bluuebunny/crossref_metadata_2025 huggingface dataset:
In Search of α1-Microglobulin on the Lymphocyte Surface
Authors: Åkerström Bo , Nilsson Kenneth , Berggård Barbro , Berggård Ingemar
Published: December 2022

α1-Microglobulin was found by immunofluorescence not to be associated with human lymphoid and nonlymphoid cell lines. No accumulation of α1-microglobulin was detected in culture media of these cell lines. A weak membrane fluorescence with anti-α1-microglobulin on peripheral lymphocytes could not be blocked by the purified protein. No release of α1-microglobulin into the growth medium was seen by normal cultured leukocytes. Treatment of normal lymphocytes, erythrocytes, and various cell lines with solubilization techniques did not yield any α1-microglobulin. α1-Microglobulin and protein HC display immunologic and biochemical identity. However, anti-protein HC stained almost all of the tested cell lines and normal lymphocytes. Blocking experiments with the purified protein were not successful. Antibodies reacting with a minor impurity (50 000 d) in the α1-microglobulin or protein HC preparations could be absorbed from anti-α1-microglobulin with normal leukocytes and a lymphoid cell line.
The Differentiation of Cytotoxic T Cells in Vitro
Authors: Okada Masaji , Klimpel Gary R. , Kuppers Rudolf C. , Henney Christopher S.
Published: December 2022

Within 15 hr of establishment of a murine mixed lymphocyte culture, a soluble mediator was produced that was capable of augmenting primary cytotoxic responses to alloantigen. The factor did not induce responsiveness in the absence of antigen, since the amplified response seen in its presence was specific for the stimulating alloantigen. The factor did not therefore appear to function by polyclonal activation of cytotoxic precursor cells.

Production of the amplifying factor(s) was induced by unfractionated spleen cells, but not by cells subjected to UV irradiation or to sonication, making it likely that this deficiency is the basis of the well-documented failure of these stimulator cells to induce primary cytotoxic responses. The amplifying effects of the factor were distinctive from, but synergistic with, those of 2-mercaptoethanol. Production of the amplifying mediator did not require cell division but was dependent upon the presence of Lyt 1+ cells. On the other hand, Lyt 2+ cells were not needed for mediator production, but served as a target cell population on which the factor exerted its action. These findings are compatible with the hypothesis that direct T-T cell collaboration can amplify the differentiation of cytotoxic cells.
Low Molecular Weight Eosinophil Chemotactic Factor in Granulomatous Liver of Murine Schistosomiasis
Authors: Tsuda Shingo , Fukuyama Kimie , Epstein William L.
Published: December 2022

An acidic peptide, preferentially chemotactic for eosinophils, was extracted from livers of mice infected with Schistosoma mansoni. Sephadex G-25 column chromatography showed that the majority of the eosinophil chemotactic activity was detected in the fractions just after elution of the molecular marker vitamin B12 (m.w. 1355.4). This activity began to appear in the livers of some mice 5 weeks after infection. Peak activity was detected at 8 to 12 weeks after infection and persisted at least until 16 weeks. It was sensitive to carboxypeptidase-A. By Dowex-1 anion exchange chromatography, the activity eluted as a narrow peak at pH 3.1 to 2.6 as shown for eosinophil chemotactic factor of anaphylaxis (ECF-A). The activity was also detected in a broad peak at pH 6.3 to 3.7. Unlike ECF-A, the activity was stable to boiling in both acid and alkali. These findings suggest that granulomatous liver of murine schistosomiasis-derived eosinophil chemotactic factor (ECF-G) may play a specific role in eosinophil accumulation in this chronic inflammation.
The Location of the C3 and GLO (Glyoxalase 1) Loci of the Ixth Linkage Group in Mice
Authors: Rubinstein Pablo , Vienne Katalin , Hoecker Gustavo F.
Published: December 2022

Electrophoretic variants of glyoxalase 1 (GLO) and C3 in the mouse were used to define the genetic location of the corresponding genes. Previous work had demonstrated that these two loci are linked to H-2, the major histocompatibility locus of the mouse. The segregation of GLO and H-2 in 880 informative meioses established that the recombination fraction for these two genes is 0.0174 ± 0.005; two intra—H-2 cross-overs position GLO on the K-side of H-2.

Similarly, the linkage of C3 to the H-2 locus was confirmed. The recombination fraction is 0.114 ± 0.022. A triple-point cross, involving C3, H-2, and GLO, discloses that the order of these genes is (from the centromere) GLO—H-2—C3.

GLO is also linked to the MHC in humans. The data given here show that the position of GLO relative to the MHC is the same in both species, although the recombination frequency is higher in humans. It is speculated that the centromeric end of the MHC may be the more ancient gene of the system, because it has remained in linkage with GLO. It is further speculated that the location of C3 in humans may be on chromosome 6, beyond the distance within which conventional family analysis might reveal linkage to HLA.
Isolation and Characterization of Actin and Myosin from B-Lymphocytic Guinea Pig Leukemia Cells
Authors: Fechheimer Marcus , Cebra John J.
Published: December 2022

Actin and myosin have been isolated from a guinea pig B cell leukemia line, L2C. The m.w. and amino acid compositions of these proteins are similar to actin and myosin from other nonmuscle cell types. L2C actin polymerizes to form filaments and activates the ATPase activity of skeletal muscle myosin. Actin in crude lymphocyte extracts does not polymerize as well as predicted from the critical concentration of purified lymphocyte actin suggesting that other factors in lymphocyte extracts regulate actin polymerization. Lymphocyte myosin polymerizes to form synthetic filaments at low ionic strength. Lymphocyte myosin binds to actin, but its ATP-ase activity is not activated by actin. Possible mechanisms for regulation of the lymphocyte contractile apparatus and its importance in a number of lymphocyte functions are discussed.
Involvement of Anti-Ig-Activated Serine Protease in the Generation of Cytoplasmic Factor(s) that Are Responsible for the Transmission of Ig-Receptor-Mediated Signals
Authors: Kishimoto Tadamitsu , Kikutani Hitoshi , Nishizawa Yoshio , Sakaguchi Nobuo , Yamamura Yuichi
Published: December 2022

Involvement of serine protease-activation in the generation of cytoplasmic factor(s) that induced NHP-specific protein kinase activity in nuclei in anti-Ig-stimulated cells was described. DFP or PMSF with anti-Ig inhibited the induction of cytoplasmic factor(s), whereas pretreatment of cells with DFP or PMSF without anti-Ig did not show any inhibitory effect on anti-Ig-induced generation of cytoplasmic factor(s). TAME or BAME with anti-Ig inhibited the generation of cytoplasmic factor(s) and the simultaneous addition of TAME or BAME with DFP protected the generation of cytoplasmic factor(s) against the inhibitory effect of DFP, showing the involvement of trypsin-like, arginine-type serine protease in anti-Ig-induced generation of cytoplasmic factor(s). Anti-Ig-stimulated membrane preparations induced cytoplasmic factor(s) in normal cytoplasm. The m.w. of precursor proteins present in resting B cells and active cytoplasmic factor(s) were approximately 150,000 and 45,000, respectively. These results showed that anti-Ig-activated membrane-bound serine protease split precursor proteins in resting B cells into active cytoplasmic factor(s) responsible for signal transmission.
Early Events in Thymocyte Activation
Authors: Facchini Andrea , Astaldi Giulia C. B. , Cocco Lucio , Wijermans Pierre , Manzoli Francesco A. , Astaldi Alberto
Published: December 2022

Previously, it has been shown that a human thymus-dependent serum factor (SF), isolated from peripheral blood and acting on precursors of mature T lymphocytes, induces an increase in the synthesis of cyclic AMP and proteins in thymocytes. We have now investigated the action of SF on the incorporation of 3H-leucine and 32P-orthophosphate into nuclear proteins of thymocytes after 15 to 240 min of culture. SF induced a rapid increase in the synthesis and phosphorylation of nuclear proteins, especially in the phosphorylated nonhistone chromatin proteins (P-NHCP). Electrophoretic patterns in polyacrylamide gels of the P-NHCP fractions, extracted from the chromatin of the stimulated cells, showed that proteins with m.w. higher than 50 × 103 were synthesized to a larger extent as compared with unstimulated cells. These data suggest that SF acts specifically on the synthesis of P-NHCP and may in this way control DNA-template activity.
Studies of Reticuloendothelial Function in the Mouse with Model Immune Complexes
Authors: Finbloom David S. , Plotz Paul H.
Published: December 2022

We have used stable immune complexes site-specifically cross-linked by way of their combining sites to characterize the elimination of immune complexes in the mouse. These complexes, prepared from rabbit or mouse anti-DNP IgG antibodies, were separated into relatively homogeneous preparations of monomers, dimers, and heavy oligomers. We found that oligomers consisting of rabbit antibodies were cleared from the serum at rates similar to oligomers of homologous mouse antibody. Serum clearance of dimeric immunoglobulin was indistinguishable from clearance of monomer, but dimer accumulated in the liver at least three times faster than monomer. In addition, dimer, like heavy oligomer, bound to circulating blood cells whereas monomer did not. Heavy oligomers were removed from the serum initially at rates about 3-fold that of monomer, principally by the liver. Eighty per cent of heavy oligomers were located within the liver at 1 hr, with less than 1% in the spleen, kidneys, or lungs. Recovery data suggest that oligomers that remain primarily within the intravascular space before liver uptake were removed considerably more rapidly than those oligomers diffusing into the interstitial space.
Antibody-Dependent Complement-Mediated Killing of Schistosomula in Intraperitoneal Diffusion Chambers in Mice
Authors: Kassis Amin I. , Warren Kenneth S. , Mahmoud Adel A. F.
Published: December 2022

The present investigations were designed to determine the biologic role of the different host defense mechanisms against schistosomiasis mansoni that are independent of direct cell-parasite contact. Schistosomula were placed in Millipore chambers of 0.22-µm pore size and were implanted i.p. into immune and nonimmune age- and sex-matched control mice. Significant degree of mortality was detected on recovery of the implanted schistosomula at 24 hr; the mean dead organisms in immune animals was 30 ± 4% as compared to 10 ± 2% in control mice (p < 0.01). This effect on the implanted schistosomula into immune mice was confirmed by injecting these organisms into normal recipients and by recovering those that mature to adult worms. The ability of immune mice to kill significant numbers of implanted schistosomula was dependent on the intensity of infection and developed only after 8 weeks, which parallels the onset of protective immunity in vivo. Adoptive transfer of immune sera to normal recipients resulted in a mean of 23 ± 2% dead schistosomula in the recipient mice, which represents approximately 65% of the protection detected in the donor animals. Depletion of the eosinophils or neutrophils in the immune mice did not alter their ability to kill significant numbers of the implanted schistosomula. Cobra venom factor treatment, on the other hand, resulted in abrogation of the significant schistosomula killing detected in immune mice or in animals adoptively transferred with immune serum. These results demonstrate that specific acquired immunity to schistosomiasis in vivo is partially mediated by humoral factors and is dependent on the presence of complement. The Millipore chambers may help elucidate the significance of the many mechanisms which have already been described for host immune responses against the multicellular parasite Schistosoma mansoni.
Effects of Indomethacin on Cyclic Nucleotide Levels and Histamine Release from Rat Serosal Mast Cells
Authors: Lewis Robert A. , Holgate Stephen T. , Jackson Roberts L. , Maguire James F. , Oates John A. , Frank Austen K.
Published: December 2022

Previous studies in homogeneous and mixed cell systems on the changes in cyclic nucleotide levels that accompany the immunologic activation of mast cells for histamine release have not considered the extent to which these changes occur secondarily to the generation of the oxidative products of arachidonate. The recognition that mast cells themselves generate oxidative products of arachidonate upon immunologic activation and that exogenous prostaglandins can modulate cellular levels of 3′,5′-cyclic adenosine monophosphate (cyclic AMP) prompted the assessment of the effects of endogenous oxidation of arachidonate on changes in cyclic nucleotide levels accompanying immunologic activation. Thus, immunologic activation of purified rat serosal mast cells was carried out in the presence of a concentration of indomethacin that completely suppressed the oxidative conversion of arachidonate to the prostaglandins D2 (PGD2) and F2a (PGF2a) without having any effect on the time course and magnitude of histamine release. The immunologic activation of rat mast cells gave an early and a late monophasic rise of both cyclic AMP and 3′,5′-cyclic guanosine monophosphate (cyclic GMP). The inhibition of cyclo-oxygenase by 10 µM indomethacin abolished both monophasic rises in cyclic GMP and the second monophasic rise in cyclic AMP, but had no significant effect on the early peak of cyclic AMP that followed immunologic activation. Thus, those changes in cyclic nucleotide levels that are suppressed by indomethacin are not part of the primary activation-secretion event of mast cells and may be secondary to the generation of oxidative products of arachidonate. In contrast, the indomethacin-resistant elevations in cyclic AMP observed in the homogeneous rat mast cell system 5 to 15 sec after activation may represent an early event in the coupled activation-secretion response.
Effects of Arachidonic Acid and Its Metabolites on Antigen-Induced Histamine Release from Human Basophils in Vitro
Authors: Marone Gianni , Kagey-Sobotka Anne , Lichtenstein Lawrence M.
Published: December 2022

We have studied the role of arachidonic acid (AA) metabolism in immediate hypersensitivity reactions. The AA analog, eicosa-5,8,11,14-tetraynoic acid (ETYA), which inhibits both the cyclo-oxygenase and lipoxygenase pathways of AA metabolism, was found to cause a dose-dependent inhibition of antigen-induced histamine release from human basophils; complete inhibition occurred at 10-4 M. In contrast, nonsteroidal anti-inflammatory drugs (NSAID) including indomethacin (10-9 to 10-6 M), meclofenamic acid (10-6 to 10-4 M), and aspirin (10-5 to 10-3 M), which block the cyclo-oxygenase pathway of AA metabolism, enhanced mediator release. AA (10-7 to 10-5 M) also enhanced basophil mediator release, an effect that was not affected by preincubation with indomethacin. Other fatty acids including linoleic, linolenic, and stearic acids were without effect. Indomethacin (and the other NSAID studied) reversed the inhibition caused by the AA metabolite, prostaglandin (PG) E2, in a dose-dependent fashion. This reversal of inhibition was not unique to PG, but was observed with all inhibitors of histamine release studied that act via activation of adenylate cyclase including isoproterenol and the H2 agonist, dimaprit. NSAID, however, did not reverse the inhibition caused by drugs that do not act on adenylate cyclase, i.e., dibutyryl cAMP, isobutylmethylxanthine, and 2-deoxyglucose. In the leukocyte preparations studied, the NSAID did not block the increase in the intracellular cAMP levels caused by agonists that act via adenylate cyclase. It appears that a product(s) of AA metabolism, generated via the lipoxygenase pathway, is involved in IgE-mediated histamine release and that the same or a different product(s) modulates the control mechanisms that are linked to adenylate cyclase.
Characterization of Isolated Guinea Pig Kupffer Cells: Accessory Cell Function in Mitogen-Induced T Lymphocyte Activation
Authors: Rogoff Thomas M. , Lipsky Peter E.
Published: December 2022

Techniques were developed to isolate and purify guinea pig Kupffer cells (KC) so that their potential to interact with immunocompetent lymphocytes could be assessed. KC were prepared from guinea pig liver by sequential enzymatic digestion with collagenase and trypsin, followed by differential centrifugation, overnight culture, and glass adherence. Peritoneal exudate macrophages (PEM), induced with mineral oil but treated in otherwise identical fashion, were used for comparison in all experiments. More than 95% of the cells in KC preparations had the morphologic features of mononuclear phagocytes (Mø) and possessed peroxidase and nonspecific esterase activity. KC also exhibited many of the physiologic characteristics of Mø, including glass adherence, fluid phase pinocytosis, and phagocytosis of both nonspecific and opsonized particles. The capacity of KC to function as accessory cells necessary for the induction of mitogen-stimulated T lymphocyte proliferation was examined. Lymph node lymphocytes (LNL) depleted of adherent cells by passage over nylon wool columns were used as the responding T cell population. LNL when cultured alone were unable to undergo proliferative responses to optimal concentrations of phytomitogens. The addition of either KC or PEM to these cultures restored mitogen responsiveness to LNL. Furthermore, modification of both KC and PEM by treatment with neuraminidase and galactose oxidase generated mitogenic moieties capable of inducing blastogenesis in adherent cell-depleted LNL populations. These results provide evidence that KC are capable of functioning as accessory cells for the activation of T lymphocytes.
Human Macrophage and Lymphocyte Responses to Mitogen Stimulation after Exposure to Influenza Virus, Ascorbic Acid, and Hyperthermia
Authors: Manzella John P. , Roberts Norbert J.
Published: December 2022

Ascorbic acid supplementation, in several clinical cold-like viral illness studies, produced a modest reduction in severity and duration of symptoms. The current studies examined effects of ascorbic acid and hyperthermia (38.5°C) on PHA-stimulated transformation responsiveness of normal and influenza A virus-exposed human mononuclear leukocytes.

PHA responsiveness was significantly depressed by exposure to influenza virus (p <0.02), but enhanced by ascorbic acid (p <0.02) or hyperthermia (p <0.001). Neither ascorbic acid nor hyperthermia stimulated the mononuclear leukocytes in the absence of PHA. The adverse effect on PHA responsiveness caused by exposure to influenza virus was counteracted by ascorbic acid (p <0.005), hyperthermia (p <0.02), or both ascorbic acid and hyperthermia (p <0.001). Highly purified normal macrophages and lymphocytes were exposed independently to ascorbic acid or hyperthermia and tested for PHA responsiveness singly and together. Ascorbic acid, and probably hyperthermia, appeared to enhance macrophage function primarily. The data demonstrated that ascorbic acid and hyperthermia affected PHA-induced transformation responses of normal human mononuclear leukocytes, and counteracted the depression of PHA transformation responsiveness caused by exposure of the mononuclear leukocytes to influenza A virus.
Maturation of Bursal Stem Cells within Allogeneic or Syngeneic Bursal Microenvironment: Acquisition of Postbursal Maturity
Authors: Vainio Olli , Toivanen Auli
Published: December 2022

Differentiation of bursal stem cells in an allogeneic or syngeneic bursal microenvironment was compared. Bursal stem cells were transplanted into CY-treated 4-day-old recipients and permitted to differentiate in these hosts for 6 weeks. Their maturity degree was thereafter assessed by transplanting them into secondary recipients by using morphologic and functional criteria. As the secondary recipients 4-day-old CY-treated or CY-treated and surgically bursectomized chicks were used. The results obtained demonstrate that bursal stem cells develop to mature postbursal cells also within an allogeneic bursa. They also indicate that although the interaction of different lymphoid cells requires histocompatibility, the interaction between stromal cells in the bursa and lymphoid progenitors is not genetically restricted.
C9 Hemolytic Activity of the Soluble C5b-9 Complex of Guinea Pig Complement, Analogous to Human SC5b-9
Authors: Kinoshita Taroh , Hong Kyongsu , Inoue Kozo
Published: December 2022

When guinea pig serum was treated with zymosan, hemolytic activity of C9 appeared in a macromolecular complex with a m.w. of about 1,000,000 as well as free C9. The complex was isolated by sequential gel filtrations and sucrose density gradient centrifugation. On immunodiffusion analysis the C5 and C9 incorporated into the complex showed loss of some antigenic determinants of the respective precursor proteins. Immunoelectrophoretic analysis revealed that the complex moved faster than free C9. Preliminary estimation of the subunit composition of the complex by sodium dodecylsulfate (SDS) polyacrylamide gel electrophoresis showed that it contained similar proteins to those found in the human SC5b-9 complex. The complex seemed to contain one molecule each of C5b, C6, C7, C8, and protein corresponding to human S protein and three molecules of C9.

The C9 hemolytic activity of the complex was found to be very similar to that of free C9, both dose dependently and kinetically. It was neutralized with antibody to free C9 in a first order fashion, but the antibody to C5 did not neutralize the hemolytic activity in spite of its ability to precipitate the complex. The complex showed C9 hemolytic activity and also slight C7 activity. However, no other hemolytic activity was found when assayed with all the intermediate cells along the classical pathway or when assayed as any other of the components of complement. On the other hand, the human counterpart, SC5b-9, showed no C9 hemolytic activity but competitively inhibited the hemolytic activity of free C9, as reported previously by Kolb and Müller-Eberhard.
Lymphocyte Adherence to High Endothelial Venules: Characterization of a Modified in Vitro Assay, and Examination of the Binding of Syngeneic and Allogeneic Lymphocyte Populations
Authors: Butcher Eugene C. , Scollay Roland G. , Weissman Irving L.
Published: December 2022

Mouse lymphocytes incubated on fresh frozen sections of mesenteric lymph nodes adhere specifically to the endothelium of high endothelial venules (HEV), specialized vessels through which lymphocytes normally enter lymph nodes from the blood. An assay of the specific binding of lymphocytes to HEV in frozen sections is characterized and is used to compare the capacity of various syngeneic and allogeneic lymphocyte populations to adhere to HEV in mouse mesenteric lymph nodes. The relative adherence of lymphocyte populations to HEV in this in vitro system correlates highly with their localization in HEV a) during single-pass perfusion through the lymph node at 37°C, or b) in vivo 15 min after i.v. injection. With this assay we have shown that the binding of mesenteric node lymphocytes (MNL) to syngeneic mesenteric node HEV is constant: no significant variation was observed between individual mice, between sexes, between mice of various ages (3 weeks to 9 months), or after treatment of the lymphocyte donor with hydrocortisone acetate. B lymphocytes adhered as well as T lymphocytes and exhibited no preference for HEV bordering on B areas in the lymph node section. In general, other syngeneic cell populations bind in proportion to their content of mature lymphocytes. MNL exhibited no preferential adherence to syngeneic as opposed to allogeneic HEV, but A/J and BALB/c MNL demonstrated greater binding than C57BL/6J and AKR/Cum MNL, regardless of the HEV donor strain.
T Lymphocyte-Mediated Suppression of Myeloma Function in Vitro
Authors: Abbas Abul K.
Published: December 2022

Alloreactive cells generated by in vitro stimulation of C57BL/6 (H-2b) spleen lymphocytes with irradiated MOPC 315 or MOPC 104E (H-2d) cells were shown to lyse 51Cr-labeled myeloma targets at high effector:target ratios under conditions of maximal cell contact. After coculture at low effector:target ratios under conditions of inefficient cell contact, the alloreactive cells cause variable and frequently minimal lysis of myeloma targets but markedly suppress antibody secretion even by viable myeloma cells. The suppressor cells are radioresistant T cells lacking I-J subregion-encoded surface determinants; their precursors are insensitive to cyclophosphamide; suppression is H-2 specific and not mediated by secreted factors; and the suppression is blocked by Cytochalasin B, a known inhibitor of T cell-mediated cytolysis. These properties are typical of cytolytic T lymphocytes (CTL) and not of defined suppressor T cells, suggesting that inhibition of myeloma function probably represents a pre-lytic effect of the alloreactive CTL, although a CTL-like suppressor cell effect cannot be definitively excluded. These results are discussed with reference to the possible relationships between suppressor and cytolytic T lymphocytes.
Studies on the Induction and Expression of T Cell-Mediated Immunity
Authors: Grimm Elizabeth A. , Bonavida Benjamin
Published: December 2022

Secondary cytotoxic thymus-derived lymphocytes (CTL) with specificity for the priming alloantigen are shown to be generated by in vitro stimulation of allosensitized memory splenocytes with NAGO-oxidized syngeneic, allogeneic, or xenogeneic cells. The cell surface aldehyde resulting from NAGO modification of stimulator cells is required for triggering both lymphocyte proliferation and secondary (2°) CTL activation, since treatment with the reducing agent, KBH4, completely inhibited these responses. Compared to alloantigen stimulation, NAGO-oxidized stimulator cells induced an earlier peak of proliferation (2 to 3 vs 4 to 5 days), whereas the peak of the cytotoxic activity was identical (days 3 to 4).

Syngeneic splenocytes, peripheral T cells, plastic and nylon nonadherent cells, and the P388D1 macrophagelike cell line were all efficient stimulators after NAGO modification. Stimulation by syngeneic cells was shown to be mitomycin C resistant, U.V. irradiation sensitive, and ineffective when heat killed cells are used. Therefore, NAGO stimulation is distinct from antigen specific activation whereby U.V. treated cells, heated cells and nonviable membrane fragments have been shown to differentiate memory cells into 2° CTL. In addition, neither NAGO-treated sheep red blood cells nor syngeneic thymocytes were found to be effective stimulators. The addition of specific anti-Ia serum to the culture fluid inhibited both the proliferative and cytotoxic response, suggesting a role for an Ia signal during NAGO stimulation. These results show that the signal(s) required for the activation of 2° CTL from memory populations can be provided by certain subpopulations of NAGO-modified cells and is also dependent on the presence of Ia or an Ia-like molecule.
Activation to IgG Secretion by Lipopolysaccharide Requires Several Proliferation Cycles
Authors: Gronowicz Eva Severinson , Doss Carol , Schröder Jim
Published: December 2022

We have investigated the relationship between IgG secretion and cell proliferation after polyclonal activation of murine spleen cells with lipopolysaccharide (LPS). It was found that IgG secretion was optimal when cells proliferated extensively. Under those conditions, DNA synthesis commenced 8 to 12 hr after exposure to LPS. Increased proliferative activity was observed up to day 3, when the majority of the lymphoblasts were mitotically active. Two inhibitors of DNA synthesis, thymidine (TdR) and hydroxyurea (HU) caused a reduction in both the IgM and the IgG response, but the latter was more severely reduced. The inhibition was strongest when TdR and HU were added to cultures early after exposure to LPS, indicating that the cells developing to Ig secretion were continuously proliferating. 5-bromo-2′-deoxyuridine (BrdU) caused a general inhibition of IgM and IgG secretion at high concentrations, and a selective inhibition at low concentrations. The selective inhibition of IgG secretion, when measured on day 4, was also observed after a pulse of BrdU on days 1 and 2. The data suggest that development to IgG secretion is a complex process, which requires several proliferation cycles.
Functional Differentiation of T Cell Precursors
Authors: Cohn Marjorie L. , Scott David W.
Published: December 2022

Irradiated mice reconstituted with bone marrow from sheep γ-globulin- (SGG) tolerant syngeneic donors display reduced IgG responsiveness to challenge with trinitrophenylated (TNP)-SGG compared with recipients of normal marrow. This effect is SGG-specific and is due neither to suppressor T cells nor to antigen carryover. “Helper T cell precursor tolerance” can be induced with as little as 40 µg tolerogen (SGG). Unlike mature helper T cells, these precursors show both a rapid induction and rapid waning patterns, suggesting a high rate of turnover. Our results imply that marrow helper T cell precursors bear antigen-specific receptors and that the T cell repertoire must be at least partially generated before residence in the thymus.
Structural Evidence for Recombination at the Igh (H Chain) Complex Locus in BAB 14 Mice
Authors: Claflin J. Latham , Wolfe Jacqueline , Ruppert Valerie J.
Published: December 2022

Anti-PC antibodies from the recombinant mouse BAB 14 were evaluated by analytical isoelectric focusing in an attempt to gain information on the crucial question of whether the recombinational event in this strain occurred in V region structural genes or in allotype-linked regulatory genes. Immune sera from BAB 14, C57BL/6 and its congenic partner CB20 (both Ighb), BALB/c and its congenic partner BC8 (both Igha) were isofocused. Antibodies bearing two distinct idiotypes, T15 Id and M511 Id, were detected by an in situ localization procedure with radiolabeled anti-T15 and anti-M511 idiotypic antibodies. The IEF spectrotypic pattern of IgG antibodies of each strain revealed separate populations of T15 Id-positive (IEF-T15) and M511 Id-positive (IEF-M511) antibodies. The IEF-T15 and IEF-M511 spectrotypes in BAB 14 most closely resembled those of C57BL/6 due to the dominant charge effect of the CH region. However, the entire set of IEF-T15 bands and a portion of the IEF-M511 bands in BAB 14 show a slight but clearly evident anodal shift in its pI relative to C57BL/6. This finding of a shift, regardless of IgG subclass, demonstrates that the difference between BAB 14 and C57BL/6 resides in the VH region and not the CH region. Thus, there is direct evidence from structural data that H chains of BAB 14 are composed of a CH segment (allotype marker) commonly expressed by C57BL/Ka (Ighb) mice and VH segment commonly expressed by BALB/c (Igha) mice. These findings are discussed in terms of recombination in structural versus regulatory genes.
Emergence of Suppressor Cells of Immunoglobulin Synthesis During Acute Epstein-Barr Virus-Induced Infectious Mononucleosis
Authors: Haynes Barton F. , Schooley Robert T. , Payling-Wright Charles R. , Grouse Jan E. , Dolin Raphael , Fauci Anthony S.
Published: December 2022

In vitro polyclonal immunoglobulin (Ig) production and immunoregulation were studied in peripheral blood lymphocytes (PBL) from 17 acute (<2 weeks after onset of symptoms) and 14 convalescent (2 to 5 weeks after onset of symptoms) Epstein-Barr virus (EBV)-induced infectious mononucleosis (IM) patients. PBL from both groups developed fewer anti-sheep red blood cell (SRBC) plaque-forming cells (PFC) (p < 0.001) and fewer intracytoplasmic (ICP) Ig-positive cells (p < 0.01) than normal PBL after in vitro stimulation with pokeweed mitogen (PWM). However, there was a significant trend toward increased PWM-induced anti-SRBC PFC with increasing time after onset of IM symptoms (R = 0.389, p < 0.025). PBL from acute but not convalescent IM patients significantly suppressed the PWM-induced anti-SRBC PFC response (15 ± 8% of expected, p < 0.001) and the ICP Ig response (21 ± 8% of expected, p < 0.001) of normal PBL in allogeneic co-cultures. The suppressor cell activity in PBL of acute IM patients was removed by treatment with anti-T cell antiserum and complement as well as by the addition of 10-5 M hydrocortisone to the cell cultures.

In addition, acute IM cells in the presence of normal T cell help were able to respond normally in vitro to PWM to generate both anti-SRBC PFC and ICP Ig. Thus, PBL from both acute and convalescent IM patients manifest decreased Ig production after in vitro polyclonal stimulation. These decreased responses were associated with the emergence of corticosteroid sensitive suppressor T cells that were demonstrable in PBL of acute IM patients, and were no longer demonstrable in PBL of convalescent IM patients. The transient presence of peripheral blood suppressor T cells in these acute IM patients was associated with an uneventful clinical course and may represent in vivo immunoregulatory cell activation in response to EBV-induced polyclonal B cell activation.
Specific 125I-Iodination of Cell Surface Lipids: Plasma Membrane Alterations Induced During Humoral Immune Attack
Authors: Schlager Seymour I.
Published: December 2022

A method whereby lactoperoxidase-catalyzed 125I-iodination of plasma membrane lipids can be achieved is described. The reaction results in a uniform and stable labeling of neutral lipids, phospholipids, lysophosphatides, free fatty acids, and triacylglycerides. By the use of this method, the action of antibody plus complement (C) on the specific release of lipid from the plasma membrane of line-10 tumor cells was studied. Within 15 min after the addition of C to antibody-sensitized cells, the enhanced release of specific lipid classes from the cell surface was observed; these lipids included sphingomyelin, phosphatidylserine, and phosphatidylcholine. The release of phosphatidylethanolamine and, in some instances, triglycerides, was reduced after antibody-C attack. Neither the specificity of the antibody used to sensitize the cells nor the ability of the antibody plus C to be cytotoxic to the cells appeared to affect the identity or amounts of lipids released from the cells.
Deficient Production of the Lymphokine Eosinophil Stimulation Promoter in Chemically Induced and Mutation Diabetes Mellitus in Mice
Authors: Kazura James W. , Gandola Carl , Rodman Harvey R. , Mahmoud Adel A. F.
Published: December 2022

The cellular defects possibly responsible for diminished in vivo granuloma formation in diabetic Schistosoma mansoni-infected mice were investigated. Diabetic and control animals develop a similar degree of eosinophilia. Eosinophils obtained from diabetic mice also respond normally to the lymphokine eosinophil stimulation promoter (ESP). Lymphoid cells of chemically induced (streptozotocin) and mutation diabetic (db/db) mice, however, have a decreased capacity to produce/secrete ESP in response to soluble egg antigens of S. mansoni. Administration of insulin to diabetic mice is associated with a partial reversal of the decreased ability of their lymphoid cells to generate ESP. These findings show that defective cellular immunity in diabetic animals may be partially explained by the failure of their lymphocytes to produce the soluble mediators involved in recruitment of target cells.
Alternative Complement Pathway-Mediated Lysis of Measles Virus Infected Cells: Induction by IgG Antibody Bound to Individual Viral Glycoproteins and Comparative Efficacy of F(ab′)2 and Fab′ Fragments
Authors: Sissons J. G. Patrick , Cooper Neil R. , Oldstone Michael B. A.
Published: December 2022

Lysis of measles virus-infected cells by human serum is dependent on IgG antibody and an intact alternative complement pathway. The binding sites of IgG measles virus antibody, or its fragments, on the surface of measles virus-infected cells were determined by cell surface radioiodination and immunoprecipitation. It was found that IgG bound to either of the two measles virus surface glycoproteins was effective in inducing cell lysis by the alternative pathway. Thus, IgG bound only to the viral hemagglutinin (HA) or only to the fusion protein (F) could induced lysis in C4-depleted human serum; the same number of IgG molecules bound to either HA or F induced equivalent dose-related lysis. Divalent antibody bound preferentially to HA on the surface of intact cells; this enabled antibody to F to be obtained free of antibody to HA by adsorption. Quantitative studies of binding and lysis showed that lysis of measles virus-infected cells by whole or C4-depleted serum required binding of at least 10 times more Fab′ than F(ab′)2 or IgG molecules per cell. In these experiments, C4-depleted serum was quantitatively and kinetically equivalent to whole serum in mediating lysis of measles-infected cells. Hence, the binding of IgG to either of the two measles viral polypeptides (HA, F) expressed on the surface of infected cells induces lysis by the alternative pathway, and there is a clear requirement for divalency for effective induction of lysis.
The Beige Mutation in the Mouse
Authors: Roder John C. , Lohmann-Matthes Marie-Luise , Domzig Wolfgang , Wigzell Hans
Published: December 2022

In the preceding paper we showed that the beige mutation in the mouse leads to a complete impairment in NK cell function, which is predetermined at the level of progenitor cells in the bone marrow. The observations in this report suggest that the beige defect is selective for NK cells since killer function in promonocytes, macrophages, and alloimmune or lectin-generated T cells is relatively normal. In addition, spleen cell-mediated ADCC against chicken erythrocytes and promocyte ADCC against tumor cells was also identical in homozygous mutants or their heterozygous littermate controls. However, spleen cell-mediated ADCC against tumor cells was impaired in bg mice as well as antibody-independent NK cytolysis. When combined with the work of others these results suggest that NK and K cells are identical and one effector can kill by an antibody-dependent or independent mechanism. Since the beige defect lies within the lytic machinery of the cell rather than at the level of target recognition, these results also suggest that the NK lytic mechanism as well as the effector cell type is distinct from that in T cells, macrophages, promonocytes, or nonlymphocytic effectors participating in ADCC. These mice should provide a valuable model for studies of in vivo resistance to tumors, grafts, and virus-infected cells in the absence of functional NK cells.
Early Damage of Herpes-Infected Cells by Antibody-Dependent Cellular Cytotoxicity: Relative Roles of Virus-Specified Cell-Surface Antigens and Input Virus
Authors: Shore Steven L. , Cromeans Theresa L. , Norrild Bodil
Published: December 2022

Previous studies in our laboratory indicated that tissue culture cells acutely infected with type 1 herpes simplex virus (HSV) could be recognized at 2 hr post-infection (PI) by antibody-dependent cellular cytotoxicity (ADCC) and damaged by ADCC at 3 hr PI. Since the design of these studies did not preculde the possibility that damage occurs even earlier in the infectious cycle, we performed the present series of experiments to assess this possibility as well as to gauge the relative contributions of virus-specified cell-surface antigens and input virus in rendering target cells susceptible to cytotoxic attack.

When present during both the first 2 hr of infection as well as during the following 4-hr ADCC assay period, both actinomycin D (1 µg/ml), an inhibitor of RNA synthesis, and cycloheximide (5 µg/ml), an inhibitor of protein synthesis, strongly reduced the cytotoxic reaction (by 70 and 90%, respectively). The inhibitory effects of these agents indicate that the bulk, but not all, of the ADCC reaction was directed at target cells synthesizing virus-specified surface antigens. Between 10 and 30% of the injured cells were attacked because of input virus that had not penetrated the target cell. This was shown by a) the small portion of ADCC that resisted actinomycin D and cycloheximide, b) the biphasic ADCC kinetic reaction curve of target cells exposed to antibody and effector cells immediately after virus adsorption, and c) the small but reproducible cytotoxic reaction against target cells exposed to virus inactivated by ultraviolet light. A small portion of the infected cell population could be injured by ADCC within the first hour after viral adsorption due to input virus.
Mechanisms of Action of Hyperimmune Serum in Mediating Protective Immunity to Rodent Malaria (Plasmodium Berghei)
Authors: Quinn Thomas C. , Wyler David J.
Published: December 2022

Little is known of the mechanisms whereby antibody can transfer protective immunity to malaria. We therefore examined in vivo the effects of the administration of hyperimmune serum to nonimmune rats on 1) the course of parasitemia after inoculation of P. berghei-infected erythrocytes, 2) the clearance of 51Cr-labeled infected erythrocytes, 3) the intraerythrocytic maturation of parasites, and 4) the invasion of erythrocytes by merozoites. Parasitemia rose in rats treated with normal serum but decreased in rats treated with hyperimmune serum. This protective effect of hyperimmune serum was not associated with a significant increase in the rate of clearance or the total reticuloendothelial organ uptake of 51Cr-labeled infected erythrocytes. Hyperimmune serum had no effect on the early intraerythrocytic development of the parasites since parasite maturation from ring form to trophozoite stage progressed normally. However, ring forms failed to increase after schizont rupture in recipients of hyperimmune serum, whereas an increase was observed in recipients of normal serum. Our in vivo study therefore supports in vitro evidence that the major protective role of antibody is to interfere with the interaction of the extracellular form of the parasite (merozoite) and the erythrocyte during the invasion process. Specifically, our data exclude opsonization of infected erythrocytes or interference with early parasite maturation as major mechanisms of antibody-mediated protection. Finally, our study suggests that the spleen may play an additional role by removing opsonized merozoites, since hyperimmune serum administered to splenectomized rats was only partially effective in preventing merozoite invasion of erythrocytes.
Isolation of the Fourth Component (C4) of Rat Complement
Authors: Daha Mohamed R. , van Es Leendert A.
Published: December 2022

The fourth component of rat complement was purified to homogeneity by sequential chromatography of rat plasma in benzamidine on QAE-A50, SP-C50, hydroxyapatite, and gel filtration on Bio-Gel A 1.5. The final material was homogeneous on SDS-PAGE analysis and had a calculated m.w. of 198,000. A monospecific antibody against rat C4 was obtained from immunized rabbits. The concentration of rat C4 in the plasma of normal 4-month-old Wistar rats was 190 ± 34 µg/ml (mean ± 1 S.D.).
Preservation of C3, C5, and C5a Functional Activities by a New Radiolabeling Method; Demonstration of C5 Products in Complement-Activated Serum
Authors: Kreutzer Donald L. , Kunkel Steven , Ward Peter A. , Showell Henry , McLean Robert H.
Published: December 2022

A new method is described for the radiolabeling of C3, C5, and C5a. Using a methylation procedure we have labeled C3, C5, and C5a with 14CH2O, achieving high specific activity, without loss of hemolytic activity in the case of C3 and C5, and in the case of C5a, full retention of chemotactic and enzyme releasing activities. The distribution of radiolabel in the α and β chains of C5 was 75 and 25%, respectively. Using intact labeled C5 the fragmentation pattern in zymosan-activated human serum, assessed by gel filtration and electrophoresis, is complex, yielding multiple fragmentation products. The labeling method described in this paper yield materials that have many advantages over the currently used labeling procedures.
Characterization of the IgE Receptor Isolated from Human Basophils
Authors: Hempstead Barbara L. , Parker Charles W. , Kulczycki Anthony
Published: December 2022

The IgE receptor of human basophils was purified by using simple and repetitive affinity chromatography on human IgE-Sepharose. Basophils were partially purified from peripheral blood of patients with chronic myelogenous or basophilic leukemia. Cells were labeled with 125I by using the lactoperoxidase method and were solubilized with nonionic detergent. Elution of IgE-Sepharose with 0.5 N acetic acid, 1% NP-40 allowed recovery of active IgE receptor. Analysis of human IgE receptor by SDS polyacrylamide gel electrophoresis with 10% gels demonstrated one major radioactive peak with an apparent m.w. of 58,000 to 68,000, somewhat larger than rat IgE receptor. The purified human IgE receptor was active since ∼10 to 42% of labeled receptor could specifically rebind to insolubilized human IgE. Rebinding was blocked by nanomolar concentrations of soluble human IgE or rat IgE but not by human or rat IgG, heat-inactivated human IgE, or heat-aggregated human IgG; thus it appears that rat IgE as well as human IgE binds specifically to human IgE receptor. The relative abilities of active rat IgE and active human IgE to inhibit human IgE receptor rebinding could not be precisely determined because of the limitations in assessing the proportion of human IgE that retains receptor-binding activity.
Presence of Circulating Anti-H-2 Antibody and Cell-Mediated Precursors in Successful DBA/2J-Plus-(DBA/2J × C3H/HeJ)F1 Parabiosed Mice
Authors: Drell Daniel W. , Carlson George A. , Wegmann Thomas G.
Published: December 2022

Joining adult DBA/2J (H-2d) mice with adult (DBA/2J × C3H/HeJ)F1 (H-2d/H-2k) mice in parabiosis results in long-term survival of 44% of such pairs, the complete takeover of the F1 lymphoid and erythropoietic systems by parent type cells, and specific unresponsiveness toward C3H/HeJ skin grafts on the parental partner. In an adoptive transfer system measuring the killing of an 131I-iododeoxyuridine-labeled tumor that is H-2 compatible with C3H/HeJ, parabiont spleen cells show full reactivity upon sensitization to C3H/HeJ alloantigens and do not inhibit the sensitization of normal DBA/2J cells to C3H/HeJ. The adoptively transferred parabiont cells do not appear to be active unless reexposed to C3H/HeJ antigen. However, intact parabionts show accelerated tumor destruction in situ even when not deliberately restimulated with C3H/HeJ spleen cells. Parabiont serum gives accelerated tumor clearance on passive transfer. Parabiont serum also shows complement-dependent antibody titers in vitro of 1:10 to 1:20 against lymphoid target cell antigens coded for by genes in the interval from K to I-E of the H-2 complex. Most of this activity is sensitive to 2-mercaptoethanol. Although circulating antibody is present in the parabionts, F1 parabiont skin shows no delay in rejection by normal DBA/2J recipients.