bluuebunny/crossref_metadata_2025 huggingface dataset:
Oxygen metabolism in cloned macrophage cell lines: glucose dependence of superoxide production, metabolic and spectral analysis.
Authors: Kiyotaki C , Peisach J , Bloom B R
Published: December 2022

The requirements of a cloned macrophage-like cell line, J774.16, for oxygen metabolism, and the nature of the defect in oxidative metabolism in a variant clone derived from it, J774.C3C, were studied. Upon stimulation with phorbol myristate acetate (PMA), the parental clone produced approximately 1 nmol O2-/min/10(6) cells, whereas the variant clone produced no detectable O2- under the same conditions. Sustained O2- production by J774.16 was totally dependent on extracellular glucose; in glucose-free medium, the cells initiated O2- production but could not sustain it. When cells were stimulated with PMA, glucose-C-1 oxidation of J774.16 cells increased 20-fold while that of J774.C3C remained at resting levels. O2- production in J774.16 cells was inhibited by some agents known to block mitochondrial electron transport before coenzyme Q, such as rotenone and tetrathiafulvalene, whereas antimycin A enhanced O2- production. A dissociation between O2- production and glucose-C-1 oxidation was observed when J774.16 was treated with certain metabolic inhibitors. Quinacrine, 2,4-dinitrophenol, chlorpromazine, and trifluoperazine inhibited O2- production completely under conditions in which glucose-C-1 oxidation was reduced only by 30%. Rotenone inhibited O2- production with no effect on glucose-C-1 oxidation whereas antimycin A augmented O2- production 50% but inhibited glucose oxidation by 20%. Glucose transport studies, with 2-deoxy-D-glucose, showed that the Km for glucose transport of both clones was about 1 mM, indicating that cells could effectively transport glucose even at low concentrations. The Vmax for glucose transport in both J774.16 and variant J774.C3C cells doubled after PMA stimulation, indicating that the variant was effectively stimulated by PMA, even though O2- was not produced. Similarly, PMA induced protein phosphorylation in both clones. No differences between clones J774.16 and J774.C3C in hexokinase, glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, glutathione reductase, or glutathione peroxidase activities could be found. When dithionite-reduced and -oxidized difference spectra of plasma membranes of these clones were compared, comparable levels of b-type cytochrome were found in both clones. However, CO difference spectra indicated that CO was bound to a b-type cytochrome (presumed to be b-245) in clone J774.16 but not in J774.C3C.(ABSTRACT TRUNCATED AT 400 WORDS)
VL-VH expression by monoclonal antibodies recognizing avian lysozyme.
Authors: Smith-Gill S J , Mainhart C R , Lavoie T B , Rudikoff S , Potter M
Published: December 2022

Seven BALB/c hybridoma antibodies directed against the protein antigen, hen egg-white lysozyme c (HEL), were characterized on the basis of their ability to bind lysozymes from 10 species of birds, and their ability to bind HEL competitively. The hybridomas were separable into three complementation groups based upon competitive interactions. The fine specificities of all antibodies were distinct, but two, HyHEL-8 and HyHEL-10, had very similar and overlapping reactivity patterns. To test the hypothesis that VL-VH pairing correlates with binding specificity, the N-terminal amino acid sequences were determined to identify the VL and VH isotopes (subgroups) of the anti-HEL antibodies. HyHEL-8 and -10 shared the VK23 light chain isotype and nearly identical heavy chains in Kabat subgroup I, whereas the heavy and light chain isotypes of all other antibodies differed from HyHEL-8 and -10 and from each other. The heavy and light chain isotypes expressed by HyHEL-8 and -10 are also expressed by XRPC-25, a DNP-binding myeloma protein that does not bind lysozyme. These results are discussed with respect to the contributions of various genetic sources of structural diversity to antibody functional diversity.
Allogeneic bone marrow transplantation in mice after total lymphoid irradiation: influence of breeding conditions and strain of recipient mice.
Authors: Waer M , Ang K K , Van der Schueren E , Vandeputte M
Published: December 2022

Different groups of C57BL/ka or BALB/c mice received a dose of 34 Gy or 42 Gy of fractionated total lymphoid irradiation (TLI) before bone marrow transplantation with 30 X 10(6) BALB/c or C57BL nucleated bone marrow cells, respectively. BALB/c mice that were not bred in specific pathogen-free conditions before TLI showed a high morbidity and mortality rate after 34 Gy of TLI and allogeneic bone marrow transplantation as compared with BALB/c or C57BL that were bred in pathogen-free conditions before irradiation. Many of the conventionally bred BALB/c mice had clinical and histologic signs of graft-vs-host disease after TLI and allogeneic bone marrow infusion. Although leucocytosis and lymphocytosis and the immunologic competence as measured with in vitro tests were equally depressed after 34 Gy TLI in BALB/c and C57BL mice, chimerism was nevertheless significantly easier to obtain in BALB/c mice. The incidence of chimerism after TLI could be enhanced in C57BL mice by increasing the total radiation dose from 34 to 42 Gy. This augmentation of chimerism was paralleled by the induction of more suppressor cells after 42 Gy of TLI in C57BL mice.
The long-lived medullary thymocyte re-visited: precise quantitation of a very small subset.
Authors: Scollay R
Published: December 2022

The cortical and medullary regions of the thymus are well defined, both histologically and in terms of the phenotype and immunocompetence of the cells found in them. The relationship between the two populations is controversial however, and we still do not know which one contains the immediate precursors of peripheral T cells. Data derived from thymus grafts have suggested that medullary cells remain in the thymus for very long periods (perhaps a year or more) and this has been used as argument against the medulla being the source of significant numbers of emigrating cells. In those experiments, however, the T6 chromosomal marker was used to distinguish graft from host cells, so only cells that were dividing or could be induced to divide were detected, leaving the possibility of a considerable bias in the sample. We have repeated these experiments, but have used donor/host combinations differing at the Thy-1 locus, so that greater than 99% of graft cells could be analyzed. We have shown that medullary cells with a life span of more than a few weeks make up considerably less than 1% of the total thymocyte population, although these long-lived cells appear to be a real and potentially interesting population. Thus a long intrathymic life span is not typical of medullary cells in general and cannot be used as an argument against the medulla as a possible source of emigrant T cells.
The role of T cell subsets in the generation of secondary cytolytic responses in vitro against class I and class II major histocompatibility complex antigens.
Authors: Vidović D , Klein J , Nagy Z A
Published: December 2022

Strain combinations generating cytotoxic T lymphocytes (CTL) specific for a single class I (K or D) or class II (A or E) MHC molecule were set up. The responder cells were separated into Ly subsets (Ly-1+2-, Ly-1-2+, and Ly-1+2+) on day 5 of culture by using lytic or non-lytic selection techniques and monoclonal Ly-specific antibodies. The separated subsets were restimulated on day 8 and tested for secondary CTL activity on day 12. Class II-specific secondary CTL could be generated from all three subsets, whereas class I-specific CTL developed only in the Ly-1+2+ and Ly-1-2+ subsets. The Ly-1+2+ cells underwent a phenotypic shift to Ly-1-2+ by day 12, whereas CTL generated from the Ly-1+2- and Ly-1-2+ subsets retained their phenotype up to the secondary effector stage. The cells separated according to their Ly phenotypes on day 5 were the progeny of unprimed progenitors expressing the same Ly phenotypes. Unprimed Ly-1+2+ cells gave rise to CTL in the absence of the other subsets, while unprimed Ly-1+2- and Ly-1-2+ cells required the help of Ly-1+2+ cells (or soluble factors) during priming to become non-lytic CTL precursors by day 5, and cytolytic cells after restimulation. The Ly-1+2- subset could generate class II-specific secondary CTL only in the absence of the other two subsets. Apparently, alloantigen-primed Ly-1+2+ and Ly-1-2+ cells suppressed the development of cytolytic activity in the Ly-1+2- subset. The combined data provide a comprehensive pathway of CTL differentiation from T cell subsets.
Epidermal Langerhans cells can function as stimulatory cells but not as accessory cells in CTL induction.
Authors: Tsuchida T , Iijima M , Fujiwara H , Pehamberger H , Shearer G M , Katz S I
Published: December 2022

In this study, we determined whether Langerhans cells (LC) can function as both stimulatory and accessory cells (ACC) in alloreactive and syngeneic modified CTL induction in vitro. LC exhibited potent Ia+ stimulatory function, comparable to spleen cells, in both allo-CTL induction and in TNP-modified CTL induction assays. In contrast to spleen cells, however, when LC were syngeneic to the responders in an Ia+-adherent cell-depleted CTL culture, they could not restore the CTL responses. This was due, in part, to the suppression of CTL induction, presumably via secretion of prostaglandin E by epidermal cells (EC), because addition of indomethacin reversed the suppression. Even in the presence of indomethacin, however, LC were unable to exhibit ACC function. Finally, we tested the possibility that both Ia+ and Ia- cells were required to fulfill ACC functions. However, when spleen cells treated with anti-Ia antibody and complement and EC and indomethacin were added to an Ia+-adherent cell-depleted CTL culture, there was no significant restoration of the CTL response. Thus, LC can clearly act as Ia+ stimulatory cells in CTL induction but cannot serve as ACC in CTL induction, whereas Ia+ spleen cells may subserve both functions. These data suggest a functional heterogeneity of Ia+ antigen-presenting cells in spleen and in epidermis.
Immunoglobulin production by human peripheral lymphocytes induced by anti-C3 receptor antibodies.
Authors: Daha M R , Bloem A C , Ballieux R E
Published: December 2022

The exact function of receptors for C3b (CR1), which are present on B lymphocytes, is not clear. The present studies were performed to determine the influence of heterologous anti-CR1 on the production of IgM, IgG, and IgA by human peripheral B lymphocytes in vitro. Anti-CR1, raised in rabbits by immunization with purified erythrocyte CR1, was rendered immunospecific and was converted to F(ab')2 and Fab' by enzyme digestion. Pre-immune F(ab')2 and Fab' served as controls. Normal human blood T lymphocytes and T cell-depleted mononuclear cells were cultured for 6 days in RPMI medium and 10% heated fetal calf serum with a low dose (7 micrograms/ml) of pokeweed mitogen (PWM) alone or with PWM and F(ab')2-anti-CR1. In control experiments the effects of F(ab')2-anti-CR1 without PWM and of pre-immune F(ab')2, Fab'-anti-CR1 and pre-immune Fab' in combination with PWM were also tested. When PWM and F(ab')2-anti-CR1 were present simultaneously, 4.3 X 10(3), 2.5 X 10(3), and 1.8 X 10(3) ng/ml of IgM, IgG, and IgA were synthesized, respectively, in cultures containing 4.5 X 10(5) mononuclear blood cells. All other combinations resulted in synthesis of less than 250 ng/ml of each class of Ig. The presence of purified CR1 in the cultures containing PWM and F(ab')2-anti-CR1 caused a dose-dependent decrease of Ig synthesis. By culturing B cells isolated by E-C3b rosetting with T cells and monocytes, the anti-CR1 responding population was demonstrated to be mainly in the CR1-rich B cell fraction. Fab'-anti-CR1 at high doses was also able to stimulate Ig production in the presence of low concentrations of PWM. These data suggest that triggering of CR1 on B cells results in modulation of antibody production and that triggering of CR1 with more than one antibody is required for an optimal response.
Induction of lymphocyte blast transformation by purified fibronectin in vitro.
Authors: Lause D B , Beezhold D H , Doran J E
Published: December 2022

Purified rat plasma fibronectin (Fn) induces a dose-dependent, nonspecific proliferation of lymphoid cells isolated from spleen and lymph nodes, but has no effect on thymocytes. Proliferation of cells occurred after 4 to 5 days of incubation and was generally 5- to 10-fold greater than control cells cultured without Fn or in the presence of the same concentrations of rat serum albumin. The responding cell appears to be the T cell and requires the presence of adherent accessory cells (macrophages). Although purified T and B cells failed to demonstrate a significant increase in blastogenesis in the presence of Fn, reconstitution of T, but not B, cells with irradiated peritoneal exudate macrophages restored the stimulatory effect of Fn. Furthermore, comparison of the effect of various concentrations of macrophages on the restored T cell response shows that proliferative activity in the presence of Fn is dependent on a critical number of macrophages. Adding higher numbers of macrophages beyond the optimal concentration results in a sharp decline in lymphocyte responsiveness to Fn, although the proliferation of control cells continues to increase at these macrophage concentrations. Macrophages pulsed with Fn for 1 hr failed to evoke an increase in T cell responsiveness after 3 or 5 days of incubation. In addition, incubation of Fn-pulsed T cells with macrophages that had been precultured with or without Fn also failed to result in T cell activation. The effect of Fn on T lymphocyte transformation appears to be mediated indirectly through its interaction with the macrophage, because 125I-Fn, which shows significant binding to peritoneal exudate macrophages, fails to interact with purified T cells. Radiolabeled Fn also demonstrated little or no binding activity for unseparated lymph node cells.
Natural and recombinant Escherichia coli-derived interferon-gamma differ in their reactivity with monoclonal antibody.
Authors: Le J , Rubin B Y , Kelker H C , Feit C , Nagler C , Vilcek J
Published: December 2022

Monoclonal antibody GIF-1 was found to neutralize human natural immune interferon (IFN-gamma), but not Escherichia coli-derived recombinant IFN-gamma. In addition, GIF-1 antibody failed to immunoprecipitate 125I-labeled recombinant IFN-gamma, whereas it precipitated natural IFN-gamma in a concentration-dependent manner. The lack of recognition of recombinant IFN-gamma by antibody GIF-1 may not be due to the absence of the oligosaccharide moiety in the molecules of recombinant IFN-gamma alone, because removal of carbohydrate from natural IFN-gamma by treatment with a mixture of glycosidases did not alter the selective binding of antibody, i.e., deglycosylated and untreated natural IFN-gamma were equally neutralized and immunoprecipitated by GIF-1 antibody. In addition, a minor monomeric component of natural IFN-gamma with the m.w. of 15,500, which apparently lacks carbohydrate, was also recognized by antibody GIF-1. These results suggest that the discriminative recognition of natural and recombinant IFN-gamma by monoclonal antibody GIF-1 may be due to a conformational difference at or near the active regions of natural and recombinant human IFN-gamma molecules.
Induction of interleukin 1 messenger RNA and translation in oocytes.
Authors: Windle J J , Shin H S , Morrow J F
Published: December 2022

Macrophages and P388D1 murine cultured cells produce interleukin 1 (lymphocyte-activating factor) when stimulated with lipopolysaccharide or other agents. Poly A+ RNA extracted from either type of stimulated cells and injected into Xenopus oocytes causes synthesis of material with the biologic and biochemical properties of interleukin 1. It potentiates lectin-mediated thymocyte proliferation, and it has the molecular dimensions of interleukin 1, as determined by gel exclusion chromatography. RNA from either cell type causes the synthesis of interleukin 1 with an isoelectric point of 4.8 to 5.0. RNA prepared from unstimulated macrophages or P388D1 cells does not cause interleukin 1 production by oocytes. We conclude that the amount of interleukin 1 mRNA increases greatly after stimulation of either cell type, and oocytes carry out any modifications of the polypeptide necessary for activity. The kinetics of interleukin 1 mRNA accumulation and of interleukin 1 production by macrophages and P388D1 cells are compared.
Neoantigens appear in human IgG upon antigen binding: detection by antibodies that react specifically with antigen-bound IgG.
Authors: Brown E J , Bekisz J
Published: December 2022

We raised rabbit antibodies that specifically recognize antigen-bound but not monomeric human IgG. These rabbit IgG antibodies (RAb) were induced in rabbits that were made tolerant to monomeric human IgG. They bound to immune complexes (IC) made with human IgG and various antigens including tetanus toxoid, sheep erythrocytes (E), rabbit E, or human Rh(D) + E, and were very poorly inhibitable with monomeric IgG compared to conventional rabbit anti-human IgG. RAb did not recognize complement components bound to the IgG containing IC. Cleavage of the Fc domain from human IgG markedly decreased binding of RAb to IC. Surprisingly, RAb did not bind to heat-aggregated human IgG (agg-IgG) better than to monomeric IgG. We conclude that human IgG expresses an Fc neoantigen when it binds its own antigen, and that this determinant is not expressed by agg-IgG. The implications of these findings for the biologic functions of antigen-complexed IgG are discussed.
Ia antigens and associated invariant chain are induced simultaneously in lines of T-dependent mast cells by recombinant interferon-gamma.
Authors: Koch N , Wong G H , Schrader J W
Published: December 2022

Metabolic labeling and immunoprecipitation with monoclonal antibodies, showed that lines of T-dependent mast cells (P cells) that were cultured in the absence of interferon-gamma (IFN-gamma) did not synthesize detectable amounts of Ia antigens or the invariant chain (Ii). After exposure for 24 hr to IFN-gamma produced from the cloned Mu IFN-gamma gene, P cells synthesized large amounts of Ia alpha- and beta-chains and of the invariant chain. The I-A and I-E antigens of these T-dependent mast cells were indistinguishable from those of B cells in terms of the binding of a panel of monoclonal antibodies and in terms of their structure and that of associated molecules as analyzed in two-dimensional gels. The effect of IFN-gamma on Ii synthesis in P cells was not due merely to inhibition of proliferation, as is the case with certain B cell lines, because treatment of P cells with mitomycin C did not induce the invariant chain. Variant lines of P cells that did not respond to IFN-gamma with the synthesis of Ia antigens, but only the enhanced expression of H-2K, H-2D, and H-2L antigens, also failed to synthesize Ii chain. Experiments using three tumor lines related to B lymphocytes, however, indicated that in these cells IFN-gamma could enhance the synthesis of Ii in the absence of Ia antigens. Whereas Ii chain may be essential for Ia expression, Ii chain may have further functions and in some lines can be regulated independently by IFN-gamma in the absence of Ia antigen expression.
Electron microscopic and hydrodynamic studies of protein A-immunoglobulin G soluble complexes.
Authors: Hanson D C , Phillips M L , Schumaker V N
Published: December 2022

The soluble complexes formed by reacting staphylococcal protein A (SpA) with rabbit immunoglobulin G (IgG) antibodies were characterized by hydrodynamic and electron microscopic methods. In moderate SpA excess, equilibrium mixtures of SpA and rabbit IgG formed four discrete complexes that sedimented at approximately 7, 10, 13, and 15S. The putative complexes were visible by electron microscopy and appeared to contain one, two, three, and approximately five molecules of IgG. Probably because of its elongated shape, SpA was not clearly visible in these mixtures or in control preparations of SpA alone. Both native IgG and IgG modified by cleavage of its single-hinge disulfide bond formed similar complexes on interaction with SpA. It was possible to resolve heterogeneous mixtures of IgG-SpA complexes by using an analytical ultracentrifuge equipped with a photoelectric scanner interfaced to a small computer. The relative concentrations and sedimentation velocities of different complexes in a mixture were determined from computer-generated integral and derivative plots. Both hydrodynamic and electron microscopic methods revealed that the distribution of complexes was sensitive to the IgG to SpA molar ratio. The relative amounts of faster complexes increased as the IgG to SpA molar ratio was increased. Surprisingly, when the IgG to SpA molar ratio was greater than or equal to 2, the complexes were converted into a unique 17S complex. This rather unprecedented transformation was reversible: the addition of excess SpA caused the dissociation of the 17S complex into a mixture of the 7, 10, 13, and 15S structures. The average translational diffusion coefficient of the 17S complex was 2.62 +/- 0.13 Ficks. In the electron microscope, the complex appeared to be exceptionally compact with an average diameter of 287 A. The stoichiometry of the 17S complex, together with sedimentation equilibrium, diffusion, and electron microscopic measurements, indicated that it is composed of four molecules of IgG and two molecules of SpA.
A model for the formation and interconversion of protein A-immunoglobulin G soluble complexes.
Authors: Hanson D C , Schumaker V N
Published: December 2022

We present a model for the formation and interconversion of the soluble complexes formed by reacting staphylococcal protein A (SpA) with rabbit immunoglobulin G (IgG) antibodies. The basic elements of the model are developed from reported hydrodynamic and electron microscopic studies of these complexes (see accompanying companion paper), together with established structural and binding properties of IgG and SpA. The model includes specific symmetry and binding requirements for IgG-SpA combination, and a steric constraint between neighboring IgG molecules. We discuss how such a constraint could influence the assembly and distribution of equilibrium complexes. After formulating a convenient symbolism for representing IgG-SpA complexes, the suggested model is used to construct plausible structures for the four predominant complexes observed in moderate SpA excess. Distributions of these stable complexes at different IgG:SpA ratios, together with LeChatelier's principle and a straightforward thermodynamic derivation, are used to predict likely arrangements of equilibrium structures. Also, a scale model of the unique IgG4-SpA2 complex formed in IgG excess is constructed from reported x-ray diffraction and amino acid sequence data. An intuitive thermodynamic argument is used to show that the suggested steric constraint could cause the rather unprecedented reversible transformation of the four 7 to 15S complexes into the unique 17S complex. A computer simulation is used to predict equilibrium concentrations of the various proposed complexes at different IgG:SpA ratios. In support of the suggested structures, the calculated thermodynamic distributions agree surprisingly well with those measured with the ultracentrifuge. We point out how the proposed arrangements of the complexes, and in particular the 17S complex, can account for many of their novel properties, such as antigen-induced conformational changes. Reported differences in complement activation and precipitate formation by SpA complexes formed with antibodies from various species are also discussed with regard to possible differences in structural arrangements of the complexes.
The fate of E. coli lipopolysaccharide after the uptake of E. coli by murine macrophages in vitro.
Authors: Duncan R L , Morrison D C
Published: December 2022

The fate of bacterial lipopolysaccharide (LPS) after the uptake of Escherichia coli by macrophages in vitro was studied. The LPS of the galactose epimerase-deficient E. coli J5 mutant was specifically radiolabeled with [3H]galactose by growing the organism in a basic salts medium containing galactose. Control bacteria were uniformly radiolabeled by growth in [14C]glucose and unlabeled galactose-containing medium. Surface constituents of E. coli were also labeled with 125I. After in vitro phagocytosis of labeled E. coli by murine peritoneal exudate macrophages, the rate of exocytosis of LPS, as assessed by release of 3H over a 72-hr period, was considerably reduced in comparison with other bacterial constituents (14C and 125I release). The [3H]galactose-labeled material exocytosed from macrophages and that remaining intracellularly (obtained from macrophage lysates) were isolated by cesium chloride (CsCl) density gradients and were shown to have altered density profiles as compared with purified E. coli LPS. The macrophage-"processed" [3H] galactose-containing fractions from CsCl density gradients of culture supernatants or macrophage lysates were capable of clotting Limulus amebocyte lysate. The [3H]galactose material obtained from 48-hr macrophage lysates and culture supernatants could also induce a lethal response in actinomycin D-treated mice. These data suggest that bacterial LPS may be selectively retained by the macrophage and that the post-phagocytic events that result in bacterial degradation are not accompanied by the degradation of LPS. Furthermore, although the LPS may be modified by the macrophage, it retains its biologic activity.
Regulation of human natural killing. III. Mechanism for interferon induction of loss of susceptibility to suppression by cyclic AMP elevating agents.
Authors: Leung K H , Koren H S
Published: December 2022

In this study we demonstrated that human NK cells activated by IFN or poly I:C were partially resistant to suppression by PGE2, PGD2, PGA2, PGI2, dibutyryl cAMP, isoproterenol, and theophylline. This partial loss of inhibition was not due to endogenous PG production because the addition of indomethacin to cultures stimulated with IFN or poly I:C did not prevent the partial loss of sensitivity to PGE2. NK cells incubated in the presence of PGE2 overnight, however, were not sensitive to inhibition. IFN or poly I:C did not stimulate PG synthesis nor elevate intracellular cAMP levels of NK cells. On the other hand, IFN or poly I:C diminished the accumulation of intracellular cAMP levels in NK cells in response to PGE2 stimulation. Dibutyryl cAMP and theophylline suppressed the cytolytic activity of the unstimulated cells more than that of the activated cells. A possible mechanism for the IFN-induced unresponsiveness to PGE2 may be a compartmentalized loss of cAMP responsiveness. Cycloheximide, puromycin, emetine, and actinomycin D blocked NK activation by IFN and poly I:C as well as the acquisition of resistance to PGE2-mediated suppression.
The role of mitogens and lymphokines in the induction of ornithine decarboxylase (ODC) in T lymphocytes.
Authors: Fidelus R K , Laughter A H , Twomey J J
Published: December 2022

Polyamine synthesis occurs early in lymphocyte activation after stimulation with antigen or mitogen. Ornithine decarboxylase (ODC) is the primary enzyme in the polyamine cascade. We have examined the induction of ODC by mitogens and/or lymphokines in human peripheral blood T lymphocytes. When isolated populations of monocytes and T lymphocytes were stimulated with phytohemagglutinin (PHA) there was little or no change in ODC activity. The combination of T lymphocytes and monocytes enhanced mitogen-induced ODC activity 10-fold. Several interleukin 1 (IL 1)-containing supernatants and fractionated human IL 1 were capable of substituting for monocytes in supporting PHA induction of ODC in T lymphocytes. Interleukin 2 (IL 2) and IL 2-containing supernatants were also capable of increasing ODC activity in T lymphocytes in the absence of monocytes. Lymphokines alone in the absence of PHA could not induce ODC. We conclude that both mitogens and monocytes are required for the induction of polyamine synthesis in T lymphocytes, and that supernatants containing IL 1 or IL 1 and IL 2 can substitute for monocytes in the induction of ODC in mitogen-stimulated T lymphocytes.
Macrophage activation to kill Leishmania tropica: defective intracellular killing of amastigotes by macrophages elicited with sterile inflammatory agents.
Authors: Hoover D L , Nacy C A
Published: December 2022

Resident peritoneal macrophages and macrophages elicited by injection of C3H/HeN mice with sterile inflammatory agents were exposed to amastigotes of Leishmania tropica in vitro and treated with lymphokines. Resident macrophages developed the capacity to kill intracellular parasites; microbicidal activity of activated resident cells ranged between 60 and 80%. In contrast, inflammatory macrophages responded poorly to lymphokines for intracellular killing of amastigotes; microbicidal activity of cells elicited with chronic inflammatory agents ranged between 0 and 45%. Defective intracellular killing of L. tropica by inflammatory macrophages was independent of the agent used to elicit the cells, but was clearly associated with the number of immature macrophages in the population. That intracellular killing capacity may reflect the presence of a killing mechanism in tissue-derived cells that is not yet developed in undifferentiated macrophages is supported by studies with peripheral blood monocytes: these cells were also incapable of eliminating intracellular amastigotes in the presence of potent activating factors. These observations on inflammatory macrophage interactions with amastigotes may provide important insights into the chronic nature of leishmanial disease.
Macrophage activation to kill Leishmania tropica: kinetics of macrophage response to lymphokines that induce antimicrobial activities against amastigotes.
Authors: Oster C N , Nacy C A
Published: December 2022

Lymphokine (LK) treatment of resident peritoneal macrophages from C3H/HeN mice induced two antimicrobial activities against Leishmania tropica: increased resistance of activated macrophages to infection with amastigotes and intracellular destruction of those parasites that entered activated cells. The onset and duration of these two antimicrobial activities were quite different. Resistance to infection was observed as early as 4 hr after the addition of LK, became maximal at 8 hr, and persisted in a subpopulation of treated cells for as long as 72 hr. In contrast, intracellular killing occurred with as little as 4 hr of LK treatment after infection, and maximal killing was observed in cultures exposed to LK 24 hr. Intracellular killing capacity of lymphokine-treated cells was progressively lost in macrophages treated longer than 12 hr before exposure to parasites. This decay in ability to destroy intracellular L. tropica was also seen in macrophages cultured longer than 12 hr before LK treatment, and may reflect loss of macrophage responsiveness to LK with increasing time in vitro. Thus, treatment of macrophages with lymphokines induced both a stable change in cell-parasite interactions, resistance to infection, and a short-lived capacity to destroy intracellular amastigotes.
Heterogeneity of infection enhancement of dengue 2 strains by monoclonal antibodies.
Authors: Halstead S B , Venkateshan C N , Gentry M K , Larsen L K
Published: December 2022

Seven dengue (DEN) 2 virus strains were studied for antibody-dependent enhancement (ADE) of infection in P388D1 mouse macrophage-like cells by using a panel of five DEN 2 monoclonal antibodies. DEN 2 strains were of diverse temporal, geographic, and disease origins. By hemagglutination inhibition and a plaque-reduction neutralization test in LLC-MK2 cells, two of the monoclonal antibodies were type specific and three were flavivirus group reactive. In LLC-MK2 cells, the seven DEN 2 viruses each were neutralized by all five monoclonal antibodies. In P388D1 cells, two DEN 2 strains were enhanced by only three monoclonal antibodies, two by four antibodies, and three by all five antibodies, demonstrating that in some instances enhancement is epitope related and not a concentration-dependent function of virus-antibody interactions. However, ADE did not segregate with determinants exhibiting either the flavivirus group or the dengue type specificity. The presence or absence of enhancement determinants on DEN 2 strains did not correlate with the geographic origin of virus or the severity of disease yielding the strain. The heterogeneous distribution of enhancement determinants may provide a valence mechanism contributing to a multiple increase of infection enhancement in macrophages.
Elucidation of the topography and determination of the protective epitopes on the E glycoprotein of Saint Louis encephalitis virus by passive transfer with monoclonal antibodies.
Authors: Mathews J H , Roehrig J T
Published: December 2022

We have identified and characterized eight antigenic epitopes on the 53,000 dalton envelope (E) glycoprotein of Saint Louis encephalitis (SLE) virus by using monoclonal antibodies. One of these epitopes (E-1c) encoded for the type-specific biologic functions of hemagglutination (HA) and neutralization (N). Injection of 50 ng of anti-E-1c antibody protected the majority of mice from peripheral challenge with 100 i.p. LD50 of SLE virus. Similar levels of protection with antibodies specific for other epitopes usually required greater than or equal to 1000-fold additional antibody. Attempts to block N or protection at the E-1c antigenic domain by using antibody to several other SLE epitopes that strongly competed for the E-1c site were unsuccessful. Enhancement of protection was observed with mixtures of the more cross-reactive antibodies. The E-1c antibody was also effective in abrogating SLE virus replication until neural invasion occurred. On the basis of these findings, the topologic arrangement and function of the eight SLE E glycoprotein epitopes on the virion spike is proposed.
Immunologic memory to phosphocholine. IV. Hybridomas representative of Group I (T15-like) and Group II (non-T15-like) antibodies utilize distinct VH genes.
Authors: Chang S P , Perlmutter R M , Brown M , Heusser C H , Hood L , Rittenberg M B
Published: December 2022

The anti-phosphocholine (PC) memory response elicited in BALB/c mice by phosphocholine-keyhole limpet hemocyanin (PC-KLH) contains two groups of antibodies distinguished by their fine specificity for PC and p-nitrophenylphosphocholine (NPPC). Group I antibodies are inhibited by both PC and NPPC, while Group II antibodies are inhibited appreciably only by NPPC; only Group I antibodies are dominated by the T15 idiotype. Anti-PC hybridomas representative of the memory response to PC-KLH were produced to examine the variable region genes expressed by memory B cells. Two IgM hybridomas were of the Group I type, because they were inhibited by both PC and NPPC and they bound to the pneumococcus R36A. However, only one of these antibodies (PCM-2) expressed a T15 idiotope, while the other (PCM-1) did not express any of three T15 idiotopes. Despite its negative T15 idiotype profile, N-terminal amino acid sequencing of PCM-1 purified heavy chain and Southern blots of the hybridoma DNA indicated that it utilizes the T15 VH and JH1 genes. Three hybridomas, IgG1, IgM, and IgE, typical of Group II antibodies, were examined; these were negative for three T15 idiotopes and displayed measurable avidity only for NPPC in a PC-protein binding inhibition assay. These three hybridoma antibodies, like serum Group II IgG1, did not measurably bind to the bacterium R36A. The heavy chain amino termini of all three of these antibodies were inaccessible for Edman degradation. Southern blots of DNA from the IgG1 hybridoma revealed the T15 VH gene to be in the germ line configuration only and unassociated with any JH segment, indicating that this Group II antibody utilizes a VH gene different from the T15 family. These results signify that, whereas some diversity of the (anti-PC) memory response may be generated by somatic diversification of variable regions important in the primary response, a significant contribution to the overall heterogeneity of memory antibodies originates in the expression of additional variable region genes.
Regulation of IgM and IgD synthesis in B lymphocytes. II. Translational and post-translational events.
Authors: Yuan D
Published: December 2022

Studies investigating the relative rates of biosynthesis of mu- and delta-polypeptide chains in normal resting B lymphocytes have shown that the translation rate of mu m is about sevenfold higher than that of delta m, thus reflecting the relative abundance of mRNA encoding these two chains. The turnover rate of cell surface IgM is faster, however, than that of cell surface IgD, resulting in higher expression of cell surface IgD relative to IgM under steady state conditions. LPS stimulation of B lymphocytes induces the complete cessation of synthesis of the delta-chain, thus accounting for the gradual disappearance of IgD from the cell surface of activated cells.
Production of large quantities of human immunoglobulin in the ascites of athymic mice: implications for the development of anti-human idiotype monoclonal antibodies.
Authors: Abrams P G , Ochs J J , Giardina S L , Morgan A C , Wilburn S B , Wilt A R , Oldham R K , Foon K A
Published: December 2022

Human-murine hybridomas, generated by fusion of human CLL cells with non-Ig-secreting murine myeloma NS-1, and secreting the IgM kappa expressed on the CLL cell surface, were grown in the peritoneal cavity of doubly pristane-primed nude mice to produce large quantities of highly concentrated human IgM kappa for the development of anti-idiotype monoclonal antibodies. Although the hybridomas secreted only 0.8 to 1.5 micrograms/ml/10(6) cells in tissue culture, they produced 8 to 12 ml of ascites from two mice containing 1.7 mg/ml of human IgM. Partial purification was then easily accomplished by euglobulin precipitation. Two weekly injections of 100 micrograms of this material into BALB/c mice resulted in the development of both anti-idiotypic and anti-human IgM antibodies. This system overcomes the major hurdle of generating sufficient quantities of the human idiotypic Ig to immunize and screen for specific murine antiidiotypes, and provides the material to produce these therapeutically promising reagents rapidly for clinical trials. This is, to our knowledge, the first report of a human Ig produced in mouse ascites.
The role of Ia molecules in the activation of T lymphocytes. IV. The basis of the thymocyte IL 1 response and its possible role in the generation of the T cell repertoire.
Authors: Rock K L , Benacerraf B
Published: December 2022

The activation requirements for thymocyte proliferation were investigated. Thymocytes proliferate in the presence of exogenous interleukin 1, which has been used as the classic assay for this factor. This response, however, is greatly decreased in cultures of purified thymic T cells. Purified thymic T cells will proliferate in the presence of IL 1 if accessory cells are added to culture. The requisite accessory cell is a non-T, adherent, radioresistant cell found in macrophage/dendritic cell-enriched fractions of both thymus and spleen. This cell bears Ia molecules, which are critically involved in the activation of thymocytes. This thymocyte-accessory cell interaction is not dependent on exogenous nominal antigens. Therefore, it appears that IL 1 allows the expansion of thymocytes with specificity for self-class II MHC antigens. This response was found to be unique to this stage of T cell development and can be observed with both mature and immature thymic T cell subsets. The implications of these findings for the physiologic expansion of self-restricted T cells in the thymus are discussed.
Allosuppressor- and allohelper-T cells in acute and chronic graft-vs-host disease. IV. Activation of donor allosuppressor cells is confined to acute GVHD.
Authors: Pals S T , Radaszkiewicz T , Gleichmann E
Published: December 2022

Groups of nonirradiated BDF1 mice were injected with unseparated spleen cells from B10, B10.D2, or DBA/2 donors. The diverse clinical and pathologic symptoms that developed during the course of the ensuing graft-vs-host reaction (GVHR) were related to the functional subsets of donor-T cells activated in the host. The activation of F1-specific donor T suppressor (TS) cells was confined to those GVH F1 mice that developed acute GVH disease (GVHD) (donor B10 or B10.D2). Moreover, activation in these GVH F1 mice of the Lyt-1-2+ donor TS cells sharply preceded the onset of and coincided with (week 2 to 6) the suppressive pathologic symptoms characteristic of acute GVHD, such as pancytopenia and suppression of splenic IgG production. The activation of these alloreactive TS effector cells was briefly preceded by the activation of F1-specific Lyt-1+-2- donor T helper (TH) cells and stimulation of the host's lymphoid tissue. Thus, in acute GVHD, a sequential alloactivation first of donor TH and then of TS cells was found. Those F1 mice that recovered from acute GVHD and developed stimulatory pathologic symptoms showed a concomitant loss of donor TS cell activity. An initial activation of F1-specific Lyt-1 +2- donor TH cells was also found in that parent----F1 combination (donor DBA/2), which failed to develop acute GVHD. Significantly in that combination, the alloactivation of donor TH cells was not followed by activation of significant numbers of donor TS cells. Instead, the DBA/2-injected BDF1 mice directly developed a persistent increase in splenic Ig formation and lupus-like GVHD.
Murine immune responses to Salmonella lipopolysaccharide: oral administration of whole bacteria to C3H/HeJ mice induces secondary anti-LPS responses, especially of the IgA isotype.
Authors: Jirillo E , Kiyono H , Michalek S M , McGhee J R
Published: December 2022

Because our past studies have shown that oral administration of thymic-dependent antigens induces higher IgA responses in lipopolysaccharide (LPS) nonresponsive C3H/HeJ mice than in syngeneic, LPS-responsive C3H/HeN animals, it was of interest to compare anti-LPS responses in these mouse strains after oral administration of particulate antigens containing LPS. C3H/HeJ and C3H/HeN mice were given smooth Salmonella typhimurium LT-2 or rough S. minnesota Rb (R345) or Re (R595) organisms by gastric intubation for 3 consecutive days/wk for 2 wk and were boosted by the i.v. route with either the same bacterial immunogen or with purified homologous LPS. Four days later, splenic anti-LPS plaque-forming cell (PFC) responses were assessed with a panel of indicator sheep erythrocytes (SRBC) coated with LPS derived from either smooth (S-LPS-SRBC) or rough (Rb-LPS-SRBC or Re-LPS-SRBC) Salmonella. In separate studies, both serum and salivary antibodies of the IgM, IgG, and IgA isotypes were determined by ELISA, with whole Salmonella cells used as the coating antigen. Oral immunization with LT-2 resulted in good IgM, IgG1 and IgA splenic anti-LPS PFC responses in C3H/HeJ mice, with the major isotype being IgA. Mice boosted i.v. with purified LPS gave five- to sixfold higher anti-S-LPS PFC responses than did mice given whole bacteria by the i.v. route. Low anti-Rb-LPS and anti-Re-LPS PFC responses were seen in both mouse strains. Enhanced immune responses in orally primed C3H/HeJ mice was not due to LPS-induced polyclonal responses, because splenic cultures from these mice gave poor mitogenic responses to LPS. A similar pattern of response was obtained when C3H/HeJ or C3H/HeN mice were given RB (R345) or Re (R595) bacteria orally and boosted i.v. with purified homologous LPS or whole cells. C3H/HeJ mice again showed higher immune responses in all isotypes than did C3H/HeN animals. Mice given Rb (R345) immunogen gave maximum responses to Rb-LPS, lower responses to Re-LPS, and no responses to S-LPS, whereas C3H/HeJ mice immunized with Re (R595) immunogen gave maximum PFC responses to Re-LPS and lower responses to Rb-LPS. Serum and salivary antibody titers closely paralleled the splenic PFC responses, and IgA antibodies were the predominant isotype observed, with higher IgA responses occurring in orally immunized C3H/HeJ mice than in C3H/HeN animals. These results clearly indicate that C3H/HeJ mice given whole Salmonella by gastric intubation elicit higher PFC and antibody responses to the three major LPS regions than do identically treated LPS-responsive C3H/HeN mice.
T cell regulation of polyclonally induced immunoglobulin secretion in humans.
Authors: Rosenkoetter M , Reder A T , Oger J J , Antel J P
Published: December 2022

We measured the pokeweed mitogen (PWM)-induced secretion of IgG by the unfractionated mononuclear cells (MNC) of young adult donors, and correlated the results with the functional activity of cell suspensions enriched for T helper (T4+) and T suppressor/cytotoxic (T8+) cells. The distribution of IgG levels secreted by MNC differs from a Gaussian curve, implying that the group is composed of distinct heterogeneous populations. When donors were compared who were judged to be very low responders or very high responders on the basis of IgG secretion levels by MNC (less than 700 ng/ml or greater than 2500 ng/ml), no differences were found in the capacity of T4+-enriched cells to support PWM-driven IgG secretion by a common B cell pool. In contrast, the addition of 0.2 X 10(5) T8+ cells from these low responders to PWM-stimulated cultures of 0.5 X 10(5) T4+ cells plus 0.5 X 10(5) B cells resulted in significantly less IgG secretion (389 +/- 121 ng/ml) than did the addition of the same number of T8+ cells from the high responders (2241 +/- 548 ng/ml, p less than 0.01). Normalized percent suppression by T8+ cells was higher in low responders than in high responders (77.0 +/- 9.9% vs 33.0 +/- 8.5%, p less than 0.01). Both high and low responders markedly suppressed IgG secretion when 0.5 X 10(5) T8+ cells were added. No correlation was found either between proportion of T3+, T8+, T4+, or M1+ cells within the MNC population and levels of IgG secretion by MNC or between T8+ numbers and levels of suppression induced by a constant number of T8+-enriched cells. Our data indicate that differences in the functional activity of T8+ cells, rather than quantitative differences, account for the wide range of PWM-induced IgG secretion by MNC.