bluuebunny/crossref_metadata_2025 huggingface dataset:
INTERACTION OF CHOLERA TOXIN AND TOXIN DERIVATIVES WITH LYMPHOCYTES
Authors: Holmgren J. , Lindholm L. , Lönnroth I.
Published: June 2004

The interaction of cholera toxin and a number of toxin derivatives, containing different proportions of light and heavy toxin-composing subunits (L and H), with mouse lymphocytes was studied. Experiments with [125I]toxin showed that a single cell can rapidly, within minutes, bind up to 40,000 molecules of toxin, the association constant was estimated to 7 ± 4 x 108 liters/mol, and binding was found to be very similar at 37°C and 5°C. Immunofluorescence studies revealed that the toxin attachment is located on the cell surface, and that purified L subunit but not H subunit binds to the cells. A natural cholera toxoid, built up by aggregated L subunits, showed almost identical binding properties as toxin to the cells. Pure GM1 ganglioside, the proposed membrane receptor structure for toxin, prevented entirely the cellular binding of both toxin and toxoid.

Cholera toxin in concentrations down to approximately 5 x 10–11 mol/liter (corresponding to 10 bound molecules/cell) inhibited thymus cells from being stimulated to DNA synthesis by concanavalin A (con A), and spleen cells from such stimulation by phytohemagglutinin. The GM1 ganglioside but not a series of other pure structurally related gangliosides and neutral glycosphingolipids neutralized this toxin activity. Toxin derivatives which, in similarity with toxin, possessed H as well as L subunits but in other proportions, were potent inhibitors of con A-induced thymocyte stimulation, whereas the natural toxoid (aggregated L subunits), purified toxin L subunit and purified toxin H subunit were up to 300-fold less active on a weight basis. The capacity of cholera proteins to inhibit con A-induced thymocyte stimulation correlated well with their activity in the rabbit intradermal toxicity assay.

The inhibitory action of cholera toxin on con A-induced thymocyte stimulation did not depend on decreased cell viability from the toxin treatment, nor was it caused by a reaction between toxin and con A. [125I]con A bound equally well to the cells when toxin was present as when it was absent, which proves that the toxin did not compete for cellular con A receptors. Nor did the toxin seem to disturb the general mobility of membrane receptors or their ability to accumulate in caps.

It is concluded that the L type of subunit confers rapid and firm binding of cholera toxin to lymphocyte membranes, probably to GM1 ganglioside receptors. For biologic activity the additional presence of H subunit is important. One manifestation of toxin action on lymphocytes is inhibition of lectin-induced DNA synthesis; probably this effect relates to the ability of cholera toxin to raise the levels of intracellular cyclic 3'5'-adenosine monophosphate.
STIMULATED LYMPHOCYTE CULTURES
Authors: Lohrmann Hans-Peter , Graw Carole M. , Graw Robert G.
Published: June 2004

Lymphocytes, stimulated with different doses of plant mitogens or allogeneic cells, incorporate varying amounts of [3H]thymidine. Theoretically, this may be due to different numbers of responding cells, to earlier proliferative response of these cells, and/or to their more or less rapid transit through the cell cycle. Dog peripheral blood lymphocytes were stimulated in vitro with different doses of phytohemagglutinin (PHA), or with allogeneic lymphocytes. After their synchronization by incubation with hydroxyurea (4 mM), the mean durations of the cell cycle, and of the different cell cycle phases were constant and unrelated to strength or type of stimulation. PHA-stimulated lymphocyte cultures were maintained in the presence of colchicine, to prevent clonal proliferation of responding lymphocytes. DNA uptake in this setting, attributed to first generation responders, was related to the strength of proliferative lymphocyte response in control cultures without colchicine. Furthermore, cell proliferation occurred earlier with greater stimulation. It is concluded that higher [3H]thymidine uptake in vitro by stimulated lymphocytes is due to greater numbers of responding cells, which are triggered into proliferative response earlier, and not to a more rapid transit of the responding cells through the cell cycle.
ONTOGENY OF B LYMPHOCYTES
Authors: Gelfand Michael C. , Sachs David H. , Lieberman Rose , Paul William E.
Published: June 2004

The frequency of lymphocytes bearing complement receptors in the spleens of 2-wk old mice appears to be controlled by two independent genes. The presence of a "high" allele at either locus leads to intermediate or high frequency of CRL at 2 wk of age. One of the genes controlling complement receptor lymphocyte (CRL) frequency (CRL-1) is linked to the H-2 complex. Thus, in progeny of (AKR x DBA/2)F1 x DBA/2, all mice with a low frequency of CRL at 2 wk of age are homozygous for the H-2 type of the low CRL parent (DBA/2). Furthermore, in the B10 series of congenic mice, CRL frequency at 2 wk of age is similar to the frequency in the donor of the H-2 region. Thus, C57BL/10, B10.BR, and B10-D2 mice are all of the low CRL type while B10.A mice are intermediate in CRL frequency at 2 wk. C57BR and DBA/2, the donors of the H-2 complex of the B10.BR and B10.D2, respectively, are of low CRL type while the A/WySn, the donor of the H-2 complex in the B10.A, is an intermediate CRL strain. Similarly in the A/WySn series of congenic mice, A.CA, A.SW, and A.BY are all low CRL strains while the A/WySn is intermediate.

Studies of CRL frequency in mice with recombinant H-2 chromosomes (B10.A(2R), (4R), and (5R); B6/TL+; and A/TL-) indicate that CRL-1 is to the right of the Ss-Slp genes and to the left of Tla.
ACTIVITY OF HOST-DERIVED T CELLS WHICH DIFFERENTIATE IN NUDE MICE GRAFTED WITH CO-ISOGENIC OR ALLOGENEIC THYMUSES
Authors: Kindred Berenice , Loor Francis
Published: June 2004

If nude mice are grafted with a neonatal thymus, host type precursor cells develop within the graft thymus and after about 6 wk the T-cell population of the thymus, spleen, and lymph nodes is of host type. However, immunological responsiveness produced in nude mice in this manner is incomplete: (a) the ability to react to T-cell mitogens in vitro is greater than in untreated nudes but lower than in normal mice; (b) the response to T-cell dependent antigens is less than normal; and (c) the rejection of skin grafts is slower than in normal animals. Whether host precursor cells which differentiate in an allogeneic thymus are able to reject skin grafts from thymus donor strain appears to depend on the strain combination used.
PROPERTIES OF PLASMINOGEN ACTIVATORS FORMED BY NEOPLASTIC HUMAN CELL CULTURES
Authors: Rifkin Daniel B. , Loeb John N. , Moore George , Reich E.
Published: June 2004

A series of human cell lines has been examined for fibrinolysis in culture. The sera that are activating for fibrinolysis by human cells are mouse, monkey, human, horse, and bovine. Individual human sera show considerable variation in the ability to activate fibrinolysis. In common with other neoplastic or transformed mammalian and avian cell cultures, human cell lines of neoplastic origin produce substantial amounts of plasminogen activator. Several cultures of nonmalignant origin also produce plasminogen activator, whereas cultures obtained from trypsinized human embryos, or from human embryonic skin do not. The human melanoma plasminogen activators are of two kinds: a major component with a mol wt of 50,000, and a minor species with a mol wt of approximately 60,000. Both are DFP sensitive, serine proteases.
TWO POPULATIONS OF T LYMPHOCYTES IMMUNE TO THE LYMPHOCYTIC CHORIOMENINGITIS VIRUS
Authors: Volkert Mogens , Marker Ole , Bro-Jørgensen Klaus
Published: June 2004

In this report seven different parameters were employed to investigate the spleen and lymph node cells from mice at the early and the late state of immunity to the lymphocytic choriomeningitis (LCM) virus. Distinct differences were observed. Morphological studies revealed a different size distribution of the cells in the preparations from the early and the late state of immunity. The cell mixtures of early immune cells contained many more large and blast-like lymphoid cells than the other. Where the cell function was concerned, the cytotoxic activity against LCM virus-infected target cells was almost entirely a function of the early immune cells, and our data strongly indicate that enhancement does not play any role for the disappearance by time of this cell activity. The antiviral effect after transfer to acutely infected animals was also predominantly a function of the early immune cells and the same was the case concerning the ability to protect against a lethal acute infection. However, the early immune cells were almost inactive after transfer to chronically infected virus carriers, whereas transplants of late immune cells to such mice had a very strong antiviral effect. The resistance to X irradiation also varied. Even high X-ray doses could not destroy the function of early immune cells, whereas the function of the late immune cells was readily impaired by X-ray treatment. The early and the late immune cells have one thing in common—both are susceptible to treatment with anti-theta serum. Because of the differences observed between the early and the late immune cells, it is concluded that they belong to different cell populations. However, because of the common susceptibility to anti-theta serum, probably both populations are T-cell lymphocytes. The implications of the results and the role of the different cells in the combat of the viral infection are discussed.
COMPLETE RESTORATION OF BURSA-DEPENDENT IMMUNE SYSTEM AFTER TRANSPLANTATION OF SEMIALLOGENEIC STEM CELLS INTO IMMUNODEFICIENT CHICKS
Authors: Toivanen Paavo , Toivanen Auli , Vainio Olli
Published: June 2004

For transplantation of semiallogeneic bursal stem cells into cyclophosphamide-treated 3-day old chicks, two lines of chickens homozygous at the major histocompatibility locus and their F1 hybrids were used in reciprocal combinations. The semiallogeneic transplantations resulted in a complete restoration of antibody formation to sheep red blood cells (SRBC) and Brucella, of microscopic morphology of bursa fabricii, and of germinal center formation in the spleen. In contrast, allogeneic bursal stem cells were not effective in restoring secondary response to SRBC and germinal center formation, while they were able to reconstitute anti-Brucella responses and bursal morphology. These findings indicate an effective cooperation of donor and host cells leading to a complete restoration of the bursa-dependent lymphoid system, when the donor and recipient share at least one haplotype determining the major histocompatibility antigen complex.
IDENTIFICATION OF A NOVEL CELL TYPE IN PERIPHERAL LYMPHOID ORGANS OF MICE
Authors: Steinman Ralph M. , Lustig Dinah S. , Cohn Zanvil A.
Published: June 2004

Several properties of lymphoid dendritic cells in situ have been determined, and contrasted to information previously established for lymphocytes and mononuclear phagocytes. Dendritic cells are not found in newborn mice, and their concentration in both spleen and mesenteric lymph node does not reach adult levels until 3–4 wk of age. Dendritic cells largely disappear from adherent populations following administration of steroids (2.5 mg hydrocortisone acetate s.c.) and ionizing radiation (Do of 100 rads for Co60). Splenic dendritic cells can originate from precursors located in both bone marrow and spleen itself, probably the red pulp. The mature splenic population does not actively divide (pulse labeling index with [3H]thymidine of 1.5–2.5%), but does turnover at substantial rate, 10+% of the total pool per day. The influx of new cells appears to be derived from a proliferating precursor compartment, but the mechanism for efflux or turnover is not known. Dendritic cells in spleen and node undergo little or moderate increase in numbers during development of a primary immune response. These in vivo characteristics, taken together, further distinguish dendritic cells as a novel cell type, distinct from mononuclear phagocytes and lymphocytes.
POTENTIATION OF T-CELL-MEDIATED IMMUNITY BY SELECTIVE SUPPRESSION OF ANTIBODY FORMATION WITH CYCLOPHOSPHAMIDE
Authors: Lagrange P. H. , Mackaness G. B. , Miller T. E.
Published: June 2004

Delayed-type hypersensitivity (DTH) appears in mice immunized with less than an optimal immunogenic dose of sheep red blood cells (SRBC), but is blocked progressively as antibody production increases in response to larger doses of SRBC. Treatment with cyclophosphamide (CY) was shown to release T cells from this inhibitory influence of the humoral response, and cause enhancement of DTH. The magnitude of this enhancing effect on T-cell activity was markedly dependent on the time of treatment relative to the time of immunization, and on the time chosen for measuring DTH. The reasons for these pronounced effects of timing are threefold: (a) CY given before antigenic stimulation has a long-lasting effect on antibody formation, but no apparent effect on the precursors of activated T cells. (b) After antigenic stimulation, T cells also become susceptible to CY. (c) The production of a nonspecific participant (monocyte) in the DTH reaction is also suppressed by CY, though the supply of circulating monocytes is not immediately affected by the drug.

The differential effect of CY on T and B lymphocytes depends on the differing physiological states of the majority of cells that make up these two populations. The former are resting cells that are insensitive to CY until exposed to specific antigen, while the latter are drawn from a rapidly replicating precursor pool and are susceptible to CY at all times.
THE MODIFYING EFFECT OF BCG ON THE IMMUNOLOGICAL INDUCTION OF T CELLS
Authors: Mackaness G. B. , Lagrange P. H. , Ishibashi T.
Published: June 2004

The inhibition of activated T cells by products of the humoral immune response is almost abolished by systemic infection with BCG. As a result, BCG-infected mice develop very high levels of delayed-type hypersensitivity (DTH) in response to doses of sheep red blood cells (SRBC) that cause complete suppression of DTH in normal mice. This systemic effect of BCG is dose-dependent, and lasts for about 3 wk. Its main effect is to counteract the inhibition of T cells by products of the humoral response. As a result, and in contrast to the T-cell-potentiating effects of cyclophosphamide (CY) which depend on a diminished production of antibodies, increased levels of DTH in BCG-infected mice are associated with increased antibody production. Since BCG and CY act in different ways, their effects are additive. Very remarkable levels of DTH are achieved when they are used in combination.
CELL-TO-CELL INTERACTION IN THE IMMUNE RESPONSE
Authors: Basten A. , Miller J. F. A. P. , Sprent J. , Cheers C.
Published: June 2004

Specific immunological tolerance was induced in CBA mice by a single injection of deaggregated fowl immunoglobulin G (FγG). The unresponsive state was stable on adoptive transfer and irreversible by pretreatment of tolerant cells with trypsin. Tolerant spleen cells could suppress the response of normal syngeneic recipients. They also suppressed the adoptive primary response of spleen cells to FγG in irradiated hosts. The inhibitory effect was on the indirect (7S) plaque-forming cell (PFC) response.

Incubation of the tolerant cell population with anti-θ serum and complement reversed the suppressor effect. Furthermore, the addition of purified T cells from normal donors restored the capacity of the anti-θ serum-treated tolerant cells to transfer an adoptive response to FγG. The existence of FγG-reactive B cells was supported by the demonstration of normal numbers of antigen-binding cells in the spleen and thoracic duct lymph from tolerant animals. Moreover, the formation of caps by these cells implied that they could bind antigen normally.

These experiments provided direct evidence for the existence of suppressor T cells in the tolerant population. Further evidence was derived from examination of the effect of antigen "suicide". Tolerant spleen cells were treated with radioactive FγG under conditions known to abrogate T-cell helper function. When these cells were transferred together with normal spleen cells into irradiated hosts, suppression of the primary adoptive response to FγG was no longer observed. Inhibition of an adoptive secondary response to FγG was obtained by transferring tolerant spleen cells with primed B cells provided high doses of tolerant cells were used. By contrast low doses exerted a helper rather than a suppressor effect in this system.
MEDIATION OF IMMUNOLOGIC DISCHARGE OF LYSOSOMAL ENZYMES FROM HUMAN NEUTROPHILS BY GUANOSINE 3',5'-MONOPHOSPHATE
Authors: Ignarro L. J. , George W. J.
Published: June 2004

The purpose of this investigation was to elucidate the relationship of cyclic GMP and calcium to the immunologic discharge of lysosomal enzymes from purified human neutrophils. Contact of neutrophils with a variety of immunologic stimuli, including zymosan particles treated with either normal or rheumatoid arthritic (RA) serum, heat-aggregated (agg) IgG, particulate and immobilized agg IgG each treated with RA serum, and zymosan-treated serum, provoked the discharge of ß-glucuronidase, but not cytoplasmic lactate dehydrogenase, and stimulated the accumulation of cyclic GMP. Both enzyme release and elevation of cyclic GMP levels required the presence of extracellular calcium as neither cellular event proceeded in its absence. Cholinergic enhancement of the immunologic secretion of ß-glucuronidase from neutrophils by acetylcholine was associated with a concomitant accumulation of cyclic GMP. These actions of acetylcholine on neutrophils did not proceed in the absence of extracellular calcium. Whereas the concentrations of cyclic GMP in neutrophils were elevated by both immune reactants and a combination of the latter and acetylcholine, cyclic AMP levels remained unaltered. Thus, cyclic GMP, but not cyclic AMP, was associated with the immunologic and pharmacologic discharge of lysosomal enzymes from neutrophils. Contrariwise, cyclic AMP, but not cyclic GMP, was associated with inhibition of lysosomal enzyme release. For example, dibutyryl cyclic AMP and epinephrine inhibited the release of ß-glucuronidase from neutrophils that was elicited by each of the immune reactants tested. Moreover, cyclic AMP levels in the cells were elevated markedly in every instance that enzyme discharge was inhibited by epinephrine. Epinephrine did not alter the neutrophil concentrations of cyclic GMP at times when those of cyclic AMP were elevated. The data in this report constitute partial evidence that the immunologic discharge of lysosomal enzymes from human neutrophils is mediated or signaled by intracellular cyclic GMP and that calcium is linked to this stimulation of enzyme secretion.
SELECTIVE ROLES OF THYMUS-DERIVED LYMPHOCYTES IN THE ANTIBODY RESPONSE
Authors: Tada Tomio , Takemori Toshitada
Published: June 2004

Passively transferred thymocytes and spleen cells from donors primed with keyhole limpet hemocyanin (KLH) exerted differential suppressive effect on IgM and IgG antibody responses of syngeneic recipients immunized with DNP-KLH depending primarily on the time when KLH-primed cells were transferred. This was demonstrated by the decrease in the numbers of DNP-specific direct and indirect PFC in the spleen of the recipients given KLH-primed cells at different times during primary and secondary immunization. Whereas the cell transfer simultaneously with or 2 days after the primary immunization produced only slight suppression of the peak IgM antibody response, it caused profound suppression of late IgM and IgG antibody responses. By contrast, the cell transfer 3 days after the immunization produced immediate suppression of the ongoing IgM antibody response resulting in its earlier termination, while being unable to prevent the induction of IgG antibody response. KLH-primed cells could moderately suppress the secondary anti-DNP antibody response, in which IgG antibody response was found to be slightly more sensitive than IgM antibody response to the suppressive influence of KLH-primed cells.

The suppressive effect of the KLH-primed spleen cells was completely eliminated by the in vitro treatment of the cells with anti-θ and C before cell transfer, indicating that cells responsible for the suppression are, in fact, T cells. The suppression of DNP-specific antibody response by KLH-primed T cells was achieved only if the recipients were immunized with DNP-KLH but not with DNP-heterologous carrier, suggesting that direct interaction between T and B cells is necessary for the suppression of the antibody response. It is concluded that susceptibility of B cells to the specific suppressive influence of T cells is inherently different depending on the differentiation stage of B cells and on the immunoglobulin class they are destined to produce.
PHYSIOCHEMICAL AND BIOLOGICAL PROPERTIES OF THE MAJOR BASIC PROTEIN FROM GUINEA PIG EOSINOPHIL GRANULES
Authors: Gleich Gerald J. , Loegering David A. , Kueppers Friedrich , Bajaj Satya P. , Mann Kenneth G.
Published: June 2004

Guinea pig eosinophil granules are characterized by the presence of a basic protein of low molecular weight which accounts for greater than 50% of granule protein. This protein, termed the major basic protein (MBP), readily aggregates and becomes insoluble, and the formation of aggregates is dependent on the establishment of disulfide bonds. Analysis of concentrated preparations of MBP often revealed a series of bands which were multiples of a monomeric unit with a mol wt of approximately 11,000. Analysis of reduced and alkylated MBP on a 10% agarose column equilibrated with 6 M guanidinium chloride revealed a single polypeptide chain with a mol wt of 10,800. Amino acid analysis of the protein revealed the presence of 13% arginine, consistent with the basic character of the molecule. Four residues of tryptophan, were present, indicating that MBP is not a histone. The MBP did not increase vascular permeability when injected into the skin of guinea pigs, nor did it antagonize the effect of histamine and bradykinin in the skin. MBP also did not contract the isolated guinea pig ileum and when mixed with histamine or bradykinin did not inhibit their activity on the gut. MBP had only weak, if any, antihistaminic activity. MBP possessed weak bactericidal activity when compared to histone and then only with one strain of E. coli. MBP precipitated DNA, neutralized heparin, and activated papain. On a molar basis MBP was more active than cysteine in activating papain. These results do not point to any unique biological activity associated with MBP other than those expected of a protein as basic as it is and one which possesses reactive sulfhydryl groups. Possible functions of eosinophils based on the properties of the MBP are discussed.
CELL-MEDIATED IMMUNE RESPONSES IN VITRO
Authors: Peavy Duane L. , Pierce Carl W.
Published: June 2004

The effects of soluble concanavalin A (Con A) or Con A-activated spleen cells on the generation of cytotoxic lymphocytes (CL) in mixed leukocyte cultures (MLC) were examined. Mitogenic concentrations of soluble Con A or small numbers of Con A-activated spleen cells substantially inhibited CL responses. The suppression was partial rather than absolute and was critically dependent upon the concentration and time of addition of soluble Con A or Con A-activated spleen cells to the MLC.

Suppressive effects of Con-A activated spleen cells were mediated by T cells since suppressor cell activity was abrogated by treatment of spleen cells with anti-θ serum and complement before or after Con A activation. X irradiation of spleen cells before Con A treatment also abrogated generation of suppressor cell activity. After activation by Con A, however, the function of suppressor cells was radioresistant.

Although the precise mechanism(s) of suppression is, as yet, unknown, the precursors of CL must be exposed to Con A-activated cells during the early phases of the immune response for suppression to occur. Kinetic studies revealed that suppression of CL responses was not due to a failure to initiate an immune response, but represented a response which developed initially, but subsequently aborted. The relevance of these observations to the concepts of T-cell-T-cell interaction and regulatory control of immune responses by T cells is discussed.
CLONAL CHARACTER OF F1 HYBRID LYMPHOCYTE SUBSET RECOGNITION OF PARENTAL CELLS IN ONE-WAY MIXED LYMPHOCYTE CULTURES
Authors: Gebhardt Bryan M. , Nakao Yoshinobu , Smith Richard T.
Published: June 2004

Proliferation of F1 hybrid lymphocytes in mixed lymphocyte cultures is stimulated by mitomycin-blocked parental cells. The demonstration of this phenomenon using F1 hybrids derived from congenic lines of mice establishes that the stimulation is controlled by genes in or closely linked to the major histocompatibility locus chromosome region. In agreement with the finding that tumor-bearing mice have an increased capacity for primary alloantigen recognition, it was observed that the F1 hybrid response to parent was also augmented by tumor bearing.

Chromosomal analysis of dividing cells in one-way mixed cultures confirms that F1 cells, and not the blocked parental cells, enter mitosis. Stimulation of F1 cells by a soluble mediator liberated by the parental cells was not observed and mitomycin blocking of parental cells seems to be a completely effective blocking agent ensuring that parental cells can not enter DNA synthesis.

The specificity and clonal nature of F1 recognition of parent was demonstrated using a 5-bromodeoxyuridine-suicide procedure. Distinct clones of lymphocytes in F1 spleen cell populations seem to recognize one or the other parent, but not both, in such experiments. These observations and others in tumor systems suggest that most or all heterozygous organisms may possess potentially self-reactive clones of lymphocytes.
REGULATION OF CELL-MEDIATED CYTOTOXICITY
Authors: Sabbadini Edris
Published: June 2004

Mice were treated with sublethal and midlethal doses of irradiation (500–700 rads) and injected intravenously with allogeneic or semiallogeneic F1 hybrid spleen cells. The cytotoxicity developed by their spleen cells was measured with the lysis of 51Cr-labeled target cells and was found to be stronger (although delayed in time) than the cytotoxic activity of spleen cells from nonirradiated mice. The injection of syngeneic thymus or spleen cells in the irradiated mice after their treatment with allogeneic spleen cells exerted a suppressor activity, i.e., reduced the level of cell-mediated cytotoxicity (CMC). The majority of effector cells involved in the modified CMC response of irradiated mice was shown to be of host origin and lysed specifically target cells of the same genotype as the donor. A small percentage of the cells obtained from the spleens of irradiated recipients of allogeneic spleen cells was composed of donor cells which lysed specifically target cells of the same genotype as the host. These results demonstrate that the precursors of the cytotoxic cells responsible for target cell lysis are relatively radioresistant and suggest that their response is regulated by radiosensitive thymus-dependent cells.
LONG-TERM ESTABLISHMENT OF A HUMAN PLASMACYTE CELL LINE DERIVED FROM A PATIENT WITH IgD MULTIPLE MYELOMA
Authors: Jobin M. E. , Fahey J. L. , Price Z.
Published: June 2004

Cell line LA-49, derived from pleural fluid cells of a patient with IgD multiple myeloma, was established in culture and maintained for more than 1 yr. The D-myeloma protein produced in culture was similar to the serum D-myeloma protein in electrophoretic mobility and in delta- and lambda-chain antigens.

The plasma cell tumor culture, LA-49, differed from numerous immunoglobulin-producing B-lymphoblastoid cell lines established in this laboratory in: (a) Morphology (revealing various stages of maturation); (b) type of immunoglobulin produced (IgD vs. IgM, IgG, and/or, rarely, IgA); (c) growth characteristics (requirement of plasmacyte-stimulating factor); and (d) chromosomal features (polyploid vs. pseudodiploid).

A growth factor was needed for cell division and maintenance of culture viability. This factor was supplied readily by irradiated feeder layers of normal human fibroblasts or conditional media from fibroblast cultures. Preliminary characterization of this factor revealed it to be a protein with a mol wt of approximately 150,000 daltons.
VARIATION IN SUSCEPTIBILITY OF A HUMAN LYMPHOID CELL LINE TO IMMUNE LYSIS DURING THE CELL CYCLE
Authors: Pellegrino Michele A. , Ferrone Soldano , Cooper Neil R. , Dierich Manfred P. , Reisfeld Ralph A.
Published: June 2004

Cultured human lymphoid cells RPMI 8866 at different stages of their growth cycle vary in their susceptibility to lysis by rabbit, human, and guinea pig complement activated by HL-A antibodies or heterologous antibodies directed to membrane antigens; cells in G1 phase are the least sensitive to lysis. To investigate the cause of differential susceptibility of cells RPMI 8866 to lysis, the expression of HL-A determinants and the ability of cells to react with complement were investigated. No change was detected in the density of HL-A antigens on RPMI 8866 cells in synchronous growth as determined by quantitative microabsorption assays, isotopic antiglobulin tests and yields of soluble HL-A antigens. Cells did not vary during the growth cycle in their ability to interact with complement components and in their capacity to activate the complement system through the classical or alternate pathway. These data suggest that variability in lytic susceptibility is due to changes in the structure of the cell membrane or in its ability to repair complement induced damage at certain intervals during the cell cycle. Therefore, this cell line constitutes a useful model to investigate the final steps of the cytolytic reaction.
ON THE RELATION OF PRODUCTS OF ACTIVATED LYMPHOCYTES TO CELL-MEDIATED CYTOLYSIS
Authors: Henney Christopher S. , Gaffney Judith , Bloom Barry R.
Published: June 2004

Experiments have been designed to test the hypothesis that soluble mediator production and T-cell-mediated cytotoxicity are necessarily related phenomena, and that soluble mediators may be involved in the mechanism of cytolysis. To this end, agents known to inhibit T-cell-mediated lysis in vitro have been studied for their effects on the production of two lymphocyte-derived mediators, lymphotoxin (LT) and migration inhibitory factor (MIF).

A clear dissociation between mediator production and cell-mediated cytolysis was found using inhibitors of protein synthesis. Pactamycin and emetine, in doses of 10–7 M to 10–6 M, suppressed production of MIF and LT with only slight effect on killing of mastocytoma cells by immune T cells. On the other hand colchicine and vinblastine inhibited T-cell-mediated cytolysis in a dose-related manner but had no significant effect on either MIF or LT production, A striking dichotomy was also observed after augmentation of intracellular cyclic 3'5' adenosine monophosphate (cAMP) levels with cholera enterotoxin. Increased cAMP levels were associated with abrogation of direct lytic activity, but were without significant effect on MIF or LT production in guinea pigs or mice. These findings indicate that mediator production and direct lymphocyte-mediated cytolysis can be experimentally dissociated and represent independent cell-mediated immune functions.
ISOLATION AND IDENTIFICATION BY SEQUENCE ANALYSIS OF EXPERIMENTALLY INDUCED GUINEA PIG AMYLOID FIBRILS
Authors: Skinner Martha , Cathcart Edgar S. , Cohen Alan S. , Benson Merrill D.
Published: June 2004

Amyloidosis was produced experimentally in guinea pigs by multiple casein injections. Amyloid fibrils were isolated and fractionated and a protein obtained that had an amino acid composition comparable with A protein, a unique nonimmunoglobulin constituent of secondary amyloid deposits. N-terminal sequence analysis demonstrated a sequence homologous with that of A proteins from human and monkey preparations but preceded by a 5-residue peptide which had an N-terminal histidine. A definite species specificity in A protein from human and guinea pig was identified on immunologic analysis.
INFECTIOUS AGAMMAGLOBULINEMIA: TRANSMISSION OF IMMUNODEFICIENCY WITH GRAFTS OF AGAMMAGLOBULINEMIC CELLS
Authors: Blaese R. Michael , Weiden Paul L. , Koski Irma , Dooley Nancy
Published: June 2004

Agammaglobulinemia induced in chickens by surgical bursectomy and irradiation at hatching is transmissible at 4 mo of age to sublethally irradiated normal chickens by bone marrow transplantation. The immunodeficiency which develops in the recipients persists for life and is characterized by inability to produce antibody and by progressive loss of detectable serum IgM and IgG.
EVIDENCE FOR LINKAGE BETWEEN HL-A HISTOCOMPATIBILITY GENES AND THOSE INVOLVED IN THE SYNTHESIS OF THE SECOND COMPONENT OF COMPLEMENT
Authors: Fu S. M. , Kunkel H. G. , Brusman H. P. , Allen F. H. , Fotino M.
Published: June 2004

HL-A analysis of a family with C2 deficiency revealed evidence for close linkage between the C2 defect and the histocompatibility HL-A loci. The propositus was homozygous both for C2 deficiency and the HL-A haplotype 10,W18. Among seven children of three double backcross matings, no recombinants were found. The possible significance of such linkage is discussed.
ISOLATION OF ANTIGEN-BINDING CELLS FROM UNPRIMED MICE
Authors: Julius Michael H. , Herzenberg Leonard A.
Published: June 2004

Cells binding DNP groups conjugated to fluoresceinated mouse gamma globulin (FDNP-MGG) were isolated from spleens of unprimed mice using a fluorescence-activated cell sorter (FACS). The isolated cells were specifically enriched at least 100-fold for anti-DNP precursor activity in an adoptive transfer assay as compared to unfractionated spleen. The fraction depleted of binding cells, although depleted of anti-DNP precursor activity, responded as well as unfractionated spleen when assayed for anticarrier (keyhole limpet hemocyanin [KLH]) precursor activity.

High avidity binding cells were stained using low concentrations of FDNP-MGG. Medium and low avidity binding cells were stained using high concentrations of FDNP-MGG in the presence of free hapten which selectively blocked staining of the high avidity binding cells. Cells were supplemented with an excess of carrier-primed (KLH), nylon-purified splenic T cells and transferred to irradiated recipients. DNP-KLH was given at transfer and 5 days later. The anti-DNP plaque-forming cell (DNP-PFC) response and the avidities of the DNP-PFC in the irradiated recipients were measured by hapten inhibition of direct PFC plaque formation 12 days after transfer. At this time, very few indirect PFC were found.

There was a positive correlation between the avidity of the DNP-binding cells and the avidity of the anti-DNP antibody secreted by their progeny. High avidity DNP-binding cells gave rise to predominantly high avidity anti-DNP-PFC. Medium and low avidity binding cells gave rise to medium and low avidity DNP-PFC.
INDUCTION OF MACROPHAGE PLASMINOGEN ACTIVATOR BY ENDOTOXIN STIMULATION AND PHAGOCYTOSIS
Authors: Gordon Saimon , Unkeless Ay C. , Cohn Z. A.
Published: June 2004

The injection of thioglycollate medium into the peritoneal cavity of the mouse induces high levels of macrophage fibrinolytic activity due to the production and secretion of a plasminogen activator, a trypsinlike serine protease, which is absent in unstimulated macrophages. Intraperitoneal injection of endotoxin or mineral oil can stimulate only a fraction (<10%) of the fibrinolytic activity of thioglycollate cells, similar to the partial stimulation (<10%) seen 1–2 days after phagocytosis of latex or SRBC by unstimulated macrophages. The endotoxin-stimulated macrophages contain and release relatively low levels of plasminogen activator, but these primed cells can be triggered to produce and secrete high levels of enzyme, by phagocytosis of latex. Under conditions where the plasminogen activator is induced and secreted, there are no effects on the production and/or release of lysozyme or intracellular acid hydrolases,

Discovery of a two-stage procedure for inducing macrophage plasminogen activator made it possible to study the role of cell priming and phagocytosis separately. Endotoxin was a more effective priming agent, weight for weight, than lipid A:BSA complex. Secretion of the plasminogen activator was induced only by thioglycollate, or endotoxin and latex. In situ fibrinolysis was induced by these agents and mineral oil, BCG, and fetal calf serum, in decreasing order of effectiveness. Phagocytosis of latex in all cases except thioglycollate stimulation, increased fibrinolytic activity from three- to sixfold.

Latex and a variety of other particles such as M. lysodeikticus, aggregated γ-globulin and immune complexes showed dose-dependent stimulation of fibrinolysis by endotoxin-primed macrophages. Although the initial phagocytic trigger was not specific for the substance employed, the ability to induce a sustained response depended on the persistence of the phagocytized particle within the cell. Fibrinolysis and secretion of plasminogen activator continued at high levels for at least 9 days after uptake of latex, a nondigestible particle, whereas plasminogen activator was secreted only transiently after ingestion of rapidly digested M. lysodeikticus. The induction of plasminogen activator secretion provides a mechanism by which the activated macrophage can exert a selective effect on its extracellular environment.
FORMATION OF SLOW-REACTING SUBSTANCE OF ANAPHYLAXIS IN HUMAN LUNG TISSUE AND CELLS BEFORE RELEASE
Authors: Lewis Robert A. , Wasserman Stephen I. , Goetzl Edward J. , Austen K. Frank
Published: June 2004

The capacity to extract slow-reacting substance of anaphylaxis (SRS-A) from human lung tissue or cells after immunologic activation, together with the measurement of SRS-A in both the extract and the surrounding fluid, permits study of total SRS-A generation. That the material extracted is SRS-A was established by both differential bioassay and purification. SRS-A accumulation was entirely intracellular after limited IgE-dependent direct or reversed anaphylactic activation. Intracellular accumulation also generally preceded release, with generation of SRS-A continuing well beyond a plateau in the cellular SRS-A level and the release of preformed mediators. The quantity of SRS-A generated after immunologic activation was modulated by the introduction of exogenous cyclic nucleotides, revealing a site of cyclic nucleotide action distinct from that on mediator release. The capacity to determine not only the release of preformed mediators but also the generation of a newly formed mediator, the sum of SRS-A in cells and supernate, adds an additional dimension to the analysis of the cellular events of immediate hypersensitivity.
RELEASE OF DNA IN CIRCULATING BLOOD AND INDUCTION OF ANTI-DNA ANTIBODIES AFTER INJECTION OF BACTERIAL LIPOPOLYSACCHARIDES
Authors: Fournié Gilbert J. , Lambert Paul H. , Miescher Peter A.
Published: June 2004

The present data demonstrate the induction of antisingle-stranded (SS) DNA and antidouble-stranded DNA antibodies in various strains of mice, including athymic C57BL/6 nude mice, after the injection of bacterial lipopolysaccharide (LPS). This anti-DNA response is dose dependent and varies quantitatively according to the strain of the injected mice. It is not correlated to the H-2 histocompatibility locus nor to the immune response to LPS. The lipid A fraction appears to be the active part of the LPS molecule for this particular effect. In addition, it was found that DNA is released in circulating blood a few hours after the injection of LPS. Most of the DNA released has physicochemical and immunochemical characteristics of SS DNA. Therefore, the anti-DNA response induced by injections of LPS may be the result of a release of DNA in a particularly immunogenic form at a time when the immune system, in particular the B lymphocytes, is rendered capable by LPS of developing an immune response to such a soluble antigen. These effects of LPS may account for the triggering or the exacerbation of ante-DNA antibodies during infections with gram-negative bacteria, and a similar mechanism may be involved in the pathogenesis of systemic lupus erythematosus.
MACROPHAGE-LYMPHOCYTE CLUSTERS IN THE IMMUNE RESPONSE TO SOLUBLE PROTEIN ANTIGEN IN VITRO
Authors: Werdelin Ole , Brændstrup Otto , Pedersen Eskild
Published: June 2004

We have studied the physical interaction between macrophages and lymphocytes during the immune response to purified protein derivative of tuberculin (PPD) in vitro. Mixtures of peritoneal macrophages and lymph node lymphocytes from guinea pigs immunized with tubercle bacilli formed cell clusters during 20 h of culture with PPD. The number of clusters produced was correlated to the number of immune lymphocytes in the cultures. Peritoneal macrophages which had been pulsed with PPD and untreated lymph node lymphocytes produced cell clusters in the absence of free PPD in numbers equivalent to those produced by the same cells in the presence of free PPD. In cultures containing a mixture of PPD-pulsed macrophages, not-pulsed macrophages, and immune lymphocytes with no free PPD, cell clusters developed mainly between the antigen-pulsed macrophages and lymphocytes. Cluster formation was antigen-specific with the specificity residing in the lymphocytes, mainly or exclusively in the T lymphocytes. These data indicate that in the process of cell cluster formation macrophages serve as antigen-binding (or -processing) cells, while a subpopulation of lymphocytes interact physically and specifically with the macrophages.
EVIDENCE FOR THE EXPRESSION OF Ia (H-2-ASSOCIATED) ANTIGENS ON THYMUS-DERIVED LYMPHOCYTES
Authors: Frelinger Jeffrey A. , Niederhuber John E. , David C. S. , Shreffler Donald C.
Published: June 2004

We have demonstrated in an anti-Ia serum the presence of specific antibodies reacting with T cells, as well as with B cells, using a highly sensitive dye exclusion test. This antiserum reacts with both spleen and lymph node in a characteristic biphasic titration curve killing up to 70% of these cells. It also reacts with cortisone-resistant thymocytes. The A.TH-α-A.TL serum can be absorbed with spleen, lymph node, cortisone-resistant thymus, or normal thymus cells. Further in vivo absorptions in BALB/c nude cannot remove all of the cytotoxic activity for normal BALB lymph node lymphocytes, while completely removing the activity for nude cells. A Thy-1 positive cell line derived from a C57Br leukemia is reactive with this anti-Ia serum.
STUDIES ON MEDIATOR PRODUCTION BY HIGHLY PURIFIED HUMAN T AND B LYMPHOCYTES
Authors: Rocklin Ross E. , MacDermott Richard P. , Chess Leonard , Schlossman Stuart F. , David John R.
Published: June 2004

Highly purified populations of T and B lymphocytes obtained by affinity column separation were stimulated by antigen and their ability to produce two mediators, migration inhibitory factor (MIF) and lymphocyte mitogenic factor (LMF) was assessed. Both T- and B-cell populations made MIF; the production of MIF was antigen-specific using purified protein derivative of tuberculin, streptokinase-streptodornase, and Candida antigens. The MIF activity from both populations could not be attributed to antigen-antibody complexes as the inhibitory activity eluted from Sephadex G-100 columns in the same region corresponding to mol wt 23,000 daltons. Further studies indicate that the T cells producing MIF are proliferating cells whereas the B cells producing this mediator are not.

In contrast, LMF was made only by T cells and not B cells when these populations were stimulated by antigen. The LMF induced the [3H]thymidine incorporation into both T and B cells obtained from donors lacking sensitivity to the antigens used to elicit the factor. Chromatographic studies indicate that LMF eluted from Sephadex G-100 in a fraction of mol wt 23,000 daltons where MIF is also found; however, since B cells produce MIF but not LMF, these two factors appear to be distinct from one another.

Some of the implications of these findings are discussed. The explanation for the production or lack of production of MIF by lymphocytes obtained from patients with immunodeficiency disorders requires reinterpretation.
RECEPTOR FOR THE FOURTH COMPONENT OF COMPLEMENT ON HUMAN B LYMPHOCYTES AND CULTURED HUMAN LYMPHOBLASTOID CELLS
Authors: Bokisch Viktor A. , Sobel Alain T.
Published: June 2004

This report describes receptors for C4b on human peripheral B lymphocytes. The simultaneous presence of C3b and C4b receptors on the same lymphocytes was demonstrated by the formation of mixed rosettes consisting of the lymphocytes, EAC14 and EAC1423. Furthermore, reduction of the number of EAC1423 rosette-forming lymphocytes in a lymphocyte population by albumin gradient centrifugation concomitantly reduced EAC14 rosette-forming lymphocytes. Binding of EAC14 intermediates to receptors on human lymphocytes and erythrocytes could be inhibited by equal amounts of soluble C3b or C4b, suggesting the presence of a single receptor for both ligands on those cells. In contrast, the results of the rosette assay with Raji cells, cultured human lymphoblastoid cells, EAC14 and EAC1423 suggested that the receptors for C4b and C3b are distinct entities, since Raji cells formed rosettes with EAC1423, but not with EAC14. Moreover, this report demonstrates a cooperation of erythrocyte-bound C4b and C3b in the binding of EAC1423 to B lymphocytes. In contrast to KAF-treated C3b, KAF-treated C4b did not bind to B lymphocytes, indicating that these cells lack a receptor for C4d.
EFFECTS OF CONCANAVALIN A ON MOUSE PERITONEAL MACROPHAGES
Authors: Edelson Paul J. , Cohn Zanvil A.
Published: June 2004

Concanavalin A (Con A) binds to saccharide residues on the mouse peritoneal macrophage plasma membrane and stimulates extensive pinocytic interiorization of the membrane. The overall pinocytic rate is increased 3.5–4.5 times by the addition of Con A, and the surface marker enzyme adenosine triphosphatase can be identified histochemically in association with the cytoplasmic vesicles generated after exposure of the cells to Con A. Once formed, these pinocytic vesicles may persist for several days and fail to show morphologic evidence of fusion with primary or preformed secondary lysosomes. There is no apparent effect on the capacity of the macrophage to ingest either latex particles or IgG-coated SRBC administered either simultaneously with or subsequent to the Con A.
GENETIC CORRELATION OF A MOUSE LIGHT CHAIN VARIABLE REGION MARKER WITH A THYMOCYTE SURFACE ANTIGEN
Authors: Gottlieb Paul D.
Published: June 2004

The inbred and congenic strain distribution of the IH-peptide marker in the variable region of mouse immunoglobulin light chains has been compared with other known genetic markers. A positive correlation was noted between the IH-peptide marker and expression of the Ly-3.1 thymocyte cell surface antigen. This suggests that the locus responsible for IH-peptide expression is genetically linked to the Ly-2 and Ly-3 loci in linkage group XI on chromsome 6 of the mouse.