bluuebunny/crossref_metadata_2025 huggingface dataset:
Antibody-independent C1 activation by E. coli.
Authors: Tenner A J , Ziccardi R J , Cooper N R
Published: December 2022

Antibody-independent interactions of C1 with several E. coli strains were examined. Purified C1 was directly activated by the semi-rough mutant E. coli J-5, its parental wild-type strain, E. coli 0111:B4, and two clinical isolates, E. coli (P) and E. coli (A), in the absence of C1 inhibitor. E. coli J-5 activated C1 about 10-fold more rapidly and bound approximately threefold more C1 than the other strains. E. coli J-5, but not the other strains, also bound C1s2, provided that the subcomponent was offered to the bacteria in the presence of C1q and calcium; such binding was thus independent of the presence or absence of C1r2. After C1 activation in the absence of C1 inhibitor, activated C1s spontaneously dissociated from E. coli 0111:B4, (P), and (A), but remained associated with E. coli J-5. The regulatory protein C1 inhibitor prevented C1 activation by the weaker activators, E. coli strains 0111:B4, (P), and (A), but had no effect on C1 activation by E. coli J-5. Although C1 inhibitor thus failed to modulate C1 activation by E. coli J-5, it did block the enzymatic activity of activated C1 bound to this strain. Analyses of the molecular processes involved revealed differences with other systems. In the presence of C1 inhibitor, the C1s subunit of C1 activated by E. coli J-5 underwent further cleavage with the release into the supernatant of C1s fragments and complexes of C1 inhibitor with light chain fragments. Such fragments were not disulfide-linked to the remainder of the C1s molecule. The bulk of the heavy chain remained adherent to the surface of E. coli J-5. This finding documents the presence of a binding site for activated C1s on the surface of E. coli J-5 and localizes this site to the heavy chain. These studies thus indicate that several E. coli strains are direct C1 activators. Furthermore, E. coli J-5 provides another example of a direct C1 activator having binding sites not only for C1q but also for dimeric C1s. The studies also show that there are multiple properties of particles which determine the ability to activate C1, the rate of activation, the possibility of regulation of the activation process by C1 inhibitor, and the fate of activated C1.
Phorbol esters inactivate the lytic apparatus of cytotoxic T lymphocytes.
Authors: Russell J H
Published: December 2022

Treatment of cloned murine CTL with phorbol myristate acetate (PMA) initiates the loss of lytic potential of these cells. The process is relatively rapid (t 1/2 = 2.2 +/- 0.7 hr), occurs independently of protein synthesis, and can be largely reversed within 3 hr by incubation in normal medium with or without the addition of supernatants from activated lymphocytes. Examination of PMA-treated or untreated CTL in assays that separate binding and post-binding events suggests that the lytic apparatus itself is a major target of the PMA-initiated event.
Stimulation of phagocytosis in bone marrow-derived mouse macrophages by bacterial lipopolysaccharide: correlation with biochemical and functional parameters.
Authors: Cooper P H , Mayer P , Baggiolini M
Published: December 2022

It has been shown that low concentrations of E. coli lipopolysaccharides (LPS) greatly and selectively stimulate phagocytosis and related functions in mouse bone marrow-derived macrophages. Culture in the presence of 50 ng/ml LPS induced on average a 10-fold enhancement of phagocytosis of IgG-coated sheep erythrocytes. Activation was in two stages--a small increase observed during the first 8 to 12 hr, and the major increase noted between 16 and 24 hr. Phagocytic activity remained at the maximal level for 24 hr and then declined progressively. Stimulation by LPS was dose-dependent; significant effects could be observed at 0.8 ng/ml and the maximum was reached at 10 ng/ml. LPS-treated cells also showed a markedly increased tendency to form colonies. All these effects could be prevented by the addition of 100 ng/ml polymyxin B together with LPS, indicating that the active principle is lipid A. The LPS-dependent increase in phagocytic activity is probably mediated by increased Fc receptor capacity because both parameters were influenced in parallel by the stimulus. Phagocytosis-related events, such as enhanced hexose monophosphate shunt activity, H2O2 formation, and nitroblue tetrazolium reduction were also stimulated by LPS. By contrast, pinocytosis was unaffected. Measurements of cell-associated enzyme activities showed that lactate dehydrogenase, acid phosphatase, and cathepsin D were significantly increased. Beta-glucuronidase, beta-galactosidase, alkaline phosphodiesterase, and aminopeptidase were unchanged and NAD nucleosidase was markedly decreased after LPS treatment. 5'-Nucleotidase and glucosamine uptake were undetectable both in control and LPS-stimulated cells. LPS treatment induced a significant increase in cell-associated protein, but did not result in cell proliferation or increased cell loss as shown by the DNA content that remained constant. LPS-induced changes were dependent on de novo protein synthesis; cycloheximide prevented enhancement of phagocytosis, Fc receptor capacity, and colony formation.
An analysis of the relationship between 5-lipoxygenase product generation and the secretion of preformed mediators from mouse bone marrow-derived mast cells.
Authors: Razin E , Romeo L C , Krilis S , Liu F T , Lewis R A , Corey E J , Austen K F
Published: December 2022

The quantitative relationships between the secretion of a granule-associated mediator, beta-hexosaminidase, and the oxidative metabolism of arachidonic acid by the 5-lipoxygenase pathway were analyzed for a homogeneous population of T cell-dependent, bone marrow-derived, murine mast cells. The mast cells were either sensitized with a monoclonal IgE and challenged with specific antigen, or to bypass a transmembrane signal, were stimulated with calcium ionophore A23187. The released products of the 5-lipoxygenase pathway were quantitated by integrated ultraviolet absorbance after resolution by reverse phase-high performance liquid chromatography in the case of 5-hydroxy-eicosatetraenoic acid (5-HETE), and by separate radioimmunoassays for leukotriene C4 (LTC4) and leukotriene B4 (LTB4). The activation-release response of the cells was perturbated by the introduction of three pharmacologic agents, each directed to different steps in the 5-lipoxygenase pathway of arachidonic acid metabolism, and the action of each agent was determined for separate cell samples while present and after its removal by washing. 5,6-Dehydroarachidonic acid (5,6-DHA), an irreversible inhibitor of 5-lipoxygenase, prevented formation of 5-HETE from exogenous [14C]arachidonic acid and from membrane-derived arachidonic acid in a dose-related fashion when sensitized mast cells, preincubated with drug, were washed before antigen activation. Release of 5-HETE, LTC4, and LTB4 was inhibited by 5,6-DHA in a corresponding dose-related fashion, with a minimal preincubation period of 1 to 5 min before the cells were washed and subjected to antigen-dependent activation. In contrast, the inhibitory effect of 5,6-DHA on beta-hexosaminidase release was lost after three washes and was not evident after one wash unless the preincubation period was extended to 15 min. The capacity of 5,6-DHA to prevent leukotriene generation without altering beta-hexosaminidase release was also observed with ionophore-activated mast cells. Preincubation of sensitized cells with diethylcarbamazine (DEC), followed by a wash before antigen-dependent activation, produced inhibition of leukotriene generation, no effect on beta-hexosaminidase release, and augmentation of 5-HETE release at the maximum dose studied; thus, DEC interrupts the pathway distal to the formation of 5-hydroperoxy-eicosatetraenoic acid (5-HPETE) from arachidonic acid by 5-lipoxygenase. Preincubation of sensitized cells with incremental amounts of the prostacyclin analog U-60,257, followed by washing and antigen challenge, inhibited LTC4 release without altering the release of LTB4 or beta-hexosaminidase.(ABSTRACT TRUNCATED AT 400 WORDS)
IgA-dependent cell-mediated activity against enteropathogenic bacteria: distribution, specificity, and characterization of the effector cells.
Authors: Tagliabue A , Boraschi D , Villa L , Keren D F , Lowell G H , Rappuoli R , Nencioni L
Published: December 2022

Antibody-dependent cellular cytotoxicity (ADCC) against murine enteropathogenic bacteria such as Salmonella typhimurium and Salmonella tel aviv or Shigella X16 was assessed by using IgG, IgA, and secretory IgA (sIgA) in a 2-hr in vitro assay where peripheral and intestinal lymphocytes were used as effector cells. It was found that IgG could arm splenocytes (SpL) better than IgA. However, IgG did not arm lymphocytes from Peyer's patches (PPL) or from mesenteric lymph nodes (MnL), whereas IgA of plasmacytoma origin against S. tel aviv and purified intestinal sIgA against Shigella X16 induced specific antibacterial ADCC with both SpL and PPL. When sIgA were tested with intestinal lymphocytes from the epithelium and the lamina propria, i.e., cells from the gut mucosa which first interact with enteric bacteria, it was found that both these lymphoid populations were able to express sIgA-dependent ADCC against Shigella X16. In parallel tests, cells from thymus and popliteal lymph nodes failed to express ADCC. Blocking studies with purified IgG and IgA of goat, rabbit, and mouse origin demonstrated that the Fc-alpha and Fc-gamma receptors were specifically involved in IgA- or IgG-dependent antibacterial ADCC. At least two effector populations, a macrophage and a Thy-1.2- lymphocyte, were observed to exert IgA-ADCC at the splenic level, whereas only lymphoid cells expressed this activity at the GALT level. Together, these results describe a new activity of IgA against enteropathogenic bacteria.
Requirements for suppressor T cell activation.
Authors: Usui M , Aoki I , Sunshine G H , Dorf M E
Published: December 2022

Third-order (Ts3) suppressor cells are generated after conventional immunization. These cells, however, will not mediate suppressor cell function unless specifically triggered by an activating signal, termed TsF2. This report analyzes the mechanism of this TsF2-mediated triggering event. TsF2-mediated suppression is genetically restricted by genes in the I-J and Igh-V regions. The target of the I-J restrictions is a firmly adherent accessory cell, which appears to express I-J-related determinants. These accessory cells are sensitive to cyclophosphamide treatment and 500 R irradiation. In contrast, the target of the Igh-V restriction of TsF2 appears to be the Ts3 cell, which carries antigen-specific, idiotype-related receptors. The mechanism of suppressor cell activation appears to involve two stages. Presentation of I-J-restricted TsF2 by I-J-compatible presenting cells and a second step involving idiotype-anti-idiotype interactions between TsF2 and the Ts3 cell. I-J compatibility is not required with the accessory cell for Ts3 activation. Finally, we hypothesize that the anti-idiotypic determinants expressed on TsF2 can serve as an internal image of antigen, thereby permitting specific targeting of the factor.
Antigen presentation by the BCL1 murine B cell line: in vitro stimulation by LPS.
Authors: Harris M R , Kindle C S , Abruzzini A F , Pierce C W , Cullen S E
Published: December 2022

We examined the antigen-presenting capacity of BCL1 tumor cells, which are capable of differentiating in vitro with respect to immunoglobulin synthesis/secretion under the influence of LPS. In vivo passaged BCL1 cells depleted of host cell contamination either by positive selection employing panning with anti-lambda reagents, or by elimination of latex-ingesting adherent cells, are capable of MHC-restricted antigen presentation to a GAT-immune T cell line. The BCL1 cells act as antigen-presenting cells when freshly explanted, but gradual loss of this function occurs, and cells cultured for 3.5 days cannot present antigen unless LPS is included during the culture period. BCL1 cells are equivalently Ia+ after the culture period with or without LPS stimulation. Other B cell lines capable of antigen presentation appear to express this trait constitutively, and the in vivo passaged BCL1 line is therefore unique among B cell lines in having antigen-presenting cell function that can be modulated. The data suggest that freshly explanted or LPS-cultured BCL1 cells are heterogeneous with respect to antigen-presenting capacity, and the basis for this heterogeneity is being sought. BCL1 offers an opportunity to study requirements for antigen presentation by B cells.
Activated T cells in vivo and in vitro: divergence in expression of Tac and Ia antigens in the nonblastoid small T cells of inflammation and normal T cells activated in vitro.
Authors: Burmester G R , Jahn B , Gramatzki M , Zacher J , Kalden J R
Published: December 2022

Elevated numbers of non-blastoid T cells expressing either the Tac or Ia antigens were found on separate cell populations in inflammatory synovial tissues and fluids of individuals with arthritis. Those synovial T cell preparations containing Tac+ cells exhibited marked proliferation upon the addition of IL 2 without concomitant mitogen stimulation; T cell eluates containing Ia+ but not Tac+ T cells did not show significantly increased levels of blastogenesis. Paired T cell preparations from blood had only minor increases in the number of Tac+ T cells and moderate increases in the number of Ia+ cells. The blood cells did not exhibit significant proliferation to IL 2. In contrast mitogen or allogeneic activation of T cells induced blastoid cells that expressed abundant per cell amount of Ia or Tac antigens. These blastoid cells resembled the small T cells of inflammation in having only very limited overlap between the population that bore Ia antigens and those with the Tac antigen; however, there was a preponderance of Tac-bearing cells.
Activation of antigen-specific suppressor T lymphocytes in man involves dual recognition of self class I MHC molecules and Leu-4/T3-associated structures on the surface of inducer T lymphocytes.
Authors: Damle N K , Mohagheghpour N , Engleman E G
Published: December 2022

We showed previously that fresh Leu-2+ T cells respond to autologous antigen-primed Leu-3+ T cells by proliferation and differentiation into suppressor T cells (Ts) that specifically inhibit the response of fresh Leu-3+ cells to the original priming antigen. This study was undertaken to characterize the role of various cell surface molecules expressed by antigen-primed Leu-3+ cells in their activation of Leu-2+ Ts cells. Alloactivated Leu-3+ blasts were treated in the absence of complement with a variety of monoclonal antibodies recognizing distinct antigens on human lymphoid cells, and then were examined for their functional effects on fresh autologous T cells. Prior treatment of Leu-3+ blasts with anti-Leu-4 or anti-HLA-A,B,C framework antibodies, but not with anti-Leu-1, anti-Leu-3, anti-Leu-5, or anti-HLA-DR framework-specific antibodies, not only blocked proliferation of fresh Leu-2+ cells, it also prevented their differentiation into Ts cells. Furthermore, after their activation by Leu-3+ blasts, Leu-2+ Ts cells inhibited the response of fresh Leu-3+ cells from only those individuals who shared HLA-A,B phenotypes with suppressor-effector cells. These results suggest that both the inductive and effector phases of suppression involve dual recognition of autologous class I MHC molecules and structures associated with the Leu-4 (T3) molecule on the surface of antigen-reactive Leu-3+ cells.
The human thymic microenvironment: cortical thymic epithelium is an antigenically distinct region of the thymic microenvironment.
Authors: McFarland E J , Scearce R M , Haynes B F
Published: December 2022

The thymic microenvironment is a complex tissue essential for normal T cell maturation. Prothymocytes in the subcapsular cortical (SCC) region of the thymus undergo cell division and migrate to the inner cortex. The majority of cortical thymocytes cease dividing and die, but a minority are exported to the periphery. We have previously shown thymic hormones in SCC and medullary thymic epithelium and have identified a monoclonal antibody (TE-4) that defines human endocrine thymic epithelium. However, no marker that selectively defines cortical thymic epithelium has been available. In this study, we have produced two monoclonal antibodies, TE-3A and TE-3B, raised against human thymic stroma that bind to an intracellular antigen in cortical but not medullary thymic epithelium. In double immunofluorescence assays in which we used anti-keratin, anti-thymosin alpha 1, and anti-endocrine thymic epithelium antibodies (TE-4, A2B5), TE-3+ SCC epithelium was TE-4+ and contained keratin and thymosin. alpha 1. In contrast, TE-3+ inner cortical epithelium was TE-4/A2B5 nonreactive and did not contain thymosin alpha 1. An ontogeny study of seven fetal and five neonatal thymuses demonstrated that expression of the TE-3 antigen was acquired at 10 wk fetal gestation. Using TE-3 antibody, we observed sequential stages of separation of cortical and medullary epithelium from 12 to 20 wk fetal gestation. In dysplastic (severe combined immunodeficiency disease) thymuses, strands of TE-3+ nonendocrine cells encircled nests of TE-4+ endocrine epithelium. Thus, human cortical thymic epithelium is antigenically distinct from endocrine medullary epithelium. Antibodies against the TE-3 antigen define an intracellular molecule that may reflect a specialized function of cortical thymic epithelium.
Mechanism of induction of mouse erythrocyte receptor switch in human B cells.
Authors: Zalewski P D , Forbes I J , Valente L , Comacchio R
Published: December 2022

An early event in phorbol ester-induced maturation of chronic lymphocytic leukemic (CLL) B cells is a membrane change characterized by the inactivation of a mouse erythrocyte receptor (MER). This event, the MER-switch, is quantified by inhibition of rosette formation. By using [3H]phorbol dibutyrate ([3H]PDBu), both to stimulate MER-switch and assay binding of PDBu to CLL cells, it was shown that MER-switch was an irreversible, time-dependent event which occurred some time after maximal binding of [3H]PDBu to cells. Two classes of binding sites, one of high affinity (Kd 1 to 2 nM) at low frequency (1.5 to 5 X 10(4) sites per cell), and a lower affinity site (Kd 33 to 50 nM) of higher frequency (2 to 3.5 X 10(5) sites per cell), were detected. Binding of [3H]PDBu was inhibited by phorbol ester analogs that stimulated MER-switch, but not by inactive analogs. This, and the similarity in shapes of the binding and rosette inhibition curves over a range of concentrations, suggests that stimulation of MER-switch by phorbol esters is due to this specific binding. The phorbol ester receptor and MER are distinct because MER-ve T cells and MER-ve atypical B cells from a patient with CLL had both classes of PDBu receptor. Solubilized MER did not bind [3H]PDBu. Time-course studies, and the irreversibility of the switch, despite removal of most of the bound [3H]PDBu, indicate that inhibition of rosetting is not due to competitive or steric hindrance by phorbol esters. Equivalent activities of soluble MER were released from fresh and phorbol ester-treated CLL cells, indicating a rearrangement of MER, rather than a loss. A supernatant of phytohemagglutinin-stimulated human spleen cells also induced MER-switch in CLL lymphocytes, suggesting that a lymphokine may be a natural inducer of this event.
Isolation and functional characteristics of FcR+ and FcR- human monocyte subsets.
Authors: Zembala M , Uracz W , Ruggiero I , Mytar B , Pryjma J
Published: December 2022

Human peripheral blood monocytes (M phi) were separated into Fc receptor positive (FcR+) and FcR- subsets by rosetting with O,Rh+ human erythrocytes coated with IgG anti-D antibody, followed by density gradient centrifugation and adherence. The FcR+M phi suppressed PHA- and PPD-induced lympho-proliferation and also exhibited a strong cytostatic effect against tumor cell lines in vitro, but were a poor source of antigen-presenting cells. In contrast, the FcR- subset did not cause suppression but was highly active in the presentation of PPD to T cells and in stimulation of both allogeneic mixed lymphocyte reaction (allo-MLR) and autologous (auto-MLR). PWM-induced immunoglobulin secretion in vitro was greatly enhanced by the FcR- but not the FcR+ subset. Nevertheless, both FcR+ and FcR- subsets showed comparable HLA-DR antigen expression. These results indicate that human M phi, like lymphocytes, may consist of several subsets that differ significantly in their immunologic functions.
Cord blood plasma lipoproteins inhibit mitogen-stimulated lymphocyte proliferation.
Authors: Curtiss L K , Forte T M , Davis P A
Published: December 2022

Lipoproteins were isolated from adult plasma and the umbilical cord blood plasma of newborn infants and were compared for their capacity to inhibit mitogen-stimulated [3H]thymidine uptake of adult peripheral blood mononuclear cells in vitro. Relative to the comparable adult lipoproteins, cord blood low density lipoproteins and high density lipoproteins inhibited mitogen stimulation at twofold to fourfold lower total protein concentrations. Apoproteins AI, B, and E were quantitated by radioimmunoassay of each of the adult and cord blood lipoprotein fractions. A strong correlation was observed between inhibitory activity and the amount of apoprotein E in the cord blood low and high density lipoproteins. Further evidence that lipoproteins containing apoprotein E accounted for the difference in suppressive activity of cord blood low and high density lipoproteins relative to the adult lipoproteins was obtained by selective removal of the apoprotein E-containing lipoproteins by using immunoaffinity chromatography or heparin-agarose adsorption. The results indicated that cord blood lipoproteins containing apoprotein E in association with apoproteins AI or B are capable of suppressing lymphocyte proliferation in vitro.
Anti-idiotype antibodies to the 9.1C3 blocking antibody used to probe the lethal hit stage of NK cell-mediated cytolysis.
Authors: Werkmeister J A , Burns G F , Triglia T
Published: December 2022

We previously described a monoclonal antibody, 9.1C3, which blocked natural killer (NK) cell-mediated cytolysis by acting on effector cells during a late step in the lethal hit stage. The present work describes the production in rabbits of anti-idiotypic (anti-id) antibodies to the 9.1C3 antibody. In addition to reacting specifically with the 9.1C3 antibody, the anti-id antibodies bound strongly to the K562 target cell. The anti-id antibodies blocked killing of K562 targets by NK, antibody-dependent cellular cytotoxicity, and NK-like cells but did not inhibit killing by cytotoxic T lymphocytes (CTL). Pretreatment of cells and washing before assay indicated that blocking occurred at the target cell level. Of particular interest, single cell assays with Percoll-enriched large granular lymphocytes demonstrated that the antibodies caused no reduction in binding. These data are consistent with a model for NK cell-mediated lysis that involves a secondary target cell receptor independent of the primary NK-target cell interaction. The anti-id antibodies immunoprecipitated cell surface proteins of relative m.w. 79K and 62K unreduced, and 94K and 79K reduced from K562 target cells. The development of anti-id antibodies may be a useful procedure to explore the structure and function of cellular receptors involved in NK cell-mediated cytolysis.
Characterization of the La (SS-B) antigen from several mammalian sources.
Authors: Hoch S O , Billings P B
Published: December 2022

The La or SS-B antigen is associated with rheumatic diseases, systemic lupus erythematosus, and Sjogren's syndrome, and is part of a larger ribonucleoprotein complex. Immunoaffinity chromatography allowed for the efficient separation of the La antigen from the bulk of the cellular proteins, with a minimum of protease exposure. Protein blot analysis of the affinity-isolated material indicated a major immunoreactive polypeptide of 50,000 m.w. A comparison of this antigen in a number of mammalian sources (human, rabbit, and rat) suggested strong conservation of the native polypeptide m.w. Likewise, in a direct comparison of this antigen from Epstein-Barr virus-infected cells in which there are distinct differences in the antigen-associated RNA species, the immunoreactive polypeptide species were of similar size. The La protein is readily susceptible to endogenous proteolysis, with the resulting generation of smaller, discrete polypeptides that still retain antigenicity. By using the La protein to monitor potential degradation, we have developed a simple two-step procedure to isolate the La-associated snRNP complex. The complexes thus isolated provide material suitable as a source of both the active antigen and of the functional ribonucleoprotein complex.
Preferential induction of specific lambda-isotypic antibodies in mice.
Authors: Ju S T , Dorf M E
Published: December 2022

A high proportion (greater than 40%) of lambda-anti-NP antibodies were induced after the administration of hapten conjugates of the relatively T-independent antigen NP-Ficoll. In 11 of 12 strains, lambda 1 anti-NP antibodies were the predominant isotype. In lambda 1-defective SJL mice, lambda 2,3 anti-NP antibodies were the major species after NP-Ficoll immunization. In contrast, the ability to elicit a high proportion of lambda-anti-NP antibodies with the T-dependent conjugate of ovalbumin, NP-OVA, varied among mouse strains. Igh-1b-bearing mice were high producers of lambda 1 anti-NP antibodies (greater than 70% of the response); DBA/2 and BALB/c mice were moderate (40 to 50%) lambda 1 anti-NP producers, and A.TL, AKR, NZB, and C3H mice were low lambda 1 anti-NP producers (less than 10%) after primary NP-OVA immunization. In the latter group, NP-OVA preferentially elicits kappa-bearing anti-NP antibodies. The parameters that influence the distribution of light chain isotypes were investigated. The preferential induction of lambda-anti-NP antibodies with NP-Ficoll was a) partially influenced by Igh-linked genes, b) adjuvant independent, and c) maintained on prolonged immunization. In contrast, induction of a high proportion of kappa-anti-NP antibodies by NP-OVA is (a) strictly regulated by Igh-linked genes and (b) enhanced after hyperimmunization. The immunochemical, genetic, and cellular bases for these observations are discussed.
Effect of catalase on the proliferation of human lymphocytes to phorbol myristate acetate.
Authors: Sagone A L , Husney R , Guter H , Clark L
Published: December 2022

Phorbol esters have been documented to stimulate the proliferation of human blood mononuclear cell cultures. In addition, these agents are also known to stimulate the production and release of reactive oxygen species by monocytes. We demonstrated previously that H2O2, one of these oxygen metabolites, impairs the proliferative capacity of human blood lymphocytes. Therefore, in these experiments, we determined whether or not the H2O2 released by monocytes after activation by PMA modifies the proliferation of lymphocytes to this agent. Human blood mononuclear cells (80% lymphocytes and 20% monocytes) were incubated with PMA, and lymphoblastic transformation (LBT) was quantitated at 3 and 5 days by pulsing the cultures with thymidine. Initial experiments established that the concentration of PMA required for optimal LBT was 50 ng/ml. We then demonstrated that this concentration of PMA also induces a burst in hexose monophosphate shunt activity and H2O2 production of mononuclear cells as indicated by the enhanced oxidation of 14C-glucose and 14C-formate, respectively. The amount of H2O2 released into the medium was substantial. Our measurements indicate that the concentration of H2O2 could reach values as high as 0.008 mM during the first 2 hr of the cultures. The addition of catalase to PMA-treated cultures in concentrations sufficient to scavenge the H2O2 released by the monocytes was associated with an enhanced thymidine uptake (mean 79%). These results indicate that the hydrogen peroxide released by the monocytes modifies the response of lymphocytes to the PMA. Paradoxically, mononuclear cell cultures depleted of monocytes also had a lower proliferation to PMA than mononuclear cell cultures. This observation indicates that monocytes also produce factors required for lymphocyte proliferation to PMA such as an interleukin. In contrast, to PMA cultures, catalase did not alter the proliferation of mononuclear cell cultures stimulated by PHA. We previously documented that PHA does not stimulate an immediate burst in the oxidative metabolism of mononuclear cultures. Therefore, the effect of catalase in these two culture systems appears to correlate with the capacity of the mitogen to stimulate the oxidative metabolism of mononuclear cells. These observations suggest that the release of reactive oxygen species by monocytes may modify the response of lymphocytes to antigens both in vitro and in vivo.
Modulation of the biologic activities of IgE binding factor. VI. The activation of phospholipase by glycosylation enhancing factor.
Authors: Iwata M , Akasaki M , Ishizaka K
Published: December 2022

Glycosylation enhancing factor (GEF) from rat T cells is a kallikrein-like enzyme and enhances the assembly of N-linked oligosaccharides to IgE binding factors during their biosynthesis, whereas another T cell factor, i.e., glycosylation inhibiting factor (GIF), is a fragment of phosphorylated lipomodulin (i.e., phospholipase inhibitor), which when dephosphorylated inhibits phospholipase and the glycosylation process. The two T cell factors compete with each other when they are added to normal mesenteric lymph node cells during the formation of IgE binding factors. The addition of GEF to T cell hybridoma 23A4 cell switches the cells from the formation of unglycosylated IgE binding factor to the formation of N-glycosylated IgE binding factor. However, GEF neither inactivated GIF nor inhibited the formation of GIF by the T cell hybridoma. Stimulation of the T cell hybridoma with either affinity-purified GEF or bradykinin resulted in the release of GIF from the cells. GIF released by GEF stimulation had a m.w. of approximately 15,000 and bound to monoclonal antibody against lipomodulin. GEF and bradykinin also induced normal mesenteric lymph node cells to release GIF. Incorporation of 14C-arachidonic acid into 23A4 cells, followed by stimulation of the cells with GEF, resulted in the release of 14C-arachidonate. The results suggest that lipomodulin, a phospholipase inhibitory protein, is present in lymphocytes, and indicate that GEF and bradykinin induce the activation of phospholipase by stimulating cells to release lipomodulin.
Histamine release in acute anaphylactic enteropathy of the rat.
Authors: Lake A M , Kagey-Sobotka A , Jakubowicz T , Lichtenstein L M
Published: December 2022

Investigations into the role of allergic enteropathy in acute and chronic intestinal inflammation have been hampered by the lack of objective confirmation for intestinal mast cell activation. Utilizing an established model of acute allergic enteropathy in the rat, we report the enhanced intraluminal recovery of the mast cell mediator histamine after in vivo antigen challenge in sensitized animals. The enhanced histamine recovery is dose dependent, antigen-specific, and restricted to that segment of bowel challenged, thus confirming local intestinal anaphylaxis. The progression of histologic enteropathy is documented and shown to correlate with the entry of mast cells into the intestinal lumen during, but not before, the anaphylactic response. Pretreatment of the sensitized animal with prostaglandin E2 or doxantrazole, but not cromolyn, significantly inhibits the anaphylactic response.
Granule-associated serine neutral proteases of the mouse bone marrow-derived mast cell that degrade fibronectin: their increase after sodium butyrate treatment of the cells.
Authors: DuBuske L , Austen K F , Czop J , Stevens R L
Published: December 2022

Mouse bone marrow-derived mast cells, differentiated in vitro with concanavalin A splenocyte-conditioned medium and sensitized with monoclonal IgE, release neutral serine proteases after activation with specific antigen. Sodium dodecyl sulfate polyacrylamide gel electrophoretic (SDS-PAGE) analysis of the supernatants from immunologically activated mast cells revealed the presence of four prominent proteins of 27,000, 29,000, 30,000 and 31,000 m.w. When the supernatants and sonicated residual cells from antigen-challenged or nonactivated IgE-sensitized mast cells were incubated with [3H]diisopropylfluorophosphate ([3H]DFP) and the proteins were subjected to SDS-PAGE followed by autoradiography, proteins of 27,000 to 31,000 m.w. were labeled with [3H]DFP. The antigen-dependent release of labeled proteins was accompanied by a corresponding depletion of similarly sized [3H]DFP-labeled proteins from these cell pellets relative to unactivated cells. The SDS gels were also stained with Coomassie Blue and were sectioned to separate the individual proteins for measurement of their incorporated radioactivity; the net percent antigen-dependent release of all four [3H]DFP-labeled proteins ranged from 64 to 68% and was comparable to that of the secretory granule markers, beta-hexosaminidase and histamine. That the [3H]DFP-labeled proteins were derived from the secretory granules of the cells was supported by studies in which mast cells were cultured for 4 days in the presence of 1 mM sodium butyrate. This treatment produced a differential increase in their cellular content of histamine (10-fold), [3H]DFP binding proteins (two- to fourfold), and beta-hexosaminidase (minimally), while the net percent antigen-dependent release of each of these constituents was unchanged. After sensitization and antigen activation, the net percent release of histamine, beta-hexosaminidase, and the four [3H]DFP-labeled proteins was 51, 59, and 53 to 61%, respectively, for sodium butyrate-treated cells, and 53, 60, and 64 to 68%, respectively, for cells not exposed to sodium butyrate. Human plasma fibronectin was used as a substrate to demonstrate that the exocytosed proteins possessed proteolytic activity. As assessed by optical density scanning of stained SDS-PAGE gels of the substrate, the proteases present in the supernatants of antigen-activated cells, but not of sensitized unchallenged cells, rapidly degraded native fibronectin at pH 7.0. This degradation was prevented by pretreatment of the exocytosed proteins from immunologically activated cells for 90 min at 37 degrees C with 2 mM DFP.(ABSTRACT TRUNCATED AT 400 WORDS)
Monoclonal antibodies specific for beta-adrenergic ligands.
Authors: Chamat S , Hoebeke J , Strosberg A D
Published: December 2022

After somatic cell fusion between splenocytes of immunized BALB/c mice and NS-1 myeloma cells, eight clones were obtained secreting anti-alprenolol antibodies as characterized by means of an ELISA. Four of these were subcloned and were studied further. The association constant for alprenolol ranged from 1.9 X 10(6) M-1 to 24 to 10(6) M-1. Competitive inhibition of [3H]-l-dihydroalprenolol binding revealed cross-reactivity with beta-adrenergic ligands, with a higher avidity for antagonists than for agonists. Two of the antibodies had a higher affinity for the l-isomer than for the d-isomer. The most stereospecific of these antibodies showed only affinity for beta-adrenergic antagonists and for the agonist isoproterenol. The other recognized both beta-adrenergic antagonists and agonists; it also showed an increase in tryptophan fluorescence after ligand binding. This property was used for the physicochemical study of the hapten-antibody interaction.
A rabbit helper T cell clone reactive against group-specific streptococcal carbohydrate.
Authors: Jackson S , Folks T M , Wetterskog D L , Kindt T J
Published: December 2022

Cloned rabbit T cell lines specific for the group-specific carbohydrate of group C streptococcus were established to study T cell responses to this antigen. T cell lines and clones of these lines were prepared from peripheral blood lymphoid cells of animals previously hyperimmunized with group C streptococcal vaccine. All cell lines studied were antigen specific in that they proliferated in the presence of group C but not group A vaccine. Three clones from one rabbit showed strong proliferation in the presence of soluble group C carbohydrate, in addition to the particulate vaccine. One of these clones, 5317.1, upon additional characterization, was shown to possess helper activity in that it induced autologous B cells to produce anti-carbohydrate antibody. Interestingly, B cells could not be stimulated by this carbohydrate-responsive clone to produce anti-carbohydrate antibody in the presence of soluble carbohydrate; the intact vaccine was required to demonstrate the helper effect of the clone. Phenotypic characterization of this clone with a panel of monoclonal antibodies indicated that it was of the T1 subpopulation of peripheral T cells. No suppressor activity was observed for any of the lines studied. Although the use of outbred animals in T cell cloning experiments has certain disadvantages, the present results indicate that cloned rabbit T cells may be useful tools for the elucidation of phenotypic and functional differences of rabbit T cell subsets.
T65 antigen modulation in a phase I monoclonal antibody trial with chronic lymphocytic leukemia patients.
Authors: Schroff R W , Farrell M M , Klein R A , Oldham R K , Foon K A
Published: December 2022

Antigenic modulation of the T65 cell surface antigen was assessed in chronic lymphocytic leukemia patients (CLL) receiving therapy with the murine monoclonal antibody T101 in a phase I clinical trial. A total of 12 patients received 1, 10, or 40 mg doses administered over 2 hr, or 50 or 100 mg doses administered over 50 hr. Decreases in T65 antigen expression (up to 90%) coincided with decreases in the circulating leukemic cell count in those patients who received T101 over a 2-hr period, indicating that clearance of circulating T65 antigen-positive cells could account for most of the observed decreases in T65 antigen expression. In contrast, analysis of bone marrow specimens from these patients indicated that decreases in T65 antigen density in this relatively stationary population resulted not from a cell decrement but rather from antigenic modulation. Pulmonary toxicity prevented administration of doses greater than 40 mg over a 2-hr period; therefore, higher doses (50 and 100 mg) were administered over a 50-hr period. This treatment schedule resulted in greater than 90% reduction in T65 antigen density of both circulating and bone marrow leukemic cells without dramatic drops in circulating leukemic cell counts, indicating that antigenic modulation accounted for most of the observed decreases in T65 antigen density under these conditions. Reexpression of T65 antigen by modulated cells was observed both in vitro and in vivo within 2 to 4 days. Immunoperoxidase staining of in vivo-modulated specimens and in vitro modulation studies with 125I-T101 suggested that T65 antigen-T101 antibody complexes were internalized during modulation. Although antigenic modulation inhibits the potential therapeutic effectiveness of unconjugated T101 antibody in CLL patients, treatment of CLL with T101 drug or toxin immunoconjugates under conditions that bring about rapid and extensive internalization of the T65 antigen may provide an effective means of therapy.
OKT4+ cytotoxic T cells can lyse targets via class I molecules and can be blocked by monoclonal antibody against T4 molecules.
Authors: Strassman G , Bach F H
Published: December 2022

Human cytotoxic T lymphocytes (CTL) have been shown to recognize either class I or class II major histocompatibility (MHC) products. This recognition has been correlated with the expression of OKT antigens on the surface of the CTL. Thus, OKT4+ CTL have been shown to be reactive with class II products, whereas OKT8+ effectors recognize class I molecules. In this study, responder cells were separated according to their OKT4 or OKT8 cell surface phenotype on a fluorescence-activated cell sorter (FACS). The OKT4+ subsets were stimulated with an LCL mutant that did not express DR and MB/MT but did express SB and class I antigens. After 7 days in culture, the activated subsets were tested on a panel of class I matched or mismatched targets. The cytotoxicity observed could be correlated with the presence of matched class I antigens. In addition, monoclonal antibody (MCA) W6/32, directed at a monomorphic determinant on HLA-A and -B molecules, blocked lysis. Furthermore, six OKT4+ CTL clones were derived from the OKT4+ bulk cultures; three clones were found to be directed at class I molecules whereas the other three recognized class II determinants. The ability of these clones to lyse their relevant targets was blocked by OKT4 MCA, raising questions as to the role of the T4 molecule in antigen class-specific CTL recognition.
Proliferative and cytotoxic immune functions in aging mice. II. Decreased generation of specific suppressor cells in alloreactive cultures.
Authors: Gottesman S R , Walford R L , Thorbecke G J
Published: December 2022

The ability of spleen cells from aged C57BL/6 mice to generate specific suppressor cells in mixed lymphocyte cultures (MLC) against allogeneic H-2 antigens was investigated. The suppressor cells from young and old mice were assayed in parallel for their ability to inhibit the proliferative response and the generation of cytotoxicity in fresh MLC. Suppressor cell generation was found to be significantly decreased in 41% of aged mice (23 to 28 mo) as compared to young controls (3-8 mo). The suppressor cells were H-2-specific, radiation-resistant (1000 R), and Thy-1+; they did not function by lysing the fresh stimulators or responder cells, or by absorbing the interleukin 2 in the fresh cultures. Suppression required very small numbers of cells to be effective. It was concluded that the effect of aging was less marked on specific suppressor cell generation than on generation of cytotoxic T cells in the MLC. However, a third type of response studied, the proliferative response, was affected earliest and most severely.
NK recognition of target structures: is the transferrin receptor the NK target structure?
Authors: Newman R A , Warner J F , Dennert G
Published: December 2022

That the transferrin receptor acts as a target antigen for human NK cells has previously been suggested. In this study we used two models to examine the hypothesis that the transferrin receptor is recognized by NK cells. In the first model, we employed mouse cloned NK cells in conjunction with the species-specific monoclonal antibody R17 217, which binds to the murine transferrin receptor. We show that there is no correlation between the amount of transferrin receptor expressed on targets and the susceptibility of these targets to NK lysis or NK binding in cold target competition assays. In the second model, we used human NK cells and transferrin receptor-positive transformants as targets. These transformants were derived from mouse L cells transfected with human DNA and selected for the presence of human transferrin receptor. Results show that, in contrast to the mouse system, there is a correlation between the expression of the human transferrin receptor on targets and the ability of these targets to competitively inhibit the lysis of K562 by NK cells. However, because inhibition is not complete, other cell surface antigens probably play a role in human NK-target interactions.
Selective suppressive effects of glucocorticoids on the early events in the human B cell activation process.
Authors: Bowen D L , Fauci A S
Published: December 2022

The effect of hydrocortisone on the induction of human B cell activation and proliferation has been described. Hydrocortisone prevents the anti-mu-induced cell enlargement of small tonsillar B cells, blocks expression of the activation markers 4F2 and 5E9 induced by anti-mu, inhibits RNA synthesis of small B cells stimulated by anti-mu with or without BCGF, and suppresses B cell proliferation in response to anti-mu and BCGF or to Staphylococcus aureus Cowan I. In contrast, hydrocortisone does not affect the proliferative response of in vitro or in vivo preactivated B cells. Therefore, hydrocortisone has a selective inhibitory effect on early events in the human B cell cycle that subsequently leads to inhibition of total RNA and DNA synthesis. Possible mechanisms of this action are discussed. These studies further define the nature of glucocorticoid-induced modulation of human B cell activation and proliferation.
Unique surface phenotype of T cells in lymphoproliferative autoimmune MRL/Mp-lpr/lpr mice.
Authors: Takei F
Published: December 2022

MRL/l mice, which carry the mutant gene lpr, develop massive T cell lymphoproliferation and an autoimmune syndrome. The surface antigen profile of MRL/l lymphocytes was analyzed with rat monoclonal antibodies. They included YE1/9.9, anti-transferrin receptor; YE1/7.1, which reacts with a population of proliferating immature T cells; and YE1/19.1, a newly identified antibody reactive with a surface antigen on EL-4 cells, which consists of two disulfide-bonded polypeptide chains, each of 115,000 m.w. Flow cytometric analysis of lymphocytes from the enlarged lymph nodes of MRL/1 mice showed the majority of them to be Lyt-1+, YE1/7.1+, YE1/19.1+, YE1/9.9-. In contrast, YE1/7.1 and YE1/19.1 antibodies did not significantly react with lymphocytes from the congenic MRL/n mice, which lack the lpr gene and do not develop lymphoproliferation. Lymphocytes from younger MRL/l mice, which have almost normal size lymph nodes, were also YE1/7.1-, YE1/19.1-. A small proportion of mitogen-stimulated MRL/n lymph node cells appeared to express these antigens at low densities, but it is not known whether they represent a unique cell population. Among the several lymphoid cell lines tested, only the T lymphoma line EL-4 had the Lyt-1+, YE1/7.1+, YE1/19.1+ phenotype. These results suggest that the lpr mutation induces the expansion of a unique T cell subset.
Antigen-dependent regulation of interleukin 2 receptor expression on cloned human cytotoxic T lymphocytes.
Authors: Kaplan D R , Braciale V L , Braciale T J
Published: December 2022

IL 2 receptor expression as a function of time after antigenic stimulation was examined on antigen-dependent human CTL clones specific for type A influenza virus. The anti-Tac monoclonal antibody was used to follow IL 2 receptor levels on the cloned cells. Shortly after antigenic stimulation, IL 2 receptor expression was maximal; by 1 wk, however, levels had decayed considerably, and by 2 wk only background expression remained. Reexpression of IL 2 receptors could be induced by exposure of quiescent clones to antigen or lectin. IL 2-driven proliferation of the cytotoxic clones was also examined, and it decayed with the same kinetics as IL 2 receptor levels. Proliferation of quiescent cells could also be obtained by antigen-specific stimulation. Thus, IL 2 receptor expression by human CTL clones at least in part regulates the antigen-specific proliferation of these cells.