant degree of protection against deep seated candidiasis in mice. Repeated attempts to transfer resistance by the transfer of sensitized lymphoid cells gave negative results, even though cutaneous delayed hypersensitivity was transferred by the cells. The results suggest that cell-mediated immunity is not of primary importance in this model of murine candidiasis, and that humoral immunity contributes to protection.
Microtubule Assembly and Function in Normal and Glutathione Synthetase-Deficient Polymorphonuclear Leukocytes
Authors: Oliver Janet M. , Spielberg Stephen P. , Pearson Christine B. , Schulman Joseph D.
Published: December 2022

Depression of glutathione (GSH) by specific and non-specific oxidants has been shown before to cause microtubule disassembly and to prevent Concanavalin A (Con A)-induced assembly in human polymorphonuclear leukocytes. These data indicate either that a critical level of GSH is required for assembly or that products of GSH oxidation such as glutathione disulfide (GSSG) cause disassembly. Neutrophils from a patient with GSH synthetase deficiency contain only 20% normal GSH (far below the depressed level associated with microtubule inhibition in normal cells) with no corresponding elevation of GSSG. These cells provide a test system to analyze the biochemical pathway connecting GSH to microtubule integrity. Con A induces a rapid and equal increase in centriole-associated microtubules in both normal and GSH-deficient cells. Thus, levels of GSH above 20% of normal are not essential to promote microtubule assembly. On the other hand, our data emphasize an important role for normal (high) GSH in protecting tubulin against direct attack by oxidants such as H2O2: in normal neutrophils phagocytosis induces microtubule assembly despite concomitant production of H2O2; in GSH-deficient cells microtubule assembly is prevented. Tertiary butylhydroperoxide (BHP) is confirmed to be a highly specific GSH oxidant, inhibiting Con A-induced microtubule assembly in normal but not GSH-deficient cells. In both cases the GSH:GSSG ratio is lowered from approximately 100:1 to below 10:1. However, the absolute level of GSSG achieved after BHP in GSH-deficient cells is only one-fifth that obtained in normals. It is suggested that a critical level of GSSG must be attained to inhibit microtubule assembly and/or cause microtubule disassembly after the specific oxidation of GSH.
Inhibition of Tumor Growth Mediated by Lymphocytes Sensitized in Vitro to a Syngeneic Murine Teratocarcinoma 402AX
Authors: Bartlett Perry F. , Fenderson Bruce A. , Edidin Michael
Published: December 2022

TerC, a cell line derived from a strain 129 teratocarcinoma 402AX, was used to sensitize syngeneic 129 (H-2bc) splenic lymphocytes in vitro. The effector cells generated inhibited in vitro growth of TerC as measured by an 125I-IUDR post-labeling technique. It was also shown, with a modified Winn assay, that the sensitized cells were effective in preventing TerC growth in vivo. The effector lymphocyte was nonadherent to nylon wool, was sensitive to anti-Thy-1·2 + C, and was phenotypically Ly 1-2+. The anti-TerC effector T lymphocytes were not functional in a 51Cr-release assay. However, this failure to lyse appears not to be due to some intrinsic membrane resistance since both BCG and ConA-activated killers were able to lyse TerC. The TerC-sensitized lymphocytes displayed no H-2 restriction and were able to growth inhibit in vitro a wide range of tumorigenic cell lines, e.g., P815 (H-2d), EL-4 (H-2b),Sal (H-2a), and BALB/c (H-2d) 3T12. Mouse blastocyst cell lines were also inhibited. BALB/c 3T3 and mouse fibroblast cell strains were not growth inhibited. Thus, it appears that oncofetal antigens expressed on TerC are capable of initiating a cell-mediated response and that these antigenic specificities are shared by many transformed cell lines.
Suppressor Cell Activity in Tumor-Bearing Mice
Authors: Farrar William L. , Elgert Klaus D.
Published: December 2022

Spleen cells from fibrosarcoma-bearing mice elaborated a factor of 30 to 50,000 daltons in m.w. that was capable of inhibiting DNA synthesis and DNA polymerase activities of mitogen-stimulated lymphocytes plus unrelated cell types. The cells responsible for producing these factors were characterized as anti-Thy 1 serum- and hydrocortisone-sensitive mildly nylon wool adherent spleen cells (suggesting a suppressor T cell). Both DNA polymerases alpha (α) and beta (β) were significantly inhibited by prepared supernatants from cultures of tumor-bearing host (TBH) spleen cells. DNA polymerase polymerase (IDP). Basal levels of TBH lymphocyte DNA synthesis and polymerase α activity were always higher than their normal counterparts; however, upon mitogen stimulation this was reversed. Normal animal spleen cell DNA polymerase α activity exhibited a direct correlation with mitogen-induced DNA synthesis. In the presence of mitogen, normal spleen cells demonstrated a significant increase in DNA synthesis with a corresponding increase in DNA polymerase α activity, whereas TBH spleen cells upon mitogen stimulation resulted in a concomitant decrease in DNA synthesis and polymerase activity. Inhibition kinetics revealed that IDP was not a competitive inhibitor of DNA polymerase α and did not bind directly to the DNA template in the absence of polymerase. The kinetics imply the production of an enzyme-substrate-inhibitor complex which cannot undergo further reaction to yield a normal product. The data suggest that substances elaborated by tumor-induced suppressor cells operate at a molecular site of DNA polymerases resulting in inhibited cellular proliferation.
X-Linked Genetic Control of Hapten-Polysaccharide-Mediated Specific Immune Unresponsiveness in CBA/N Mice
Authors: Merchant Bruce , Snippe Harm , Lizzio Elaine F. , Inman John K.
Published: December 2022

CBA/N mice carry an X-linked recessive B cell defect which prevents them from making antibody responses to thymic independent antigens such as arsonate-, sulfonate-, or DNP-conjugated Ficolls. They do, however, make good plaque-forming cell (PFC) responses to the same haptens coupled to hemocyanin; thus, they do not selectively lack Ig receptors reactive with these haptens. When CBA/N mice are injected with DNP-Ficoll 1 or 2 hr before immunization with DNP-hemocyanin, their otherwise normal PFC response to this thymic dependent antigen is virtually abolished. As little as 10 ng of DNP-Ficoll can profoundly suppress the fourth day, direct PFC response to 50 µg of DNP-hemocyanin. Hence, the B cell defect that causes failure of CBA/N mice to respond to DNP-Ficoll is not associated with a failure to recognize this haptenated thymic independent antigen. The CBA/N susceptibility to haptenated Ficoll blockade is hapten specific. A similar type of hapten-specific blockade can be mediated by DNP-conjugated pneumococcal polysaccharide. This susceptibility to hapten-specific polysaccharide-mediated blockade occurs in the male but not the female hybrid offspring of defective CBA/N female mice. Hence, it is transmitted via the X chromosome and is recessive or has reduced penetrance in females. Our findings indicate that this X-linked vulnerability to hapten-specific, polysaccharide-mediated blockade probably represents a separate functional aspect of the basic X-linked recessive CBA/N defect which heretofore was recognized mainly as an inability to respond to most thymic independent antigens.
Impairment of Cell-Mediated Immunity in Mutation Diabetic Mice (db/db)
Authors: Mandel Mark A. , Mahmoud Adel A. F.
Published: December 2022

Mutation diabetes in the mouse occurs in the C57BL/Ks strain. All homozygous animals (db/db) develop obesity, elevated blood sugar levels and increased or normal blood insulin concentration. The defects in cellular immunity in db/db mice and their littermate controls were examined both in vivo and in vitro. Significant suppression of delayed footpad swelling and first and second set skin allograft rejection time were observed. In addition, DNA synthesis in spleen cells after nonspecific mitogen stimulation was markedly inhibited. Diabetic animals with a mean blood sugar of 512 ± 101 mg/100 ml did not respond to exogenous insulin therapy by lowering their blood sugar levels or reversing the defect in DNA synthesis. Adding insulin to spleen cell culture in vitro had no demonstrable effect on their response to mitogens. Thus, mutation diabetic mice with their known defect in the peripheral utilization of insulin have markedly suppressed cell-mediated immune mechanisms.
The Subclass Specificity for the Binding of Murine Myeloma Proteins to Macrophage and Lymphocyte Cell Lines and to Normal Spleen Cells
Authors: Segal David M. , Titus Julie A.
Published: December 2022

Murine myeloma proteins of the four IgG subclasses and of the IgA and IgM classes have been examined for their ability to inhibit the binding of affinity cross-linked trimeric rabbit IgG to various types of murine cells. In addition, proteins of the IgG1, IgG2a, and IgG2b subclasses have been cross-linked with dimethyl suberimidate and the purified oligomers have been tested for direct binding. With a macrophage line (P388D1), a lymphocyte line (AKTB-1), and normal spleen cells, proteins of the IgG1, IgG2a, and IgG2b subclasses all compete for binding at the same site. Differences in binding affinities between proteins of the IgG1, IgG2a, and IgG2b subclasses are small and are not greater than differences in affinities when proteins within a subclass are compared. Proteins of the IgG3 subclass and the IgM and IgA classes do not significantly inhibit IgG binding at the highest concentrations tested.

P388D1 cells are also able to bind iodinated monomeric IgG2a proteins with higher affinity than expected from inhibition studies. This binding is inhibitable by proteins of the IgG1, 2a, and 2b subclasses, and is trypsin sensitive. These results could be explained by the existence of a second type of receptor, or by assuming that the iodination procedure increases the affinity of IgG2a proteins for P388D1 cells.
Studies on Cytotoxicity Generated in Human Mixed Lymphocyte Cultures
Authors: Seeley Janet K. , Golub Sidney H.
Published: December 2022

Human mixed lymphocyte cultures (MLC) have been observed to generate cytotoxicity not only against allospecific targets but also against autochthonous control target cells. This work investigates the types of target cells susceptible to the MLC-activated effectors and also analyzes the time course of the generation of these activities. Normal human peripheral blood lymphocytes (PBL) were sensitized to normal allogeneic mitomycin C-treated PBL in MLC and subsequently were tested for cytotoxicity against various targets in a 51Cr-release cell-mediated lympholysis (CML) assay. Two types of cytotoxic activities were clearly distinguishable by the types of target cells susceptible to the effectors: a) The allocytotoxicity was directed at the relevant allogeneic Con A blasts (alloblasts), but not to the autochthonous control Con A blast (autoblast) targets. b) In contrast, the “anomalous” cytotoxicity, which was originally defined as alloactivated cytotoxicity against the autochthonous lymphoblastoid cell line (auto-LCL) target, was shown to have a broad range of cytotoxic activity against all cultured cell line targets tested. The allocytotoxicity was further distinguished from the “anomalous” reactivity by studies on the time of development in culture of these activities. The peak anomalous activity always occurred 1 or 2 days earlier and declined more rapidly than the peak allocytotoxicity. Experiments to find common antigenic determinants between the sensitizing cells and targets susceptible to anomalous cytotoxicity revealed no obvious antigenic relationships.
Activation of Human B Lymphocytes
Authors: Dimitriu Alexandra , Fauci Anthony S.
Published: December 2022

Human monocytes, after in vitro activation by mixed lymphocyte culture (MLC) superantants produce a monokine (MK) that enhances the plaque-forming cell (PFC) response of pokeweed mitogen (PWM)-stimulated human B lymphocytes. Technical conditions and kinetics of MK production were established. Irradiation of monocytes (5000 rads) does not abolish MK production but heat-killed cells are unable to release the factor. Highly T cell-deplated monocyte populations still produced the PFC-enhancing factor. The same MK has an inconsistent enhancing effect on the PFC responses of lipopolysaccharide (LPS) and nocardia water-soluble mitogen (NWSM)-stimulated B cells. Other macrophage activators such as LPS, phytohemagglutinin (PHA), and latex particles failed to induce consistently the liberation of the PFC-enhancing MK. The target cell for the MK activity on PWM-stimulated B cells appears to be the B lymphocyte itself. These studies demonstrate that soluble monocyte products can have substantial modulatory effects on human B cell function.
Hapten-Specific Tolerance Induced by Hapten Conjugates of d-Glutamic Acid, d-Lysine (D-GL) or Isologous γ-Globulin: Evidence for Central B Cell Tolerance in the Presence of Carrier-Primed Helper T Cells
Authors: Katz David H. , Borel Yves
Published: December 2022

A comparison has been made of the well known hapten-specific tolerance systems induced, respectively, by hapten-D-GL or hapten-isologous γ-globulin conjugates. The principal question addressed in this study concerned the comparative maintenance of B cell tolerance, induced by one or the other method, after adoptive transfer into carrier-primed, irradiated recipient animals and, in addition, what role, if any, might be played by T lymphocytes in the tolerant donor cell population in maintaining such tolerance. The results clearly show that insofar as the hapten-specific B cell is concerned, no obvious difference exists in the capacity to maintain tolerance adoptive transfer between the hapten-D-GL and hapten-isologous γ-globulin systems; such cells remained tolerant even in the presence of excess helper T cell activity. Moreover, under the conditions employed, depletion of T lymphocytes from the tolerant donor cell population did not affect the maintenance of hapten-specific B cell tolerance after adoptive transfer to irradiated recipients.
In Situ Lymphoid Cells of Mouse Mammary Tumors
Authors: Blazar Beverly A. , Heppner Gloria H.
Published: December 2022

A method of separating lymphoid cells from solid mouse mammary tumors was developed and evaluated. In this method the tumors are digested with 0.01% collagenase, 0.01% DNAase, and 0.025% trypsin in Dulbecco's PBS into suspensions of cells with a viability of 90%. The suspensions are fractionated on a continuous gradient of Ficoll in tissue culture medium. In model experiments this gradient was found to separate, cleanly, admixed cells of an established mammary tumor cell line and dissociated thymus glands. Recovery rates were 50% for the tumor cells and 80% for the thymocytes. The preparation of the cell suspensions and the gradient separation procedure are not harmful to the cells as indicated by trypan blue exclusion and the ability to grow in cell culture.
In Situ Lymphoid Cells of Mouse Mammary Tumors
Authors: Blazar Beverly A. , Miller Fred R. , Heppner Gloria H.
Published: December 2022

Lymphoid cells were isolated by isokinetic gradient centrifugation from mouse mammary tumors and evaluated for their ability to affect tumor cell survival with the microcytotoxicity assay. In all experiments stimulation of the growth or survival of tumor cells was seen. This stimulation was not seen with separated lymphoid cells treated with anti-lymphocyte serum and complement. The lymphoid cells separated from tumors were more stimulatory than were lymph node cells; even when lymph node cells of the tumor bearer were cytotoxic to the tumor cells, the separated tumor-associated lymphoid cells were markedly stimulatory. The gradient procedure had no effect on the activity of sensitized lymph node cells, but it did separate cytotoxicity and stimulation into different fractions.
Immunosuppressive Activity of Antibody Directed Against Endogenous C-Type Virus Interferes with Early Events of the Immune Response
Authors: Schumann Gebhard , Moroni Christoph
Published: December 2022

We have analyzed the effects of an antiserum prepared against BALB/c endogenous xenotropic C-type virus on the humoral immune response of mice. Both in vivo and in vitro, this serum suppresses the response to sheep red blood cells, an effect that can be absorbed out by purified BALB/c xenotropic C-type virus or Friend leukemia virus, but not by Rous sarcoma virus. The serum produces its maximum effect when administered together with or before the antigen, but not 24 hr later. This suggests that it acts on an early event of the immune response. Evidence is presented to show that the critical viral antigen is expressed before the spleen cells are experimentally stimulated by antigen. The same immunosuppressive effect was observed in a variety of mouse strains, including the high-leukemia incidence AKR strain and virusfree 129/J mice, indicating that it is independent of the expression of endogenous virus. The finding that a viral antigen is involved in the transition from a resting to a dividing lymphocyte is discussed with respect to viral involvement in leukemia.
Effects of Costimulator on Immune Responses in Vitro
Authors: Shaw Jennifer , Monticone Vicky , Mills Gordon , Paetkau Verner
Published: December 2022

We recently described a factor, costimulator, that is required for the concavalin A-induced proliferation of CBA mouse thymocytes in vitro (see Reference 1). Using the costimulator dependence of mouse thymocytes as an assay, we have now determined that spleen cells from congenitally athymic (nude) BALB/c mice do not produce costimulator in response to Con A, and spleen cells depleted of Thy 1-positive cells do not respond to it in the presence of Con A. Thus, costimulator both requires thymus-derived (Thy 1+) lymphocytes for its production and has an effect on this type of cell. (However, the costimulator-producing and responsive cells may be different.) Purified costimulator preparations are a source of the required second component for the stimulation of adult, CBA/J thymic lymphocytes by PHA, normally a poor mitogen for these cells. They also enhance the level of DNA synthesis in a mixed leukocyte reaction, and the specific generation of cytotoxic lymphocytes to allogeneic tumor cells in vitro. Costimulator is not H-2 restricted in its effects, and it is produced in mixed leukocyte reactions. Finally, it has been possible to grow normal, primary thymic lymphocytes in culture for about 20 days by adding partially purified costimulator to the cultures.
Complement-Induced Modifications in Membrane Fluidity: Studies with Resealed and Glutaraldehyde-Treated Erythrocyte Membrane Ghosts
Authors: Giavedoni Elda B. , Mason Ronald P. , Dalmasso Agustin P.
Published: December 2022

We investigated the effect of antibody and complement (C) on the fluidity of the lipid bilayer of resealed membrane ghosts by means of electron spin resonance spectroscopy. In contrast to intact red cells, resealed membrane ghosts do not rupture after treatment with C and thus C-induced changes in membrane properties can be directly ascribed to the action of C. Membranes treated with antibody alone (MA) had no modifications in the fluidity parameters. Treatment of MA with normocomplementemic rabbit serum, but not with C6-deficient rabbit serum, produced a significant increase in T‖' and a decrease in Δl and Δh, which are indicative of a more rigid structure. These modifications were absent from MA treated with the first eight C components (MAC1–8) but became apparent upon addition of C9 to the MAC1–8. Cross-linkage of the membrane proteins with glutaraldehyde increased the rigidity of the lipid bilayer. Membranes pretreated with 0.025% glutaraldehyde still responded to C with an additional increment in rigidity. However, the spin label became insensitive to C when the membranes were pretreated with 0.25% glutaraldehyde. Treatment with glutaraldehyde did not impair the development of C lesions as evaluated by the release of entrapped marker molecules. Therefore, the C-induced changes in membrane fluidity, when observed, are only the concomitant of the C lesion and not an essential component of their mechanism of production.
The Complex of C5b and C6: Isolation, Characterization, and Identification of a Modified form of C5b Consisting of Three Polypeptide Chains
Authors: Yamamoto Ken-Ichi , Gewurz Henry
Published: December 2022

A new method for isolation of the complex of C5b and C6 (C5b-6) which yielded milligram quantities of C5b-6 of high purity was devised. C5 and C6 first were separated from C7 by column chromatography on DEAE-cellulose. C5b-6 then was generated from the C7-free reagent by incubation with serum-treated zymosan, and purified by DEAE-cellulose and Bio-Gel A 1.5 m column chromatography. Upon analysis by sucrose density gradient ultracentrifugation and gel filtration, two forms of the C5b-6 complex were recognized. The first maintained its complexed state (10.4S) during ultracentrifugation, and formed aggregates (>16S) upon reaction with C7. The second and predominant form dissociated into C5b (7.6S) and C6 (5.7S) during ultracentrifugation and did not form aggregates with C7. Both forms retained their elution characteristics on gel filtration over a 1-month period at 4°C. SDS-polyacrylamide gel electrophoretic analysis also revealed two different forms of the C5b-6 complex. C5b-6 capable of aggregating with C7 was a complex of unmodified C5b and C6. The predominant form was, however, a complex of modified C5b (designated C5b′) and C6. In the absence of a reducing agent, both forms of C5b had an apparent m.w. of 180,000 daltons. Upon reduction, C5b yielded two polypeptide chains designated α′ (103,000 daltons) and β (75,000 daltons), whereas C5b′ dissociated into three polypeptide chains, i.e., two fragments of the α′ chain designated α′1 (75,000 daltons) and α′2 (27,500 daltons), and the β chain (75,000 daltons). The possible significance of this proteolytic modification of C5b in relation to the hemolytic efficiency of lysis mediated by C5b-6 is discussed.
T Cell Growth Factor: Parameters of Production and a Quantitative Microassay for Activity
Authors: Gillis Steven , Ferm Mary M. , Ou Winny , Smith Kendall A.
Published: December 2022

Several soluble factors have recently been associated with the proliferation and differentiation of thymus-derived lymphocytes. One of these factors present in medium conditioned by T cell mitogen-stimulated lymphocytes has the ability to promote the long-term culture of normal and antigen-specific cytotoxic T cells. We report a method to test for this proliferative stimulus in the form of a sensitive microassay based upon the tritiated thymidine incorporation of continuous murine tumorspecific cytotoxic T cell lines (CTLL). The microassay requires microliter quantities of sample fluid and is amenable to quantitative analysis. This highly reproducible, quantitative assay for T cell growth factor (TCGF) has allowed investigation as to the kinetics of TCGF generation and has revealed that T lymphocytes are required for its production. Further investigation has supported the notion that this nonspecies-specific factor is actively removed from tissue culture medium by the proliferation of either T cell mitogen-activated lymphocytes or CTLL.
Glucocorticoid Receptors and Glucocorticoid-Sensitive Secretion of Neutral Proteinases in a Macrophage Line
Authors: Werb Zena , Foley Rosemary , Munck Allan
Published: December 2022

A continuous line of mouse macrophages (P388D1) has been shown to secrete elastase, collagenase, and plasminogen activator at activities comparable to those of macrophages elicited by an inflammatory stimulus in vivo. At physiologic concentrations anti-inflammatory glucocorticoids selectively and reversibly inhibited secretion of the three proteinases but did not inhibit secretion of lysozyme, a constitutive enzyme produced by the P388D1 cells. The secretion of the neutral proteinases was inhibited 50% by 2 to 10 nM dexamethasone. Proliferation of the macrophages was also glucocorticoid sensitive. The P388D1 macrophages contained about 4000 saturable glucocorticoid-binding sites per cell. Concentrations of hormone saturating the high affinity receptor site (for dexamethasone the dissociation constant for steroid-receptor binding, Kd, was 4 nM) correlated well with concentrations inhibiting secretion of the proteinases. Only glucocorticoids and progesterone competed for binding to the specific receptors. Temperature-sensitive translocation of hormone-receptor complexes from “cytoplasm” to nucleus similar to that found with rat thymocytes was demonstrated. Thus, the interaction between glucocorticoids and the P388D1 cell line provides a model for the regulation of macrophage secretion of neutral proteinases under normal and stress conditions.
Development of B Cell Subsets: Effect of Priming on in Vitro TNP-LPS Responsiveness
Authors: Diaz-Espada Fernando , Martinez-Alonso Carlos , Bernabe Rosa R.
Published: December 2022

Mice were injected with DNP-derivatives of thymus-dependent (DNP-OA) and thymus-independent (DNP-Ficoll and TNP-LPS) antigens, and the response to TNP-LPS in vitro was studied after several priming periods. DNP-OA priming decreases the amount of cells responding to TNP-LPS in vitro. In the case of DNP-Ficoll and TNP-LPS-primed cells, there is an initial burst of responsiveness to TNP-LPS, which progressively decreases until an abolishment of TNP-LPS responsiveness is found at day 40 after immunization. The sensitivity to TNP-LPS reappears as new precursor cells differentiate into mature cells. We suggest that B cells progressively gain the capacity to respond to thymus-independent and thymus-dependent antigens (B1-B2 differentiation) and that challenge with a particle antigen increases the ratio of maturation throughtout the pathway.
Polymorphism of Tissue and Serum Amyloid A (AA and SAA) Proteins in the Mouse
Authors: Gorevic Peter D. , Levo Yoram , Frangione Blas , Franklin Edward C.
Published: December 2022

Amino acid sequence studies of the amino terminal 25 residues of amyloid A (AA) protein and the serum precursor (SAA) induced with casein or LPS indicate differences in the sequence at position 6 and significant heterogeneity at several other positions in SAA. These findings suggest that SAA is a polymorphic serum protein and raise the possibility that only certain forms of SAA are processed to the tissue amyloid fibril.