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Various immunologic functions of rabbit lymphocytes were studied during ontogeny. Liver cells of 17-day-old fetuses responded to concanavalin A and only after 22 days responded to phytohemagglutinin. The thymus cells of 22-day-old fetuses and the spleen cells of 29-day-old fetuses were stimulated by concanavalin A but not by phytohemagglutinin. The response to phytohemagglutinin in thymus and spleen cells was obtianed only after the birth. These results suggest that, in the rabbit, there are different subsets of T lymphocytes that respond either to concanavalin A or phytohemagglutinin. An alternative hypothesis is that the receptor for concanavalin A and the receptor for phytohemagglutinin appear on the same cells at different stages of the development. No stimulation of fetal liver was obtained with B mitogens, i.e., Nocardia water soluble mitogen (NWSM) and antisera against allotypes of the b series whereas neonatal spleen cells were stimulated. These results indicate that the ability of lymphocytes to respond to a nonspecific rabbit B cell mitogen, namely NWSM, and to a specific rabbit B cell mitogen, namely anti-allotypic antibody, matured only in peripheral lymphoid organs, such as spleen, in spite of the fact that these two mitogens stimulate different subsets of B lymphocytes. Significant synthesis of immunoglobulins by liver cells was obtained in 22-day-old fetuses. The liver cells possessed Fc receptors after 17 days of gestation and a significant antibody dependent cell-mediated cytotoxicity after 22 days. Both functions decreased after birth in these cells. These results showed a sequential acquisition of immunologic functions of rabbit lymphocytes.

Lymphoid cells bearing Fc receptors are able to lyse antibody-coated animal target cells in an antibody-dependent cell-mediated cytotoxicity reaction. The results presented here show that liposomes consisting of sphingomyelin, cholesterol, and cardiolipid and coated by cardiolipidic antibodies could be destroyed by spleen or thymus cells. No alteration of liposomes was observed when normal rabbit serum was used or when the effector cell population was depleted of cells bearing Fc receptors. The lysis of antibody-coated liposomes by effector cells could be carried out in two steps. In the first step, the fixation of antibodies on the cardiolipidic antigens could lead to a reorganization of the liposomal membrane. In the second step, the effector Fc-receptor-bearing cells might amplify this alteration of the liposomes.

The membrane expression of the J558 idiotype on B and T lymphocytes was studied in (A/J x BALB/c)F1 and (CAL.20 x BALB/c)F1 mice who were maternally suppressed for this idiotype. Animals suppressed in this fashion exhibit a chronic inability to synthesize J558 Id bearing antibodies in response to an antigenic challenge by alpha 1,3 linked dextran. However, by immunofluorescence, we observed that these maternally suppressed animals actually exhibited a considerable increase of B lymphocytes, which bear endogenously synthesized membrane immunoglobulin expressing the J558-IdI. Furthermore, a subset of T cells emerges that we have previously described as Lyt2.2+ cells exhibiting the J558-IdI determinant, which are capable of transferring the suppression to naïve recipients [1]. The goal of the experiments described in this communication was to thoroughly characterize those B cell clones that are expanded during idiotype suppression. Therefore, we immortalized these J558 Id+ B cells from idiotype suppressed mice as hybridomas. Four hybridomas secreting monoclonal antibodies bearing the J558 IdI but devoid of any specificity for alpha 1,3 linked dextran were obtained. It was established that these monoclonal antibodies bear the J558-Id by two criteria; namely, their direct binding to a panel of monoclonal anti-J558 IdI and a single anti-J558 IdX antibody, as well as by blocking the interaction of J558 with these antibodies. Notable was the fact that they all bore kappa light chains, even though in anti-dextran antibodies the expression of the lambda light chain is required for the expression of this idiotype. These results reaffirm the notion that idiotypic determinants are three dimensional structures which can be reconstructed by a variety of heavy and light chain sequences. Furthermore, an idiotope used in the regulation of an immune response (as our data implicate the J558 IdI is) is borne by antibodies displaying diverse specificities, which are coordinately regulated through this common idiotope.

The lymphocytes are coated by an external glycoprotein layer the so-called cell coat. There is some difference between the structure of the cell coat of T- and B-derived lymphocytes. The cell coat represents the chemical basis of some lymphocytic properties, i.e. their homing and electrophoretic mobility and it plays and important role in the expression of antigenicity of lymphocytes and the triggering of blast transformation.

Inhibition of circulating antibody production by steroids has been repeatedly shown in steroid-sensitive animals such as mice or rabbits. It is well known that after a single injection of 5 mg of cortisone acetate, mouse spleen cells loose their ability to be transformed by B cell mitogens such as lipopolysaccharides or Nocardia water soluble mitogen. Therefore it was of interest to study in the rabbit the influence of cortisone on the reactivity of spleen cells to a B cell mitogen (blast transformation and polyclonal strimulation) and their ability to be cytotoxic in the antibody-dependent cell-mediated cytotoxicity assay. Our data demonstrate that the cells which mediate such a cytotoxicity are cortisone-resistant whereas the B lymphocytes--which can be transformed and polyclonally activated by these mitogens--are sensitive to cortisone treatment.

Five cell lines were isolated after prolonged culture of human buffy coat leukocytes. The cells consisted for the most part of lymphoblasts morphologically similar to those produced by in vivo or in vitro stimulation of small lymphocytes by antigens or mitogens. One of the lines possessed virus-like particles of the Epstein-Barr type. All the established lines were capable of mediating antibody-dependent cytotoxicity (LADC) quantitatively at a level not significantly different from that mediated by freshly isolated, normal human peripheral blood lymphocytes (PBL). The LADC of the lymphoblasts was inhibited by all the human IgG subclasses tested (IgG1, IgG2, IgG3, IgG4) as was the LADC of PBL. It is concluded that, at least so far, the lymphoblasts have retained the properties of the lymphocytes from which they were originally derived as regards their ability to mediate LADC.

The Freeman polysaccharide - the hapten of lipopolysaccharide - induced a blast transformation of mouse and rabbit spleen lymphocytes. The magnitude of the response in rabbit was quite similar to that obtained by lipopolysaccharide whereas in mice it was weaker. The stimulation of lymphocytes of both species did not require the presence of macrophages. Similar results were obtained with four LPS and their correspondent haptens prepared from S. typhi-murium, S. abortus-equi, S. johannesburg R+ and R-.

Nocardia water soluble mitogen (NWSM) is known to stimulate mouse and rabbit lymphoid cells and to act selectively on murine B-derived lymphocytes. In this paper, evidence is presented that NWSM is also a mitogen for rabbit bursal equivalent cells and does not bring about blast transformation of thymus-derived rabbit cells. Pretreatment of rabbit spleen lymphocytes with antibody directed against rabbit thymus lymphocyte antigen (anti RTLA serum) and with complement did not affect the strong increase of thymidine incorporation which follows stimulation with NWSM. The mitogen-induced polyclonal activation of antibody-forming cells and resulted in the presence of 30% of cells with the ultrastructural characteristics of plasmocytes. These observations led to the conclusion that NWSM is a mitogen for a B-derived lymphocyte of the rabbit.

Macrophage inhibition factor (MIF) like substances were found in the supernatants of human lymphoid cultures stimulated by concanavalin A, tuberculin and allogeneic lymphocytes. These agents are usually regarded as elective stimulators of T lymphocytes. No such a phenomenon was shown when lymphocytes were stimulated by lipopolysaccharide or by goat anti-human F(ab')2 serum, which are known as preferential B-cell stimulators.

The in vitro synthesis of allotypes of b4/b5 offspring obtained from b4/b4 mothers immunized against paternal allotype b5/b5 was studied in comparison to similar offspring that had escaped from suppression and normal heterozygous b4/b5 rabbits. Nocardia water-soluble mitogen-a rabbit B-cell mitogen which is known to induce the differentiation of small lymphocytes into plasma cells and polyclonal activation of Ig, was able to break in vitro the allotypic suppression induced in vivo.

Following immunization with bacteria (i.e. Salmonella johannesburg), rabbit spleen lymphocytes developed a specific blast response when the lymphocytes were stimulated with polysaccharide, the haptenic moiety of lipopolysaccharide. A clear cut dissociation was noted in the blast response induced by polysaccharide compared with those induced by lipopolysaccharide and lipid A. There was no correlation between the magnitude of the cellular responses and that of the antibody response. Moreover, there was less specificity at the cellular level than at the level of antibody secreted by cells. A decrease of 3H-thymidine incorporation was often observed after immunization, at the level of peripheral blood lymphocytes. An inhibitory effect of these cells was shown on the blast response of spleen lymphocytes with polysaccharide. A high blast response to Salmonella polysaccharide which could be observed in some non-immunized rabbits might be related to a natural sensitization of animals with the same or related unknown antigens which could not be recognized by anti-S. johannesburg antibodies.

Immunization of humans with tetanus toxoid (TT) results in an inhibition of the reactivity of peripheral lymphocytes to TT as well as to one-way mixed lymphocyte reaction. The strongest inhibition was observed in cells taken 3 or 7 days after immunization. These reactivities reappeared on the 14th day and reached their initial levels by the 21st day after immunization. The role of the various cell types was studied by culturing separately purified T or B lymphocytes taken on day 7 after immunization. B cells alone are not stimulated by TT or allogeneic cells. Purified T cells are stimulated much more than the initial total cell population. When B cells were mixed and cultured along with T cells, a suppressor effect upon stimulation appeared which reduced 3H-thymidine incorporation to the levels of the initial population. Treatment with 5-bromodeoxyuridine followed by exposure to light of cells stimulated with TT abrogated their ability to respond to subsequent stimulation with TT; whereas the responsiveness of treated cells to allogeneic lymphocytes was not significantly affected and vice versa. These results suggest that each stimulator activated a separate population of T cells but that in vivo immunization provoked the development of both a specific and nonspecific suppressor activity. Most probably this suppressor activity was brought about by adherent, surface Ig-bearing (B) cells.