Arthur H. Hale

H-2 antigens incorporated into phospholipid vesicles elicit specific allogeneic cytotoxic T lymphocytes

Abstract Different H-2 antigen-containing subcellular fractions were tested for their ability to elicit specific anti-H-2 cytotoxic T lymphocytes (CTLs). Intact cells and membrane vesicles were capable of eliciting strong anti-H-2 primary and secondary CTL responses. However, detergent-solubilized H-2 antigens partially purified with lentil lectin were greatly reduced in their capacity to elicit secondary anti-H-2 (CTLs) and at all amounts tested did not elicit a primary CTL response. Lentil lectin-purified H-2 antigens incorporated into egg lecithin plus cholesterol (30% w/w) vesicles elicited a strong secondary anti-H-2 CTL response and a low but significant primary anti-H-2 CTL response. The results also indicate that T-cell-defined specificities are closely associated with serologically defined private and public specificities.

A study of the inability of subcellular fractions to elicit primary anti-h-2 cytotoxic t lymphocytes.

Abstract An analysis was undertaken to understand the inability of H-2 containing plasma membranes and partially purified H-2 antigens incorporated into lipid vesicles to elicit primary anti-H-2 CTLs. It was found that in the presence of supernatants from concanavalin A stimulated spleen cells H-2 containing-subcellular fractions could elicit anti-H-2 CTLs. The result suggests that cytotoxic T lymphocyte precursor cells are available for interaction with alloantigens within the subcellular fractions and that the defect is the inability of these H-2 antigen-containing subcellular fractions to stimulate T helper activity.

The spatial relationship of the viral and H-2 antigens recognized by anti-viral CTLs

With the use of monospecific rabbit anti-G protein and mouse monoclonal anti-H-2Kk, we have analyzed the spatial relationship of the serologically defined H-2Kk antigens and the major surface glycoprotein (G protein) of vesicular stomatitis virus (VSV) to those antigens recognized by B10.A (k, d) anti-VSV cytotoxic T lymphocytes (CTLs). The ability of monoclonal anti-H-2Kk or rabbit anti-G protein to inhibit specifically the cytolytic activity of B10.A anti-VSV CTLs indicates that the G protein and the H-2Kk molecules are in close proximity to the viral and H-2Kk antigens recognized by the anti-VSV (CTLs. By the method of sequential immunoprecipitation, we also demonstrated that only 10–30 percent of the serologically defined G and H-2Kk molecules are in theG-H-2Kk complexes.

H-2 antigens incorporated into phospholipid vesicles interact specifically with allogeneic cytotoxic T lymphocytes

Abstract Subcellular fractions containing different H-2 antigens were tested for their ability to inhibit specific T cell-target cell conjugate formation. H-2-containing membrane vesicles, lentil-lectin-purified H-2 antigens solubilized with detergent (referred to in the text as high-density fraction) or incorporated into lipid vesicles, inhibited T cell-target cell conjugate formation effectively and specifically. However, two- to threefold more protein was required to inhibit T cell-target cell conjugate formation when detergent-solubilized lentil-lectin-purified H-2 antigens were tested. This suggests that a lipid matrix is advantageous for interaction with anti-H-2 T-cell receptors. Experiments were also undertaken to demonstrate specific binding of liposomes containing 125I-labeled H-2 antigen to anti-H-2-specific cytotoxic T lymphocytes (CTLs). The binding of the 125I-labeled H-2-containing liposomes was saturable and was specifically inhibited by unlabeled H-2 antigens. Monospecific anti-H-2 sera specifically inhibited the binding of liposomes containing H-2 antigen to the CTLs. The results suggest that a specific interaction can occur between serologically defined H-2 antigens and the receptor of anti-H-2 CTLs.

Molecular requirements for trinitrophenyl recognition by antihapten cytotoxic t lymphocytes.

Abstract The requirement of direct covalent association of trinitrophenyl (Tnp) groups with cell surface components for functional interactions with anti-Tnp cytotoxic T lymphocytes (CTLs) was analyzed. This question was approached by comparing the ability of two methods of trinitrophenylation to render cells susceptible to lysis by anti-Tnp CTLs. As previously shown, cells modified with trinitrobenzene sulfonic acid were susceptible to H-2-restricted lysis by anti-Tnp CTLs. However, cells incubated with Sendai virus covalently associated with Tnp groups, were not rendered susceptible to lysis by anti-Tnp CTLs. These same target cells, however, were susceptible to H-2-restricted lysis by anti-Sendai virus CTLs. Direct analysis of the number of Tnp groups on cells modified by either method indicates no significant difference in the number of Tnp molecules associated with the different target cells. The results suggest that direct covalent association of Tnp groups with cell surface specific components is a minimal molecular requirement for recognition and lysis by anti-Tnp CTLs.

Elicitation of primary cytotoxic T lymphocytes in nude mice

Abstract The presence of cytotoxic T-lymphocyte precursor cells (CTLp) within BALB/c nude mice was analyzed. Allogeneic T lymphocytes were injected intravenously into BALB/c nude mice. The resulting graft versus host reaction circumvented the thymus defect of the nude mice and induced the differentiation of CTLp into anti-H-2 cytotoxic T lymphocytes.

Effect of prostaglandins on elicitation of anti-viral cytolytic activity

We have studied the modulating effect of prostaglandins (PG) on the elicitation of secondary anti-vesicular stomatitis virus (VSV) cytotoxic T-lymphocytes (CTL) with viral antigens. The results indicate that PGE1 and PGE2 both inhibit the induction of anti-VSV CTLs by ultraviolet-inactivated VSV. Consistent with the result was the augmentation of anti-VSV CTL activity when VSV-primed spleen cells were cultured in the presence of indomethacin. These results suggest that PGs may modulate anti-viral CTL responses.

A study of the ability of H-2Kk-Iak containing subcellular fractions to elicit primary anti-H-2 cytotoxic T lymphocytes.

Abstract In order to identify an antigen required for elicitation of anti-H-2 cytotoxic T lymphocytes (CTLs), we have purified the H-2-Kk glycoprotein, incorporated it into a lipid vesicle, and tested it for its ability to elicit anti-H-2Kk CTLs. The results indicate that a lipid vesicle containing purified H-2Kk can elicit specific secondary anti-H-2Kk CTLs. In addition it was shown that in association with a partially purified Iak fraction the primary anti-H-2Kk CTL response was enhanced. It was also shown that Iak antigens alone could elicit an anti-H-2k CTL response. Although a low primary response was found with purified H-2Kk, it was observed that lipid vesicles containing both H-2Kk and Iak glycoproteins could elicit a significantly enhanced primary anti-H-2Kk CTL response. Lipid vesicles containing H-2Kk-Iak were tested for their enhanced capacity to elicit anti-H-2 CTLs as well as for their ability to elicit anti-H-2Kk CTLs in the presence of supernatants from concanavalin A-stimulated spleen cells.

Cytotoxic T lymphocytes specific for vesicular stomatitis virus recognize the major surface glycoprotein of VSV

Abstract The major surface glycoprotein (G) of vesicular stomatitis virus (VSV) was purified and incorporated into lipid vesicles containing the purified hemagglutinin-neuraminidase (HN) and fusion (F) glycoproteins of Sendai virus. These lipid vesicles were then used to modify and render target cells susceptible to lysis by anti-VSV cytotoxic T lymphocytes (CTLs). This result provides direct evidence that the G protein is a target antigen of anti-VSV CTLs.

Loss of reactivity of a myeloma tumor with H-2 compatible thymus-derived lymphocytes.

Abstract It was previously shown that MOPC-315-EL, a subline of the BALB/c myeloma tumor MOPC-315, reversibly alters its reactivity with T cells that recognize H-2d, 2,4,6-trinitrophenyl, and minor histocompatibility antigens. This report demonstrates (a) that CTLs directed against vesicular stomatitis virus and Sendai virus are unable to recognize viral antigens associated with the unreactive tumor cell, and (b) that incorporation of Sendai virus antigens into the same membrane as the H-2 gene products is required for effective recognition by cytotoxic T lymphocytes.