bluuebunny/crossref_metadata_2025 huggingface dataset:
HLA class II restriction governing cell cooperation between antigen‐specific helper T lymphocytes, B lymphocytes and monocytes for in vitro antibody production to influenza virus
Authors: Fischer Alain , Sterkers Ghislaine , Charron Dominique , Durandy Anne
Published: February 2007

To study HLA class II compatibility requirement for in vitro antibody production to influenza virus, semipurified T lymphocytes, B lymphocytes and monocytes from HLA‐typed responder donors were used. The presence of the three subpopulations was required for antibody production while a mixture of only two of those was ineffective. When using fresh T lymphocytes which exert an allogeneic suppressive effect and may also exhibit allogeneic helper activity, it was not possible to conclude an HLA class II‐linked restriction of T‐B cell cooperation although there was a suggestion of it. However, a grown H3 hemagglutinin‐specific T cell line (L2), previously shown to be restricted by HLA‐DR molecule (DR1) for interaction with antigen‐presenting cells and devoid of allogeneic reactivity, exerts an HLA class II‐restricted helper activity. This was demonstrated by various combinations of HLA‐DR semi‐compatible or incompatible B lymphocytes and/or monocytes with L2 T cells.

The restriction element was identified as an HLA‐DR determined since HLA‐DC‐compatible, HLA‐DR‐incompatible B lymphocytes were not helped by L2 T cells. In addition, monoclonal anti‐HLA‐DR but not anti‐HLA‐DC antibodies directed to the relevant specificity did inhibit the antigen‐specific helper activity.

We present evidence that not only T monocyte but also T‐B and/or T‐B‐monocyte interactions are HLA class II restricted.
Stromal cell types in the developing thymus of the normal and nude mouse embryo
Authors: van Vliet Els , Jenkinson Eric J. , Ton Rosetta Kings , Owen John J. T. , van Ewrjk Willem
Published: February 2007

The anatomical distribution of various nonlymphoid cell types in the embryonic mouse thymus in vivo and in vitro, as well as in the thymic rudiment of the nude mouse embryo, has been studied. For this purpose a panel of monoclonal antibodies, ER‐TR3, 4, 5, 6 and 7, directed to various types of stromal cells of the mouse thymus, was used in combination with immunoperoxidase labeling on frozen sections. It was shown that as early as day 13 in thymic ontogeny distinction of TR4+ cortical epithelial cells and TR5+ medullary epithelial cells is possible. Thus, as far as stromal components are concerned, the thymus at day 13 in ontogeny is already subdivided into cortex and medulla. At day 13, Ia (TR3) was expressed in a focal pattern in the medulla subsequently appearing throughout both cortex and medulla by day 16. The thymic rudiment of the nude mouse embryo differs markedly from the normal embryonic thymus in its lack of demonstrable Ia antigen. Furthermore, TR4 and TR5 were only expressed on occasional epithelial cells lining the cysts of the nude thymus in a mutually exclusive fashion. The majority of stromal cells of the nude thymus, however, is negative for all ER‐TR antibodies tested.

In addition, we have shown that in organ cultures the organization of the stroma of thymic lobes remains intact, at least for a period of 11 days. Embryonic thymi cultured in the presence of deoxyguanosine, which causes depletion of lymphoid cells, also contain cortical and medullary areas as identified by the presence of TR3,4+ and TR5+ stromal cells. This indicates that the lack of organization in the nude thymus is not simply due to the absence of lymphoid cells.
The selective localization of B lymphocytes in the spleen and the role of complement receptors
Authors: Kraal Georg , van Hoogstraten Ingrid , Klerx Jac P. A. M. , van Dijk Hans
Published: February 2007

The role of complement receptors on the localization of T and B cells in the spleen of mice was studied using short‐term homing experiments in cobra venom factor (CoF)‐ treated animals. The localization ratio of B and T cells in the spleen of CoF‐treated mice decreased significantly compared to control recipients. No changes could be found in the relative distribution of resident T and B cells in the spleen or other lymphoid organs of CoF‐treated animals and when their spleen or lymph node cells were transferred, the localization pattern was normal. When cells were incubated in serum prior to transfer a disturbed localization ratio in the spleen of untreated recipients was observed. This was due to a blockade of complement receptors as determined by the inability of the incubated cells to form EAC rosettes. No blockade of EAC rosettes and no changes in localization ratios upon transfer could be observed when the cells were incubated in functionally C3‐depleted serum. The results suggest a role for the complement‐receptor on B cells in the initial localization in the spleen, whereas no influence upon the selective localization in high endothelial venules‐ bearing organs was found.
Involvement of T3 and T8 molecules in human T cell receptor function. Anti‐T3 or anti‐T8 antibodies do not block the antigen‐dependent activation of a subset of alloreactive cytolytic T cell precursors
Authors: Moretta Alessandro , Pantaleo Giuseppe , Mingari Maria‐Cristina , Moretta Lorenzo
Published: February 2007

In the present study, we have analyzed the effects of anti‐T8 and anti‐T3 monoclonal antibodies (mAb) on the generation of human allospecific cytolytic T lymphocytes (CTL). A sensitive limiting dilution microculture system, in which graded numbers of responder T cells were added to 105 irradiated allogeneic spleen cells and saturating amounts of exogeneous interleukin 2, has been employed to provide minimal estimates of the frequencies of alloreactive CTL precursors (CTL‐P) present in T lymphocyte populations freshly isolated from peripheral blood. Addition of anti‐T8 or anti‐T3 mAb at the onset of micro mixed lymphocyte culture containing peripheral blood T lymphocytes as responding cells caused a 65–80% reduction in the frequency of specific CTL‐P which proliferate under these conditions. Moreover, in most instances (> 80%) the cytolytic activity of those CTL‐P which underwent clonal expansion in the presence of either anti‐T8 or anti‐T3 mAb was found to be resistant to inhibition by the corresponding mAb added during the cytolytic assay. In addition, a great degree of overlap existed between microcultures resistant to anti‐T3 and those resistant to anti‐T8 mAb. In contrast, neither antibody had any inhibitory effect when added later to the cultures, presumably because at this stage CTL‐P had been already triggered by alloantigens. Taken together our data indicate that the heterogeneity in the susceptibility to inhibition by anti‐T3 or anti‐TS mAb, previously observed at the level of CTL clones, is predetermined already at the level of peripheral blood CTL‐P.
Involvement of T11 molecules in antigen receptormediated T lymphocyte functions: effect of anti‐T11 monoclonal antibody on functional capabilities of alloreactive T cell clones
Authors: Moretta Alessandro , Pantaleo Guiseppe , Lopez‐Botet Miguel , Mingari Maria‐Cristina , Carrel Stephane , Moretta Lorenzo
Published: February 2007

As shown by previous studies, the sheep erythrocyte‐binding T11 molecule is involved in T cell activation, as well as in mechanisms of specific allogeneic target cell lysis. In this study, we utilized two anti‐T11 monoclonal antibodies (mAb) that inhibited the specific cytolytic activity of mixed lymphocyte culture (MLC)‐activated T cells to analyze, at the clonal level, the involvement of T11 molecules in (a) antigen‐specific vs. nonspecific mechanisms of target cell lysis, and (b) antigen‐driven T cell proliferation and interleukin 2 (IL2) production vs. IL2‐induced cell proliferation. In contrast to anti‐T3 or anti‐T8 mAb, antibodies to T11 molecules inhibited the cytolytic activity of MLC‐derived allospecific clones in a uniform manner. In addition, anti‐T11 antibodies inhibited the specific activity of cytotoxic T lymphocyte clones resistant to anti‐T3 antibodies, even after antibody‐induced modulation of T3 molecules (while anti‐T3 mAb had no effect). Similarly, anti‐T11 antibodies inhibited the alloantigen‐induced proliferation and IL2 release of alloreactive clones independent of their T4+ or T8+ phenotype. The inhibitory activity of anti‐T11 antibodies appears to be confined to antigen‐specific T cell functions since neither natural killer‐like activity of cytotoxic T lymphocyte clones nor the IL 2‐induced clonal proliferation was affected. Thus, our results indicate that T11 molecules are functionally involved in antigen recognition by T cell regardless of their function and T4/T8 phenotype. The possible mechanisms of anti‐T11 antibody‐mediated inhibition are discussed.
Phytohemagglutinin binds to the 20‐kDa molecule of the T3 complex
Authors: Valentine Mary A. , Tsoukas Constantine D. , Rhodes Gary , Vaughan John H. , Carson Dennis A.
Published: February 2007

Current findings have suggested that the T3 molecular complex is an essential antigenic signal transducer during T cell activation. Lectins, such as phytohemagglutinin (PHA), activate T cells nonspecifically. Concesivably, lectins may mediate their stimulatory action by affecting the T3 complex. In the present investigation we have studied the involvement of the T3 molecular complex in the PHA‐mediated activation of T cells. We selectively modulated the surface expression of T3 molecules by anti‐T3 antibody and subsequently tested the ability of the modulated cells to respond to PHA. Reduction of T3 expression by 70% resulted in 80% inhibition of the PHA response. This effect was specific for T3 since modulation of other T cell surface molecules (T4, T8) did not affect the PHA‐induced mitogenesis. To determine if PHA could interact directly with the T3 complex, immunoblotting (Western blot) analyses of anti‐T3 immunoprecipitates were performed. A 20‐kDa member of the T3 complex reacted not only with the anti‐T3 antibody, but also with PHA itself. These results provide the first evidence for direct binding of PHA to one of the molecules of the T3 complex. The combined data suggest that a major pathway of PHA‐induced T cell activation involves the T3 complex. Possible activation mechanisms are discussed.
Binding of Coprococcus comes to the Fc portion of IgG. A possible role in the pathogenesis of Crohn's disease?
Authors: van de Merwe Joop P. , Stegeman Jeanette H.
Published: February 2007

Previous studies have shown that the fecal flora of Patients with Crohn's disease (CD) differed from the flora of healthy subjects by a higher number of anaerobic grampositive coccoid rods. Sera from patients with CD agglutinated four strains of coccoid rods (Me44, C18, Me46 and Me47) more frequently and stronger than sera from healthy subjects and patients with other diseases. One of these bacteria, Coprococcus comes strain Me46, was not ingested by neutrophils after coating with specific IgG. In the present study, therefore, the binding of IgG and of Fab and Fc fragments to the four coccoid rods was investigated using immunofluorescence and absorption techniques. Results were compared with those obtained with Staphylococcus aureus strain Cowan I and showed that Me46, like S. aureus, bound (nonspecifically) IgG through its Fc portion whereas strain Me47 bound IgG through the Fab portion. Possible implications of the findings for CD are discussed.
Activation of human T lymphocytes III. Triggering of bystander cytotoxicity in cytotoxic T cell clones by antibodies against the T3 antigen or by a calcium ionophore
Authors: Schrezenmeier Hubert , Kurrle Roland , Wagner Hermann , Fleischer Bernhard
Published: February 2007

The role of T cell differentiation antigens in antigen‐specific and nonspecific cytotoxicity by human cytotoxic T lymphocyte (CTL) clones was investigated. In contrast to other reports, several monoclonal antibodies (mAb) against the T3 antigen only marginally blocked antigen‐specific cytotoxicity at high concentrations but induced cytotoxicity against third party cells at concentrations from 10 to 0.001 μg/ml. Susceptibility to anti‐T3‐induced lysis was variable but was found with all target cells, Incubation of CTL with anti‐T3 mAb even led to self‐destruction of the CTL. The effect was independent of the presence of Fc receptors on the target cell and could be obtained with F(ab')2 fragments of the antibody as well. Only activated but not resting T cells could be induced to lyse by anti‐T3. Furthermore, this type of bystander killing of target cells could also be induced by the Ca2+ ionophore A23187. Antibodies against the T8 differentiation antigen inhibited antigen‐specific, oxidation‐induced and antGT3‐induced cytotoxicity by T8+ CTL clones, whereas triggering by the ionophore A23187 was not inhibited. These results show that undirected killing can be triggered in CTL by activating a transducing molecule directly without involving the antigen receptor. Since this triggering of the lethal hit can still be inhibited by mAb against the T8 molecule, the T8 molecule probably has a regulatory role in a late phase of CTL triggering.
Role of protein synthesis in induction of interferon‐γ by mitogens in human lymphocytes
Authors: Siggens Kenneth W. , Tinsley Jonathan M. , Morris Alan
Published: February 2007

The effect of inhibition of protein synthesis on interferon gamma (IFN‐γ) mRNA induction has been examined in human peripheral blood leukocytes and growing T lymphoblasts. Inhibition of protein synthesis by cycloheximide or puromycin when lymphocytes were stimulated with mitogen alone had little effect on the steady‐state levels of IFN‐γ mRNA induced. Activation of the IFN‐γ gene appears to occur independently of synthesis of protein factors.

Synergistic induction of IFN‐γ by mitogen plus the phorbol ester mezerein was at least in part accounted for by increased levels of IFN‐γ mRNA in both fresh lymphocytes and growing lymphoblasts. Inhibition of protein synthesis abolished the synergistic effect on mRNA leading to levels similar to those observed in cells treated with mitogen alone. Synergistic induction is dependent upon synthesis of protein factors either within the cells or produced as soluble mediators; these factors are not simply lymphokines since addition of exogenous factors to the cultures did not reverse the block on synergy by the inhibition. These data suggest that protein factors though they may be important in exerting qualitative control on the level of production of IFN‐γ have no role in the initial activation of the gene.
Human T cell recognition of cloned HL A class I gene products expressed on DNA transfectants of mouse mastocytoma P815
Authors: Maryanski Janet L. , Moretta Alessandro , Jordan Bertrand , de Plaen Etienne , van Pel Aline , Boon Thierry , Cerottini Jean‐Charles
Published: February 2007

Cloned genes for human major histocompatibility complex (MHC) class I antigens were introduced by DNA‐mediated gene transfer into a high‐efficiency transfection recipient (HTR) cell line previously derived from mouse mastocytoma P815. Cell surface expression of HLA‐A3, AW24 and CW3 gene products on P815 transfectants was demonstrated by radioimmune assay and by flow cytometry. The human MHC class I gene products were apparently expressed on P815 transfectants in a form recognized by human cytolytic T lymphocytes (CTL). Human CTL generated in unidirectional mixed lymphocyte culture against AW24+ donor lymphocytes clearly lysed P815‐HLA‐AW24+ transfectant target cells, but not untransfected P815(HTR) controls. Moreover, P815‐HLA transfectants could stimulate in vitro a significant alloreactive human CTL response. Lysis of P815‐HLA transfectant target cells by human CTL was inhibited by a monoclonal antibody directed against human MHC class I gene products. These mouse cell transfectants may be useful for the study of human T cell responses.
Transformation of B and non‐B cell lines with the 2,4,6‐ trinitrophenyl (TNP)‐specific immunoglobulin genes
Authors: McCubrey James , McKearn John P. , Köhler Georges
Published: February 2007

The rearranged μ and x genes from the 2,4,6‐trinitrophenyl (TNP)‐specific hybridoma Sp6 have been introduced into B cells from three different stages of differentiation as well as 5 non‐B cell lines to determine the levels and modes of immunoglobulin (Ig) gene expression. In pre‐B cells transformed with the μ and ϰ genes, low levels of Sp6‐ specific μ RNA were produced and approximately 210‐fold less μ and 800‐fold less ϰ proteins were produced than in the hybridoma Sp6. The Ig proteins were present intracellularly, but were not detected on the cell membrane. In mature surface sIg+ B cell transformants, higher levels of μSp6 and ϰSp6 proteins and RNA were produced than in the pre‐B cell transformants (12 × μ, 70 × ϰ). These transformants displayed the μSp6 and ϰSp6 proteins on the cell membrane and also secreted the transfected Ig product. Plasma cell transformants produced the highest amounts of μSp6 and ϰSp6proteins. These transformants secreted pentameric IgM but did not display detectable amounts of these proteins on the cell membrane.

T cell and one fibroblast transformant produced Ig as normal sized μSp6 and ϰSp6proteins. All other μSp6 and ϰSp6 non‐B cell transformants (melanoma, teratoma and macrophage) failed to produce enough Ig to determine whether the Ig proteins were of the correct molecular weights. The T cell and fibroblast transformants that produced Ig proteins did not secrete or display detectable Ig on the cell membrane. The expression of Ig did not inhibit the expression of the T cell antigen Thy‐1 in the T cell transformants.
Oscillation of antibody production and regulatory T cells in response to antigenic stimulation
Authors: Zöller Margot , Lopatta Dagmar , Benato Benjamino , Andrighetto Giancarlo
Published: February 2007

Priming with the T‐dependent antigen trinitrophenylated horse red blood cells led to an oscillatory response of plaque‐forming cells (PFC) with peaks of response after 4, 8, 12 and 16 days. Limiting dilution (LD) analysis of regulatory T cells indicated that the response is partly due to activation of helper T cells (Th), but mainly is the result of release from suppression by activation of a third regulatory T cell (T3) population (Zöller, H. and Andrighetto, G., Immunology 1985. 55: 703). Priming resulted in consecutive activation of suppressor T cells (Ts; day 1‐3), a population regulating Ts(T3; day 2‐4) and of Th (day 4). Furthermore, regulatory elements displayed a cyclical behavior like PFC. Th and T3 oscillated within a period of 4 days in phase with PFC, while Ts oscillated phase‐shifted to PFC, Th and T3. Hence, oscillation of PFC after priming with a T‐dependent antigen is the result of interactions among regulatory T cells.

The data are interpreted as follows. Th are the primum movens of the oscillatory behavior of PFC response, since Th oscillate in phase with PFC and display the same pattern of decreasing amplitudes as PFC. However, the basic regulatory elements are Ts and T3 which are interlinked and oscillate phase shifted. As revealed by the time‐ course analysis of LD curves, T3 function as suppressor cells for Ts. Hence the status of mutual interactions of Ts and T3 determines suppression of Th or release from suppression, i.e. when T3 are found at high frequencies, Th are released from suppression, since T3 suppresses Ts. Instead, the increase in the frequency of Ts and their predation on Th determine the decrease of the frequency of Th. Thus, these data support the hypothesis of a circular network, the central regulatory core being composed of two elements, Ts and T3, with mutual suppressive activity.
Several V genes participate in the early phenyloxazolone response in various combinations
Authors: Kaartinen Matti , Pelkonen Jukka , Mäkelä Olli
Published: February 2007

Seventeen monoclonal anti‐2‐phenyloxazolone antibodies from the early (day 7) primary response were partially sequenced with an mRNA method. Ten antibodies expressed the VH−Ox1 gene. The remaining seven express at least four but probably six different germ‐line VH genes belonging to Dildrop's groups 1, 5, 6 and 7 (Immunol. Today 1984. 5: 85). Two of them have been met before in other antibodies, one (group 6) in J606 and the other (group 7) in antibodies to the influenza virus hemagglutinin.

Eleven kappa chains were partially sequenced and five of them (all VH−Ox1 antibodies) express the Vx−Ox1 gene. One expresses another germ‐line gene of the Vx−Ox1 family, one the Vx89.4 gene, three the Vx45.1 gene and one a new Vx gene. The Vx45.1 gene was found to form anti‐phox antibodies with two new VH genes.

The frequency of somatic mutations in day 7 antibodies was estimated by comparing germ‐line sequences and antibody sequences. It is low (one mutation per 2500 nucleotides sequenced), twenty times lower than in antibodies obtained a week later. Two anti‐idiotype antisera (495 and 260) are useful in the typing of monoclonal antibodies. 260 bound only to antibodies coded by both VH−Ox1 and Vx−Ox1 genes. 495 bound strongly to antibodies coded by the VH−Ox1 gene and weakly to antibodies coded by the (related) VH101 gene regardless of the light chain partner.
Quantitative representation of two germ‐line V genes in the early antibody response to 2‐phenyloxazolone
Authors: Pelkonen Jukka , Kaartinen Matti , Mäkelä Olli
Published: February 2007

Early anti‐phenyloxazolone antibodies of B ALB/c mice include a subset that bears the nearly or totally unmutated VH−Ox1 sequence. A fraction of this subset also bears the nearly or totally unmutated Vx−Ox1 sequence. The whole subset is recognized by an anti‐idiotype serum 495 and the VH−Ox1/Vx−Ox1 fraction also by another anti‐idiotype serum 260.

Frequencies of the VH−Ox1 subset and its Vx−Ox1 fraction in early IgM and IgG antibodies were determined by typing hybridomas with anti‐idiotype antisera. The whole subset appears to make up approximately one half of IgG antibodies, two thirds of this being 495+/260+ in both isotypes. IgM data are less certain but the same frequencies may be valid.

Affinities of 495+/260+ antibodies ranged from 1.3 × 106 to 11 × 106, affinities of 495+/260− antibodies from 0.47 × 106 to 1.1 × 106, and affinities of doubly negative antibodies were less than 0.41 × 106. High affinity is probably an explanation for the high proportion (one third) of the 495+/260+ antibodies in the early response. Doubly negative (and low‐affinity) hybridomas may not have been classified as producers of phenyloxazolone antibodies in earlier studies, and this could explain the still higher reported frequency (73%) of VH−Ox1/Vx−Ox1 antibodies.
Electronmicroscopical observations on monocyte‐lymphocyte interactions upon stimulation with pokeweed mitogen latex conjugate
Authors: De Vries Egbert , Van Weij Johan P. Der , Van Der Veen Corrie J. P. , Doekes Gert , Steven Malcolm M. , Cats Arnold
Published: February 2007

In this study we report on the preparation and application of pokeweed mitogen (PWM) conjugated to latex particles. This conjugate (PWM‐latex) was prepared by incubation of PWM with latex particles in the presence of glutaraldehyde. The effect of the addition of PWM‐latex to human peripheral blood mononuclear cells (PBMC) was fully comparable to the addition of PWM alone i.e. differentiation of lymphocytes into blast cells followed by proliferation of these blast cells as measured by DNA synthesis and total cell number. Electronmicroscopically PWM‐latex was found to be taken up by monocytes within the first 24 h after addition. Although no direct interaction could be observed between PWM‐latex and lymphocytes, the latter were found to differentiate into blast cells. Due to interactions of these blast cells with latex‐containing monocytes, latex particles were obviously released from the phagocytes and close contacts between latex particles and blast cells were regularly seen. In addition, it was found that blast cells of T cell origin, as judged by their positive reaction with anti‐T cell monoclonal antibodies, had taken up latex particles. Based on enzyme‐cytochemical, functional and light microscopical studies, monocytes could not be detected in the PWM‐latex‐driven PBMC stimulation after 6 days. At the electronmicroscopical level evidence was found that the inability to demonstrate macrophages could be due to the release of the lysosomal enzyme content and of parts of the cytoplasm.
Control of Ia antigen expression on phagocytic cells of the thymic reticulum by interferon‐gamma and prostaglandins
Authors: Papiernik Martine , Dombret Hervé , Stefanos Simon , Wietzerbin Juana
Published: February 2007

Phagocytic cells of the thymic reticulum (P‐TR) spawned in the microenvironment of primary thymic stroma cultures express class II antigens on their membranes. We can show that Ia‐antigen expression is regulated by the local secretion of interferon‐gamma (IFN‐γ): (a) early primary culture supernatants contain IFN as tested by a cytopathogenic inhibition test after concentration of IFN activity; (b) the addition of a specific anti‐IFN‐γ antiserum to primary cultures reduces Ia expression on P‐TR suggesting that IFN‐γ produced in situ is implicated in Ia expression; (c) the percentage of Ia+ cells declines as primary cultures age. This phenomenon parallels the disappearance of thymocytes which are the supposed candidate for IFN‐γ secretion. (d) When isolated from their stromal environment and replated in secondary cultures, P‐TR lose their membrane Ia antigen which can be re‐expressed in the presence of recombinant IFN‐γ, but not IFN‐α/β. The decline of Ia expression on P‐TR during the aging of primary cultures may be delayed by the inhibition of prostaglandin secretion.
Functional and mechanistic studies on the toxicity of deoxyguanosine for the in vitro proliferation and differentiation of human peripheral blood B lymphocytes
Authors: Scharenberg John G. M. , Spaapen Leo J. M. , Rijkers Ger T. , Duran Marinus , Staal Gerard E. J. , Zegers Ben J. M.
Published: February 2007

Deoxyguanosine (dGuo) has been implicated as the toxic metabolite causing a severe impairment of cellular immunity in children with a genetic deficiency of purine nucleoside phosphorylase (PNP). In peripheral blood T cells of normal donors both the pathway which leads to phosphorylation of dGuo (ultimately resulting in deoxyguanosine triphosphate, dGTP) and the salvage pathway which starts with degradation of dGuo by PNP (resulting in the formation of guanosine triphosphate, GTP) contribute to the inhibition of proliferation. In normal peripheral blood B cells, addition of dGuo leads to an inhibition of proliferation and differentiation. The concentrations of dGuo needed to cause a 50% inhibition are equivalent for peripheral blood T cells and B cells. Inhibition of B cell differentiation can be observed at the level of intracytoplasmic as well as secreted Ig and concerns all Ig isotypes. The early phase of B cell activation which takes place during a 24‐h preculture with formalinized Cowan I Staphylococci is not affected by dGuo; it is not until proliferation and differentiation of B cells, brought about by culturing in the presence of crude concanavalin A supernatant, occurs that inhibitory effects of dGuo become evident. Addition of dGuo to B cell cultures results in an intracellular accumulation of GTP and dGTP. Addition of 8‐aminoguanosine, a PNP inhibitor, next to dGuo, completely prevents the dGuo‐mediated inhibition. Under these circumstances the dGuo‐mediated increase in intracellular GTP is abrogated while dGTP accumulation still occurs. This indicates that the inhibitory effect of dGuo on the proliferation and differentiation of peripheral blood B lymphocytes of normal donors is independent of dGTP accumulation.
Analysis of immunoglobulins expressing the Vx21E and Vx21D gene products. Evidence for multigenic control of the expression of VH−VL combinations
Authors: Primi Daniele , Devaux Christian , Pierres Michel , Barbier Eliane , Cazenave Pierre‐André
Published: February 2007

The aim of this work was to study the genetic control of VH−VL combined expression. To this end immunoglobulins expressing the Vx21E and Vx21D gene products were isolated from the normal sera of several inbred strains of mice using a monoclonal antibody that selectively reacts with Vx21E and Vx21D subgroups. Analysis of the isoelectric focusing (IEF) pattern of the IgG heavy chains of these immunoglobulins obtained from H‐2‐congeneic mice revealed as expected that Igh‐linked loci strongly influence VH−VL combined expression. The IEF pattern of Vx21D and Vx21E‐associated heavy chains isolated from recombinant inbred mouse strains, however, revealed different phenotypes from either parental strains in strains in which both H‐2 and Igh loci segregated from the same parent. These data, therefore, strongly suggest that the expression of VH−VL combinations is controlled in part by genetic loci which segregate independently from those linked to H‐2 and Igh.
Interferon‐γ induces light chain synthesis in interleukin 2 stimulated human B cells
Authors: Bieh‐Thuy Lě Thi , Queen Cary , Fauci Anthony S.
Published: February 2007

Human B cells were cultured without added lymphokines, with interleukin 2 (IL 2) or interferon‐γ (IFN‐γ) alone, or with a combination of IL2 and IFN‐γ. Treatment with IL2 alone induced differentiation of the B cells, as shown by an increase in the number of immunoglobulin (Ig)‐containing cells and plaque‐forming cells, and by a larger amount of Ig secreted into the medium. In most of the induced Ig‐containing cells, heavy chains but not light chains were detectable by cytoplasmic staining. Treatment with IFN‐γ alone did not stimulate B cell differentiation. However, a combination of IFN‐γ and IL2 was more effective than IL 2 alone in inducing B cell differentiation, as measured by further increases in the number of plaque‐forming cells and in total Ig secretion. Furthermore, after treatment with both 1L 2 and IFN‐γ, most cells that contained cytoplasmic heavy chains also contained cytoplasmic light chains. We conclude that IFN‐γ acts in synergy with IL2 in B cell differentiation by enhancing light chain synthesis, leading to secretion of Ig molecules.
Is the antibody response specific?
Authors: Etlinger Howard M.
Published: February 2007

Introduction of heterologous immunoglobulin (Ig) or red blood cells into mice results in an increased production of Ig and immunogen‐reactive antibody. Since the increase in both total Ig and specific antibody is similar, it is concluded that the antibody response is specific. This result, which contrasts with those of previous studies in which the production of large amounts of nonspecific Ig was reported, suggests that in vivo T‐dependent B cell activation and differentiation to plasma cells requires the presence of immunogen and linked recognition.
The human immune response against major allergens from house dust mite, Dermatophagoides pteronyssinus: II. Idiotypic cross‐reactions of allergen‐specific antibodies
Authors: Saint‐Remy Jean‐Marie R. , Lebrun Philippe M. , Lebecque Serge J. , Masson Pierre L.
Published: February 2007

Antisera were prepared in rabbits against the idiotypic (Id) determinants of anti‐allergen antibodies. These antibodies were isolated from the plasma of 10 unrelated patients by immunoabsorption. Three major allergens isolated from the house dust mite, Dermatophagoides pteronyssinus (DPT), were used. The anti‐Id antisera were rendered specific by successive absorption on insolubilized allergens, IgG and IgM from a pool of human sera and Ig from the donor after removal of anti‐DPT antibodies by absorption.

Anti‐Id antibodies raised essentially against IgG antibodies were found to inhibit the agglutinating activity of both IgG and IgM anti‐DPT antibodies toward allergen‐coated latex. These anti‐Id antibodies were also able to inhibit partly the binding of radiolabeled allergen to IgE anti‐DPT antibodies. Matching the anti‐Id antisera with the various individual anti‐DPT antibodies revealed an Id cross‐reactivity of about 71% for IgG, 61% for IgM and 62% for IgE. A second and predominant type of anti‐Id antibodies recognized bystander idiotopes as the allergen did not inhibit Id‐anti‐Id reaction. These idiotopes were apparently more specific to individuals (private) as the anti‐DPT antibodies of a given patient did not agglutinate latex particles coated with anti‐Id antibodies prepared against Id of another patient.
Alteration of the T cell self‐specificity repertoire by treatment with anti‐la antibody during embryonic life
Authors: Kimoto Masao , Kishimoto Susumu
Published: February 2007

The effect of anti‐la antibody on the development of murine neonatal self‐Ia‐reactive T lymphocytes in the thymus was studied. A C57BL/6 (B6) mouse pregnant with (B6 ± C3H/He)F1 (B6C3F1) embryos was injected with monoclonal anti‐I‐Ek(mαI‐Ek) antibody every other day starting from the 12th day of gestation. The mixed leukocyte culture reaction (MLR) responses of thymocytes from mαI‐Ek‐treated neonatal mice were assayed against adult B6 (H‐2b), C3H/H3 (H‐2k) or BALB/c (H‐2d) splenic stimulator cells. The syngeneic MLR response of mαI‐Ek‐treated B6C3F1 neonatal thymocytes against C3H stimulator cells was augmented compared to that of untreated neonatal thymocytes. Experiments utilizing recombinant inbred strains of mice as well as monoclonal anti‐Ia antibody blocking experiments suggested that the antigen(s) responsible for this augmentation of syngeneic MLR resided in the I‐Ek molecule. The MLR response against B6 or BALB/c stimulator cells was not influenced by mαI‐Ek treatment. We speculate that mαI‐Ek antibody blocked the expression of I‐Ek molecules in B6C3F1 embryos and this resulted in the alteration of T cell self‐specificity repertoire in the thymus. Our results support the concept that Ia molecules play important roles for the elimination of self‐reactive T lymphocyte clones in the thymus.
B cell activation by the nontransforming P3HR‐1 substrain of the Epstein‐Barr virus (EBV)
Authors: Hu Cheng‐Po , Aman Pierre , Masucci Maria‐Grazia , Klein Eva , Klein George
Published: February 2007

The P3HR‐1 substrain of Epstein‐Barr virus does not transform B cells. This defect is known to be determined by the loss of the coding sequence for the nuclear antigen EBNA‐2. The virus can attach to and enter resting B cells. The initial events after EBV infection are reminiscent of those induced by polyclonal B cell activators. Similar to the effect of these, P3HR‐1 virus lowers membrane IgD expression on B cells and abrogates the transient elevation of activation markers BB‐1 and LB‐1 induced by the culture conditions. An important event of B cell activation is the acquisition of competence to respond to specific growth factors produced by T cells. This was induced by the P3HR‐1 virus. The infected B cells had elevated [3H]thymidine incorporation when exposed to the supernatant of PHA‐treated T cells. The EBV receptor is identical with the complement receptor CR2. Ligand binding to CR2 has been shown both with mouse and human B cells to deliver certain activation signals. Therefore, it is possible that the early step of activation by EBV is initiated through the binding to the receptor and is thus a cell surface event.
A novel approach for preparing anti‐T cell receptor constant region antibodies
Authors: Traunecker Andre , Dolder Beatrice , Karjalainen Klaus
Published: February 2007

To obtain antibodies against the individual chains of the T cell receptor (TCR) complex, we have produced chimeric proteins containing domains from immunoglobulin (Ig) and TCR polypeptide chains. Basically, the Ig light chains were used as carriers for the TCR constant (C) region domains. The exons which encode the main body of the C regions of the α, β and the related γ polypeptide chains were “engineered” into the intronic region between the rearranged Ig variable (V) region and Cx region genes. All three chimeric genes were expressed in myeloma cells, and the proteins of expected apparent molecular weight were produced. Secreted proteins containing the Cβ domain were purified from the culture supernatant by using anti‐x antibody affinity columns, and two rabbits were then immunized with the purified protein. Both rabbits produced antibodies able to immunoprecipitate the heterodimeric TCR protein.
Repertoire of murine λ‐positive variable domains: polyclonal induction of λ isotypes and their associated pattern of antibody specificities
Authors: Sanchez Pierre , Cazenave Pierre‐André
Published: February 2007

The diversity of λ variable (Vλ) domains is extremely restricted when compared to those of VH and Vλ. In addition, each Vλ domain is determined by the Cλ domain, For these reasons, the λ system is an excellent model for the study of the associated VH region repertoire. The study of Vλ domain diversity has been limited by the small contribution (approximately 5%) of λ‐bearing proteins to the total Ig pool. We now show that treatment of BALB/c mice with rabbit anti‐λ1 antibodies coupled to lipopolysaccharide induces a production of polyclonal λ1 light chain‐bearing Ig whereas, conversely, treatment with rabbit anti‐λ2 antibodies induces a production of polyclonal λ2 + λ3 light chain‐bearing Ig. The antigenic specificities of these two distinct λ populations were then studied using B1355 dextran, (4‐hydroxy‐3‐nitrophenyl)acetyl (NP) and 2,4‐dinitrophenyl (DNP) as antigens. The anti‐α(1‐3)dextran antibody specificity was found to be mediated exclusively by antibodies bearing the λ1 isotype whereas the anti‐NP and anti‐DNP antibody specificities are mediated by both the λ1 and λ2 + λ3 isotypes. In addition, various mouse strains with the VHa or VHb allotypic haplotype and the r or r phenotype were treated with rabbit anti‐λ1 antibodies. The λ1 anti‐NP and anti‐DNP antibody specificities were similar in all strains whereas the λ1 anti‐α(1‐3)dextran specificity was linked to the presence of the VHa allotypic haplotype. The mouse strains with the r or r phenotype did not differ in terms of their patterns of λ1 antibody specificities.
Differentiation antigens on rhesus monkey lymphocytes I. Identification of T cells bearing CD3 and CD8, and of a subset of CD8‐bearing cells
Authors: Nooij Frans J. M. , Borst Jannie G. , Van Meurs Gerard J. E. , Jonker Margreet , Balner Hans
Published: February 2007

Rhesus monkeys provide an excellent preclinical model to test the effect of monoclonal antibodies (mAb) in vitro and in vivo. So far, mostly mAb have been used which were originally raised against human cell surface antigens but cross‐reacted reasonably well with homologous antigens on rhesus monkey cells. However, to optimize the model, it was necessary to produce mAb which react specifically with subsets of rhesus monkey lymphocytes. In this report, three mouse anti‐rhesus monkey mAb are described, specific for different subsets of rhesus monkey T lymphocytes. None of the reagents cross‐reacts with human lymphocytes. Characterization of these mAb was based upon indirect immunofluorescence, using a simultaneous staining technique, and immunoprecipitation of the specific target antigens. One antibody (GM9) reacts with the same subset as is recognized by mAb specific for human CD8+cells. The second mAb (GM13) is specific for a subset of CD8+ cells. A third mAb (FN18) was of particular interest: it identifies a cell surface complex, RhT3, expressed on mature T lymphocytes, of which the polypeptide chains have a molecular mass of 22 and 27 kDa. The data strongly suggest that RhT3 is a CD3‐like determinant, so far unidentified in the rhesus monkey.
Heterogeneity of responsiveness of chronic lymphocytic leukemic B cells to B cell growth factor or interleukin 2
Authors: Hivroz Claire , Grillot‐Courvalin Catherine , Brouet Jean‐Claude , Seligmann Maxime
Published: February 2007

We compared the proliferative responses of purified human leukemic B cells from 12 cases of chronic lymphocytic leukemia to highly purified B cell growth factor (BCGF) and recombinant interleukin 2 (rIL 2) spontaneously, and in a coactivation assay using anti‐μ monoclonal antibodies. Heterogeneity of response to one or the other lymphokine was observed from case to case. Spontaneous responses were observed to BCGF in one case, to rIL 2 in 3 cases and to both lymphokines in one other case. Costimulation with anti‐μ monoclonal antibody induced a proliferative response to BCGF in 3 additional cases and to rIL 2 in 4 cases. Interestingly, cells from BCGF‐ responsive cases also proliferated in response to rIL 2. The leukemic B cells from 4 chronic lymphocytic leukemia patients were unresponsive. Cells from 5 cases expressed IL 2 receptors, although rIL 2 induced a direct proliferative response in only 3 of these cases. Expression of the B cell activation antigen B5 was associated to BCGF responsiveness.
Presence of IgE suppressor factors in human colostrum
Authors: Sarfati Marie , Vanderbeeken Yves , Rubio‐Trujillo Manuel , Duncan Doug , Delespesse Guy
Published: February 2007

In spite of intensive investigations, the ability of breast feeding to delay and to attenuate atopic diseases in children remains debatable. This study documents a mechanism whereby breast feeding might interfere with the synthesis of IgE by breast‐fed infants. Indeed, we show that colostrum contains IgE‐binding factors (IgE‐BF) capable of suppressing the in vitro synthesis of human IgE. Colostrum obtained from 15 donors was successively depleted of lipids and casein, filtered through Amicon XM50 membrane (mol. mass cut‐off 50 kDa) and lyophilized. IgE‐BF was demonstrated in such preparations by two different approaches, i.e. a classical rosette inhibition assay and Western blot analysis. In the first instance, lyophilized preparations of colostrum inhibited the binding of IgE‐coated bovine erythrocytes to IgE receptors on the surface of RPMI 8866 lymphoblastoid cells. The rosette‐inhibiting activity could be absorbed on IgE‐ but not on IgG‐Sepharose 4B and it could be recovered in the eluate of IgE‐Sepharose 4B. The molecular mass of IgE‐BF was comprised between 10 to 20 kDa as estimated by gel filtration through a calibrated Sephadex G‐75 column. After fractionation on 12% sodium dodecyl sulfate‐polyacrylamide gel electrophoresis and transfer to nitrocellulose membrane, colostrum displayed one band of 14 kDa and reacted with radiolabeled IgE but not with IgG nor IgM. This 14‐kDa band could be removed by absorbing colostrum with IgE‐ but not with IgG‐Sepharose 4B. Most importantly, the colostrum IgE‐BF suppressed the spontaneous in vitro synthesis of IgE by B lymphocytes derived from allergic donors without altering the production of IgM.
Administration of recombinant interleukin 2 to mice enhances production of hemopoietic and natural killer cells
Authors: Piguet Pierre‐Fragois , Grau Georges , Irle Claude , Vassalli' P.
Published: February 2007

Seven days of continuous perfusion of mice with human recombinant interleukin 2 (rIL 2) (∼ 3 × 104 U/day) increased the percentage of large mononuclear leukocytes (LML) among bone marrow, spleen, lymph node cells and liver interstitial cells (LIC). An increase in the lymphokine‐activated killer (LAK) activity was evident in these organs. The greatest increase in the number of LML and in the LAK activity was observed among the liver interstitial cells (about 500‐fold increase). The LML were nonphagocytic, Thy‐l+, sIg−, Ly 2+, L3T4− and asialo Gm1+. Perfusion of athymic nude mice, or of thymectomized, irradiated radiation chimera, showed that the Thy‐l+, LAK+ LML were the thymus and T lymphocyte‐independent progeny of Thy‐1−marrow precursors. The LML had no T cell function in a graft‐ vs. ‐host reactivity assay, neither did they have an inhibitory effect on T lymphocyte function in vivo. rIL 2 perfusion did not significantly affect the medullary hemopoiesis but did strongly enhance the extramedullary hemopoiesis, particularly within the interstice of the liver: the number of erythroid and myeloid cell was increased as well as the number of colony‐forming units per spleen and colony‐forming units per culture for various lineages (20‐50‐fold increment). These results show that in vivo, rIL 2 has a global enhancing effect on hemopoiesis together with a more selective influence on the production of LML.
Accessory molecules and T cell activation I. Antigen receptor avidity differentially influences T cell sensitivity to inhibition by monoclonal antibodies to LFA‐1 and L3T4
Authors: Regnier‐Vigouroux Anne , Blanc Dominique , Pont Suzanne , Marchetto Sylvie , Pierres Michel
Published: February 2007

A series of BALB/c‐derived T hybridoma cells, capable of producing interleukin 2 (IL2) in response to poly(Glu60, Ala30, Tyr10) (GAT) presented by syngeneic B lymphoma cells in the context of Ad restriction determinants, was used as a model system to evaluate the roles of LFA‐1 and L3T4 accessory molecules in antigen‐specific T cell activation. Examination of the antigen requirement for optimal IL2 responses revealed marked differences in the apparent avidity of these cells for GAT/Ad complexes. A relationship was observed between this parameter and the susceptibility of T hybridoma cells to inhibition by monoclonal antibodies directed at 5 distinct epitopes of LFA‐1, and at Aβd allodeterminants. In contrast, L3T4a‐specific monoclonal antibodies were found to block in a similar fashion the antigen‐specific IL2 responses of T hybridoma cells, regardless of the apparent avidity of their antigen receptors. It was also shown that both L3T4+ and L3T4− T hybridoma cells were capable of recognizing GAT plus Ad with high avidity. Thus, the quality of T cell antigen recognition appears to critically influence the involvement of LFA‐1, and only to a marginal extent that of L3T4, in antigen‐specific T cell activation. The implications of these findings are discussed in the context of recent data indicating that L3T4 may not only be an la‐binding protein.
An attempt to produce “pre‐T” cell hybridomas and to identify their antigens
Authors: Aihara Yukoh , Bühring Hans‐Jörg , Aihara Michiko , Klein Jan
Published: February 2007

With the aim of identifying some of the stages in the development of pre‐T cells (cells of the T cell lineage before they enter the thymus), we have produced a large number of hybridomas by the fusion of BALB/c bone marrow cells, bone marrow cells from BALB/c‐nu/nu mice, BALB/c fetal liver cells and BALB/c fetal thymocytes with the AKR thymoma BW5147. The hybridomas were selected for the expression of the Thy‐1.2 antigen of th3 normal celldonor and for their ability t9produce interleukin 2 (IL 2) upon co‐culture with irradiated normal spleen cells. A set of these hybridomas is described in this communication. The hybridomas were then used to immunize rats and to generate monoclonal antibody‐producing B cell hybridomas. Most, if not all, of the immunizing hybridomas were derived from pre‐T cells as evidenced by the fact that they produce IL 2, and express some of the T cell markers (the Thy‐1.2, Ly‐1, Ly‐2 or L3T4 antigens). The monoclonal antibodies were tested on a panel of pre‐T cell hybridomas and on normal cells obtained from spleen, lymph nodes, thymus and bone marrow. The testing was carried out by the microcytotoxicity assay and flow cytometric analysis. Three groups of antibodies could be distinguished. Some antibodies were broadly reactive, being positive with virtually all the clones in the pre‐T cell panel and with a substantial fraction of normal lymphoid cells. The identity of the antigens detected by these antibodies remains unknown but they do not seem to correspond to any of the known cell surface markers. Other antibodies reacted only with some of the pre‐T cell clones and did not react at all with normal lymphoid cells obtained from adult animals. Finally, other antibodies still reacted only with a minor subpopulation of thymocytes or of thymocytes and bone marrow cells, as well as some of the pre‐T cell clones; they did not react with spleen and lymph node cells. These antibodies might be specific for cells in the prethymic phase of the T cell differentiation pathway. They should prove useful for the identification of pre‐T cell markers and hence for the isolation of pre‐T cells and their functional analysis.