bluuebunny/crossref_metadata_2025 huggingface dataset:
The proliferative capacity of human T lymphocyte subpopulations in a continuous culture system
Authors: Pang Gerald T. M. , Booth Roger J.
Published: February 2007

The continuous growth of subpopulations of human T lymphocytes was investigated using a culture system containing conditioning factors from mitogen‐stimulated lymphocytes. Cultures of purified peripheral blood lymphocytes rapidly became enriched for T lymphocytes, detected as sheep erythrocyte rosette‐forming cells, and could be maintained for up to a month in an actively growing state. Subpopulations of T lymphocytes bearing Fc receptors for IgM (T.M) or IgG(T.G) could not be detected in these cultures unless the cells were first washed and cultured for 3 days in medium devoid of conditioning factors. During this second culture step, many of the cells reverted from a large, blast‐like state to a small lymphocyte morphology and a proportion of the T lymphocytes re‐expressed Fc receptors. The proportions of T.M and T.G lymphocytes so detected remained constant throughout the continuous culture period indicating that the system permitted the proliferation of all T lymphocytes. Fractionation studies supported this conclusion by demonstrating that purified T, T.M and T.G but not B lymphocyte populations proliferated when cultured in the presence of conditioned medium. The majority of cells in cultures of purified T.M lymphocytes re‐expressed IgM Fc receptors following reculture in unconditioned medium. However, the re‐expression of IgG Fc receptors by cultured T.G lymphocytes could not be achieved.
Antigen‐specific suppressor factors produced by T cell hybridomas for delayed‐type hypersensitivity
Authors: Hewitt Jane , Liew Foo Y.
Published: February 2007

This study describes the generation of T hybridoma lines which secrete factors specifically suppressing delayed‐type hypersensitivity (DTH) to sheep red blood cells (SRBC). AKR strain‐derived T lymphoma BW 5147 cells were fused with spleen cells from mice primed with SRBC and containing antigen‐specific T suppressor cells for DTH. Supernatants from the derived hybridomas were tested for suppression of either expression or induction of DTH to SRBC. Six lines produced specific suppressor activity for the expression of DTH; 4 lines produced suppressor activity for the induction of DTH of which only one line was antigen‐specific. These lines were passaged in normal AKR mice, and the serum obtained had activity up to 10−4dilution. The factor was effective across the H‐2 barrier.
Lipopolysaccharide and lipid A‐induced human blood lymphocyte activation as detected by a protein A plaque assay
Authors: Smith C. I. Edvard , Hammarström Lennart , Bird A. Graham , Kunori Takao , Gustafsson Björn , Holme Tord
Published: February 2007

Various purified cell wall lipopolysaccharides (LPS) from gram‐negative bacteria and derivatives of these LPS were tested for their stimulatory capacity for human peripheral blood cells. Immunoglobulin (Ig) production was tested by an indirect plaque‐forming cell assay using Staphylococcus aureus protein A‐coupled erythrocytes and specific anti‐Ig as developing serum. This method allows the detection of the majority of cells secreting Ig of a single class, and the number of plaque‐forming cells detected are approximately 100–1000 times the amount obtained using normal sheep red cells as targets. LPS containing the O antigen‐specific chain, as well as mutant products only containing lipid A and ketodeoxyoctonate trisaccharide, could induce cell division and antibody synthesis. The polypeptide antibiotic polymyxin B was found to inhibit LPS‐induced activation. Furthermore, purified lipid A, complexed with bovine serum albumin, was also found to activate human peripheral blood B cells. These findings demonstrate that human peripheral blood lymphocytes can be activated by LPS and also indicate that lipid A is the active part of these molecules.
Monoclonal antibodies in cell‐mediated cytotoxicity against human melanoma and colorectal carcinoma
Authors: Herlyn Dorothee , Herlyn Meenhard , Steplewski Zenon , Koprowski Hilary
Published: February 2007

Hybridoma‐derived monoclonal anti‐melanoma antibodies and anti‐colorectal carcinoma antibodies were found to mediate in vitro antibody‐dependent cell‐mediated cytotoxicity (ADCC) reactions against melanoma and colorectal carcinoma cells, respectively.

The antigen(s) detected in ADCC on melanoma cells maintained for more than one hundred passages in tissue culture were also found on two recently established melanoma cell lines. These antigens were not detected on skin fibroblasts of the same patients from whom the melanomas were obtained.

The ADCC reactivities of anti‐melanoma and anti‐colorectal carcinoma antibodies were found to be specific for melanoma cells and colorectal carcinoma cells, respectively.
Generation of virus‐specific cytotoxic T cells in vitro I. Induction conditions of primary and secondary Sendai virus‐specific cytotoxic T cells
Authors: Koszinowski Ulrich H. , Simon Markus M.
Published: February 2007

H‐2‐restricted cytotoxic T cells specific for Sendai virus were generated in vitro in a primary response from normal mouse lymphocytes cultured in the presence of infective as well as inactivated Sendai virus. Antigen‐presenting cells of different origin, including T cells, were found to be effective stimulators. Antibodies to Sendai virus were shown to inhibit the activation of specific precursor killer cells when added to cultures before, but not after, the addition of viral antigen. Data obtained by Lyt phenotyping, revealed that precursor killer cells specific for Sendai virus reside in the Lyt‐2,3+ T cell population and that Lyt‐l,2,3+ T cells are not required for the generation of cytotoxic lymphocytes. Different activation kinetics were demonstrated for primary and secondary antiviral cytotoxic responses, and the analysis of the proliferation and stimulation requirements suggests qualitative differences.
H‐2 restriction for lymphocyte homing into lymph nodes
Authors: Degos Laurent , Pla Marika , Colombani Jacques M.
Published: February 2007

Migratory patterns into lymph nodes of labeled spleen lymphocytes, injected intravenously, showed considerable differences according to the genetic identity or disparity of donor and recipient mice. The use of mice carrying different H‐2 haplo‐types in the same B 10 background, or, conversely, the same H‐2b haplotype in different backgrounds, showed that no homing was observed in the case of complete disparity at the H‐2 complex, contrasting with optimal homing in the case of H‐2 identity. Homing was not influenced by disparity at the genetic background.

Irradiation of recipient mice did not change the results and suggested that no immunological host‐vs.‐graft reaction was involved in the H‐2 restriction observed. The use of donor‐recipient pairs carrying recombinant H‐2 haplotypes showed that a single H‐2 K and/or H‐2 D identity allowed almost optimal homing. The results indirectly suggested that an immunological graft‐vs. ‐host reaction was not involved in the phenomenon but rather a positive recognition of H‐2K or H‐2D identity between donor and recipient cells.
Production of interferon in the murine mixed lymphocyte culture. II. Interferon production is a T cell‐dependent function, independent of proliferation
Authors: Kirchner Holger , Zawatzky Rainer , Engler Helmut , Schirrmacher Volker , Becker Hans , von Wussow Peter
Published: February 2007

Interferon production occurs after two days of culture in murine mixed lymphocyte cultures (MLC). This was demonstrated in various combinations of mouse spleen cells differing at the major histocompatibility (H‐2) locus. Interferon production could be demonstrated in one‐way MLC when F1/parent combinations were used and in reactions in which one partner was treated by puromycin. After treatment of both cell populations with mitomycin C, interferon production occurred in the absence of lymphoproliferation. Interferon production in response to alloantigen did not occur in spleen cell cultures of nude mice and in cultures treated by anti‐Θ antiserum plus complement indicating that interferon production is a T cell‐dependent function.
The idiotypy of the MOPC 173 (IgG2a) mouse myeloma protein: Characterization of syngeneic, allogeneic and xenogeneic anti‐idiotypic antibodies. Contribution of the H and L chains to the idiotypic determinants
Authors: Schiff Claudine , Boyer Claude , Milili Michèle , Fougereau Michel
Published: February 2007

Anti‐idiotypic antibodies were raised against the mouse (γ2a, κ) myeloma protein MOPC 173 in syngeneic, allogeneic and xenogeneic conditions. Syngeneic immunization resulted in individual hemagglutination titers ranging from 1:40 to 1:160000 with about 30% of nonresponders, whereas upon allogeneic immunization (in A/J mice) almost all animals responded, with titers ranging from 1:1250 to 1:320000. Anti‐idiotypic antisera were also obtained in the rabbit.

Inhibition of a radioimmunoassay using 125I‐labeled MOPC 173 Fab fragments as the reference antigen indicated that the native Ig and the Fab competed on a similar molar basis. No inhibition was observed with the Fc or with other mouse monoclonal Ig, of the same or different classes or subclasses. Standard BALB/c serum did not inhibit either.

Whatever the anti‐idiotypic antiserum used, idiotypic determinants were not found on isolated heavy (H) and light (L) chains provided they were carefully purified. Reoccurrence of idiotypic determinants was obtained in high yield whenever Ig molecules were reformed from the homologous H and L chains, whereas no potentiation of the low inhibition of isolated chains was observed when hybrid molecules were formed using heterologous complementary chains isolated from the MOPC 21A (γl, ϰ) or the LPC 1 (γ2a, ϰ) protein. These results suggest that MOPC 173 idiotypic determinants rely on the specific H‐L interaction and that they must be largely conformational in nature.

Heterogeneity of the anti‐idiotypic antibodies was analyzed by isoelectric focusing. A major pattern was observed for the A/J antisera, whereas, in the case of BALB/c, expression of a few clones, characteristic of each “high responder” animal, was strongly suggested, superimposed to faint production of antibodies by clones that were common to all mice.
The ontogeny of B lymphocytes V. Lipopolysaccharide‐induced changes of IgD expression on murine B lymphocytes
Authors: Sitia Roberto , Abbott Joan , Hämmerling Ulrich
Published: February 2007

Murine splenocytes contain two minor subpopulations of B cells, one inducible by lipopolysaccharide to convert within 2 h from IgD− to IgD+ and the other to change from IgD+ to IgD−. These two subpopulations can be separated by density centrifugation. Their relative proportions show a marked age dependency: IgD− → IgD+ cells are more frequent in suckling mice, while IgD+ → IgD− inducible cells become predominant in mice older than 3 weeks. The age dependency observed in the relative proportions between the two cell types suggests that they are ontogenetically related as progenitor‐successor. This hypothesis is corroborated by phenotype analysis of the two subsets, revealing IgD− → IgD+ cells as IgM+, Ia+, complement receptor−(CR−) and IgD+ → IgD− cells as IgM+, Ia+, CR+.

Our data show that IgD and CR are expressed concomitantly during B cell differentiation. On further differentiation, induced by lipopolysaccharide, both markers are lost from the cell surface at different rates: IgD decreases significantly in a very short period (<2.5 h) while induction of a decline in CR requires longer culture periods (>8 h). The loss of IgD may thus herald an early differentiation event toward antibody‐producing cells.
Antibody response to embryonal carcinoma cells in syngeneic mice
Authors: Damonneville Marline , Morello Dominique , Gachelin Gabriel , Stanislawski Marc
Published: February 2007

The immunoglobulin (Ig) classes and subclasses of the specific antibodies contained in antisera raised in male 129/Sv mice against cells of a syngeneic clonal line of embryonal carcinoma (F9) have been determined. Cytotoxic activity was found associated almost exclusively with anti‐F9 IgM (μχ) antibodies. A large part of anti‐F9 activity was found associated with IgG1 (γ1χ and presumably also γ1λ) antibodies, and was detectable only by indirect immunofluorescence. Traces of specific IgG2a and IgG2b antibodies were also found. No IgG3 and IgA antibodies reacting with embryonal carcinoma cells were detectable under these conditions. The serum of F9 tumor‐bearing animals had a very similar Ig composition. fürthermore, IgM and IgG1, but not IgG2 antibodies, were detected at the surface of in vivo growing F9 tumor cells.
IgG‐binding sites on macrophage cell membrane. II. Mobility of Fc receptors induced by the interaction with their corresponding IgG ligands
Authors: Sulica Andrei , Gherman Maria , Medeşan Cornel , Gheţie Victor , Sjöquist John
Published: February 2007

Mouse peritoneal macrophages were charged with IgG molecules in monomeric (mIgG), heat‐aggregated (agIgG) or antigen‐complexed (acIgG) form. Upon exposure to 37 °C, all bound IgG ligand types are redistributed on the cell surface due to the mobilization of their corresponding Fc receptor (FcR). The major findings regarding the fate of FcR on macrophages bearing IgG ligands are as follows: (a) the FcR involved in the binding of cytophilic molecules has a slow movement on the cell membrane and forms patches but never caps, while the opsonic type of FcR is rapidly capped; (b) the mobility of IgG‐binding sites was temperature‐dependent and was affected differently by sodium azide; this metabolic inhibitor enhances the disappearance of mIgG from the cell surface but decreases the capping and the disappearance of polymeric ligands; (c) both FcR types are probably ingested when complexed with specific ligand, and consequently, the rebinding of homologous IgG molecules is reduced, the clearing induced by agIgG or acIgG binding being much more extensive; and (d) cells cleared of their opsonic type of FcR are able to regenerate the receptor molecules within 8 h of incubation at 37 °C.
Suppression of phosphorylcholine‐specific IgE antibody formation in BALB/c mice by isologous anti‐T 15 antiserum
Authors: Blaser Kurt , Geiser Marianne , de Week Alain L.
Published: February 2007

In order to study the regulation of IgE antibody formation, isologous anti‐idiotypic antisera against the phosphoryl choline (PC)‐specific BALB/c myeloma proteins T15 and M167 were passively administered to BALB/c mice in the course of an anti‐PC IgE response. Isologous anti‐T 15 antiserum had a long‐lasting suppressive effect on the formation of IgE antibodies with PC specificity, whereas administration of anti‐ M167 antiserum had no or only little effect, similar to that of normal BALB/c serum. This indicates that anti‐PC IgE antibodies consist mainly of the T15 idiotype or of cross‐reacting idiotypes, and that IgE response is accessible to regulation with anti‐ idiotypic antibodies. This murine model may permit the study of regulation of an IgE response largely restricted to few defined idiotypes characterized as tumor proteins.
Generation of virus‐specific cytotoxic T cells in vitro II. Induction requirements with functionally inactivated virus preparations
Authors: Koszinowski Ulrich H. , Gething Mary‐Jane
Published: February 2007

Using noninfectious Sendai virus preparations after selective enzymatic digestion of either of the two viral envelope glycoproteins, it was possible to study the effect of different virion‐cell membrane interactions on virus‐specific cytotoxic T lymphocyte (CTL) induction in vitro. Three different virus preparations having capacity for virus‐ cell fusion, for virus‐cell adsorption or lacking the ability to bind to cell membranes, were all active in the generation of virus‐specific primary and secondary cytotoxic T cells, when added to the culture. Investigations on the responder cell requirements during CTL induction revealed that activation by addition of virions lacking the capacity to bind to cells was sensitive to the depletion of adherent cells. When virions with fusion and binding capacity were presented on tumor stimulator cells, different requirements with respect to adherent cells were obtained in the primary and secondary CTL response to Sendai virus. The data indicate that different viral antigen‐cell membrane interactions govern the activation phase and effector phase of antigen‐ primed T cell populations, while sensitization of unprimed cells is dependent on the presence of adherent, perhaps antigen‐presenting cells.
Inhibition of cell‐mediated lysis by xenoantibodies reactive with effector T lymphocytes
Authors: Rabinowitz Ruth , Laskov Reuven , Schlesinger Michael
Published: February 2007

The aim of the present study was to determine whether rabbit antisera against various murine tissue antigens contain antibodies that affect the activity of cytotoxic T lymphocytes in the absence of complement. Following heat inactivation, the xenoantisera were absorbed with mouse liver and kidney and tested for their capacity to inhibit cell‐mediated lysis (CML). In the presence of complement (C), rabbit anti‐brain antiserum (RABr), rabbit anti‐thymus antiserum (RAT) and rabbit anti‐boiled thy‐ mus antiserum (RABT) lysed effector T cells, and CML was abolished. In the absence of C, RAT maintained its strong inhibitory activity on CML, RABr had no inhibitory effect, whereas only some batches of RABT were effective. Monovalent Fab fragments of RAT inhibited CML as effectively as intact antibody molecules. The activity of RAT could be removed by absorption with T and B lymphocytes of various sources. Inhibitory antibodies could be removed by absorption with C3H B lymphoma cells and upon elution from antibody‐coated cells proved again highly effective in inhibiting CML. These results show that RAT contains antibodies which react with determinants shared by T and B lymphocytes and which block the activity of cytotoxic T cells.
Phosphorylcholine‐binding hybridoma proteins of normal and idiotypically suppressed BALB/c mice. I. Characterization and idiotypic analysis
Authors: Berek Claudia , Schreier Max H. , Sidman Charles L. , Jaton Jean‐Claude , Kocher Hans P. , Cosenza Humberto
Published: February 2007

The immune response of BALB/c mice to phosphorylcholine (PC) is dominated by an antibody bearing the idiotype of the myeloma protein TEPC 15 (MP T15). Anti‐PC antibody of a different idiotype can be elicited in BALB/c mice by neonatal suppression with an anti‐T15 idiotypic antiserum (anti‐Id‐T15).

In an attempt to further characterize the immune response to PC, spleen cells of normal and idiotypically suppressed mice have been fused with the myeloma line X63‐Ag8, and hybrid lines secreting anti‐PC antibodies have been isolated. The antibodies were characterized by two‐dimensional gel electrophoresis and their affinity for Pneumococcus pneumoniae was measured. Using both an A/J and a rabbit anti‐Id‐T15 serum, we could show that 4 out of 5 hybridomas obtained from normal BALB/c spleen cells secreted antibodies which, within the limits of the analysis, were idiotypically and structurally indistinguishable from MP T15 (T15+). Hybridomas obtained from suppressed BALB/c mice, on the other hand, secreted anti‐PC antibodies which differed idiotypically from MP T 15: of the six lines, two were negative for T15 idiotopes (T15−), while four showed some cross‐reactivity with the T15 idiotype (T15cr). All six, however, showed structural differences in one or more immunoglobulin chains.

Inhibition of anti‐PC plaque‐forming cells induced in neonatally suppressed mice revealed that the idiotypic spectrum of the hybridoma proteins was representative of clones arising under such conditions in BALB/c mice. The majority of the anti‐PC antibodies in idiotypically suppressed mice still bear idiotopes resembling MP T15. Only a minority, less than 20%, are truly T15−.
H‐2 genetic control of the response of T lymphocytes to insulins. Priming of nonresponder mice by forbidden variants of specific antigenic determinants
Authors: Cohen Irun R. , Talmon Janet
Published: February 2007

H‐2 gene control of the response of T lymphocytes to antigenic determinants of ungulate insulins has been studied. We injected H‐2‐congenic mice with different insulins emulsified in complete Freund's adjuvant and measured the proliferative response in vitro of the draining lymph node cells to various related insulins. Two groups of determinants were antigenic: the A chain loop determinant present on bovine and sheep insulins, but absent on pig insulin, and a pig‐associated determinant(s) also shared by bovine and sheep insulins. The responses to these two sets of determinants were controlled by H‐2 genes as follows: (a) The pig‐associated determinant(s) was antigenic for H‐2d but not for H‐2′ or H‐2k mice. (b) The A chain loop determinant was immunodominant over the pig‐associated determinant(s) for H‐2dmice. In the presence of the A chain loop determinant, H‐2d mice did not respond to the pig‐associated determinant(s) on the same molecule. (c) H‐2d mice did not distinguish between the bovine and sheep variants of the A chain loop that differed by substitution of one amino acid. (d) In contrast, H‐2k mice responded only to the sheep variant, while H‐2b mice responded primarily to the bovine variant of the A chain loop determinant. (e) However, injection of H‐2b mice with “forbidden” sheep insulin primed them for an in vitro response to the “permitted” bovine variant of the A chain loop determinant. Similarly, some H‐2k mice injected with forbidden bovine insulin responded in vitro to the permitted sheep variant of the A chain loop determinant. Thus, H‐2 gene products decided whether the immune response would express an affinity for the specific immunizing antigen itself and/or for a closely related variant.
Idiotypic profile of the response to phosphorylcholine induced in the absence of the homologous antigen
Authors: Cammisuli Salvatore , Cosenza Humberto
Published: February 2007

In the absence of the homologous antigen, an anti‐phosphorylcholine (PC) plaque‐ forming cell (PFC) response has been induced in vitro which is restricted to the TEPC 15 idiotype (T15). Anti‐T15 antibodies were used to focus either the keyhole limpet hemocyanin (KLH) or the fowl gamma‐globulin (FGG) carrier molecules on the membrane of B cells carrying the T15 idiotype on their immunoglobulin receptors; thereafter, these cells were allowed to cooperate in vitro for 5 days with T cells primed to the appropriate carrier molecule. A response, virtually 100% T15+, could be induced both in normal BALB/c and in T15 neonatally suppressed mice which had lost the T15 + clonal dominance. The magnitude of this response is comparable to that obtained in the presence of the PC antigen. The role of the membrane‐bound immunoglobulin receptor (sIg) in the expression of different anti‐PC clones was also investigated. We have focused either the KLH or the FGG carrier molecules on the membrane of B cells via anti‐H‐2 antibodies and then cultured these cells for 5 days with the appropriate carrier‐specific T cells. Under these conditions, B cells are activated in the absence of interaction at the sIg. The idiotypic profile of the anti‐PC PFC obtained with the anti‐H‐2‐mediated activation was then compared with the profile of the anti‐PC response obtained in the presence of PC antigen. Since similar idiotypic profiles were obtained in both cases, it can be excluded that sIg plays a direct role in favoring the expression of T15+ over T15− anti‐PC clones.
Partial amino acid sequence of the precursors of blocked heavy and light chains from C 3 H mouse myeloma 5563
Authors: Singer Helen H. , Gates Frederick T. , Kindt Thomas J. , Williamson Alan R.
Published: February 2007

A messenger RNA fraction from the C 3 H mouse myeloma 5563 was used to direct the synthesis of heavy (γ2J) and light (χ) chain precursors. The synthetic products were radiolabeled by inclusion of [3H] or [35S] amino acids in a wheat‐germ cell‐free translation system. Precursor peptides for both H and L chains were indicated by comparison by polyacrylamide gel analysis of apparent molecular weights of chains synthesized in vitro vs. in vivo. The nonglycosylated H chain synthesized in vivo in the presence of tunicamycin was used for comparison. This approach should be generally applicable for the demonstration of precursor forms of N‐glycosylated polypeptides. Following immunoprecipitation and preparative polyacrylamide gel electrophoresis, the isolated chains were subjected to automated Edman degradation. The amino acid sequence data obtained for these precursors were significantly different from those reported for BALB/c immunoglobulins. The 5563 H chain precursor bears little homology to the previously reported H chain precursor from MOPC 315 (IgA) (Jilka, J. A. and Pestka, S., Proc. Nat. Acad. Sci. USA 1977. 74: 5692). The 5563 H chain can be assigned to a heretofore undescribed H chain variable region subgroup on the basis of partial sequence data. The amino terminal sequence of the 5563 L chain precursor has maximum homology (25‐55%) to the sequence of the precursor peptide of MOPC41 χ chain.
Molecular identification of T cell‐specific antigens on human T lymphocytes and thymocytes
Authors: Andersson Leif C. , Karhi Kimmo K. , Gahmberg Carl G. , Rodt Hans
Published: February 2007

We have identified membrane glycoproteins which carry T cell‐specific antigens on human T lymphocytes and thymocytes. Purified cells were surface‐labeled with NaB3H4 after treatment with neuraminidase and galactose oxidase. Immunoprecipitations were performed with rabbit anti‐human T cell‐specific antibodies using co precipitation with protein A‐containing staphylococci strain Cowan I. The labeled membrane glycoproteins and the precipitates were subjected to polyacrylamide slab gel electrophoresis and visualized by fluorography.

The antibodies specifically precipitated 4 proteins called GP200, GP180, GP165 and GP160 (mol. wts. = 200000, 180000, 165000 and 160000) from surface‐labeled T lymphocytes and low‐density (medullary) thymocytes. The GP200 and GP180 were not labeled on high‐density (cortical) thymocytes. A protein with a mol. wt. of 45000 was precipitated from thymocytes. Another glycoprotein on T lymphocytes and thymocytes with a mol. wt. similar to that of mouse and rat Thy‐1 or Θ antigen (mol. wt. 25000) reacted with the antibodies.
T‐T cell interactions during in vitro cytotoxic T lymphocyte responses. III. Antigen‐specific T helper cells release nonspecific mediator(s) able to help induction of H‐2‐restricted cytotoxic T lymphocyte responses across cell‐impermeable membranes
Authors: Pfizenmaier Klaus , Delzeit Rosemarie , Röllinghoff Martin , Wagner Hermann
Published: February 2007

T helper cell induction and the specificity of T cell‐mediated help as generated during alloreactive and H‐2‐restricted, virus‐ or hapten‐specific cytotoxic T lymphocyte (CTL) responses have been compared. With the use of a double‐chamber culture system, it was possible to dissect and separately analyze the induction phase of T helper cells from the T helper cell effector function. The data obtained revealed that during alloreactive as well as H‐2‐restricted T cell responses, antigen‐specific T helper cells are induced. Upon specific restimulation of T helper cells, helper cell function is mediated across a cell‐impermeable membrane via soluble products in an apparently nonspecific and nonrestricted manner. The data suggest that similar rules govern T‐T cell interactions in alloreactive and H‐2‐restricted CTL responses.
Original H‐2d and alien H‐2k‐like antigens of a BALB/c fibrosarcoma as defined by in vitro and in vivo studies of cell‐mediated immunity
Authors: Sensi Marialuisa , Carbone Giusi , Parmiani Giorgio
Published: February 2007

The present study examines the immunosensitivity and the immunogenicity of both original H‐2d and alien H‐2k‐like antigens of the BALB/c (H‐2d) fibrosarcoma C‐1 as detected by in vitro and in vivo cell‐mediated cytotoxicity (CMC) assays. It was found that 51Cr‐labeled C‐1 cells were lysed in vitro by C 57 BL/6 anti‐H‐2d lymphocytes. The specificity of this reaction was shown by cold inhibition experiments in which the anti‐H‐2d cytotoxic activity on YC8 (H‐2d) targets was inhibited by unlabeled YC8 or C‐1 but not by C3UR11 (H‐2k) tumor cells. Both Dd‐ and Kd‐encoded antigens were recognized by appropriate cytotoxic effectors. The immunogenicity of H‐2d antigens of C‐1 was revealed by the ability of C 57 BL/6 anti‐C‐1 lymphocytes to lyse YC8 targets. The expression of H‐2k‐like alien alloantigens on C‐1 was indicated by the finding that anti‐H‐2k cytotoxic T lymphocytes (CTL), generated by culturing BALB/c spleen cells immune to BALB.K (H‐2k), C3Hf (H‐2k) or A (H‐2a = H‐2k/d) tissues with the cells of the same strain used for immunization, lysed C‐1 targets. The cytotoxicity of these anti‐H‐2k CTL against C 3 UR11 (H‐2k) targets could be specifically inhibited by cold C 3 UR 11 or C‐1 cells but not by two other BALB/c tumors. Using recombinant H‐2‐congenic mice, it was shown that both Dk and Kk antigens were recognized by CTL on C‐1 cells. The immunogenicity of the H‐2k‐like antigens, however, could not be detected in vitro. In fact, effector spleen cells from BALB/c mice immune to C‐1 did not develop any detectable cytotoxicity against C 3 UR 11 targets as assayed either by a direct in vitro test or after in vitro restimulation with C‐1 sarcoma cells. A similar experimental design was adopted in Winn assays carried out by mixing spleen cells of BALB/c immune mice with either C‐1 or C3 UR 11 targets and injecting the mixtures in BALB/c or hybrid recipients. These in vivo tests revealed the presence of both H‐2d (Kd and Dd) and H‐2k‐like (Kk and Dk) antigens on C‐1. At variance with the in vitro CMC assays, however, the Winn assay also detected the immunogenicity of the H‐2k alien antigens, since BALB/c anti‐C‐1 spleen cells were able to significantly reduce the growth of C 3 UR 11 lymphoma cells in (BALB/c × C 3 Hf)F1 hosts.
Change in mixed lymphocyte culture reactivity following allosensitization between DLA‐identical dog sibs
Authors: Gluckman Jean‐Claude
Published: February 2007

The mixed lymphocyte culture (MLC) is known to react to major histocompatibility complex disparities but is insensitive to minor histocompatibility antigens. To determine which modifications could amplify reactivity to these antigens, we have studied the effects of presensitization on normally nonresponsive MLC in dogs. Lymphocytes, primed in vitro to and restimulated with cells of a DLA‐identical sib, did not react to these specific cells, in contrast to normal proliferation to other stimulators. However, lymphocytes obtained after in vivo sensitization against a DLA‐identical donor showed increased reactivity to the donor in approximately half of the recipients. In comparison to primary MLC responses between DLA‐mismatched individuals, the [3H] thymidine uptake was less important in this reaction, and peak incorporation was usually not accelerated. This response appeared to be specific in that responsiveness to third‐party cells was not increased, and, alternatively, immunization against a DLA haplo‐identical donor did not modify reactivity to DLA‐identical cells. This increased reactivity after alloimmunization may reflect differences in minor histocompatibility antigens although the skin graft survival time was not significantly shorter in MLC responders than in nonresponder recipients. It seems that in vivo priming is required to expand the number of cells specific for these antigens and to enable detection of proliferation in MLC.
Murine embryonic blood between day 10 and 13 of gestation as a source of immature precursor B cells
Authors: Melchers Fritz , Abramczuk Jan
Published: February 2007

Pre‐B cells which show a delayed development of responsiveness to the B cell mitogen lipopolysaccharide (LPS) are found in murine embryonic blood between day 10 and 14 of gestation. The highest frequency of pre‐B cells in embryonic blood is found at day 12 of gestation. All pre‐B cells of embryonic blood, isolated at different days of gestation, develop responsiveness in vitro to LPS at the same time, i.e. around day 14 of gestation, yielding a peak of LPS‐induced IgM plaque‐forming cells at the equivalent of day 19 of gestation. Dextran sulfate supports the development of pre‐B cells into LPS‐reactive B cells; by itself, however, it does not stimulate these B cells into IgM plaque‐forming cells. The wave of pre‐B cells in embryonic blood occurs parallel in time to that found previously in placenta. Fetal liver and yolk sac, at this time of embryonic development, do not contain significant numbers of pre‐B cells, although they are connected with the placenta and in contact with embryonic blood by circulation.
The in vitro immune response in the absence of serum and its proteins: stimulating activity of zymosan
Authors: Burger Mikulas , Hess Max W. , Cottier Hans
Published: February 2007

In the absence of serum and its components, zymosan, together with 2‐mercaptoethanol present in the medium, stimulated the in vitro primary immune response. Zymosan was effective at a concentration of as low as 2.5 μg/ml. Removal of macrophages using iron powder resulted in higher recoveries of plaque‐forming cells in the presence of zymosan but not of fetuin. Since insoluble zymosan may be easily removed from the culture, the test system described might be suitable for the identification of specific control factors secreted by cells during the in vitro immune response.
Characterization of mouse thymocyte and peripheral lymphocyte xenoantigens by two‐dimensional electrophoresis
Authors: Hoessli Daniel C. , Vassalli Pierre , Pink J. Richard L.
Published: February 2007

Two‐dimensional electrophoresis was used to analyze the surface glycoproteins of murine thymocytes and lymph node cells. Two‐dimensional maps of unselected, radioiodinated lymphocyte surface proteins were complex, showing at least 20 different components, but simpler patterns were obtained by using rabbit antibodies directed against the surface proteins of a T lymphoma cell line, to precipitate xenoantigens from lysates of radioiodinated or biosynthetically labeled thymocytes and lymph node cells. These xenoantibodies precipitated 12–13 distinct components from each cell type, of which all but 3 were sialoglycoproteins. Two types of difference between the surface glycoproteins of thymocytes and peripheral lymphocytes could be detected. First, higher mol. wt. glycoproteins and Thy‐1 are more acidic in peripheral lymphocytes than in thymocytes, and this difference disappears after neuraminidase treatment. One additional high mol. wt. glycoprotein is also detectable in peripheral lymphocytes, probably reflecting the greater carbohydrate complexity of these molecules, when expressed on such cells. Second, 3 glycoproteins are strongly labeled only on thymocytes, and 3 others only on peripheral lymphocytes. These 6 glycoproteins might represent genuine differentiation antigens.
Plasma membrane cholesterol regulates human lymphocyte cytotoxic function
Authors: Dabrowski Marek P. , Peel William E. , Thomson Alasdair E. R.
Published: February 2007

Cell cholesterol is believed to be confined mainly to the plasma membrane. Treatment here of human peripheral blood lymphocytes with cholesterol‐free and cholesterol‐containing liposomes to effect, respectively, decreases or increases in cholesterol content measurable by chemical analysis, markedly altered effector functions of the cells. Depletion of cholesterol evoked inhibition of spontaneous and phytobemag‐glutinin‐dependent lymphocyte cytotoxicity against allogeneic target cells. Opposite effects resulted from cholesterol enrichment, with PHA‐dependent and antibody‐dependent cytotoxicities increasing significantly. Treatment, instead, with the known inhibitor of cholesterol biosynthesis, 25‐hydroxycholesterol, had suppressive effects like those resulting from lowering the cholesterol level physically by liposome treatment. Our data suggest that the plasma membrane cholesterol content of different categories of lymphocytes in man is both essential and regulatory for their cytotoxic function.
Interferon induced in human leukocytes by mitogens: production, partial purification and characterization
Authors: de Ley Marc , van Damme Joseph , Claeys Hendrik , Weening Hans , Heine Jochen W. , Billiau Alfons , Vermylen Carl , de Somer Piet
Published: February 2007

Interferon was induced in leukocyte suspensions from human buffy coats by exposure to phytohemagglutinin P, concanavalin A (Con A) and staphylococcal enterotoxin A, under a variety of cell culture conditions. Con A was found to rapidly (within 12 h) induce high yields of antiviral activity (1.5 units/1000 cells). Lesser yields were obtained with the other two mitogens studied. The interferon was partially purified to a spec. act. around 105.3 units/mg protein, by batch adsorption on controlled‐pore glass (CPG) beads and desorption by ethylene glycol. This material was characterized as containing mainly γ‐type interferon. Specifically on gel filtration, a fraction of 45 000 daltons was obtained which could account for virtually all antiviral activity present in the starting material. Furthermore, the ethylene glycol‐eluted antiviral activity was acid‐labile, serologically distinct from a and β‐type interferon and strictly species‐specific (no activity detectable on any of the available cell species sensitive to α and β‐type interferon). The crude culture supernatant also contained some antiviral activity which resembled β‐type interferon in that it adsorbed to CPG, could be desorbed by pH 2 buffer, was acid‐resistant and could be neutralized by a specific anti‐fibroblast interferon antiserum. The CPG/ethylene glycol‐purified γ‐type interferon preparation was found to inhibit the growth of certain lymphoblastoid cells (Daudi and Molt‐4). It also potentiated natural killer activity of fresh donor lymphocytes. In both respects, the γ‐type interferon preparation was not significantly more active than preparations of a and β‐interferon of similar antiviral potency.
Characterization of human lymphocytes which proliferate “spontaneously” in vitro
Authors: Treves Abraham J. , Barak Vivian , Fuks Zvi
Published: February 2007

Human peripheral blood lymphocytes proliferate “spontaneously” in vitro following the reduction in the proportion of monocytes in culture. This cell proliferation, which takes place in the absence of any extrinsic antigenic stimulation, was found to reside in a small fraction of a non‐T non‐B cell population which was also Fc receptor‐negative. Concomitant with the in vitro proliferation of the monocyte‐depleted cells, an increase in the proportion of E rosette‐forming cells was observed. Hence, in addition to the regulation of spontaneous proliferation of lymphocytes, it is suggested that monocytes may also be involved in the regulation of the proportion of T cells or in expression of the E‐rosetting marker.
Helper cell analysis in cytotoxic T lymphocyte responses I. Activation of specific and cross‐reactive helper effects in two hapten‐modified self responses
Authors: Levy Robert B. , Fujiwara Hiromi , Shearer Gene M.
Published: February 2007

C 3 H/HeN mice were immunized by skin painting with trinitrochlorobenzene or by inoculation of fluorescein isothiocyanate (FITC)‐conjugated syngeneic cells. Approximately four weeks later, spleen cells were removed, irradiated, and cocultured with spleen cells from normal mice. These cultures were stimulated with syngeneic cells conjugated with different concentrations of trinitrobenzene sulfonate (TNP‐self) or FITC (FITC‐self). The results demonstrate (a) appreciable helper cell activity from TNP‐self and FITC‐self‐primed mice, as determined by enhanced hapten‐specific cytotoxic T lymphocyte (CTL) responses, (b) that FITC helper activity enhanced the FITC‐self CTL more efficiently than the TNP‐self CTL response, and vice versa, (c) an alloantigenic stimulating population failed to activate TNP‐self helper activity and (d) that the helper activity was not exclusively specific, since FITC helpers did enhance the anti TNP CTL response, and TNP helpers could enhance the anti‐FITC‐self CTL response. The results of this study indicate that these cell‐mediated lympholysis models permit an analysis of hapten specificity and cross‐reactivity at the different levels of cellular interaction involved in the generation of an H‐2‐restricted CTL response.
The enumeration of mouse IgE‐secreting cells using plaque‐forming cell assays
Authors: Rector Edward S. , Lang Glen M. , Carter Brian G. , Kelly Ken A. , Bundesen Peter G. , Böttcher Irmgard , Sehon Alec H.
Published: February 2007

With the aid of a specific rabbit antibody preparation to purified monoclonal murine IgE, two plaque‐forming cell (PFC) assays have been developed for the detection and enumeration of mouse IgE‐secreting cells. The first assay, utilizing protein A‐coated sheep red cells (protein A‐SRC), detected antibody‐secreting cells on the basis of the class of the secreted Ig irrespective of antigen specificity. With this assay, 30% of the viable cells of two distinct IgE‐secreting hybridoma cell lines were scored as PFC. Under these conditions, plaques were not obtained with IgG1 or IgG2a‐secreting hybridoma cells. The second PFC assay, which utilized SRC coated with ovalbumin (OA‐SRC), enumerated cells secreting anti‐OA IgE antibodies. Similar kinetic patterns were observed for the cellular (IgE PFC/spleen) and humoral (IgE serum levels) responses of (C 57 BL/6 × DBA/2)F1 mice following immunization with 10 μg of OA adsorbed to 1 mg of Al(OH)3. Thus, it is concluded that the reverse plaque assay detecting all IgE‐secreting cells, as well as the antigen‐specific IgE PFC assay, can be used for the quantitation of IgE responses at the cellular level.
Hormonal regulation of the expression of la antigens on mammary gland epithelium
Authors: Klareskog Lars , Forsum Urban , Peterson Per A.
Published: February 2007

Guinea pig and mouse mammary gland epithelial cells express Ia antigen‐like molecules that react specifically with a rabbit anti‐la antigen antiserum. The murine Ia antigen‐like molecules were shown to share alloantigenic determinants with regular spleen cell Ia antigens. The expression of the mammary gland Ia antigens is under hormonal control, i.e. the Ia antigen expression is induced by pregnancy and lactation and can also be induced by the exogenous administration of lactotropic hormones.
T cell‐mediated immunity to oncornavirus‐induced tumors III. Specific and nonspecific suppression in tumor‐bearing mice
Authors: Plater Christine , Debré Patrice , Leclerc Jean‐Claude
Published: February 2007

A strong cell‐mediated immune response against Friend, Moloney, Rauscher virus‐induced (FMR) cell surface antigens has been demonstrated previously in mice which reject oncornavirus‐induced tumors. In order to identify an eventual suppressor mechanism in animals with progressively growing tumors, experiments were initiated in C 57 BL/6 mice bearing either a murine sarcoma virus (MSV) tumor or Moloney virus‐induced lymphoma (MBL2). Progressive tumor growth was induced (a) in viremic animals first infected with Moloney murine leukemia virus (M.MuLV), then inoculated as adult with MSV; (b) in nonviremic animals injected with MBL2 lymphoma cells. In the absence of tumor cells, viremia induces specific tolerance for which there is no evidence for suppressor cells. In tumor‐bearing mice, specific suppressor T cells are detected which are able to inhibit the generation of anti‐FMR cytolytic T lymphocytes in vitro and enhance the tumor growth in vivo. In addition to the specific suppressor T cells, a nonspecific suppressive activity mediated by metastatic T lymphoma cells is demonstrated in the spleens of lymphoma‐bearing animals. The respective role of the virus and tumor cells in the induction of tolerance to M.MuLV‐induced antigens, and their relationship to other components of the specific cell‐mediated immune response is discussed.
Involvement of the spleen in murine B cell differentiation
Authors: Ron Yacov , de Baetselier Patrick , Segal Shraga
Published: February 2007

Experiments were performed to study the influence of neonatal and adult splenectomy on B cell differentiation in mice. Lymph node cells of both groups were found to contain a significantly higher proportion of Ig‐bearing cells. Moreover, these cells expressed a higher density of membrane Ig and a higher IgM/IgD ratio. In addition, the response of these cells to different polyclonal B cell activators, especially dextran sulfate, was much higher than the response of the sham‐operated controls. Similar experiments were carried out in splenectomized and control animals that were lethally irradiated and reconstituted with different hemopoietic cell sources. The same pattern of results was obtained, except when reconstituting the irradiated animals with fetal liver cells.

These experiments, which were performed at long time intervals post splenectomy, show that in the absence of the spleen, there is an accumulation of less mature B cells in the periphery implying an active role of the spleen in at least certain differentiation steps of the B cell lineage.
Lymphocyte specificity to protein antigens III. Capacity of low responder mice to beef cytochrome c to respond to a peptide fragment of the molecule
Authors: Corradin Giampietro , Chiller Jacques M.
Published: February 2007

Lymph node cells derived from A.TH or A.TL mice primed with beef cytochrome c show striking patterns of reactivity when assayed in vitro for antigen‐induced T cell proliferation. Whereas cells from A.TH mice respond specifically to beef cytochrome c or peptides composed of amino acids 1‐65 and 81‐104, cells from A.TL mice respond neither to beef cytochrome c nor to peptide 1‐65, but proliferate following exposure to either peptide 81‐104 or to a cytochrome c hybrid molecule in which the N‐terminal peptide of beef (1‐65) was substituted by a similar peptide obtained from rabbit cytochrome c. Thus, T cells from mice phenotypically unresponsive to beef cytochrome may, in fact, contain populations of lymphocytes capable of responding to a unique peptide, the response to which is totally inhibited when the same fragment is presented in the sequence of the intact protein.
Hapten‐specific helper T cells III. Fine specificity of the (4‐hydroxy‐3‐nitro‐phenyl)‐acetyl (NP)‐specific response in Igh‐1b mice
Authors: Martinez‐Alonso Carlos , Coutinho Antonio , von Boehmer Harald , Bernabé Rosa
Published: February 2007

Helper cells, with specificity for the haptens (4‐hydroxy‐3‐nitro‐phenyl)acetyl (NP) or (4‐hydroxy‐5‐iodo‐3‐nitro‐phenyl)acetyl (NIP), were raised in B10.BR mice by in vivo priming and in vitro long‐term enrichment with hapten‐derivatized syngeneic spleen cells. Upon co‐culture with the homologous antigen (NP or NIP self), selected helper cells specifically responded by proliferation and by inducing large numbers of B cells to clonal expansion and immunoglobulin secretion. Criss‐cross experiments demonstrated the nonheteroclitic nature of antigen recognition by helper cells, as the proliferative and helper cell activites were in every case one order of magnitude higher when confronted with the homologous hapten used for immunization.