bluuebunny/crossref_metadata_2025 huggingface dataset:
Susceptibility of Adenovirus 2-transformed rat cell lines to natural killer (NK) cells: direct correlation between NK resistance and in vivo tumorigenesis.
Authors: Sheil J M , Gallimore P H , Zimmer S G , Sopori M L
Published: December 2022

The susceptibility to natural killer (NK)-mediated cell lysis of Adenovirus type 2 (Ad2)-transformed rat embryo fibroblast cell lines, which differed markedly in tumorigenic potential in vivo (T2C4 greater than F19 greater than F17), was investigated by using NK effector cells from F344 rat or athymic nude rat spleens. A comparison of the degree of NK-mediated lysis obtained with these tumor cell targets suggested a direct relationship between the resistance of a cell to NK cell lysis and its potential to form tumors in vivo. The cells were lysed in the following order of increasing susceptibility: T2C4 less than F4 less than F19 less than F17. Whether T cells or macrophages played a significant role in the observed lytic activity was determined by treating the NK effector cell population with anti-rat T cell serum (alpha T) and complement or by depletion of macrophages after binding to a glass bead column and treatment with carbonyl iron. A series of clonal sublines derived from the parental F17 and F4 cell lines further strengthened this relationship between tumorigenesis and resistance to NK-mediated cell lysis. Tumorigenic subclones from the non-tumorigenic F17 parental cells were demonstrated to be comparatively resistant to NK-mediated lysis. Tumorigenic subclones from tumorigenic F4 parental cell population showed a susceptibility to NK-mediated cell lysis virtually identical to the parental F4 cells. The implication of these results are discussed.
A multistage model for the action of cytotoxic T lymphocytes in multicellular conjugates.
Authors: Macken C A , Perelson A S
Published: December 2022

We propose a multistage stochastic model to explain data on the kinetics of target cell lysis by cytotoxic T lymphocytes in multicellular conjugates. A novel feature of our model is that we explicitly consider both the lethal hitting stage and the target cell disintegration stage of the cytolytic process. Further, we allow for the possibility that target cell disintegration is itself a complex process composed of many events. The comparison of our model with the data of other investigators suggests that cytotoxic T cells deliver lethal hits at random to undamaged target cells. Having received a lethal hit, the target cell disintegrates over a variable length of time. The disintegration times of target cells from different conjugates appear to be randomly distributed and to be consistent with a model in which disintegration occurs by at least two major, sequential, rate-limiting events. For conjugates containing one lymphocyte and multiple target cells, the mean rate at which a lethally hit target cell disintegrates is found to be independent of the total number of target cells in the conjugate. Our model predicts that in such multicellular conjugates, individual target cells lyse one by one, on average at approximately 30-min intervals, thus agreeing closely with previously reported experimental observations.
Regulation of anti-hapten antibody responses in vivo: memory, carrier-specific Lyt-2+ T cells can terminate antibody production without altering the peak of response.
Authors: Kanellopoulos-Langevin C , Mathieson B J , Perkins A , Maynard A , Asofsky R
Published: December 2022

By using the classical hapten-carrier antigenic system (dinitrophenyl-keyhole limpet hemocyanin) (DNP-KLH), and positive immunoselective techniques for purification of lymphocyte subsets, we studied the kinetics and role of KLH-primed T cells in the termination of a secondary in vivo plaque-forming cell (PFC) response against DNP in the mouse. Separation of T cells by Lyt-2 phenotype and recombination of T cell subsets demonstrated that a single injection of KLH in complete Freund's adjuvant (CFA) concomitantly generated both helper and suppressor T cells in the spleens of primed mice. When these KLH-primed, purified T cells were transferred into irradiated recipients along with DNP-primed, purified B cells and DNP-KLH, helper activity was demonstrable first, with the early production of a high anti-DNP PFC response. Suppression of the PFC response was observed only after the response had fully developed; the PFC response was then "terminated" within a few days. Helper activity was obtained with the carrier-primed Lyt-2- T cell population, whereas suppression required the addition of Lyt-2+ T cells. Both activities were generated over a wide dose range, were specific for the priming antigen KLH, and could be detected as early as 8 days or as late as 9 mo after a single priming injection of KLH. Lyt-2+ T cells from unprimed donors or from animals primed with a different carrier did not have the capacity to terminate the response when added with the primed Lyt-2- helper T cells. Thus, we have now demonstrated the existence of carrier-specific, long-lived, "memory" suppressor T cells, which are initially generated in parallel with specific helper T cells. Whereas these memory suppressor T cells do not inhibit the induction of a secondary antibody response, they can effectively terminate it once it has developed, and may therefore account for the normal termination of antibody responses.
Alpha-fetoprotein inhibits macrophage expression of Ia antigens.
Authors: Lu C Y , Changelian P S , Unanue E R
Published: December 2022

Murine alpha-fetoprotein (AFP) is a major protein in amnionic fluid and perinatal sera. AFP has been postulated to contribute to the immunologic hyporesponsiveness of the fetus and neonate, as well as protecting the fetus from rejection by the mother. We now report that AFP acts in vitro to inhibit macrophage expression of cell surface Ia antigens, the class II glycoproteins of the major histocompatibility gene complex. In culture, macrophages incubated with a T cell lymphokine developed Ia as detected by either immunofluorescence or a cell radioimmunoassay. Both mouse amnionic fluid and AFP inhibited the expression of Ia in a dose-dependent manner. Five preparations of AFP derived from three sources--day-old neonates, amnionic fluid, and a hepatoma cell line--were effective. The concentration of AFP that inhibited by 50% the expression of Ia was about 10(-6) M. Mouse amnionic fluid and AFP did not affect the viability of the macrophage nor was the surface expression of H-2K and C3 receptors affected. The inhibitory effect of AFP did not depend on the presence of T cells in the culture. AFP did not appear to inhibit the direct interaction of lymphokine with macrophages; AFP did inhibit if given days after a pulse of lymphokine. The concentrations of prostaglandins carried by AFP were insignificant and could not explain the inhibitory effects.
A role for macrophages in suppressor cell induction.
Authors: Usui M , Aoki I , Sunshine G H , Dorf M E
Published: December 2022

A mechanism responsible for the induction of NP-specific first order (inducer) suppressor cells (TS1) is described. TS1 cells are induced by i.v. administration of hapten-coupled splenic cells. Their activity is assessed by the adoptive transfer of NP-specific suppression during the afferent phase of the contact sensitivity response. NP-coupled firmly adherent, FcR+, I-A-bearing macrophages induce TS1. The antigen-presenting cells required for TS1 induction lack the Thy-1 and Lyt-1 markers, and are resistant to 500 R irradiation and to cyclophosphamide treatment. NP-coupled dendritic cells fail to induce TS1 activity. The induction of TS1 cells is genetically restricted by genes that map in the I-J region of the H-2 complex. The NP-coupled antigen-presenting cells must share at least one I-J allele with the TS1 donor for effective induction of TS1 activity. To minimize allogeneic effects in these studies, the activity of the TS1 population was assessed by adoptive transfer into syngeneic recipients. The present results are compared with the mechanisms required for the induction of second and third order suppressor cells.
Macrophage-hybridomas: generation, structure, and function.
Authors: Tzehoval E , Segal S , Zinberg N , Feldman M
Published: December 2022

We report the generation of macrophage-hybridomas, obtained by somatic cell fusion between macrophage-enriched C3H.eB spleen cell population, and a drug-resistant MPC-11 myeloma cell line, designated as 4T00.1L1 clone. Screening for hybridomas possessing macrophage properties was carried out by assaying the presence of two macrophage-specific enzymes: lysozyme and nonspecific esterase. Two hybridomas, E2-7 and E2-10, were selected for further studies. We found that clones of E2-7 (E2-7.7) did not express Fc receptors but possessed cell-surface Ia molecules. In contrast, clones of E2-10 (E2-10.20) possessed Fc receptors but were devoid of Ia molecules. E2-7.7 did, however, express Fc receptors after mitomycin treatment, whereas E2-10.20 eliminated the expression of Fc receptors after treatment with mitomycin C. Opsonized erythrocytes were phagocytized by E2-10.20 cells, but not by E2-7.7. Phagocytosis was thus correlated with the possession of Fc receptors. Testing the response of KLH-primed lymph node cells to KLH-pulsed hybridoma cells, we found that E2-7.7 cells caused antigen-specific lymphoproliferative response, whereas E2-10.20 did not. Thus, antigens could be presented by E2-7.7 but not by E2-10.20 cells. The response was shown to be mediated by T but not by B lymphocytes. The difference in antigen-presenting capacity could not be attributed to differences in antigen uptake by the different hybridomas, because the two hybridomas manifested the same level of pinocytosis. Both hybridomas produced IL1. The differences in the properties of the two hybridomas may indicate that the normal partners represent two distinct subpopulations of macrophages. The segregation of functional properties among the hybridoma clones may lead to a clarification of the dependence of distinct functions on defined molecular structures.
Production of lymphokines by circulating human T lymphocytes that express or lack receptors for interleukin 2.
Authors: Palacios R
Published: December 2022

The capacity for circulating human T cells which have or lack receptors for interleukin 2 (IL 2) to produce IL 2, interleukin 3 (IL 3), and interferon-gamma (IFN-gamma) under the stimulus of phytohemagglutinin was studied. By using the monoclonal anti-Tac antibody which reacts against IL 2 receptors on human T cells, concanavalin A-treated T cells were separated into IL 2 receptor-positive (Tac+ T cells) and IL 2 receptor-negative (Tac- T cells) lymphocytes. The results show that Tac+ T cells secreted IL 2 and IFN-gamma but not IL 3. Tac- T cells produced IL 2 and IL 3 but not IFN-gamma. It is concluded that: 1) both T cells lacking and T cells having receptors for IL 2 produce IL 2, but only IL 2 receptor-negative T cells appear to secrete IL 3; and 2) virtually all of the T cells that produce IFN-gamma after PHA stimulation express receptors for IL 2.
The mechanisms of inhibition of human IL 2 production. II. PGE2 induction of suppressor T lymphocytes.
Authors: Chouaib S , Chatenoud L , Klatzmann D , Fradelizi D
Published: December 2022

In vitro and in vivo experiments indicate that the production of interleukin 2 (IL 2) by T lymphocytes is critical for the development of the effector phase of immunity. Complex cellular interactions are involved for the induction of IL 2 production. We have shown in a previous study that in humans monocytes can transmit opposite signals to the IL 2-producing cells. In addition to the positive signal delivered through the release of interleukin 1, human monocytes can deliver a negative signal through the release of prostaglandin E2 (PGE2). This monokine, known to activate suppressor mechanisms in several systems, was shown to inhibit IL 2 production. The data presented in this paper show that this PGE2-dependent inhibition is strictly dependent upon the presence of radiosensitive T cells in the culture, suggesting that PGE2 induces the activation of suppressor T cells modulating IL 2 production. Kinetics experiments indicate that these suppressor cells are radiosensitive during their induction phase but become radioresistant after 18 hr of incubation in the presence of PGE2. Successful in vitro induction of suppressor cells by incubation of enriched T cells with PGE2 was decisive for the analysis of the phenomenon. The induced suppressors were capable of inhibiting IL 2 production by fresh autologous T cells as well as inhibiting PHA proliferative response by these cells. A quantitative evaluation of IL 2 receptors on PGE2-treated cells has indicated that this absorption capacity was similar to the capacity of PBL known to express a low number of IL 2 receptors, thus excluding a suppression via absorption or competition for IL 2. No detectable killing of IL 2-producing cells by PGE2-induced suppressors was observed. The OKT4 and OKT8 phenotype of suppressor cells was examined. T cells were purified at two stages of differentiation before or after induction by PGE2 in vitro treatment. We conclude from these experiments that PGE2 activates suppressor cells among precursors segregating predominantly with the OKT8 subset and fewer cells with the OKT4 subset. After differentiation, however, the suppressor cells segregate with the OKT8 subset only. Such results were obtained by using positive selection (cellular affinity columns) and negative selection (monoclonal antibodies plus complement).
Limiting dilution analysis of interleukin 2 and colony-stimulating factor producer cells in normal and autoimmune mice.
Authors: Hefeneider S H , Conlon P J , Dower S K , Henney C S , Gillis S
Published: December 2022

MRL/MP lpr-lpr (MRL-lpr) mice spontaneously develop an age-related disease characteristic of human systemic lupus erythematosus (SLE). Old MRL-lpr mice (4 mo of age) develop antibodies to nucleic acids, display immune complex glomerulonephritis, and have a massive T cell-associated lymphadenopathy. In concert with disease development is data showing an age-related loss of interleukin 2 (IL 2) production by mitogen-stimulated lymphoid cells from these mice. The loss of IL 2 production has been suggested to be involved in the onset and/or development of autoimmune disease seen in these animals. In this report, we examined the frequency of both IL 2 and colony-stimulating factor (CSF) producer T cells in the MRL-lpr mouse by using a limiting dilution analysis assay. Our results show that the number of IL 2 and CSF producer cells present in autoimmune animals is similar to the number found in normal control mice. In addition, IL 2 and CSF producer T cells from autoimmune MRL-lpr mice make similar levels of lymphokine activity, as do producer T cells from normal mice. Our data argue against the previously hypothesized role that a paucity of IL 2 production may be involved in the etiology of autoimmune disease.
Genetic polymorphism of human factor H (beta 1H).
Authors: Rodríguez de Córdoba S , Rubinstein P
Published: December 2022

Human Factor H (beta 1H) was found to be polymorphic after neuraminidase treatment and isoelectric focusing (IEF) under completely denaturing conditions. Three variants, FH 1, FH 2, and FH 3, were identified in a sample population of 81 unrelated caucasoid individuals. Family studies demonstrated correct mendelian segregation of FH 1, FH 2, and FH 3. Our data indicate that these genetic variants of human Factor H are encoded by three codominant alleles, FH*1, FH*2, and FH*3, at a single autosomal locus FH. In the sample analyzed, the gene frequencies of FH*1, FH*2, and FH*3 were, respectively, 0.691, 0.302, and 0.006.
Biochemical and functional characterization of proteoglycans isolated from basophils of patients with chronic myelogenous leukemia.
Authors: Metcalfe D D , Bland C E , Wasserman S I
Published: December 2022

Human basophils were obtained from three donors with myelogenous leukemia. Proteoglycans were labeled by using [35S]sulfate as precursor and were extracted in 1 M NaCl with protease inhibitors to preserve their native structure. [35S]proteoglycans filtered on Sepharose 4B with an average m.w. similar to that of a rat heparin proteoglycan that has an estimated m.w. of 750,000. The [35S]glycosaminoglycan side chains filtered with an average m.w. slightly smaller than a 60,000-m.w. glycosaminoglycan marker. The [35S]glycosaminoglycans were resistant to heparinase and susceptible to degradation by chondroitin AC lyase and chondroitin ABC lyase. The intact [35S]glycosaminoglycans chromatographed on DEAE Sepharose as a single peak eluting just before an internal heparin marker. These findings indicate that the [35S]glycosaminoglycans were made up only of chondroitin sulfates. No heparin was identified. The chondroitin sulfate disaccharides that resulted from the action of chondroitin ABC lyase on the basophil glycosaminoglycans consisted of 92% delta Di-4S, 6% delta Di-6S, and 2% disulfated disaccharides. The [35S]chondroitin sulfate proteoglycans were susceptible to cleavage with proteases and could be shown to be released intact from basophils during degranulation initiated by the calcium ionophore A23187. The basophil proteoglycans and glycosaminoglycans were capable of binding histamine in water, but not in phosphate-buffered saline, and had no anticoagulant activity.
Increasing extracellular potassium causes calcium-dependent shape change and facilitates concanavalin A capping in human neutrophils.
Authors: Roberts R L , Mounessa N L , Gallin J I
Published: December 2022

Stimulation of human polymorphonuclear leukocytes (PMN) by chemoattractants is associated with changes in membrane potential, and it has been speculated that these changes are important in PMN function. In the present study, human PMN isolated on Percoll gradients were exposed to extracellular K+ concentrations, which have been shown to depolarize PMN. Incubation of PMN at 37 degrees C for 15 min in 125 mM K+ increased the number that capped fluoresceinated concanavalin A (Con A) to 78% +/- 5.1 with 6.9% +/- 0.8 forming Con A caps in 5 mM K+ (p less than 0.001). In the absence of lectins, high K+ induced polarization of PMN shape with pseudopod and uropod formation (2.4% +/- 0.1 polarized in 5 mM K+ vs 83.4% +/- 1.0 polarized in 125 mM K+ p less than 0.001). Control studies, in which Na+ was decreased by substituting choline chloride or sucrose, indicated that the K+ effects were specific and not due to lowering of Na+. Increasing K+ also stimulated preferential secretion of the specific (secondary) granule constituents, caused significant mobilization of receptors for the peptide N-formyl-methionyl-leucyl-phenylalanine (fmet-leu-phe), and increased the number of PMN migrating in chemotactic assays. Unlike the capping of Con A and shape change, however, the increase in fmet-leu-phe binding, degranulation, and locomotion was not specific for raising extracellular K+ and occurred when choline chloride was substituted for sodium chloride, suggesting that these effects may be due to the lowering of Na+ or the changing of the Na+ to K+ ratio. In related studies designed to examine the mechanism of these K+-induced effects, shape change and Con A cap formation were inhibited by the absence of extracellular calcium or the presence of the calcium channel blockers, methoxyverapamil (100 microM), manganese (2 mM), or cobalt (2 mM), suggesting that the K+-induced changes depend on an influx of calcium. Thus, the increase of extracellular K+ induces calcium-dependent membrane changes in PMN similar to those seen when PMN are stimulated by a chemoattractant.
Auranofin affects early events in human polymorphonuclear neutrophil activation by receptor-mediated stimuli.
Authors: Hafström I , Seligmann B E , Friedman M M , Gallin J I
Published: December 2022

Auranofin, a new oral antirheumatic gold compound, in concentrations achieved therapeutically, inhibits neutrophil phagocytosis, chemotaxis, chemiluminescence, reduction of cytochrome c, and release of lysosomal enzymes. To further characterize the mechanism by which auranofin affects neutrophils, we studied the effects of auranofin on unstimulated properties and functions of neutrophils as well as on rapidly stimulated functions. When examined by electron microscopy, 4 micrograms/ml of auranofin significantly decreased the number of visualized centriole-associated microtubules in resting cells. Furthermore, auranofin inhibited neutrophil spreading on glass and caused a decrease in negative surface charge (electrophoretic mobility). In addition, auranofin inhibited several fmet-leu-phe-stimulated responses such as shape change, increases in centriole-associated microtubules, decreases in surface charge, and elicited membrane potential changes (di-O-C5(3) dye response). Auranofin (1 micrograms/ml) inhibited fmet-leu-phe-stimulated superoxide and hydrogen peroxide production by 80% (p less than 0.05), and also increased the affinity of receptors for fmet-leu-phe (from Ka 0.035 to Ka 0.48, p less than 0.001). Auranofin also affected neutrophil responses to phorbol myristic acetate (PMA). The total amount of PMA-stimulated superoxide production was suppressed by as little as 0.4 micrograms/ml of auranofin, but the lag time for activation was shortened by low concentrations of auranofin (0.5 to 1 microgram/ml). Four micrograms per milliliter of auranofin suppressed the decrease in surface charge induced by PMA. However, auranofin did not influence superoxide production elicited by the ionophore A23187. The results indicate that auranofin affects the earliest detected responses in neutrophil activation by certain receptor-mediated stimuli.
Deoxyadenosine (AdR) inhibition of newly activated lymphocytes: blockade at the G0-G1 interface.
Authors: Redelman D , Bluestein H G , Cohen A H , Depper J M , Wormsley S
Published: December 2022

The mechanism responsible for the lymphocytotoxicity associated with congenital adenosine deaminase (ADA) deficiency has been ascribed to an accumulation of dATP. Elevated levels of dATP can then lead to inhibition of DNA synthesis by inhibiting ribonucleotide reductase and causing a depletion of the other deoxynucleotide triphosphates (dNTP). This hypothesis was derived principally from studies with murine and human lymphoblastoid cell lines (LCL) and apparently confirmed in a limited number of investigations with lectin-stimulated lymphocytes. Our biochemical studies of lectin-stimulated mouse and human lymphocytes were not consistent with the dATP model and suggested that AdR exerted effects on lymphocyte activation that preceded the initiation of DNA synthesis. In the current studies, we focused on the effects of AdR on the early events in T lymphocyte activation, because we found they were the most sensitive to AdR toxicity. AdR blocked neither the production of T cell growth factor (TCGF) by lectin-stimulated lymphocytes nor the expression of TCGF receptors as detected by the anti-Tac monoclonal antibody that recognizes the human TCGF receptor. AdR did, however, block the early TCGF-dependent events leading to the entry into the cell cycle. By using the metachromatic fluorescence stain acridine orange, we found that AdR blocked the increased synthesis of RNA that characterizes the entry into the G1 phase of the cell cycle from the G0, resting state. Because these early effects were caused by the lowest doses of AdR, and because they preceded the synthesis of DNA by 15 to 20 hr, it suggested that these effects may be principally responsible for the in vivo toxicity associated with ADA deficiency. Furthermore, none of the other proposed biochemical mechanisms, e.g., inhibition of methylation, diminution of ATP levels, or incorporation of AdR into polyadenylated RNA, appeared adequate to explain AdR toxicity during T lymphocyte activation.
Characterization of monoclonal antibody 155.8 and partial characterization of its proteoglycan antigen on human melanoma cells.
Authors: Harper J R , Bumol T F , Reisfeld R A
Published: December 2022

Immunization of mice with a plasma membrane-enriched fraction from human malignant melanoma cells and subsequent generation of hybridomas resulted in the isolation of an IgG1 monoclonal antibody, 155.8, that recognizes chondroitin sulfate proteoglycans. By cell binding analysis, 155.8 was shown to react with seven of eight cultured melanoma cell lines, but not with a variety of lymphoblastoid cell lines or cultured tumor cells derived from other solid tumor types. Indirect immunoprecipitation of the 155.8 antigen from intrinsically labeled melanoma cells revealed a glycoprotein of Mr = 250,000 and a sulfated molecule of Mr greater than 400,000. The antigen was identified as a chondroitin sulfate type A/C proteoglycan synthesized by melanoma cells on the basis of its sensitivity to chondroitinase ABC digestion and the identification of sulfated glycosaminoglycans released from the antigen immunoprecipitated by 155.8. The determinants recognized by antibodies 155.8 and 9.2.27, another anti-chondroitin sulfate proteoglycan, immunoprecipitate only a proteoglycan from high density cesium chloride gradient fractions, (1.487 g/liter); however, they immunoprecipitate a free glycoprotein of Mr = 250,000 from low density fractions (1.317 g/liter). This demonstrated that the 155.8 and 9.2.27 determinants, both of which reside on the glycoprotein of Mr = 250,000, are also present in the proteoglycan, suggesting that this glycoprotein is the proteoglycan core protein. Monoclonal antibody 155.8 reacts with a determinant on the core protein distinct from that recognized by 9.2.27. Proteoglycans bearing 155.8 determinants are distributed on the surface of cultured melanoma cells in a punctated fashion that apparently resolves to short, filamentous structures at high magnification. Immunohistochemical analysis demonstrated that 155.8-defined proteoglycans are found in freshly biopsied melanoma tissue, suggesting that these antigens are also synthesized in vivo by melanoma cells.
Potentiating effect of muramyl dipeptide and its lipophilic analog encapsulated in liposomes on tumor cell killing by human monocytes.
Authors: Sone S , Mutsuura S , Ogawara M , Tsubura E
Published: December 2022

Human monocytes harvested from healthy donors and separated by discontinuous gradient centrifugation and adherence were highly cytotoxic to allogeneic melanoma cells, but did not affect nontumorigenic cells, as measured by lysis of the cells. After 4 days incubation of these monocytes in medium, they showed little, if any, cytolytic activity. Hydrophilic muramyl dipeptide (MDP) or lipophilic muramyl tripeptide (MTP-PE) was encapsulated within multilamellar (MLV) liposomes composed of phosphatidylserine-phosphatidylcholine. The cultured monocytes were rendered tumoricidal by interaction for 24 hr with MDP or liposomes containing soluble MDP or lipophilic MTP-PE. Moreover, freshly isolated monocytes treated for 24 hr with liposomes containing MDP or MTP-PE remained tumoricidal during culture for up to 5 days. About 1,600-times lower concentration of MDP entrapped in liposomes than of free MDP in the medium was effective for rendering monocytes tumoricidal. Similarly, about 80-times lower concentration of MTP-PE in liposomes than of free MDP was effective for the activation of monocytes. It is concluded that MLV liposomes containing MDP or MTP-PE are far more efficient in potentiating the tumoricidal activity of human monocytes than unencapsulated, free MDP.
Induction of NKCF-like activity in mixed lymphocyte-tumor cell culture: direct involvement of mycoplasma infection of tumor cells.
Authors: Wayner E A , Brooks C G
Published: December 2022

Co-culture of CBA/J spleen cells and certain lines of YAC-1 stimulators resulted in the appearance of NKCF-like activity in 24- to 48-hr supernatants. Numerous other in vitro cell lines were effective stimulators of this splenic cytotoxic factor (SCF). The cells participating in SCF production were absent from normal thymocytes and were present in BALB/c nu/nu spleen, were nonadherent, asialo GM1+, and bore low levels of Thy-1.2. SCF could mediate lysis of certain NK-sensitive tumor targets in an 18-hr 51Cr-release assay. However, the induction of SCF was not correlated with the ability of a particular cell line to be lysed by NK cells, but showed an absolute correlation with the presence of mycoplasma contamination in cultured tumor cell lines. Mycoplasma negative cell lines, including an uninfected but NK-sensitive subline of YAC-1, were unable to induce SCF. Decontamination of mycoplasma-infected lines with antibiotics or by passage through syngeneic mice abrogated the ability of infected tumor cells to stimulate SCF. The ability to induce SCF could be restored by reinfection with mycoplasma. Tumor cell-free supernatants from contaminated cultures were mitogenic for CBA spleen cells and could themselves induce SCF activity in spleen cell supernatants. SCF production and the agent responsible could be removed by passing such supernatants through 0.1-micron filters. The organism apparently responsible for SCF induction from CBA spleen cells was typed and found to be Mycoplasma orale, a nonfermentative, arginine-dependent, common tissue culture contaminant. About 50 to 60% of SCF activity could be removed by 0.1-micron filters, suggesting that SCF is composed of two components: mycoplasma organisms themselves and a soluble cytotoxic factor produced in response to mycoplasma.
T cell-accessory cell interactions that initiate allospecific cytotoxic T lymphocyte responses: existence of both Ia-restricted and Ia-unrestricted cellular interaction pathways.
Authors: Singer A , Kruisbeek A M , Andrysiak P M
Published: December 2022

The specificity of the T-accessory cell interactions that initiate primary allospecific cytotoxic T lymphocyte (CTL) responses were found to be surprisingly diverse and of three distinct major histocompatibility complex (MHC) specificities, involving responder T cell recognition of: a) self-Ia accessory cell determinants, b) allo-Ia accessory cell determinants, or c) allo-K/D accessory cell determinants. Any one of these T-accessory cell interactions was sufficient to initiate allospecific CTL responses. It was observed that when accessory cells did not express foreign class I MHC determinants, primary allospecific CTL responses were invariably initiated by Ia-restricted T-accessory cell interactions. In contrast, it was observed that when accessory cells did express foreign class I MHC determinants, primary allospecific CTL responses could be initiated by Ia-independent T-accessory cell interactions that were specific for allogeneic, but not self, K/D determinants and that did not involve recognition of polymorphic Ia determinants. The MHC specificities of the T-accessory cell interactions that initiate primary allospecific and primary trinitrophenyl (TNP)-self CTL responses were also compared. It was observed that primary allospecific and primary TNP-self CTL responses could be initiated by self-Ia-restricted T-accessory cell interactions, and that in both responses the Ia determinants that the responding T cells recognized as self-specificities on the accessory cell surface were those that their precursors had encountered on radiation-resistant thymic elements in their differentiation environment. In contrast to the initiation of primary TNP-self CTL responses that required the activation by accessory cells of Ia-restricted T helper (TH) cells, allospecific CTL responses could also be initiated by class I-restricted T cells specific for accessory cell K/D determinants. Interestingly, such class I-restricted T cells present in primary responder cell populations were triggered only by recognition of allogeneic, but not self, K/D accessory cell determinants, even when the accessory cells were modified with TNP. Thus, the present study demonstrates that primary allospecific CTL responses, but not TNP-self CTL responses, are initiated by Ia-restricted or Ia-independent cellular interaction pathways. These results raise the possibility that unprimed class I-restricted TH cells that mediate the Ia-independent cellular interaction pathway may predominantly express an allospecific, but not a self + X-specific, receptor repertoire. Possible mechanisms by which these distinct T-accessory cell interactions initiate primary allospecific CTL responses are discuss
Modulation of F1 cytotoxic potentials by GvHR: role and mode of action of non-MHC genes that determine the hybrid resistance to GvHR-associated suppression of F1 cytotoxic potential.
Authors: Ishikawa H , Kubota E , Saito K
Published: December 2022

The resistance of unirradiated F1 mice against graft-vs-host reaction (GvHR) induced by lymphocytes from certain parental strains is apparently a violation of the basic law in classical transplantation immunity. To explore genetic mechanisms of this peculiar phenomenon, GvHR-associated immunosuppression was examined on various kinds of F1 mice undergoing GvHR induced by parental lymphocytes. In F1 mice raised by crossing DBA/2 mice with various H-2-congeneic B10-series strains, parental lymphocytes having non-H-2 genetic background of DBA (DBA/2 and DBA/1) invariably could not induce GvHR-associated immunosuppression, irrespective of the H-2 haplotype incompatibility involved, whereas lymphocytes of the partner parental strain induced the immunosuppression. The number of the relevant loci in the DBA non-H-2 was assessed to be three recessive loci by examination of the capability to induce the GvHR-associated immunosuppression on lymphocytes from individual (B 10.D2 X DBA/2)F1 X DBA/2 backcross mice. On the other hand, in F1 mice raised by crossing C3H/He or AKR/J mice with various H-2-congeneic B10-series strains, parental lymphocytes of H-2k haplotype, irrespective of their non-H-2 haplotype, invariably could not induce the GvHR-associated immunosuppression. Furthermore, it was revealed that non-H-2 genes of parental C3H or AKR incorporated in the F1 mice determine the resistance of the F1 mice against the H-2k-induced GvHR. The results of examination of the resistance on individual (B10 X [B10.BR X C3H/He]F1) and (B10 X [B10.BR X AKR/J]F1) mice suggested that three non-H-2 loci of C3H/He or two non-2 loci of AKR/J incorporated in F1 hybrids could determine the resistance of the respective F1 mice.
The immunocompetence of murine stromal cell-associated thymocytes.
Authors: Fink P J , Weissman I L , Kaplan H S , Kyewski B A
Published: December 2022

Thymocyte subpopulations that associate in vivo with distinct nonlymphoid cells of the thymus have been isolated, and their immunocompetence was analyzed. Previous studies have indicated that greater than 95% of such cells bear a surface antigen phenotype representative of immature thymocytes, and are among the earliest thymic compartments repopulated by bone marrow-derived cells after lethal and sub-lethal irradiation. Stromal cell-associated thymocytes may be activated in vivo because they proliferate well in vitro with no additional stimulus, and show little increase in proliferation with the addition of T cell mitogens or allogeneic spleen cells. Stromal cell-associated T cells contain cytotoxic T lymphocyte (CTL) precursors that are indistinguishable from mature peripheral T cells by the parameters of self tolerance, alloreactivity, H-2 restriction, and stringency of self H-2 preference. CTL precursor frequencies and the cytotoxic activity of cells further separated on the basis of high levels of Thy-1 expression argue against the possibility that stromal cell-associated CTL activity is due solely to contaminating mature lymphocytes. Our data suggest that stromal cell-associated thymocytes represent an intermediate subpopulation of thymocytes that is functionally mature and that expresses an immature surface phenotype. Furthermore, the imposition of self tolerance and MHC restriction specificity appears to be tightly associated with the acquisition of immunocompetence in these thymocyte subpopulations.
Conversion of a tolerogenic to an immunogenic signal by P388AD.2 cells, a lymphoid dendritic cell-like tumor line.
Authors: Phipps R P , Pillai P S , Scott D W
Published: December 2022

The lymphoid dendritic cell-like tumor P388AD.2 is capable of converting a tolerogenic signal into an immunogenic one. In the present study, adherent P388AD.2 cells were pulsed with the tolerogen, fluoresceinated (FL) sheep gamma-globulin (SGG), washed, and incubated with normal spleen cells. After 24 hr, the spleen cells were harvested and challenged in secondary cultures with immunogenic doses of FL-thymic independent (TI) antigens. The plaque-forming cell response on day 3 of secondary culture was increased as much as 400% compared with control spleen cultures exposed to untreated P388AD.2 cells. The increased response was specific for the FL-hapten and occurred only when P388AD.2 cells were pulsed with FL-Ig or FL-F(ab')2 fragments, but not with FL-synthetic tolerogens or other FL-antigens. Furthermore, the augmentation required histocompatible T cells in the primary cultures. Additional experiments showed that if cultures were devoid of Lyt-1+ cells but not Lyt-2+ cells, no augmentation occurred. A variety of other macrophage-like tumor cells, some with known antigen presenting properties, were tested for the ability to present tolerogen in an immunogenic fashion. Only an I-A+ J774 clone was able to present tolerogen in an immunogenic fashion. However, we failed to find a correlation between the presence of surface Ia antigen and tolerogen-presenting ability. The results suggest that certain types of cells may play a role in immune regulation by abrogating the tolerogenicity of Ig tolerogens via their presentation in an immunogenic mode.
IgG-protein A complexes modulate thymidine incorporation into DNA of antibody and complement-stimulated cells.
Authors: Shearer W T , Green C G , Patel P , Langone J J
Published: December 2022

Incubation of protein A (PA) and molar excess of whole rabbit immune IgG in solution favored the formation of high m.w. complexes with the molecular formula of [(IgG)2(PA)]2, which enhanced the incorporation of [3H]thymidine ([3H]dThd) into DNA of L cells, especially in the presence of rabbit or human complement (C). In contrast, complexes with the molecular formula (IgG)(PA), formed by incubation of IgG with molar excess of PA, failed to enhance, or inhibited, the incorporation of [3H]dThd into cells treated with immune IgG alone or with C present, respectively. Concanavalin A (Con A), which is known to bind to critical exposed oligosaccharide structures present in the Fc region of IgG in the [(IgG)2(PA)]2 complexes, reversed the enhancement of [3H]dThd incorporation, and alpha-methyl mannoside, but not D-galactose, inhibited the Con A reversal. The [(IgG)2(PA)]2 complexes efficiently activated the C cascade, caused significantly higher levels of aggregation of cells, and increased the binding of immune IgG to the cells when compared to the (IgG)(PA) complexes. Both types of complexes, isolated by sucrose density ultracentrifugation, were precipitable in a radioimmunoassay with chicken anti-PA antibodies and were capable of binding to intact L cells.
The effect of deoxyguanosine on human lymphocyte function. II. Analysis of the interference with B lymphocyte differentiation in vitro.
Authors: Spaapen L J , Rijkers G T , Staal G E , Rijksen G , Duran M , Stoop J W , Zegers B J
Published: December 2022

The differentiation of normal human peripheral blood B lymphocytes into plasma cells in vitro, studied in mononuclear cells stimulated with PWM or in purified B cells stimulated with a T cell-replacing factor (TRF), can be inhibited by both deoxyguanosine (dGuo) and guanosine. The mechanism underlying this effect, which differs from the in vivo findings in PNP deficiency, was analyzed. dGuo toxicity can be antagonized by hypoxanthine but not by deoxycytidine. PNP-deficient and HGPRT-deficient B lymphocytes are not sensitive to the intoxicating properties of (deoxy)guanosine. Inhibition of PNP activity in normal B lymphocytes by 8-aminoguanosine decreases the sensitivity for dGuo intoxication. Incubation of purified B cells (stimulated with TRF) with dGuo leads to increased intracellular levels of guanosine di- and triphosphate (GDP and GTP), whereas deoxyguanosine triphosphate (dGTP) levels remain low. These observations lead to the conclusion that inhibition of B lymphocyte differentiation by dGuo is brought about by one of the end products of the pathway starting with degradation of dGuo by PNP, followed by guanine salvage by HGPRT, and possibly further phosphorylation of GMP into GDP and GTP. According to this mechanism, B lymphocyte differentiation in PNP deficiency is not sensitive to (deoxy)guanosine; because of the absence of PNP activity, these cells cannot accumulate GMP, GDP, and GTP, and therefore escape dGuo intoxication.
Discordant expression of human Ia-like antigens on hematopoietic progenitor cells.
Authors: Linch D C , Nadler L M , Luther E A , Lipton J M
Published: December 2022

The expression of HLA-DR, SB, MT2, and DC antigens on human hematopoietic progenitor cells has been determined by using monoclonal antibodies with complement (C)-mediated cell lysis and immune separation techniques. HLA-DR was detected on greater than 85% of CFU-G/M, myeloid clones (MyCl), BFU-E, and CFU-E. CFU-E were less susceptible to C-mediated lysis at suboptimal C concentrations. The polymorphic MT2 and SB antigens were also present on all categories of progenitor cells, although a lesser proportion of cells were positive. Because in most individuals the antigen density of MT2 and SB, as determined by monoclonal antibody staining, was also lower on B cells and monocytes when compared to HLA-DR expression, the lower number of positive progenitor cells probably reflects lower antigen density rather than distinct positive and negative progenitor cell populations. The DC antigen is expressed weakly on monocytes and B cells, although there is considerable individual variation. In some individuals, distinct DC-positive and -negative monocyte populations are detectable. The DC antigen was not detected on myeloid progenitor cells, even in those individuals with moderate DC expression on their monocytes and B cells. This discordant expression of DC and other Ia-like antigens on hematopoietic progenitor cells may be of physiologic significance and may assist in the purification of progenitor cells from blood and marrow.
A group of bactericidal factors conserved by vertebrates for more than 300 million years.
Authors: Kawakami M , Ihara I , Ihara S , Suzuki A , Fukui K
Published: December 2022

A complement-dependent bactericidal factor (RaRF) specific for the Ra chemotype strains of Salmonella and for the Ra-like strains of other enterobacteria has been found in sera of mice. We show here that the anti-Ra bactericidal factors are present in sera of all species, so far tested, of mammals, birds, reptiles, amphibians, and bony and cartilaginous fish. Certain properties, such as binding specificity, requirement of divalent cations for binding, and sensitivities to heat and reducing agents, of the factor in the representative animals were the same as or similar to those of the mouse RaRF. These results indicate that these factors have been conserved by vertebrates for more than 300 million years as a result of the necessity for the resistance to rough mutants of Gram-negative bacteria.
Inhibition of growth of Histoplasma capsulatum by lymphokine-stimulated macrophages.
Authors: Wu-Hsieh B , Howard D H
Published: December 2022

Peritoneal cells from mice immunized by sublethal infection inhibit the intracellular growth of Histoplasma capsulatum in vitro. Lymphokines generated in cultures of immune splenocytes stimulated with Histoplasma antigen activate normal macrophages to inhibit the intracellular growth of the fungus. Such lymphokines may inhibit the intracellular growth of H. capsulatum by 60 to 80% but have no direct effect on the viability of the fungus extracellularly. The lymphokine preparations have high interferon activity that is heat stable and acid labile. The cells in the spleen that are responsible for lymphokine production are T lymphocytes, but the help of other cells such as macrophages is essential for maximal production.
A membrane antigen (HC1) selectively present on hairy cell leukemia cells, endothelial cells, and epidermal basal cells.
Authors: Posnett D N , Marboe C C , Knowles D M , Jaffe E A , Kunkel H G
Published: December 2022

A monoclonal antibody (alpha HC1) raised against HCL cells reacted specifically with HCL cells and not with other normal or leukemic B cell types. It also reacted with BC and with vascular EC. One case of Kaposi's sarcoma, a tumor thought to derive from EC, was also stained by this antibody. Sinusoidal cells of liver and spleen and EC in the brain did not stain with alpha HC1, but EC from all other tissues and EC in culture were positive. Because the exact normal cell equivalent of the leukemic hairy cell is unknown, similarities in phenotype and function between EC and hairy cells are of particular interest.
Differential recognition and lysis of EL4 target cells by cytotoxic T cells: differences in H-2Kb antigenic density and cytoskeletal proteins.
Authors: Flores R V , Gilmer P J
Published: December 2022

Direct binding of 125I-iodinated anti-H-2Kb monoclonal antibody (B8-24-3) to EL4 cells indicated a similar association constant but a 2.7-fold higher H-2Kb antigenic density when the EL4 cells were grown in ascites (2.0 X 10(5) sites/cell) than in tissue culture (7.5 X 10(4) sites/cell). The membrane fluidity of the isolated plasma membrane fractions, measured by using a membrane-sensitive spin label, was essentially identical in the two differently cultured EL4 cells. There were some differences in the cytoskeletal proteins that were isolated from the plasma membrane fractions of the EL4 lines and analyzed by two-dimensional gel electrophoresis. These differences in the H-2Kb antigenic density and the cytoskeletal proteins may contribute to the 2.3-fold higher Vmax and the increased binding of the EL4 ascites target cells to allogeneically primed anti-H-2Kb cytotoxic T cells.
A rat anti-mouse T4 monoclonal antibody (H129.19) inhibits the proliferation of Ia-reactive T cell clones and delineates two phenotypically distinct (T4+, Lyt-2,3-, and T4-, Lyt-2,3+) subsets among anti-Ia cytolytic T cell clones.
Authors: Pierres A , Naquet P , Van Agthoven A , Bekkhoucha F , Denizot F , Mishal Z , Schmitt-Verhulst A M , Pierres M
Published: December 2022

Hybridoma H129 .19 was derived by fusion between spleen cells of a Lou / Ws1 rat immunized with an Lyt-1+,2- anti-I-Ak cytolytic T lymphocyte (CTL) clone and the nonsecreting myeloma X63-Ag8.653. The monoclonal antibody (mAb) H129 .19 (IgG2a, kappa) was selected for its capacity to inhibit the lytic potential of the immunizing clone. H129 .19 identified a monomorphic determinant on a 55 m.w. murine T cell differentiation antigen, which appeared to be homologous to the human T4 molecule in that: 1) H129 .19 reacted with 80% adult thymocytes, with a subset of splenic T cells, and with the interleukin 2 (IL 2)-producing EL4 thymoma; 2) The mAb bound to and inhibited the IL 2 production and the proliferation of various allo- or soluble antigen-reactive T cell clones that recognized restriction or activating determinants on the I-A or I-E molecules, respectively; 3) H129 .19 did not inhibit the proliferation and/or cytolysis of Lyt-2,3+ T cells specific for class I MHC antigen; and 4) Among six anti-Iak CTL clones examined in this study, the mAb H129 .19 reacted with two I-Ak-specific, Lyt-2,3- clones on which it exerted strong cytolysis inhibiting effect at the effector cell level. By contrast, two other anti-I-Ak and two anti-I-Ek CTL clones were found to express the Lyt-2,3+,T4- cell surface phenotype. The cytolytic potential of the latter clones was not inhibited by anti-Lyt-2,3 mAb. These studies strongly suggest that the mouse T4 molecule facilitates the recognition of class II MHC antigen by most but not all T cells.