bluuebunny/crossref_metadata_2025 huggingface dataset:
Immunologic Release of β-Hexosaminidase and β-Glucuronidase from Purified Rat Serosal Mast Cells
Authors: Schwartz Lawrence B. , Austen K. Frank , Wasserman Stephen I.
Published: December 2022

The acid exoglycosidases β-hexosaminidase and β-glucuronidase are present in rat serosal mast cells at concentrations of 1.2 ± 0.5 units and 0.24 ± 0.11 units/106 cells, respectively, and are released in a dose- and time-dependent fashion in response to immunologic challenge with rabbit anti-rat F(ab′)2. The sum of the released and residual β-hexosaminidase and β-glucuronidase was no different from that of resting unchallenged cells, thereby indicating that release in physiologic buffer was sufficient for complete bioavailability of the secreted enzymes when assessed with small synthetic substrates. A comparison of the release of the acid exoglycosidases with that of histamine yields regression lines that have correlation coefficients of ≥ 0.97 for each enzyme and that intersect at the point of origin, suggesting absence of a threshold effect on the release of these mediators. The slopes of these regression lines indicate that at least 86 and 65% of the cellular content of β-hexosaminidase and of β-glucuronidase, respectively, are available for immunologic release and are therefore localized in mast cell secretory granules. The predominant isomeric form of β-hexosaminidase in the rat mast cell, as indicated by its chromatographic behavior, is type A and the A isomer is the predominant form released on immunologic activation of the cells. The only isomeric form of β-glucuronidase identified in rat serosal mast cells based upon its electrophoretic mobility is the lysosomal form, and this form was the only one released after immunologic activation of these cells. The mast cell secretory granule, based upon its content of acid hydrolases and its capacity to form phagolysosomes, is considered a modified lysosome, which also contains novel chemical mediators not generally observed in lysosomes of other cells.
Age-Related Changes in Suppressor Functions of Human T Cells
Authors: Kishimoto Susumu , Tomino Shinhachiro , Mitsuya Hiroaki , Fujiwara Hirokazu
Published: December 2022

Functions of both naturally occurring suppressor and Con A-activated suppressor cels were compared among newborns, young adults, and aged subjects. When T cells were irradiated to remove suppressor activity and added to PWM-stimulated allogeneic B cell cultures, an enhanced Ig production resulted. This irradiation-induced enhancement for IgG production was significantly lower in aged group than in newborns or young adults (p ≦ 0.01). With regard to IgM production, the enhancement was significantly depressed in aged subjects as compared with newborns (p = 0.05). The radiation dose for the generation of T cells capable of exerting the greatest enhancement was between 900 and 1200 rads in both young adults and aged subjects, but as high as 1600 rads in newborns. Furthermore, newborns were shown to be enriched for T suppressor activity by co-culturing of allogeneic PBL with T cells in the presence of PWM as compared with young adults.

T cells isolated from young adults, but not from aged subjects, enhanced IgM and IgG production by allogeneic B cells beyond the synergy obtained by control T cells when pretreated with mitomycin and then co-cultured with B cells and PWM. The means of maximal enhancements were significantly greater in young adults than in the aged (p = 0.02 for IgG, p < 0.05 for IgM).

The study on the kinetics of generation of Con A-activated suppressor cells revealed that T cells isolated from young adults mediated maximal suppression of Ig production when activated by incubation with Con A for 2 days and added to PWM-stimulated PBL cultures. By contrast, T cells from aged humans required 1 or 2 more days of incubation with Con A to be activated to obtain the maximal suppressor activity. In addition, the means of maximal suppression of IgM and IgG production mediated by Con A-activated T cells were significantly greater in young adults than in the aged (p = 0.02 for IgG and p = 0.05 for IgM).

Impaired function of both naturally occurring suppressor and Con A-activated suppressor cells is discussed in relation to altered immunocompetence seen in aged humans.
Generation of H-2 Restricted Cytotoxic T Cells by Trinitrophenylated Proteins in Vitro: Specificity and Requirements
Authors: Ballas Zuhair K. , Henney Christopher S.
Published: December 2022

Addition of TNBS-conjugated soluble proteins to cultures of murine spleen cells, of the H-2k haplotype, led to the generation of MHC-restricted cytotoxic T cells. This phenomenon was effected by a wide range of proteins including ovalbumin, bovine γ-globulin, bovine serum albumin, human γ-globulin, human serum albumin (TNP-HSA), and keyhold limpet hemocyanin. Cytotoxic T lymphocytes (CTL) generated in such manner showed specificity for the hapten rather than for the protein carrier, in that they lysed cells bearing hapten coupled to either heterologous or homologous carriers.

Spleen cell populations incubated with soluble HSA substituted to various degrees (8–40 moles TNP/mole HSA) displayed TNP on their surface as evidenced by a) C-mediated lysis, and b) spectrophotometric measurement of solubilized cell membranes. However, only those cells bearing HSA substituted at more than 26 moles TNP/mole HSA were susceptible to lysis by hapten-specific H-2 restricted CTL. Furthermore, TNP-HSA with lower substitution (<26 mole/mole) did not stimulate hapten-specific cytotoxic responses.

These observations illustrate several features of target cell recognition by hapten-specific, H-2 restricted, cytotoxic T-cells: i) These CTL do not show specificity to the protein carrier. In this regard cytotoxic T cells differ from T cells responsible for delayed-type hypersensitivity (DTH). ii) Hapten and H-2 need not be covalently linked. iii) Hapten display on the cell surface, even in relatively large amounts, is not in itself sufficient. Rather, it seems that some stringent requirements of protein derivatization and/or distribution have to be met irrespective of whether the protein substituted is intrinsic to the cell membrane or exogenously added. These findings represent a reproducible system for further investigation of the exact molecular nature of the antigenic determinants recognized by CTL.
Rapid Magnetic Purification of Rosette-Forming Lymphocytes
Authors: Owen Charles S. , Winger Larry A. , Symington Frank W. , Nowell Peter C.
Published: December 2022

High gradient magnetic separation, which has previously been shown effective in extracting erythrocytes from a flowing cell suspension, has been used to separate rosetted and unrosetted human peripheral blood lymphocytes. The hemoglobin in the sheep red cells used to form rosettes was first oxidized to the paramagnetic methemoglobin form. Samples of 50 × 106 lymphocytes could be processed in 10 min under sterile conditions with greater than 90% purity of the rosetted cell fraction and maintenance of T cell function in mixed lymphocyte cultures.
Functional Characterization of Mouse T Lymphocytes with IgM-Fc Receptors
Authors: Andersson Birger , Skoglund Ann-Charlotte , Rosen Anders
Published: December 2022

The function of mouse T lymphocytes with receptors for IgM-Fc was analyzed by using immunofluorescence and a column separation method. The cellular uptake of FITC-IgM was inhibited with unlabeled Fc5 µ. IgM-FcR+ cells were active in ADCC against IgM-sensitized target cells, but not against IgG-coated target cells. The cells mediating ADCC with IgM antibody were shown to be distinct and physically separable from those mediating ADCC with IgG antibody. Helper T lymphocytes for humoral antibody formation in a hapten-carrier system were shown to be IgM-FcR+ and Fc-IgG-.
Immunotherapy of a Murine Ovarian Carcinoma with Corynebacterium Parvum and Specific Heteroantiserum
Authors: Bast Robert C. , Knapp Robert C. , Mitchell Agota K. , Thurston Joan G. , Tucker Robert W. , Schlossman Stuart F.
Published: December 2022

Immunotherapy with a combination of Corynebacterium parvum and specific heteroantiserum is significantly more effective than treatment with either single agent in prolonging the survival of mice that have received an i.p. injection of syngeneic murine ovarian carcinoma (MOT) cells. In vitro, a combination of C. parvum-activated peritoneal cells and specific heteroantiserum has proven significantly more effective than either single component in destroying 51Cr-labeled MOT cells in the absence of complement. Activation of peritoneal cells to produce lysis of tumor in the presence of specific antiserum peaked 3 to 7 days after a single injection of C. parvum and declined to baseline over 3 to 4 weeks. With repeated i.p. injections of C. parvum at appropriate intervals, activation of peritoneal cells could be prolonged and augmented. Among the routes tested, only i.p. administration of C. parvum was effective, although activation of peritoneal cells for cooperation with heteroantiserum was observed over a broad range of i.p. dosage (0.45 to 4.20 mg). These data suggest that the administration of C. parvum by appropriate doses, routes, and schedules can attract and activate a population of peritoneal effectors that mediate antibody-dependent cytotoxicity more effectively than resident peritoneal cells.
Differential Growth of Allogeneic Bone Marrow and Leukemia Cells in Irradiated Guinea Pigs
Authors: Bhan Atul K. , Kumar Vinay , Bennett Michael
Published: December 2022

Growth of normal bone marrow and L2C leukemia cell grafts was studied in lethally irradiated strain 2 and strain 13 guinea pigs by measuring 125IUdR uptake in the spleen and bone marrow and by histologic examination of these tissues. Allogeneic bone marrow cells proliferated as well as syngeneic cells in both strain 2 and 13 animals. Since strains 2 and 13 guinea pigs differ only at the I region of the major histocompatibility complex, this observation indicates that Ia disparities are not relevant to marrow graft rejection in the guinea pig.

Both Ia positive and Ia negative L2C leukemia cells of strain 2 origin grew well in the spleen of irradiated strain 2 animals. However, irradiated strain 13 animals showed considerable resistance to the growth of both leukemia cell lines. F1 hybrids (2 × 13) also showed resistance to the growth of the leukemia cells, although resistance was less than in strain 13 animals. These observations suggest the existence of an effector system capable of mediating natural resistance to L2C cells in unimmunized strain 13 and F1 guinea pigs. The nature of antigens recognized by these radiation resistant effector cells are not entirely clear. However, Ia antigens, or tumor-associated antigens dependent upon Ia antigens for immunogenicity, do not seem to be the primary targets in this phenomenon.
Avian Antigen-Binding Cells
Authors: Higgins Gloria , Choi Yong Sung
Published: December 2022

Antigen-binding cells (ABC) from spleens of HGG-immunized, bursectomized agammaglobulinemic (Bx) chickens were detected by direct autoradiography with 125I-HGG and by sandwich autoradiography with HGG plus 125I-goat-anti-HGG. The specificity of antigen binding was demonstrated by 1) inhibition of binding of 125I-HGG by preincubation with unlabeled HGG and 2) a specific increase in ABC after immunization. The ABC from Bx chickens were not B cells, as shown by the virtual absence of immunoglobulin-bearing cells in this population and by the lack of inhibition of antigen binding by anti-immunoglobulin sera. The ABC were not macrophages and did not bind HGG via Fc receptors because their frequency was unchanged after passage over nylon wool or incubation with antigen-antibody complexes. The temperature dependence and azide stabilization of the ABC were characteristic of antigenbinding T cells. Therefore, T cells capable of binding soluble antigen were demonstrated in Bx chicken spleen, which is free of contamination by B cells and passively adsorbed antibody.
Prebursal Stem Cells in the Intraembryonic Mesenchyme of the Chick Embryo at 7 Days of Incubation
Authors: Lassila Olli , Eskola Jussi , Toivanen Paavo
Published: December 2022

Cyclophosphamide-treated 18-day-old chick embryos were transplanted with cells from 7-day intraembryonic mesenchyme; the recipients and donors were identical at the major histocompatibility locus. At the age of 35 days, the cell recipients were studied to determine the reconstitution capacity of the transplanted cells. The transplantation resulted in a complete restoration of IgM and IgG class antibody production against human gammaglobulin and Brucella abortus, and of microscopic morphology of the bursa of Fabricius and of the germinal center formation in the spleen. These findings demonstrate that 7-day intraembryonic mesenchyme of the chick embryo harbor prebursal stem cells. These findings confirm our previous observations in the yolk sacembryo chimeras indicating that lymphoid stem cells originate in the intraembryonic hematopoietic sites.
Inhibition of Autoimmune Tubulointerstitial Nephritis in Guinea Pigs by Heterologous Antisera Containing Anti-Idiotype Antibodies
Authors: Brown C. Alan , Carey Kathleen , Colvin Robert B.
Published: December 2022

Strain XIII guinea pigs immunized with rabbit tubular basement membrane (TBM) develop a severe tubulointerstitial nephritis mediated by anti-TBM antibodies. Experiments were performed to determine whether heterologous antibodies to the anti-TBM antibodies (anti-idiotypes) could suppress this autoimmune disease. Anti-TBM antibodies were eluted from strain XIII kidneys with anti-TBM disease and were partially purified by ion exchange chromatography. Antisera to various fractions (I, IV, VI; corresponding to IgG2, IgG1+2, and IgG1, respectively) were raised in rabbits. The antisera were repeatedly absorbed with normal strain XIII serum to remove reactivity to normal IgG. Ten milliliters of the absorbed rabbit antisera or control sera were given i.p. to strain XIII animals at the same time they were sensitized with rabbit TBM and ovalbumin in complete Freund's adjuvant. After 14 days, control animals immunized with TBM, but given either no serum, normal rabbit serum, or antiserum raised against eluates from normal strain XIII kidneys, developed severe tubulointerstitial nephritis with 84.4 ± 5.8, 64.6 ± 9.8, and 96.2 ± 3.5% of the renal cortex involved, respectively. Animals given the anti-idiotype sera had significantly less disease, with 12.6 ± 8.3 (VI), 16.5 ± 5.1 (IV), and 31.2 ± 10.4% (I) of the cortex affected. These animals also had significantly reduced levels of anti-TBM antibodies as measured by a solid phase radioimmunoassay. The antibody response to ovalbumin and tuberculin skin test reactions were not affected. A radioimmunoassay was developed to determine whether the most active rabbit antisera (VI) had anti-idiotypic activity. These sera bound specifically to the iodinated fraction VI anti-TBM antibody eluate, and pre-absorption of the fraction VI with either anti-guinea pig IgG or by guinea pig TBM removed the reactivity with the rabbit antisera.

These results indicate that specific inhibition of an autoantibody response and its consequent disease can be achieved by means of antisera raised to the autoantibodies. The mechanism has not been established, but it is believed to be due to idiotype suppression. A similar approach might prove suitable in other autoimmune diseases.
Specific Stimulation and Suppression of a Primary in Vitro Plaque-Forming Cell Response by Monovalent Lipid Haptens in Fluid Liposomal Membranes
Authors: Humphries Gillian M. K.
Published: December 2022

Multilamellar, “fluid” liposomes containing an appropriate concentration of a synthetic dinitrophenylated lipid hapten behave as T-independent (TI), adherent cell-dependent antigens for primary, in vitro, stimulation of hapten-specific IgM plaque-forming cells (PFC). The appropriate concentration of hapten is approximately 1 mol% with respect to the other liposomal lipids, cholesterol, and dimyristoylphosphatidylcholine (DMPC). At a higher hapten concentration, 4 mol%, liposomes will suppress the anti-dinitrophenyl PFC response to co-cultured immunogenic liposomes, but not the anti-sheep cell response to co-cultured sheep cells. This suppression is also TI. The ability of “fluid,” haptenated liposomes specifically to stimulate or to suppress the TI PFC response is determined by the ratio of hapten to other lipids in a manner that is independent of the total hapten concentration in culture.
Inhibition of the in Vitro Outgrowth of Epstein-Barr Virus-Infected Lymphocytes by TG Lymphocytes
Authors: Shope Thomas C. , Kaplan Joseph
Published: December 2022

Human T lymphocyte subpopulations from subjects not acutely infected with EBV have been selected and examined for ability to inhibit the outgrowth of autologous B lymphocytes that have undergone in vitro infection with EB virus. The results show that only TG lymphocytes are inhibitory. TG lymphocytes from subjects who are EBV antibody-negative inhibit as well as those from subjects who have EBV antibody. TG lymphocyte populations, as well as other T cell fractions obtained from neonatal subjects, fail to inhibit the outgrowth of infected, autologous lymphocytes under the conditions tested. We propose that NK cells are responsible for the inhibitory effects described in this report.
Local Xenogeneic Graft-Vs-Host Reaction: A Practical Assessment of T Cell Function Among Cancer Patients
Authors: Mavligit Giora , Ishii Yoshifumi , Patt Yehuda , Wong Wendy
Published: December 2022

The local xenogeneic graft-vs-host reaction (XGVHR) was used as a practical bioassay to assess T lymphocyte function and immunocompetence among cancer patients. Positive XGVHR was found in 99.5% of normal donors, 70% of cancer patients with early stage disease, and 30% of cancer patients with metastatic disease (p < 0.001). A minimum of 4.5 × 106 immunocompetent T lymphocytes are necessary in order to elicit a positive XGVHR. Negative reactions among cancer patients are characterized by the lack of edema fluid accumulation and the appearance of the host basophils at the test site. This suggests that insufficient amounts of lymphokines are being released by the incompetent T lymphocytes, whereas the host is capable of mounting a rejection reaction as evidenced by the appearance of the basophils. Preliminary evidence suggests that the immunologic defect detected by the XGVHR cannot be corrected by monocyte depletion. The identification of putative suppressor T cell subsets may bear immunotherapeutic implications in the future.
Humoral and Cellular Regulation of Alloimmunity in Pregnancy
Authors: Pavia Charles S. , Stites Daniel P.
Published: December 2022

By using the cell-mediated lympholysis (CML) test, it was found that lymphoid cells from pregnant BALB/c females (gravid by C57BL/6 males) were not directly cytotoxic for 51Cr-labeled target cells posessing the same haplotype as the paternal parent. During various stages of gestation and degrees of parity, splenic lymphocytes from interstrain (BALB/c × C57BL/6) pregnant mice were equivalent to those from virgin BALB/c females in their in vitro proliferative responses to paternal strain cells (C57BL/6) in the one-way mixed lymphocyte reaction (MLR) and also in their ability to develop in vitro into cytotoxic effector cells toward paternal strain alloantigens. Suppressor activity on the part of splenocytes from pregnant mice could not be detected in the MLR and CML assay. Heat-inactivated serum from allogeneically pregnant BALB/c mice, when compared to serum from virgin or syngeneically pregnant females, regularly depressed both proliferative and effector phases of the MLR between related parental strain cells. The degree of inhibition was related to the final concentration of pregnant serum in culture with 1 to 3% serum producing greater than 75% inhibition. The inhibitory serum was not cytotoxic by using a sensitive 51Cr release assay, and the suppressive effect was associated with histocompatibility differences between the BALB/c and C57BL/6 mating combination. Thus although pregnancy across strong H-2 differences acts as a poor in vivo inducer of the effector component of cellular immunity, the in vitro proliferative functions of spleen cells from pregnant mice are intrinsically normal, but may be modulated by factors present in pregnancy serum.
On the Interaction of Rabbit C1q with Sheep and Rabbit Immune Complexes
Authors: Liberti Paul A. , Baillie Richard D. , Milligan Kathleen F. , Bausch Dulcie M.
Published: December 2022

The reversibility of interaction between rabbit C1q and a calcium dependent antigen-antibody system [GAT-anti-GAT-Ca+2], as well as reaction parameters thereof, have been carefully examined. By utilizing the reversibility of the GAT-anti-GAT-Ca+2 reaction by the addition of EDTA, it has been found that 1) C1q-IgG interaction is totally reversed by the disruption of the immune complex, independent of antigen-antibody ratios, 2) the amount of C1q bound to immune complexes mirrors antibody precipitated in the precipitin reaction, 3) the amount of C1q bound to “pre-formed” immune complexes is almost identical to that amount bound when immune complexes are formed in the presence of C1q, i.e., “dynamic complexes,” 4) C1q specifically bound to immune complexes can rebind as many as three times without loss of binding activity, and 5) C1q multiply reacted with immune complexes suffers no loss of hemolytic activity. Items 4 and 5 provide strong evidence that C1q remains native upon binding and that the labile nature of C1q is probably not related to its triggering actions in the complement phenomenon. From quantitative binding studies of C1q with pre-formed immune complexes formed at antibody excess, equivalence and antigen excess, the shapes of the binding curves and Scatchard plots are not suggestive of co-operative interactions between C1q and IgG. Association constants obtained with sheep or rabbit pre-formed immune complexes were significantly greater in antibody excess than at equivalence, as were amounts of C1q bound. For dynamic immune complexes formed in the presence of C1q, more C1q is bound at equivalence. Since more C1q was found to bind to these immune complexes at equivalence than in antibody excess, i.e., similar to an in vivo situation, it appears that quantities of C1q bound, rather than the energy of interaction, may be more related to the degree of complement fixation.
The Cellular Mechanism of Leukocyte Adherence Inhibition
Authors: Koppi Thelma A. , Maluish Annette E. , Halliday William J.
Published: December 2022

Human blood leukocytes from three subjects who had been contact sensitized to dinitrochlorobenzene were used in direct and indirect leukocyte-adherence-inhibition (LAI) reactions in an attempt to elucidate the cellular mechanism of reactivity. The leukocytes were separated and purified by standard procedures. In direct LAI, only T cells or populations containing T cells gave positive reactions (significantly reduced adherence) with the antigen. Supernatants from suitable leukocyte-antigen mixtures contained a soluble leukocyte-adherence-inhibition-factor (LAIF) that reduced the adherence of normal leukocytes. Only T cells or populations containing T cells were active in LAIF production; B cells, granulocytes, and monocytes were inactive. The cellular requirement for the action of preformed LAIF was not restricted: all major types of blood leukocytes were susceptible to its effect.
Response of a Continuous Macrophage-Like Cell Line to Mif
Authors: Newman Walter , Diamond Betty , Flomenberg Phyllis , Scharff Matthew D. , Bloom Barry R.
Published: December 2022

A cloned murine macrophage-like cell line, FC-1, which arose during fusion of a drug-marked myeloma cell line with primary splenic cells was shown to migrate well from capillary tubes both in medium containing 10% horse serum or in serum-free medium containing 0.5% BSA. This cell line was responsive to MIF produced by mouse spleen cells stimulated with either insolubilized Con A or with alloantigens.

Bio Gel fractionation of MIF supernatants showed the same samples to contain inhibitory material for both FC-1 cells and for primary macrophages and that this product had a m.w. of 25,000 to 50,000.

Because the migration of FC-1 was inhibited by MIF prepared and tested in serum-free medium or in plasminogen-depleted serum, migration inhibition must result from the direct interaction of MIF with macrophages rather than indirectly through an activation of serum enzymes. This was confirmed when primary macrophages that require serum were shown both to migrate well and to respond to MIF in plasminogen-depleted serum.

Because of the homogeneity of the cloned FC-1 cell line and its ability to migrate and respond to MIF in serumfree medium, this macrophage-like cell line offers unique advantages for studying the regulation of macrophage functions by products of activated lymphocytes.
Inhibition of Specific Immune Responses by Feeding Protein Antigens
Authors: Miller Stephen D. , Hanson Donald G.
Published: December 2022

We studied the effect of a single intragastric administration of ovalbumin (OVA) on the subsequent development of OVA-specific cell-mediated immune (CMI) responses in BDF1 mice. In animals fed OVA 7 days before subcutaneous sensitization with OVA-CFA, we observed a concomitant dose-dependent decrease in both the humoral and CMI responses specific for OVA. The CMI tolerance was found to be antigen-specific when assayed in vivo by ear swelling or in vitro by an antigen-induced T cell proliferation assay because OVA-fed mice responded normally to sensitization with horse γ-globulin. It was also shown that either spleen or lymph node cells, but not serum, from OVA-fed donors transferred suppression to normal recipients. The transfer was mediated by antigen-specific suppressor T cells (Ts) that appeared to inhibit the induction phase (afferent limb) of the CMI response, since the Ts were only effective when transferred before or shortly after the onset of sensitization.
Biosynthesis of a Single Chain PRO-C5 by Normal Mouse Liver mRNA: Analysis of the Molecular Basis of C5 Deficiency in AKR/J Mice
Authors: Patel Feisal , Minta Joe O.
Published: December 2022

An in vivo labeling technique has been used to probe the molecular lesions precipitating in C5 deficiency in the AKR/J mouse. 14C-labeled amino acids were administered i.p. into normal and C5-deficient mice, and the plasma was harvested 4 hr later. By using monospecific anti-C5, newly synthesized 14C-C5 was immunoprecipitated from the plasma and postmitochondrial supernatants (PMS) of a liver homogenate. By SDS-PAGE analysis it was demonstrated that normal mouse plasma (apparent m.w. 205,000) was composed of two dissimilar subunits, an α-chain (115,000 daltons) and a β-chain (82,000). However, nonsecreted C5 immunoprecipitated from the PMS was resolved into two nonreducible polypeptide chains of m.w. 200,000 and 170,000, respectively. By comparison to plasma C5, the 170,000 dalton peak polypeptide chain probably represents incompletely synthesized, partially degraded, or unglycosylated pro-C5. 14C-C5 immunoprecipitates from the plasma, and the PMS of AKR/J C5-deficient mice contained insignificant radioactivity and on SDS gels did not resolve into any distinct peaks, suggesting that C5 is not synthesized in this strain.

14C-C3 immunoprecipitated from the plasma of normal and AKR/J mice in each case was composed of covalently-linked α- and β-chains (m.w. 130,000 and 85,000, respectively), 14C-C3 immunoprecipitated from the PMS of normal and C5-deficient liver homogenates in each case migrated on SDS gels as a single polypeptide chain, pro-C3 (m.w. 200,000).

Confirmation of these findings have been achieved by cell-free translation studies. Poly(A)-mRNA isolated from normal mouse liver stimulated the incorporation of 3H-leucine into protein in a time-dependent fashion in a reticulocyte lysate system under optimal conditions. 3H-C5 immunoprecipitated from the translation reaction mixture behaved as a single nonreducible polypeptide chain (m.w. 170,000). Poly A-mRNA from the liver of the AKR/J mouse displayed similar kinetics and dose-response stimulation of protein synthesis upon translation in the cell-free system, but failed to direct the synthesis of C5 or C5 immunoreactive peptides, although C3 was synthesized normally as pro-C3. Since the intact machinery for carbohydrate synthesis is not present in the reticulocyte cell-free system, it is envisaged that the 170,000-dalton C5 polypeptide chain is possibly unglycosylated pro-C5. Thus, C5 is synthesized as a single-chain pro-C5 and post-translationally converted to a two-subunit C5 molecule by limited proteolysis. In the AKR/J C5-deficient mouse C5 is not synthesized at all, suggesting the lack of a functional mRNA for C5 in this strain.
Activation of Murine Complement by Cobra Venom Factor in the Presence of EDTA
Authors: Goldman John N. , Bangalore Shrikar , Goldman Margaret B.
Published: December 2022

The anticomplementary factor of cobra venom (CoVF) was found to activate the terminal complement (C) components (C5 through C9) in mouse or rat serum or plasma in the presence of EDTA. The same concentrations of EDTA would block C activation by EA, zymosan, and inulin and would block C activation in human and guinea pig serum by all of these activators, including CoVF. Murine serum was found to contain two factors that allowed CoVF-induced C activation and bystander hemolysis in the presence of EDTA. The first was factor B or a related component of the alternative pathway, which allowed formation of a functionally active complex of CoVF and factor B in the presence of EDTA. The second was murine C5, which led to augmented hemolysis of unsensitized bystander erythrocytes and activation of the other terminal components. Preformed complexes of CoVF and rat factor B were active in leading to C5 and C6 consumption in human or guinea pig serum that had been fortified with rat C5. In contrast to native CoVF, these complexes did not lead to electrophoretic conversion of C3 in the human or guinea pig serum. It was further shown that in the presence of rat C5, purified CoVF led to moderate activation of the terminal components in human serum-EDTA in the absence of electrophoretic conversion of C3, indicating that in these situations, CoVF activated C5 and the other terminal components with little or no utilization of C3.
A Monoclonal Antibody to Viral Glycoprotein Blocks Virus-Immune Effector T Cells Operating at H-2Dd but Not at H-2Kd
Authors: Frankel Mark E. , Effros Rita B. , Doherty Peter C. , Gerhard Walter U.
Published: December 2022

A particular monoclonal antibody that binds to the influenza virus HA molecule inhibits HA-specific thymus-derived lymphocytes mediating cytotoxicity in the context of H-2Dd but not of H-2Kd. Another monoclonal antibody blocks both sets of HA-specific effector T cells. This observation, together with related findings from other laboratories, is considered to support the idea that T cell recognition is directed against some association of viral and H-2 glycoproteins, as proposed in the original formulation of the “altered self” concept.
Ecto-5′-Nucleotidase Activity in T and B Lymphocytes from Normal Subjects and Patients with Congenital X-Linked Agammaglobulinemia
Authors: Thompson Linda F. , Boss Gerry R. , Spiegelberg Hans L. , Jansen Inga V. , O'Connor Richard D. , Waldmann Thomas A. , Hamburger Robert N. , Seegmiller J. Edwin
Published: December 2022

Ecto-5′-nucleotidase activity was measured in lymphocyte subpopulations isolated from normal subjects and patients with congenital X-linked agammaglobulinemia. B lymphocytes from normal subjects have at least three times more ecto-5′-nucleotidase activity than T lymphocytes. Patients with X-linked agammaglobulinemia have 56% of normal activity in their T cells, and lack a lymphocyte subpopulation high in nucleotidase activity. High activity of ecto-5′-nucleotidase may be a biochemical marker for mature surface immunoglobulin-bearing B cells.
Demonstration in vitro of Cytotoxic T Cells with Apparent Specificity Toward Tumor-Specific Transplantation Antigens on Chemically Induced Tumors
Authors: Chauvenet Paula H. , McArthur Carole P. , Smith Richard T.
Published: December 2022

Chemically induced tumors of mice exhibit apparently unique antigenicity upon syngeneic transplantation into appropriately immunized hosts. An in vitro counterpart of this pattern in terms of specificity has not been reported. Data are presented that demonstrate that immune peritoneal exudates contain cells cytotoxic for the specific immunogen tumor but with rare exceptions, not toward other syngeneic methylcholanthrene-induced fibrosarcomas. Only tumors highly immunogenic by transplantation criteria induce cytotoxic PEC regularly; nonimmunogenic tumors consistently fail to do so. The effector cell responsible is eliminated by pretreatment with anti-Thy.1 but not anti-Ig plus complement. In concomitant experiments, PEC populations cytotoxic in vitro also conveyed adoptive protection against the specific tumor in syngeneic hosts. This in vitro assay appears to provide a tool for studying T cell-mediated cytotoxicity toward a set of unique surface antigens present on chemically induced tumors.
Serum Opsonic Activity in Rodent Malaria: Functional and Immunochemical Characteristics in Vitro
Authors: Hunter Kenneth W. , Winkelstein Jerry A. , Simpson Thomas W.
Published: December 2022

The functional and immunochemical characteristics of serum opsonic activity in rodent malaria were examined in the present study. Schizont- and late trophozoite-enriched populations of Plasmodium berghei-infected red blood cells (IRBC) were isolated on a Ficoll density-gradient and used in an in vitro phagocytosis system composed of serum and monolayer cultures of rat peritoneal macrophages. Hyperimmune serum augmented the phagocytosis of IRBC to a greater degree than did nonimmune serum. When either IRBC or macrophages were pre-incubated with serum, the phagocytosis-promoting factors acted on the IRBC rather than on the macrophages in a manner characteristic of serum opsonins. The opsonic activity was specific for IRBC since noninfected red blood cells were rarely phagocytized and were unable to absorb opsonic activity from serum. The opsonic activity of both hyperimmune and nonimmune sera was heat stable, and unaffected by agents known to inactivate or inhibit complement (cobra venom factor and ethylenediaminetetraacetic acid). Finally, the opsonic activity was identified in preparations of purified IgG isolated from both hyperimmune and nonimmune sera.
Active Suppression of B Lymphocyte Maturation by Two Different Newborn T Lymphocyte Subsets
Authors: Durandy Anne , Fischer Alain , Griscelli Claude
Published: December 2022

Leukocytes from newborns and pregnant women were shown to be unable to mature into plasma cells in cultures stimulated by pokeweed mitogen (PWM). This is in contrast with the generation of IgM, IgG, and IgA-containing cells constantly observed in adult leukocytes. This defective B cell maturation was due to an active suppression mediated by T lymphocytes. T lymphocytes bearing receptors for IgG (Tγ) and Tγ-depleted populations were both able to exert this effect. Tγ lymphocytes inhibited B cell maturation without significantly affecting T and B cell proliferation, and their effect persisted after an incubation of 18 hr at 37°C. Tγ-depleted lymphocytes suppressed lymphocyte proliferation and, subsequently, B cell maturation into plasma cells. The suppressor activity of this second cell type was markedly diminished after incubation. Co-culture experiments showed that PWM-induced newborn suppressor T lymphocytes suppressed maturation of adult B cells only when T lymphocytes were present, suggesting that suppressor cells acted upon a target T cell population rather than directly upon B cells. Indirect inhibition of B cell maturation resulted in a lack of appearance of membrane IgG and IgA expression, although proliferation and loss of membrane IgD were observed. Although similar suppressor effects can be induced in vitro in adult leukocytes by fixation of immune complexes on Tγ populations or after concanavalin A activation of Tγ-depleted populations, the peculiarity of the perinatal period appears to be the spontaneous activation of the two types of T suppressor lymphocytes.
Immune Suppression in Vivo with Antigen-Modified Syngeneic Cells
Authors: Sherr David H. , Heghinian Krikor M. , Benacerraf Baruj , Dorf Martin E.
Published: December 2022

Intravenous administration of syngeneic spleen cells coupled with the palmitoyl derivative of fowl γ-globulin (p-FγG) results in a profound state of FγG carrier-specific tolerance in C57BL/6 mice. This nonresponsiveness can be attributed to two independent mechanisms, i.e., T cell tolerance and T cell suppression. The two phenomena could be distinguished on the basis of kinetics of induction, inducer cell dose requirements, and cyclophosphamide sensitivity. Thus, a 3-day period is required for the induction of T cell tolerance, and 7 days are required to induce immune suppression. Furthermore, at least 106 p-FγG-coupled cells must be administered to induce tolerance, whereas suppression could be induced with as few as 103 cells. In addition, the T cell tolerance was not sensitive to cyclophosphamide pretreatment, whereas the T cell suppression was cyclophosphamide sensitive. In contrast to T cell tolerance, suppressor T cells exerted their effects nonspecifically in the effector phase once they were stimulated with the inducing antigen.

In addition, a state of split responsiveness was demonstrated in animals given a subcutaneous (s.c.) injection of antigen-coupled cells. The s.c. route of induction resulted in a strong state of PFC nonresponsiveness while simultaneously priming for delayed type hypersensitivity (DTH) responses as assayed by footpad swelling. Thus, p-FγG-coupled syngeneic spleen cells are capable of inducing: 1) T cell tolerance, 2) immune suppression, and 3) priming of DTH-reactive cells, depending on the conditions employed.
Ly Phenotypes and MHC Recognition: The Allohelper That Recognizes K or D Is a Mature Ly123 Cell
Authors: Swain Susan L. , Bakke Anthony , English Michele , Dutton Richard W.
Published: December 2022

This paper describes the further characterization of the T cell activity that is induced by allogeneic MHC gene products and provides help to mouse spleen B cells in a primary in vitro response to sheep erythrocytes.

We have previously shown that the helper activity induced by antigens controlled by the K or D regions was dependent on the presence of an Ly123 cell and that a single cell, present in high frequency, was required. In the present paper we show that activity induced by a difference at H-2K is unaffected by adult thymectomy but is diminished by prior injection of anti thymocyte serum. The spleen cell population positively selected for Ly2 (Ly2 and Ly123 cells) and hence depleted of Ly1 cells retained full helper activity. Ly1 cells were inactive. In contrast, helper activity induced by differences at the whole haplotype or at Mls were present in the Ly1 population and reduced or missing from the Ly2 and Ly123 populations. We concluded that the helper activity induced by allogeneic K or D is the function of a long lived recirculating Ly123 cell present in high frequency. We were unable to find any evidence that the Ly123 matured into an Ly1 cell before performing its effector function. The Ly phenotype of the effector cell is thus more closely linked to the MHC subregion that is recognized than the T cell function performed. Possible interpretations of the findings are discussed.
Spontaneous and Induced in Vitro F1 Anti-Parental Cytotoxic T Lymphocyte Responses: Target Specificities of Cytotoxic Effectors
Authors: Ishikawa Hiromichi , Dutton Richard W.
Published: December 2022

In vitro cultures of spleen cells from about 5-week-old (AKR/J♀ × BALB/c) or (BALB/c♀ × AKR/J)F1 mice spontaneously generate CTL that lyse targets from AKR/J, BALB/c, and their F1 but not allogeneic strains. These spontaneous CTL are not generated in cultures of parental spleen cells and are only detectable in spleen cell cultures from F1 mice that were heterozygous at both the MHC and background loci.

The same F1 mice older than 20 weeks of age can respond to parental BALB/c stimulators in an in vitro culture system and generate cytotoxic T lymphocytes (CTL) that lyse BALB/c targets. This response is significantly higher than that seen in unstimulated cultures. The target antigen of these induced F1 anti-BALB/c CTL maps to H-2Dd but not to H-2Kd, and the CTL lyse not only parental targets but also heterozygous F1 targets syngeneic to the responder mice. In contrast, anti-H-2Kk CTL induced in the same adult F1 mice by stimulation with parental AKR/J cells lyse only homozygous H-2Kk targets.

Target antigen mapping as well as cold target competition analyses showed that the target specificities for these spontaneous CTL of the young F1 mice are indistinguishable from those seen for the induced responses of older mice; i.e., one target maps to Kk, and only homozygous Kk targets are lysed; the other maps to Dd, and homozygous Dd and the F1 targets are not distinguished.
Role of Nominal Antigen and Ia Antigen in the Binding of Antigen-Specific T Lymphocytes to Macrophages
Authors: Werdelin Ole , Shevach Ethan M.
Published: December 2022

We have previously demonstrated that when primed T lymphocytes were repeatedly incubated on monolayers of antigen-pulsed macrophages (Mø), the cells that failed to adhere to the monolayer demonstrated a marked depletion of their proliferative response that was specific both for the antigen used for pulsing the Mø and for Ia determinants on the Mø. In order to further analyze the contribution of the nominal antigen and Ia antigens to the physical binding of T lymphocytes to Mø, we have attempted to block the absorption of T lymphocytes to Mø with a large excess of soluble antigen and with anti-Ia sera. Our results demonstrate that anti-Ia sera inhibit but that soluble antigen augments the binding of specific T lymphocytes to Mø. The implications of these findings for “dual recognition” and “linked recognition” models of T lymphocyte receptors are discussed.
Structural Studies on the Murine Ia Alloantigens
Authors: Cook Richard G. , Capra J. Donald , Bednarczyk John L. , Uhr Jonathan W. , Vitetta Ellen S.
Published: December 2022

The α and β subunits of the murine I-A alloantigens from several H-2 haplotypes were examined by comparative tryptic peptide mapping by using double label (3H and 14C) techniques. Significant structural variation between alleles was detected in both subunits. Tryptic digests of the α polypeptides from s, b, and d showed only 65% co-elution with k; β-chains from s, b, d, and r were about 50% similar to the k β subunit. Peptide analysis of the Ak subunits from intra-H-2 recombinant strains indicated that both the α and β polypeptides are encoded within the I-A subregion.
Mechanism of Cell-Mediated Cytotoxicity at the Single Cell Level
Authors: Grimm Elizabeth A. , Thoma John A. , Bonavida Benjamin
Published: December 2022

The kinetics and rate of T cell-mediated cytolysis was assessed by measuring the times required for lysis of isolated target cells by single cytotoxic lymphocytes. Single target cell lysis was determined microscopically by observing trypan blue uptake as a function of time of incubation of effector-target conjugates in agarose. Lysis of EL-4 target cells by alloimmune peritoneal exudate lymphocytes was initiated without a lag and was essentially complete at 2 hr. Both zero-order and first-order kinetics equations were analyzed for fit to the 0 to 2 hr lysis values. Statistically, the zero-order kinetic function could be rejected (p > 0.05), but the first-order kinetics function (p < 0.01) could not. This strong evidence for first-order kinetics of T cell-mediated cytolysis implies that within each CTL-target cell population, cytolysis occurs exponentially as a random decay process and that one event in the entire process of cytolysis is rate limiting.

The first-order equation was then applied to measurements of the rate of cytolysis in many different individual effector-target cell combinations. Significant differences in the lytic rate were apparent when either the effector or target cells were varied, with the rate constants spanning a 5-fold range. The heterogeneity of lytic rates is consistent with the hypothesis that lytic efficiency is a function of both the effector and target cells used.
Brucella Abortus Lipopolysaccharide Is Mitogenic for Spleen Cells of Endotoxin-Resistant C3H/HeJ Mice
Authors: Moreno Edgardo , Berman David T.
Published: December 2022

Preparations of lipopolysaccharide (LPS) from rough and smooth strains of Brucella abortus were mitogenic for spleen cells of athymic nude mice, C3H/HeAU mice, and the endotoxin-resistant C3H/HeJ mice. The mitogenic response induced by crude smooth-LPS (f5) was greater than that produced by purified smooth-LPS (f5p); however, the dose-response curves were similar for both preparations. The mitogenic activity of mouse spleen cells to both f5 and f5p was higher than that produced by stimulation with purified rough-LPS. The dose-response curves with rough-LPS were also qualitatively different from those produced with the preparations of smooth-LPS.
Determinant specific suppression of antigen-induced T cell proliferation in the guinea pig. I. Quantitation of suppressed antigen-specific T cell responses as a consequence of prior exposure to antigen in incomplete Freund's adjuvant.
Authors: Baskin B L , Rosenthal A S
Published: December 2022

We have asked whether a correlation exists between T cell proliferation and the in vivo suppression of delayed type hypersensitivity observed after administration of antigen in incomplete Freund's adjuvant before exposure to antigen in complete Freund's adjuvant. We find that in vivo suppression is indeed paralleled by diminished in vitro responsiveness to the immunogen. Suppression of T cell proliferation is antigen-specific, dependent upon prior immunization of antigen in IFA, and can be transferred adaptively into unprimed but not primed animals by lymphoid cells from actively suppressed syngeneic donors.
Determinant specific suppression of antigen-induced T cell proliferation in the guinea pig. II. Determinant specific suppression of in vitro T cell responsiveness parallels a selective suppression of anti-hapten but not anti-carrier antibody responses.
Authors: Baskin B L , Blake J T , Rosenthal A S
Published: December 2022

Using an antigen of defined physical structure with precisely mapped immunogenic sites, we asked whether those molecular sites previously shown to be critical for immune response gene-mediated initiation of T cell proliferation and T help correspond to the same molecular regions capable of inducing antigen-specific suppression of T cell proliferation and antibody production. Inbred strain 2, 13, and 2 x 13 F1 hybrid guinea pigs were immunized with various species variants or fragments of insulin adjuvant before subsequent immunization with antigen in complete Freund's adjuvant. Analysis of the patterns of depressed T cell responsiveness showed a striking correspondence to the Ir gene-dependent mechanism that controls the recognition of discrete regions within the insulin molecule observed in T cell help in antibody production. In addition, suppression of carrier-specific T cells parallels suppression of anti-hapten antibody responses when hapten is presented on the suppressed carrier without a concomitant suppression of the anti-carrier antibody response.