bluuebunny/crossref_metadata_2025 huggingface dataset:
Separation of Lymphocyte Mitogen from Lymphotoxin and Experiments on the Production of Lymphotoxin by Lymphoid Cells Stimulated with the Partially Purified Mitogen: A Possible Amplification Mechanism of Cellular Immunity and Allergy
Authors: Gately Celia L. , Gately Maurice K. , Mayer Manfred M.
Published: December 2022

A mitogenic factor (MF) from guinea pig lymph node cells (LNC), which was produced by stimulating immune LNC with the specific antigen, ovalbumin, was partially purified by the sequential use of Sephadex G-200 gel filtration, CM-cellulose ion exchange chromatography, DEAE-cellulose ion exchange chromatography, and polyacrylamide disc gel electrophoresis. The product was purified at least 100-fold with regard to protein content. In addition, the purified MF was functionally pure with respect to the absence of lymphotoxin (LT), another guinea pig lymphokine.

The possibility that MF, or a substance closely associated with this factor, might serve to amplify cellular immune and allergic reactions by nonspecifically inducing lymphoid cells to produce other lymphokines was then tested by stimulating cells from various lymphoid tissues with the purified MF and assaying the culture fluids for LT activity as a measure of lymphokine production. All the cell types studied incorporated 3H-thymidine in response to purified MF, and some of them produced LT. Thus, spleen cells from normal guinea pigs produced some LT when stimulated with a large dose of MF, but mesenteric LNC produced none. However, mesenteric LNC from guinea pigs that had been immunized by footpad injection of complete Freund's adjuvant (CFA) 10 or 16 days previously produced LT in amounts that were dependent on the stimulating dose of MF. Substantial quantities of LT were produced by cells from peripheral lymph nodes draining the sites of immunization, as well as by spleen cells, the degree of response being dependent on the stimulating dose of MF and the time span after injection of CFA. These results indicate that purified MF, or a substance closely associated with this factor, is capable of inducing LT production by a subset of lymphoid cells that appears to be expanded after immunization. Electrophoretic observations indicated that the induction of LT production may be due to a factor which is distinct from MF although closely similar in molecular size and charge.
Cytotoxic Activity of Lymphocytes
Authors: Walker Sharyn M. , Lee Sheng-Chung , Lucas Zoltan J.
Published: December 2022

Two different lymphotoxins synthesized by human blood lymphocytes stimulated with phytohemagglutin (PHA) are described. The two toxins are called α-LT and β-LT relative to their elution order on gel chromatography. Their m.w. are 75,000 and 45,000 daltons, respectively. Both toxins appear as early as 7 hr after the addition of PHA, with the amount of β-toxin exceeding that of α-LT initially. Both toxins are differentiated from a third toxin (adherent cell toxin, ACT) made by plastic-adherent cells without requiring mitogen-stimulation. Depletion of macrophages or B cells does not affect the synthesis of either lymphotoxin. Monospecific antisera were elicited to α-LT. Antisera elicited to β-LT also neutralized α-LT but to a significantly lesser degree. α-LT appears to be the lymphotoxin described by most other workers. β-LT is unstable at 37°C which may explain why the low m.w. lymphotoxin has not been described previously.
Phytohemagglutinin-Induced Proliferation of Guinea Pig Thymus-Derived Lymphocytes
Authors: Ellner Jerrold J. , Lipsky Peter E. , Rosenthal Alan S.
Published: December 2022

Accessory cell participation in PHA-induced thymus-derived lymphocyte DNA synthesis encompasses two distinct functions. The first consists of maintenance of the functional integrity of resting lymphocytes, and the second involves the direct induction and/or support of T cell proliferation in response to this mitogen. Whereas the reducing agent 2-mercaptoethanol can support an Mø-depleted population of resting lymphocytes so that the latent biologic activity is maintained, it is not itself sufficient to allow the induction of lymphocyte proliferation in response to PHA. This latter function requires intact accessory cells.
Effects of Bcg on Lymphocyte Trapping
Authors: Zatz Marion M.
Published: December 2022

The effects of Bacillus-Calmette Guerin (BCG) administration upon lymphocyte traffic was studied in BALB/cJ mice. Intravenous or subcutaneous injection of BCG resulted in increased localization (trapping) of 51Cr-labeled syngeneic lymphocytes in the spleen or draining nodes. BCG-induced splenic trapping was biphasic, occurring 1 to 2 days after BCG injection, and reappearing 3 weeks later. Trapping in draining lymph nodes persisted and increased during the 24 days of study. These results are in contrast to those obtained with SRBC, where trapping was evident only within the first few days of antigen injection.

Additional studies revealed that prior injection of BCG influenced the ensuing magnitude of SRBC-induced trapping. Intravenous administration of BCG suppressed SRBC-induced splenic trapping; both intravenous and subcutaneous injection of BCG enhanced trapping to SRBC in the draining lymph nodes.

These studies suggest that one mechanism for potentiation of immune responses by BCG may operate via BCG's effects upon regulation of lymphocyte traffic to the lymphoid tissues.
Continuously Proliferating T Killer Cells Specific for H-2b Targets: Selection and Characterization
Authors: Dennert Gunther , De Rose Margaret
Published: December 2022

BALB/c (H-2d) thymus-derived lymphocytes sensitized to C57BL/6 (H-2b) alloantigens have been propagated in vitro for over 9 months. These T lymphocytes are specifically cytotoxic to H-2b target cells but are stimulated to proliferate by both H-2b and H-2k spleen cells. This indicates that for these selected cells the antigen requirements for cell proliferation are different from those for cell-mediated cytotoxicity. If not continuously stimulated with allogeneic spleen cells, the cytotoxic cultures fail to divide and rapidly lose their cytotoxic activity. Allogeneic erythrocytes do not stimulate cell proliferation in “quiescent” cell cultures and allogeneic tumor cells do so only in the presence of spleen cells. However, “quiescent” cell cultures display cytotoxicity in the presence of phytohemagglutinin A as do cell cultures which have lost their cytotoxic activity although they proliferate upon allogeneic stimulation. The significance of these findings is discussed.
The Merrit Alloantigenic System of Human B Lymphocytes: Evidence for Thirteen Possible Factors including one Six-Member Segregant Series
Authors: Walford Roy L. , Gossett Thomas , Troup G. M. , Gatti Richard A. , Mittal Kay K. , Robins A. , Ferrara G. B. , Zeller Edith
Published: December 2022

An analysis of the typing results of a 70-member chronic lymphatic leukemia B cell panel revealed evidence for 13 possible groups of the Merrit alloantigenic system. Six of these appeared possibly allelic and may represent a segregant series. The CLL panel was also fully typed for HLA and some degree of linkage dysequilibrium between Merrit and HLA seemed apparent from the data. Merrit antibodies can be absorbed out with selected surface membrane immunoglobulin (SMIG)-positive normal lymphocytes and less so or not at all with E rosette-forming T cells or Fc-positive SMIG-negative lymphocytes
Chemical, Physical and Electron Microscopy Studies of Fragments Produced by Limited Proteolysis of Human C1q by Pepsin
Authors: Reid K. B. M. , Doyle B. B. , Leonard K.
Published: December 2022

Digestion of human C1q with pepsin at pH 4.4 fragmented most of the non-collagen-like sequences in the molecule to small peptides whereas the regions of collagen-like sequence were left intact. The collagen-like fraction which accounted for 38%, by weight, of the digest contained 28% glycine, all the hydroxyproline and hydroxylysine, and was shown to be composed of two major fragments which could be separated on CM-cellulose in buffers containing 7.5 M urea. Human C1q is composed of equimolar amounts of the A and B chains, each of which has a m.w. of approximately 24,000. One of the pepsin fragments, of m.w. 20,000, was shown to be composed of the N-terminal 95 residues of the A chain linked via a single disulfide bond to the N-terminal 91 residues of the B chain. The second pepsin fragment of m.w. 18,200 was shown to be composed of a disulfide-linked dimer of the N-terminal 94 residues of the C chain.
Studies on the Mechanism of Action and Purification of Factor X, an Early Acting Component of the Properdin System
Authors: Smith M. C. , Steiner C. , Novak M. A. , Todd E. W. , Naff G. B.
Published: December 2022

We have identified a rabbit serum protein, tentatively designated as factor X, that interacts with zymosan (Z) to form a complex (ZX complex) which initiates activation of the properdin (P) system. Factor X was required to measure purified P preparations by the Z assay and reacted directly with Z in the absence of other known constituents of the P system.

We have shown that some mono and disaccharides such as glucose, fructose, ribose, galactose, and sucrose inhibited ZX complex formation whereas lactose did not. Once formed, ZX complexes were dissociated by ribose, sucrose, and lactose, but not by glucose or galactose. Concanavalin A prevented the uptake of Factor X by Z. Wheat germ and castor bean agglutinins, however, did not appreciably inhibit ZX complex formation.

Rabbit factor X was partially purified by fractionating serum euglobulin preparations on DEAE-cellulose columns. These preparations were further purified on dextran gels (Sephadex G-25).
The Role of Properdin Convertase and Properdin in C3NeF Activity: Requirement or Side Reaction?
Authors: Spitzer R. E. , Stitzel A. E. , Urmson J. R.
Published: December 2022

The sera of 10 patients with membranoproliferative glomerulonephritis (MPGN) demonstrate levels of properdin (7/10) and properdin convertase (7/10) which are 3 standard deviations below the normal mean. These paired levels are significantly related (p < 0.001) in these patients as they are in acute glomerulonephritis, lupus glomerulonephritis and immune complex nephritis. C3NeF was detected in 6 of 10 patients but failed to correlate significantly with either value. In vitro, when C3NeF is added to normal serum, properdin is altered in a time-dependent sequence manifested by a shift in electrophoretic mobility and a redution in the extent of diffusion by Mancini technique. These identical findings also occur on the addition of isolated and activated properdin convertase () to serum. Further, during the reaction of C3NeF, PC is altered in a manner parallel to that of properdin. Finally, sera depleted of PC by antibody adsorption will not support C3–C9 consumption on incubation with C3NeF.
Direct and Cooperative Mechanisms of Lymphocyte Triggering in Liquid and Solid Cultures
Authors: Kondracki Esteban , Milgrom Felix
Published: December 2022

The role of cell interactions in lymphocyte stimulation was analyzed by studying the kinetics of lymphocyte proliferation at different cell concentrations, and also by a lymphocyte microculture technique in solid medium. An absolute requirement for cell interactions was found in lymphocyte responses to concanavalin A, pokeweed mitogen, sodium periodate, purified protein derivative from Mycobacterium tuberculosis, and zinc chloride. No requirement for cell interactions was found in lymphocyte responses to calcium ionophore A23187. The existence of lymphocyte subpopulations with different requirements for cell interactions was observed in lymphocyte responses to phytohemagglutinin P, phytohemagglutinin HA 17, tetradecanoyl-phorbolacetate, antiserum to MOLT-4 lymphoblasts, antiserum to B411-4 lymphoblasts, antiserum to human embryo lung fibroblasts, and antiserum to HeLa cells infected with Herpes simplex virus.

Lymphocyte responses to phytohemagglutinin P were potentiated by incorporation into the solid cultures of red blood cells or their membrane preparations suggesting that membrane-membrane interactions, either directly, or through soluble mediators are likely to be the basis of cell cooperation in this system. In solid cultures, phytohemagglutinin P, phytohemagglutinin P plus red blood cells, phytohemagglutinin HA 17, tetradecanoyl-phorbol-acetate and antiserum to MOLT-4 lymphoblasts were found to stimulate mainly thymus-dependent lymphocytes, whereas antiserum to HeLa cells infected with Herpes simplex virus stimulated mainly non-thymus-dependent lymphocytes. Antiserum to B411-4 lymphoblasts stimulated both thymus-dependent and non-thymus dependent lymphocytes.
Inhibition of C1s̄-Induced Vascular Leakage in Guinea Pigs by Substituted Benzamidine and Pyridinium Compounds
Authors: Andrews Judith M. , Rosen Fred S. , Silverberg Shonni J. , Cory Michael , Schneeberger Eveline E. , Bing David H.
Published: December 2022

A variety of benzamidine and pyridinium compounds were examined for their ability to inhibit irreversibly C1s̄-induced vascular leakage in guinea pig skin. Vascular leakage was compared with esterolysis of N-Z-L-Tyr-Np and catalysis of EAC formation by C1s̄. Vascular leakage correlated significantly better with esterolytic activity than with EAC formation. The presence of a sulfonyl fluoride moiety in the compounds is important in the inhibition of C1s̄-induced vascular leakage.
Antibody-Mediated Suppression of Tumor Growth
Authors: Johnson Robert J. , Pasternack Gary R. , Drysdale B-E. , Shin Hyun S.
Published: December 2022

A radioimmunoassay was used to quantitate the number of tumor cell-bound IgG1 anti-tumor antibody molecules required to cause suppression of tumor growth in C3H mice. Radiolabeled anti-IgG1 was used to detect cell-bound IgG1 antibody. The assay was calibrated by using TNP-coupled tumor cells that had a known number of bound IgG1 anti DNP molecules. Between 70,000 and 130,000 IgG1 antitumor antibody molecules per tumor cell were sufficient to cause 50% suppression of tumor growth in mice inoculated with approximately 50,000 tumor cells.
Suppressor T Cells Arising in Mice Undergoing a Graft-vs-Host Response
Authors: Pickel Karin , Hoffmann Michael K.
Published: December 2022

We investigated the ability of mice to generate antibody-forming cells when undergoing a graft-vs-host reaction. (C57BL/6 × DBA/2)F1 mice (BDF1) injected with C57BL/6 spleen cells generated suppressor T cells which inhibit antibody synthesis by BDF1 spleen cells in vitro. These T cells arose from the donor inoculum. They differ from helper T cells in size and they act directly on antigen reactive B cells. The suppressor T cells were specifically directed against components of the H-2 region of the reciprocal parental strain (DBA/2 = H-2d) in the hybrid F1 mouse.
Isolation of an Fcγ-Binding Protein from the Cell Membrane of a Macrophage-Like Cell Line (P388D1) after Detergent Solubilization
Authors: Loube Susan R. , McNabb Trudy C. , Dorrington Keith J.
Published: December 2022

Lactoperoxidase-catalyzed iodination, NP-40 lysis, and subsequent affinity chromatography on IgG-Sepharose were used in an attempt to define some of the molecular properties of the Fc receptor of P388D1, a macrophage-like mouse tumor line. Radioiodinated material retained on columns of Sepharose coupled either to monomeric mouse IgG2a or monomeric human IgG1 appeared on SDS polyacrylamide gel electrophoresis to contain principally three labeled components, a major band of about 57,000 m.w., and two minor bands of 28,000 and 24,000 m.w. The mobilities of these components changed little on reduction, which suggested that they represented single polypeptide chains. An identical pattern was obtained with Sepharose-linked Fc fragments of human IgG1, but neither Fab fragments of IgG1 nor IgM appeared to bind these components. Since the specificity of binding to the immobilized proteins is the same as that observed in vivo, it is postulated that these proteins represent either all or some portion of the P388D1 Fc receptor.
Enumeration and Ultrastructure of C4-Producing Free Alveolar Cells from Guinea Pig Lung
Authors: Barber Thomas A. , Burkholder Peter M.
Published: December 2022

Free alveolar cells from guinea pig lung producing the fourth component of C (C4) were identified, enumerated, and characterized by using anti-C4 Fab-peroxidase conjugates in conjunction with transmission electron microscopy. The C4-producing cell population consisted of: 1) alveolar macrophages (AM); 2) less well differentiated phagocytes similar in morphology to exudate macrophages; and 3) weakly phagocytic secretory cells with numerous profiles of rough-surfaced endoplasmic reticulum (ER). Internal immunolabeling allowed the visualization of C4 in the ER, perinuclear space, and Golgi complex of producer cells and its release at cell surfaces; synthesis of C4 in vitro was sensitive to inhibitors both of protein synthesis and messenger RNA function. The percentage of free alveolar cells from normal animals competent for C4 production as indicated by cell surface immunolabeling was approximately 1% of the total cells obtained by lavage. Transnasal infection with Listeria monocytogenes, generation of a pulmonary granulomatous reaction by i.v. injection of heat-killed BCG, and aerosol infection of nonvaccinated animals with Mycobacterium tuberculosis each resulted in an increase in numbers of AM and exudate macrophage-like free alveolar cells competent for C4-production.
Polyclonal Activation of CR+ and CR- B Lymphocytes: The Kinetics of Initiation of DNA and Immunoglobulin Synthesis by Lipopolysaccharide
Authors: Hämmerling Ulrich , Chua Ramon , Hoffmann Michael K.
Published: December 2022

B cells that carry the complement receptor (CR+) were separated from B cells that lack the complement receptor (CR-) by velocity sedimentation or by passage through C-coated Sephadex columns. The kinetics of responses to bacterial lipopolysaccharide (LPS) in both B cell subpopulations were determined in three assay procedures: 1) incorporation of radioactive thymidine into DNA; 2) incorporation of radioactive leucine into immunoglobulin; 3) enumeration of cells forming polyclonal antibody to the 2,4,6-trinitrophenyl hapten.

Although both subpopulations of B cells responded to LPS, they differed in the time course. CR- B cells responded with a delay of approximately 24 hr as compared with the response of CR+ B cells. The implications to the ontogenetic status of CR+ and CR- B subpopulations are discussed.
Refractoriness to Migration Inhibitory Factor of Macrophages of LPS Nonresponder Mouse Strains
Authors: Tagliabue Aldo , McCoy James L. , Herberman Ronald B.
Published: December 2022

The responsiveness to macrophage migration inhibitory factor (MIF) of peritoneal exudate cells (PEC) from the LPS unresponsive C3H/HeJ and C57BL/10ScCR mice was assessed by the indirect agarose microdroplet macrophage migration inhibition assay. No migration inhibition with PEC from C3H/HeJ nor C57BL/10ScCR mice was detected, whereas PEC from both C3H/HeN and C57BL/10Sn mice were significantly inhibited by even a 1/32 dilution of MIF-containing supernatants. Responsiveness to MIF of C3H/HeJ PEC could, however, be induced. In vivo inoculations of Mycobacterium bovis, strain BCG, 7 days before in vitro assay rendered C3H/HeJ PEC responsive to MIF. The lack of responsiveness to MIF by C3H/HeJ PEC appeared related to some form of suppression, since a mixture of PEC from C3H/HeN mice with 10 to 15% PEC from C3H/HeJ mice resulted in undetectable migration inhibition at any MIF dilution. In contrast to the usual lack of responsiveness of their macrophage to MIF, C3H/HeJ mice were able to produce MIF in response to PPD as well as their counterpart C3H/HeN mice after BCG sensitization. These results demonstrate that macrophages from C3H/HeJ and C57BL/10ScCR mice are unable to be inhibited in their in vitro migration of MIF (possibly being directly or indirectly influenced by a suppressor cell), whereas lymphoid cells from at least one of these strains, the C3H/HeJ mice, can produce MIF in response to antigenic stimulation.
Nonimmunologic Complement Activation in Normal Human Serum Induced by Radiographic Contrast Media
Authors: Kolb William P. , Lang Joseph H. , Lasser Elliott C.
Published: December 2022

Two different radiographic contrast media (RCM), iothalamate and iodipamide, induced the activation of several complement (C) components in normal, genetically C2-deficient and agammaglobulinemic human sera in vitro. This activation was dose dependent and demonstrable by a reduction in whole C as well as C4, C2, C3, and C5 hemolytic activities. C6, C8, and C9 hemolytic activities were unaffected. Concommitant with the loss of C3 hemolytic activity was the appearance of C3 proteolytic cleavage products that were identified by immunoelectrophoresis. Both the loss of C3 hemolytic activity and the production of C3 fragments occurred in the presence of 10 mM EDTA, indicating RCM-induced C3 cleavage occurred without participation of the multicomponent C3/C5 convertases of either the classical or alternative C pathways. Furthermore, loss of C3 hemolytic activity was not due to the direct alteration of the C3 molecule by RCM because purified C3 was unaffected upon incubation with RCM at a concentration that induced 80% reduction in the C3 hemolytic activity in normal human serum.

Serum samples obtained from 40 patients, before and 30 min after undergoing i.v. pyelography, revealed no significant change in total hemolytic C activity; 34 patients received sodium and methylglucamine diatrizoate and six received sodium iothalamate. Hemolytic C3 levels were also determined for the six patients before and 30 min after administration of sodium iothalamate and no significant change in activity was detectable.
Cytotoxic T Lymphocyte Sequential Killing of Immobilized Allogeneic Tumor Target Cells Measured by Time-Lapse Microcinematography
Authors: Rothstein Thomas L. , Mage Michael , Jones Gary , McHugh Louise L.
Published: December 2022

Sequential killing of allogeneic target cells by immune cytotoxic T lymphocytes (CTL) was directly observed by time-lapse microcinematography. Target cells (EL4 lymphoma cells from C56BL/6 mice), coated with Fab fragments of goat antibody to EL4, were immobilized by binding to the floor of a polystyrene tissue culture flask that had been precoated with specifically purified antigoat Fab. On adding immune BALB/c spleen CTL to such target cell monolayers it could be verified by direct observation that individual CTL could sequentially kill several target cells, that the CTL usually separated from the target cell before target cell death, that not all contacted target cells were killed, and that duration of contact was variable and not correlated with subsequent target cell death.
Regulation of Self Tolerance in Experimental Allergic Encephalomyelitis
Authors: Welch Andrew M. , Swierkosz James E. , Swanborg Robert H.
Published: December 2022

We studied the in vitro proliferative responses of lymph node cells (LNC) and spleen cells (SpC) from Lewis rats rendered unresponsive to experimental allergic encephalomyelitis (EAE) by pretreatment with guinea pig myelin basic protein (BP) in incomplete Freund's adjuvant (IFA). These LNC and SpC have suppressor function, since they protect cell recipients against EAE. LNC from unresponsive rats responded to Con A and PHA, indicating that T cell function was not nonspecifically impaired. In contrast, SpC responses were significantly suppressed. Mitogenic responses were restored by removing glass wool-adherent SpC. Moreover, when cocultured, SpC inhibited the LNC responses to Con A and PHA. Suppression of proliferative responses was again observed when normal peritoneal cells were added back to nonadherent SpC, suggesting that the suppressor SpC are macrophages. LNC and nonadherent SpC from unresponsive rats also responded in vitro to intact heterologous (guinea pig) BP, provided the rats had also been challenged with BP in complete Freund's adjuvant (CFA).

In contrast to the EAE suppressor LNC, which are nonadherent T lymphocytes, removal of adherent SpC abrogated adoptive transfer of unresponsiveness to EAE. These studies suggest that both antigen-specific suppressor T lymphocytes and nonspecific splenic adherent suppressor cells (presumably macrophages) may play a role in the maintenance of immunologic self-tolerance.
Null Cells in the Mouse Possess Membrane Immunoglobulins
Authors: Warr Gregory W. , Lee John C. , Marchalonis John J.
Published: December 2022

Null cells lacking typical T and B cell surface markers were isolated from the spleens of congenitally athymic mice by using either nylon wool or Sepharose macrobeads conjugated with rabbit antibody to mouse IgM to remove B lymphocytes. Although these null cells were negative for surface immunoglobulin by the criterion of immunofluorescence by using rabbit antisera to mouse heavy or light immunoglobulin chains, surface immunoglobulins were readily demonstrable by two alternative and independent techniques. First, by using chicken antibody to the (Fab′)2 fragment of mouse IgG, nearly all null cells were positive for immunofluorescence. Second, immunoglobulins could be detected in lysates of null cells radioiodinated by the lactoperoxidase-catalyzed reaction. Analysis of the surface immunoglobulins of null cells by radioimmunoassay and polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate suggested that they are 1) qualitatively similar to those of B cells and 2) present in amounts between 10 and 30% of those of B cells.
Identification and Cytotoxic Reactivity of Inflammatory Cells Recovered from Progressing or Regressing Syngeneic UV-Induced Murine Tumors
Authors: Lill Patsy H. , William Fortner G.
Published: December 2022

Most tumors induced in C3H mice by ultraviolet (UV) light are immunologically rejected by normal syngeneic recipients, but will grow progressively in immunosuppressed mice and in mice treated with UV light. In this study we compared the composition and cytotoxic activity of the inflammatory cell infiltrate from tumors transplanted into syngeneic UV-irradiated or unirradiated mice. Tumor fragments were implanted in either normal (regressor) or UV-treated (progressor) mice, and removed on various days after implantation and mechanically dissociated. The cells were examined by immuno-fluorescence for θ and immunoglobulin markers, stained for morphologic examination, and tested for cytotoxicity against the tumor. No significant differences were noted in numbers of macrophages, granulocytes, or B cells recovered from progressing or regressing tumors on day 6, the time of greatest activity. However, the numbers of T cells recovered from tumor fragments implanted in normal mice was approximately 3-fold that recovered from tumor fragments implanted in UV-treated mice. Lymphocytes recovered from regressing tumor fragments were specifically cytotoxic for that tumor in a microcytotoxicity test; those from progressing tumor fragments were not cytotoxic.
Nonspecific Activation of Murine Lymphocytes
Authors: Goodman Michael G. , Fidler John M. , Weigle William O.
Published: December 2022

The lymphocyte subpopulations activated by 2-ME in C3H/St spleen cell cultures were characterized by means of kinetic and cell separation techniques. The current experiments were designed to determine whether 2-ME potentiates the maintenance of an activated state in cells that had been triggered by environmental stimuli encountered by the intact animal, or whether this thiol compound is able to transform typical resting lymphocytes to an activated state. Kinetic experiments demonstrated that the high rate of nucleotide turnover seen initially in cultured cells was not directly accentuated. Rather, this turnover was found to subside normally before 2-ME-initiated activation of lymphocyte [3H] TdR uptake began. Studies in which cells were separated according to sedimentation velocity showed clearly that 2-ME responsiveness is a quality of small lymphocytes (s = 3.9 mm/hr) and not of large, activated cells. Moreover, when cells were separated on the basis of buoyant density, reactivity to this thiol compound was found to reside within cells of high density, strongly supporting the thesis that 2-ME activates a resting lymphocyte subpopulation and does not simply maintain an activated state acquired in vivo. The influence of antigenic milieu was evaluated directly in mice reared under germfree, specific pathogen-free, and conventional conditions. Lack or limitation of environmental antigenic exposure resulted in a significantly larger DNA synthetic response to 2-ME. This pattern was presumed to reflect a greater preponderance of unstimulated lymphocytes in the spleens of germfree and SPF mice.
The Influence of α-Fetoprotein on Moloney Sarcoma Virus Oncogenesis: Evidence for Generation of Antigen Nonspecific Suppressor T Cells
Authors: Gershwin M. Eric , Castles James J. , Ahmed Aftab , Makishima Ronald
Published: December 2022

The influence of murine α-fetoprotein (AFP) on the threshold dose, mean tumor size, regression time, and number of progressors for Moloney sarcoma virus-(MSV) induced tumors in BALB/c mice was studied. AFP-treated mice developed larger tumors, required a longer period for regression, and had a significantly higher mortality. Furthermore, AFP, but not murine albumin or transferrin, allowed growth of tumors when normally subthreshold doses of virus were injected. These effects of AFP could be removed by pretreatment and precipitation of AFP by anti-AFP; on the other hand, they were not altered by dialysis and were noted at sera levels of approximately 170 to 440 µg/ml. The mechanism of this heightened susceptibility in AFP treated mice to MSV appears to be the generation of a cortisone, radiation sensitive splenic, and not lymph node, suppressor T cell. Indeed, the suppression produced by AFP was antigen nonspecific and could be demonstrated in unmanipulated mice after transfer of spleen cells from AFP-treated donors. This transfer of suppression was noted in IgG and IgA secondary responses to sheep red blood cells as well as by quantitating the response of T cell mitogens on spleen cells. These data suggest that AFP, in pharmacologic doses, either alters a subpopulation of spleen cells to exhibit suppressive function or else that AFP is involved in differential homing patterns of lymphoid cells. The relationships of AFP and immune competence appear broad and suggest the need for comparative studies with other oncofetal antigens.
Recognition by Cytolytic T and K Cells: Identification in Both Systems of a Divalent Cation-Independent, Cytochalasin A-Sensitive Step
Authors: MacLennan Ian C. M. , Golstein Pierre
Published: December 2022

Most studies of recognition events in cell-mediated cytolysis have analyzed the conditions that are required to establish mechanically stable binding between effector cells and target cells. This approach has shown that cytolytic T cells require a temperature above 20°C and free divalent cations to bind specifically to target cells. K cells on the other hand were shown to bind to IgG on target cells in the absence of divalent cations at 4°C. In this paper we shown that both K cell- and T cell-mediated cytolysis are inhibited by cytochalasin A, if this drug is added to cultures before the effector and target cells have interacted. However, the drug does not block the calcium-dependent lethal hit stage of cytolysis in either system. We then show that in both T cell- and K cell-mediated cytolysis, the cytochalasin A-inhibitable stage can proceed in the absence of divalent cations. We conclude from these data that: 1) both K cell- and T cell-mediated cytolysis have a divalent cation-independent stage, which is inhibitable by cytochalasin A; 2) firm mechanical binding of T effectors to target cells requires divalent cations. It is uncertain if the latter is necessary for cytolysis or whether K cells also have to go through a divalent cation phase before the lethal hit stage. The implications of these findings are discussed, especially as to the possibility that strong binding may not be necessary for T cell-mediated cytolysis.
Synergistic Cytotoxicity
Authors: Muchmore A. V. , Decker J. M. , Blaese R. M.
Published: December 2022

The mechanism by which non-immune mononuclear cells recognize invading foreign material and are activated for cytotoxic attack was studied in a model system employing human mononuclear cells, fresh plasma, and 51Cr-labeled xenogeneic target erythrocytes. In these experiments, fresh antibody-depleted plasma or mononuclear leukocytes alone were poorly cytotoxic to xenogeneic erythrocytes. However, these target cells were rapidly lysed when both fresh antibody-depleted plasma and mononuclear cells were present in the assay. The plasma factor could not be removed by extensive absorption with the target cells, was present in plasma from hypogammaglobulinemic patients, was heat labile, and was sensitive to incubation with zymosan and cobra venom factor. The “antigen” specificity of this reaction was directed by the serum factor inasmuch as target cells autologous to the effector cells could be killed in the presence of antibody-depleted xenogeneic plasma, but not autologous plasma. These data suggest that an important mechanism for the recognition of “foreignness” by non-immune mononuclear cells is via interaction with a plasma component, possibly a factor related to serum complement.
Immunoadjuvant Treatment of Primary Grafted and Spontaneous AKR-Leukemia
Authors: Olsson Lennart , Ebbesen Peter
Published: December 2022

Young AKR mice grafted i.v. with 101 or 103 cells from spontaneous AKR thymomas were treated with repeated i.v. injections of BCG or subcutaneous injections of irradiated AKR thymoma cells. BCG often cured mice from graft leukemia, whereas the effect of irradiated thymoma cells was less effective. Mice that did not develop graft-leukemia after graft of 101 leukemia cells and BCG treatment showed a spontaneous leukemia in 30% of the cases later. Ninety percent of nongrafted mice developed spontaneous leukemia whether BCG-treated or not. General immune reactivity as assessed in individual mice by T and B lymphocyte mitogen tests as well as the hemolytic plaque-forming cell assay had no clear correlation to the effects of immune adjuvants in respect to survival. In contrast, occurrence of self-directed immune reactions were clearly correlated to survival and cure of grafted and BCG-treated mice as revealed by assays both in vitro and in vivo. However, 13 to 25% of the mice apparently cured of leukemia developed a wasting-like syndrome that sometimes terminated in death. The implications of self-directed immune reactions as mediators of the anti-neoplastic effects of immunoadjuvants are discussed.
The Genetics of PLT Response
Authors: DeWolf William C. , Carroll Peter G. , Yunis Edmond J.
Published: December 2022

The primed lymphocyte test (PLT) has been used as a method to identify HLA-D or HLA-D-related (HLA-DRw) specificities; however, the assignment of HLA-D or HLA-DRw specificities by this procedure is often nondiscriminatory with false positive and false negative results. We have investigated three approaches to improve PLT discrimination: 1) restimulation of primed cells after various times in primary culture; 2) restimulation using a small number of responding PLT cells per well, and 3) nylon wool purification of responder cells before initial priming. The results show that when PLT cells are restimulated and the response is plotted on a log/log scale in relation to a decreasing number of responding cells per well, a characteristic “linear” relationship is seen when less than 1.0 × 104 PLT cells are used (stimulators = 5.0 × 104 cells per well). When greater than 1.0 × 104 PLT cells per well are used, a linear response is seen only when the priming culture is prolonged to 17 days or more. It is only when the PLT cell is in this “linear phase” of response that the responses to various stimulating cells can be compared to yield the most discriminatory % relative response (RR). Finally, nylon wool treatment of responder cells before sensitization improves subsequent discrimination in PLT when large numbers of primed responders per well are used. Thus, optimum discrimination in PLT is obtained: 1) by using a low number (less than 1.0 × 104) responding PLT cells per well, 2) by prolonging the priming MLC to at least 17 days, and 3) by pretreating the responder cells with nylon wool. These experiments suggest that during allogeneic stimulation in MLC, there is activation of a population of lymphocytes that react with HLA-D or HLA-DRw determinants and that other nonactivated or unprimed cells within the PLT population may alter a discriminatory response; such cells must be functionally decreased by these procedures for optimum results.
A Monoclonal IgG4 (λ) with Factor V Inhibitory Activity
Authors: Hurtubise Paul E. , Coots Macie C. , Jacob Donald J. , Muhleman Albert F. , Glueck Helen I.
Published: December 2022

A monoclonal IgG4 (λ) with inhibitory activity to human coagulation factor V was isolated from the serum of a patient with a fatal hemorrhagic diathesis by using a combination of DE-52 ion exchange chromatography and isoelectric focusing techniques. Using the criteria for defining a monoclonal immunoglobulin of restricted mobility on protein electrophoresis, immunoelectrophoresis, and isoelectric focusing, as well as neutralization with class, subclass, and light chain type antisera, we are the first to demonstrate a factor V inhibitor as a monoclonal IgG4 (λ) detectable in serum or plasma.
Release of Immature Cells from the Thymus during Solid Tumor Growth: Identification by Assay of TdT Activity
Authors: Small Myra , Lasser-Weiss Meira , Daniel Violet
Published: December 2022

In vivo anti-tumor activity of spleen cells from C3H/eb mice bearing a syngeneic fibrosarcoma was shown previously to decline to an undetectable level and be replaced by tumor-enhancing activity as tumor growth proceeds. In the light of our findings that thymocytes in the early stages of thymic processing can bring about tumor enhancement, we postulated that premature release of thymocytes and their accumulation in the spleen might account for the loss of the anti-tumor response. In the present experiments an injection of thymocytes did in fact cancel the anti-tumor response of reactive splenocytes from tumor-bearing mice. In order to determine whether premature thymocyte release occurs naturally in the tumor-bearing animals, we assayed activity of the enzyme TdT (as a marker for thymus cells) in the spleens of these mice during progressive tumor growth. Cells with TdT activity were clearly evident in the spleens of the tumor-bearing animals, were derived from the thymus, and accumulated in parallel to the loss of antitumor reactivity.
Cytotoxic Macrophages Induced in Hamsters by Vaccinia Virus: Selective Cytotoxicity for Virus-Infected Targets by Macrophages Collected Late After Immunization
Authors: Chapes Stephen K. , Tompkins Wayne A. F.
Published: December 2022

Peritoneal macrophages from LHC/Lak hamsters infected with vaccinia virus were tested for cytotoxicity against uninfected and vaccinia or herpes simplex virus-infected semiallogeneic target cells (BHK and PARA-7) in a 51Cr cytotoxicity assay. Macrophages were tested between 3 and 34 days after immunization of hamsters with 1 × 107 PFU of vaccinia virus. Peak cytotoxicity against BHK infected with vaccinia virus (BHKV) or herpes simplex virus (BHKH) occurred at 5 days after immunization and declined to baseline by 34 days. At no time after immunization was there evidence of preferential killing of the specific BHKV targets. However, macrophages tested 3 to 5 days after immunization differed from those tested later, both in their kinetics of cytotoxicity and the type of target cell recognized. Thus, 5-day immune macrophages were cytotoxic for uninfected BHK and PARA-7 hamster tumor cells as well as BHKV and BHKH. In contrast, 9-day immune macrophages were cytotoxic for BHKV and BHKH, but not uninfected BHK or PARA-7. Also, in vitro assay kinetics showed that early 5-day immune macrophages were more rapidly cytotoxic for all target cells than late 9-day immune macrophages. This study also showed that nonadherent peritoneal exudate cells (NPEC) were induced by vaccinia immunization that were nonspecific in their cytotoxicity. Although both adherent macrophages and NPEC showed preferential lysis of virus-infected targets, the cytotoxicity mediated by NPEC was abrogated by treatment with NH4Cl, whereas macrophage cytotoxicity was not affected by such treatment. In summary, this study demonstrates that with time after immunization with vaccinia virus peritoneal macrophages differ markedly in their cytotoxicity characteristics with respect to target cell recognition and rate of target cell killing. Possible models to explain preferential cytotoxicity against virus-infected targets by late vaccinia immune macrophages are discussed.
Surface Properties and in vitro Functional Capacities of Human T Lymphocyte Subsets Separated by Linear Density Gradients
Authors: Huber Christoph , Rodt Hans , Thierfelder Stefan , Braunsteiner Herbert
Published: December 2022

Human lymphocytes derived from various central and peripheral sources were separated on linear density gradients (LDG). Cells from individual density fractions were tested in parallel for: the capacity to form nonimmune rosettes with neuraminidase-treated SRBC, the number of surface-associated HTLA, and in vitro proliferative responses to mitogenic lectins and alloantigens.

Heterogeneous density distribution profiles were obtained for all sources of human T cells and revealed an organ specificity. The various T cell density classes obtained from identical organs as well as the identical density classes of different sources revealed to some extent differences in their surface marker patterns and/or their in vitro reactivities. On the basis of the combined techniques at least two major subsets among thymocytes were identified that differed in both surface properties and functional capacities. Density classes of T cells from all peripheral sources were distinguished from thymocytes by a homogeneous lowered HTLA expression. Whereas clear-cut differences in the in vitro functional capacity were observed between the two thymocyte subsets, less striking but still significant differences were found to exist among the various density classes of peripheral T cells.
Modulation of Natural Cytotoxicity by Alloantibodies
Authors: Saxena Rajiv K. , Adler William H.
Published: December 2022

Natural killer activity of spleen cells obtained from different strains of mice against the human myeloid leukemia cell line, K562, and two mouse cell lines P815 and L1210 was measured by using the 4-hr chromium release assay. The level of cytotoxic activity of spleen cells against the K562 target was usually less than 4% lysis. However, treatment of the spleen cells with a specific anti-H-2 antiserum resulted in a dose-dependent augmentation of the degree of lysis of K562 cells. The augmentation of cytotoxic activity could be obtained by pretreatment of the spleen cells with antisera or by directly adding the antisera to the cytotox-incubation medium. Anti-thy-1 and anti-immunoglobulin antisera had no enhancing effect under similar conditions. The specific alloantisera-treated spleen cells did not show any increase in cytotoxicity against P815 and L1210 target cells. Spleen cells responsible for the alloantiserum-mediated augmentation of cytotoxicity against K562 cells appear to be different from T or B cells as indicated by their resistance to anti-thy-1 and complement treatment and lack of adherence to nylon wool columns.
Demonstration of Phosphorylcholine-Specific IgE B Cells in CAB/N Mice
Authors: Kishimoto Tadamitsu , Shigemoto Shozo , Watanabe Takeshi , Yamamura Yuichi
Published: December 2022

CBA/N mice, which did not make anti-PC IgM or IgG antibody against PC-conjugated T-dependent or T-independent antigens, produced IgE antibody to PC-determinant when they were immunized with PC-KLH. PC-specificity of IgE antibody produced in CBA/N mice was determined by inhibition of PCA reaction with free PC-hapten or C-polysaccharide or by absorption of reaginic activity in the serum with C-polysaccharide. The presence of T15 idiotype on anti-PC IgE antibody produced in CBA/N × BALB/c F1 males also showed that anti-PC IgE antibody in defective mice was PC-specific. The results suggest that PC-specific Bε cells may belong to a subpopulation distinct from PC-specific precursors for IgM and IgG responses.
Markers of Human T Cell Subsets Identified by Alloantisera
Authors: Ferrara Giovanni Battista , Strelkauskas Anthony J. , Longo Anna , McDowell Joan , Yunis Edmond J. , Schlossman Stuart F.
Published: December 2022

Analysis by the indirect fluorescence test followed by fluorescence-activated cell sorter (FACS) analysis has shown that antisera recognizing subsets of human T lymphocytes can be produced by planned immunizations involving HLA-A and HLA-B compatible donors. The reactivity of these antisera against some individuals of a population but not others shows that they recognize a polymorphic cell surface component. The reactive subpopulation largely overlaps with the JRA+ subset, which was previously shown to possess regulatory properties in functional assays. The specificity of the antisera for a T cell subset and the unrelatedness of the anti-B cell activity of the same antisera has been confirmed by two-color fluorescence tests.
The Immune Response of Lewis Rats to Peptide 68–88 of Guinea Pig Myelin Basic Protein
Authors: Fritz Robert B. , Chou Frank C.-H. , Jen Chou C.-H. , Kibler Robert F.
Published: December 2022

A series of peptides, produced from peptide 68–88 by selective enzyme cleavage, were used to define the amino acid sequences reactive with Lewis rat anti-peptide 68–88 antibody. One hundred per cent of 38 rats immunized with peptide 68–88 recognized a B cell determinant residing in the region of residues 80–88. The three carboxyterminal amino acids, residues 86–88, appeared to confer conformation on this determinant. In roughly two–thirds of the rats, the reactivity with this major B cell determinant was equal to the reactivity with the immunizing peptide. In these rats the anti-peptide 68–88 antibodies were of the IgG2 class. A third of the rats recognized an additional determinant(s) and in these the predominant class of antibody was IgG1.
Involvement of Host Fc Receptors in Antibody-Mediated Cutaneous Basophil Hypersensitivity Reactions
Authors: Graziano Frank M. , Askenase Philip W.
Published: December 2022

Cutaneous basophil hypersensitivity (CBH) reactions are heterogeneous delayed time course basophil-rich responses that can be mediated by either T cells, B cells, or serum antibodies. The current study examined the mechanism by which antibodies mediate CBH in guinea pigs. Fc competition experiments were constructed by passively transferring mixtures of anti-KLH serum and normal heterologous γ-globulins. It was found that rabbit IgG and its isolated and purified Fc fragment [but not the (Fab′)2 fragment] inhibited the ability of guinea pig immune serum to transfer CBH. Concurrent inhibition of transferred KLH-specific CBH and systemic passive cutaneous anaphylaxis (PCA) reactions by rabbit IgG or its Fc fragment, and not by sheep or bovine γ-globulins, indicated that Fc receptors on cutaneous mast cells were probably involved in both CBH and PCA.

It was also found that the basophil aspect of delayed cutaneous responses elicited by PHA was inhibited by Fc competition maneuvers. This could mean that some forms of apparently T cell-mediated CBH may be T cell dependent, but via secretion of molecules that bind to Fc receptors, as seems required in antibody-mediated CBH.