Gunther Dennert

Effects of cytotoxic monoclonal antibody specific for T200 glycoprotein on functional lymphoid cell populations

Abstract A monoclonal antibody against T200 glycoprotein is selectively cytotoxic for thymocytes and mature thymus-dependent (T) lymphocytes. All T-cell functions assayed, cell-mediated cytotoxicity, helper cell activity, and proliferation in response to T-cell mitogens or allogeneic cells were abolished by prior treatment of spleen cells with anti-T200 antibodies and complement. In contrast, thymus-independent (B) cell responses to lipopolysaccharide (LPS) and other B-cell mitogens were unaffected. Although treatment of spleen cells with anti-T200 antibodies and complement markedly reduced their capacity to mount an in vitro antibody response to sheep red blood cells (SRBC), responsiveness could be restored by the addition of SRBC-primed T-helper cells. Treatment of bone marrow cells with anti-T200 antibodies and complement did not eliminate either in vivo colony-forming units-spleen (CFU-S) or prothymocytes. It is concluded that T lymphocytes become sensitive to complement-mediated lysis by anti-T200 antibodies as a consequence of cell-surface modifications occurring shortly before or just after their entry into the thymus. In contrast to Thy-1 antigen, the selective killing by antibody against T200 glycoprotein cannot be readily accounted for by quantitative differences in the expression of T200 glycoprotein on the cell surface. Fluorescence-activated cell analysis showed that T200 glycoprotein was expressed in similar amount on the majority of all thymocytes, spleen, and bone marrow cells.

Retinoids and the Immune System: Immunostimulation by Vitamin A

Publisher Summary This chapter discusses retinoids and the immune system and describes immunostimulation by vitamin A. Retinol, given orally to mice, can significantly reduce the incidence of skin papillomas after 9,10-dimethyl-1,2-benzanthracene induction. Inhibition of carcinogenesis by retinoids in vivo can result from mechanisms similar to the ones operative in vitro. Transplantable tumors are used frequently to examine the protective effects of retinoids. Retinoids can also influence the cellularity of lymphoid organs by either decreasing cell numbers at toxic doses or increasing them at subtoxic doses. The histological changes in lymphoid organs, caused by retinoids, are reflected in their inhibitory or stimulatory effects on various immune responses. Thus, high doses of retinoids can inhibit both humoral and cell-mediated immunity, whereas subtoxic doses can stimulate them. Retinoids act in the induction phase of immunity. Retinoids stimulate the induction of T killer activity in vivo and in vitro both to allogeneic and syngeneic cells.

CLONED CELL LINES WITH NATURAL KILLER ACTIVITY

Publisher Summary This chapter examines the cloned cell lines with natural killer (NK) activity. The characterization of NK cells and the respective factors that modulate their cytolytic activity has been seriously hampered by inadequate methodology to separate NK cells from other classes of lymphocytes. While NK cell lines grow very rapidly in tissue-culture media containing T-cell growth factor (TCGF), they stop synthesizing DNA in normal medium and eventually die. Lymphokines contained in conditioned media, therefore, appear to be essential for the continuous proliferation and well being of these cells. In an attempt to delineate the lymphokines responsible for NK-cell proliferation and expression of cytolytic activity, a number of lymphokine preparations of various purities were tested. Results showed that preparations containing TCGF, like Con A conditioned media, supernatant of the cell line LBRM 33, or TCGF purified from LBRM 33 SN, stimulate NK-cell proliferation. In contrast, supernatants containing lymphocyte activating factor or interferon did not stimulate. Cell supernatants depleted of TCGF by absorption with cells failed to stimulate DNA synthesis.

Effects of IgM on the in Vivo and in Vitro Immune Response

Summary The effect of sheep erythrocyte specific IgM on the in vitro and in vivo immune response to SRBC is described. Kinetic data resulting from separate injection of antibody and antigen suggest that the stimulatory effect is mediated by an antigen-antibody complex formed in the peripheral circulation, and subsequently trapped in the spleen. If an optimal antigen dose is chosen, then IgM given with it does not improve the response, thus supporting the view that IgM works by increasing the spleen concentration of antigen given at suboptimal doses. In vitro studies with normal spleen cells show that high concentrations of IgM inhibit the sheep erythrocyte response. In experiments performed in vivo with the thymectomized animals, IgM does not substitute for the T cell component. However, in vitro experiments with T cell deficient spleen cells do show a small improvement of the SRBC response when low doses of IgM are given. This IgM stimulated response is much lower than the response obtained when B spleen cells are mixed with sensitized T cells. This work was supported by the Deutsche Forschungsgemeinschraft and SFB 74 as well as by a grant from the National Institute of Allergy and Infectious Diseases, No. AI-06544 to Dr. E. S. Lennox. I thank Miss S. Nase, Miss G. V. Hesberg and R. Austin for excellent assistance and Dr. K. Rajewsky and Dr. E. S. Lennox for stimulating discussions.

A study of Schistosoma mansoni reinfection in thymectomized rats.

Abstract Thymectomized irradiated rats were reconstituted with bone marrow from thymectomized donors subjected to chronic thoracic duct drainage. These animals (“B rats”) showed a profound suppression of T-cell activity in various in vitro and in vivo tests. B rats of the Fisher strain infected twice with S. mansoni did not exhibit any resistance to reinfection, while intact controls showed a significant reduction in the number of challenge worms. High levels of peripheral eosinophils were present in infected intact rats, while B rats showed only modest eosinophilia under the same conditions of infection. Granuloma formation and other cellular inflammatory reactions in the liver of infected animals were almost completely abolished in B rats.

Isolation and partial characterization of human T-cell growth factor

Human T-cell growth factor (TCGF) has been isolated from conditioned media of the Jurkat T-leukemia cell line. Using a high-efficiency isolation procedure involving hollow fiber concentration, gel filtration and 3 steps of reverse-phase HPLC we obtained 100 to 600 pmol TCGF per liter of conditioned medium. Jurkat cell-derived TCGF (jTCGF) has a molecular weight of 15,750. The amino acid composition of jTCGF agrees well with that derived from the cDNA sequence coding for this protein (Taniguchi et al, Nature 302, 305, 1983). jTCGF is highly active in vitro in stimulating the proliferation of T-cells as measured by 3H-thymidine incorporation into DNA (half-maximal stimulation with 3 fmol/100 microliters well).

H-2-restricted T-cell-mediated cytotoxicity: rediscovering allo-H-2 reactivity?

Abstract The phenomenon that strong syngeneic T-cell-mediated cytotoxicity is observed if killer, stimulator, and target cells share H-2 histocompatibility antigens is called H-2 restriction. Here a syngeneic model system making use of hapten-coupled stimulator and target cells is used to explore whether H-2 restriction is absolute or not. Using TNP-coupled spleen or tumor cells as stimulator or target cells in syngeneic and allogeneic situations, it is shown that neither the induction step nor the effector step of TNP-dependent killing is H-2 restricted. By varying the experimental assay conditions more or less H-2-restricted, TNP-dependent killing can be observed. For instance, suboptimal coupling of TNP to targets may result in H-2-restricted killing. Similarly, the use of spleen cell targets as opposed to spleen blast cells or tumor cells may result in H-2-restricted lysis. In contrast optimal coupling of TNP to sensitive target cells and coupling of TNP to cells with certain H-2 haplotypes may lead to significant TNP-dependent killing which is not H-2 restricted. Since hapten-coupled cells lacking H-2 are neither stimulators nor targets these results suggest that the T-cell receptor recognizes TNP-modified H-2 antigens simply as nonself-H-2. Thus hapten coupling of syngeneic cells appears to lead to a histocompatibility antigen change similar to the situation in an allogeneic cytotoxic reaction. Experiments are presented which support this view showing that TNP-coupled and uncoupled syngeneic or allogeneic stimulator and target cells cross-react. For instance allogeneic sensitization may lead to killing on TNP-coupled targets syngeneic to the effector cells and TNP-coupled stimulator cells syngeneic to the effector cells may induce killing on uncoupled syngeneic targets. TNP-dependent cytotoxicity can therefore be envisaged as a kind of allogeneic reactivity due to modification of H-2 antigens by the TNP coupling. This conclusion may have bearing on other model systems in which syngeneic killing appears to be H-2 restricted. In support of this possibility it is shown that allogeneic sensitization may lead to priming of memory cells able to respond to minor histocompatibility antigens.

FUNCTIONAL SPECIFICITY OF A PERMANENT T CELL LINE

ABSTRACT Mouse T cells selected in a one way mixed lymphocyte culture were maintained in tissue culture for three years by restimulation. These T cells proliferate only when stimulated with cells sharing H-2 antigens with the strain used for selection of the cell line. Intra H-2 mapping revealed that the IA k subregion codes for the majority of stimulating determinants(s). Cytotoxic effector function could be induced by specific alloconfrontation and was directed against targets carrying IA k coded antigens. This cell line was also able to induce a positive allogeneic effect by activating B cells in an in vitro primary humoral response to sheep erythrocytes. Helper activity was again specific for cells carrying IA subregion coded antigens. Supernatant factor(s) capable of substituting for the cell line could be generated by stimulation with cells expressing H-2 k region coded determinants. Such supernatants, however, were not strain specific in their activation of an in vitro primary humoral response to sheep erythrocytes.

Continuously Proliferating Allospecific T-Cell Lines

Studies of the function and specificity of thymus-derived lymphocytes (T cells) have met with increasing difficulties in recent years. The main reason is that T cells perform many different regulatory and effector functions in both cell-mediated and humoral immunity, each of which functions is apparently executed by a distinct subset of T cells. This was, for instance, shown by functional assays in which T cells primed for one function were assayed for other functions (Dennert, 1976). By combining functional assays with serological procedures, it was established that T cells with specific functions may display specific Ly antigens (Cantor and Boyse, 1975). This very promising approach to the separation of T-cell subsets has yet to be fully exploited, since it is difficult to prepare sufficient amounts of Ly-specific antibody for experimentation. But the recent success in preparing hybridomas producing specific antibody has opened new avenues for the preparation of unlimited amounts of specific antibodies (Kohler and Milstein, 1976). Other procedures aimed at enriching or even purifying T-cell populations have had little success. For instance, fractionation of T cells by adsorption to and elution from matrices to which antigen was attached have yielded disappointing results (Rubin and Wigzell, 1973), with the exception of alloantigen-specific cytotoxic T cells (Golstein et al., 1971).