Biomedical subjects

D C Roopenian

Publications and source records attributed to D C Roopenian.

68 records · Page 4Linked to original sources

PubMed1989

Responses against antigens encoded by the H-3 histocompatibility locus: antigens stimulating class I MHC- and class II MHC-restricted T cells are encoded by separate genes.

The purpose of this work was to study the genetic basis of histocompatibility antigens encoded by the mouse minor histocompatibility (H) locus H-3. Both class I major histocompatibility complex (MHC)-restricted cytotoxic T lymphocytes (CTL) and class II MHC-restricted helper T cells (TH) specific for antigens encoded by genes within the H-3 locus were isolated and analyzed. Typing a number of mouse strains for expression of antigens recognized by these TH and CTL suggested that there was a different strain distribution pattern of expression of the antigens recognized by TH compared with those recognized by CTL. Separation of the genes whose products stimulate TH from those whose products stimulate CTL was suggested by: (1) analysis of the strain B10.FS(92NX)/Grf that has undergone recombination within the H-3 region; (2) genetic segregation studies of (B10.UW-H-3b/Sn x C57BL/10Sn)F2 mice; and (3) F1 complementation studies in which CTL specific for products of the TH-defined gene(s) could not be detected, even in the absence of immune responses to products of the CTL-defined genes. Taken together, these data suggest that in addition to two genes (B2m and Cd-1) within the H-3 region whose products typically stimulate class I MHC-restricted CTL, there is at least one additional gene whose product selectively stimulates class II MHC-restricted TH. This new gene is located telomeric from the CTL-defined genes and between the loci we and un on chromosome 2. These data demonstrate a novel degree of complexity of the H-3 "locus" and suggest selective presentation of minor H gene products in the context of class I or class II MHC proteins.

Animals↗

PubMed1988

Generation of helper cell-independent cytotoxic T lymphocytes is dependent upon L3T4+ helper T cells.

The role of L3T4+ (CD4+) Th cells in generation of CTL specific for discrete minor histocompatibility Ag was investigated. Suppression of the function of Th cells in vivo by chronic treatment with anti-L3T4 mAb prevented congenic strains of mice from being primed and from generating CTL specific for Ag encoded by the minor histocompatibility loci--H-3, H-1, and B2m. Analysis of proliferative responses and lymphokine secretion of cells from animals primed with one of these minor H Ag, beta 2-microglobulin, but not treated with anti-L3T4 antibodies, indicated that L3T4- class I MHC-restricted T cells were themselves responsible for the very great majority of the observed minor H Ag-specific proliferation and secretion of lymphokines associated with both T cell proliferation and activation of CTL. All together, the data indicate that in responses against discrete minor H Ag, L3T4+Th-independent CTL are generated through an L3T4+Th-dependent pathway.

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PubMed1988

Adoptive immunity in immune-deficient scid/scid mice. I. Differential requirements of naive and primed lymphocytes for CD4+ T cells during rejection of minor histocompatibility antigen-disparate skin grafts.

Using a model system in which mature lymphocytes were adoptively transferred into immunodeficient C.B17-scid/scid recipients, the requirement for CD4+ T cells in rejection of previously healed-in multiple minor-H-antigen-disparate skin grafts was investigated. Depletion of functional CD4+ cells was accomplished by anti-CD4 antibody + complement treatment prior to adoptive transfer followed by chronic anti-CD4 serotherapy in vivo. Homozygous scid mice harboring nondepleted naive donor cells effectively rejected minor-H-antigen-disparate grafts, whereas scid/scid mice harboring CD4+ T-cell-depleted naive cells did not. Homozygous scid mice harboring minor-H-antigen-primed cells rejected the minor-H-antigen-disparate allografts regardless of whether or not the animals had received anti-CD4 treatment, although rejection by the mice receiving anti-CD4 treatment was retarded compared to mice not receiving anti-CD4 treatment. All the mice that received viable donor cells rejected minor + H-2 disparate allografts, demonstrating effective immunity in this case was not dependent upon the activity of CD4+ T cells. These data suggest that in responses against multiple minor-H-antigen-disparate tissue, primary allograft rejection is absolutely dependent upon CD4+ cells, and secondary allograft rejection is not. The implications of these findings in understanding interactions of T cell subsets in vivo and of the utility of using homozygous scid mice as recipients for exploring the function of transferred lymphoid cell populations are discussed.

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PubMed1987

Increased expression of major histocompatibility complex antigens on lymphocytes from aged mice.

Many studies have reported age-related changes in immune responses that could be due to alterations in lymphoid cell numbers or functions. Here we report the results of studies using immunofluorescent staining and in vitro assays of cellular function to compare the expression of cell surface antigens on lymphocytes from mice up to 2 years of age. No significant changes were observed in the frequencies of spleen cells bearing class I or class II major histocompatibility complex (MHC) antigens, surface immunoglobulin, or Thy-1, Ly-1, Ly-2, or L3T4 antigens. However, the densities (per cell) of both class I and class II MHC antigens were increased significantly on cells from aged as compared to young mice, whereas the densities of the other cell surface antigens studied were unchanged or slightly decreased. The increased levels of MHC antigen expression in old relative to young mice were shown to be functionally significant regarding immunological stimulation. These data suggest that T-cell clones silent in young individuals may be activated in comparable situations in older animals, leading to immunological alterations perhaps including increased autoreactivity.

Aging↗

PubMed1985

Phorbol myristate acetate and in vitro T lymphocyte function. III. Selective impairment by PMA of lethal hit delivery by cloned CTL.

Phorbol myristate acetate (PMA) has little immediate effect on the lysis of antigenic tumor targets by the representative cytotoxic T lymphocyte (CTL) clones B6D/2-7c, 5MB6-5, and 5MB10-31. However, prolonged contact (24-48 hr) with PMA (10(-6)M) can profoundly depress the lytic activity of these and other cloned T lymphocytes. This concentration of PMA is neither toxic nor mitogenic for cloned T lymphocytes. B6D/2-7c cells that are treated with PMA lose some ability to bind tumor targets; however, the primary defect in PMA-treated B6D/2-7c cells appears to be at the level of lethal hit delivery, because cells remain essentially ineffective at tumor cell lysis in the presence of agglutinating lectin. Nonetheless, PMA-treated 5MB10-31 and B6D/2-7c cells continue to respond to the proliferative stimuli associated with alloantigens, especially in the presence of exogenous lymphokines. B6D/2-7c cells treated with PMA neither acquire the ability to suppress the cytolytic activity of untreated B6D/2-7c cells, nor undergo any significant alteration of Lyt-2 expression. PMA-induced loss of lytic activity is reversible, and cytolysis is reexpressed by PMA-treated B6D/2-7c cells if they are incubated with 2 degrees MLC SN, but not WEHI-3 SN. The reexpression of cytolysis occurs in the presence of cytostatic concentrations of cytosine arabinoside (ARA-C), indicating that cell proliferation is not required for this process. These data show that cloned CTL are capable of reversible cytotoxic function, and they establish the utility of PMA to probe requirements for expression of CTL function.

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PubMed1984

Responses against single minor histocompatibility antigens. II. Analysis of cloned helper T cells.

We have utilized a clonal approach to investigate functional and immunogenetic characteristics of T cells responsive to the miHA H-1.3. Our data prove the existence of H-2 I region-restricted helper T cells that are specific for H-1.3, or for determinants encoded by closely linked loci. These conclusions are based upon the following observations: 1) the cloned Th are driven to proliferate only by H-2 I-compatible stimulator cells that bear the appropriate (H-1.3) miHA; 2) antigenic stimulation causes the cloned Th to secrete lymphokine with high levels of IL 2-like activity; and 3) they are not specifically lytic for relevant target cells. These cloned Th were isolated at a rather low frequency from an H-1.3-immune MLC population compared with the observed frequency of H-1.3-specific Tc that proliferate autonomously after antigen stimulation; i.e., HITc isolated from the same MLC. They were, however, capable of promoting the clonal expansion of an H-1.3-specific HITc, suggesting that Th can collaborate with HITc in an anti-miHA response.

Animals↗

PubMed1984

Influence of PMA on T-lymphocyte responses to mitogenic lymphokines.

Phorbol myristate acetate (PMA), a reagent frequently used in vitro to bypass macrophage function and to elicit cytokines, has a variety of effects on the behavior of T lymphocytes in tissue culture. For example, PMA in concentrations greater than 10(-10)M is highly comitogenic with phytohemagglutinin (PHA) for C3H/HeJ thymocytes, and significantly potentiates lymphokine-induced proliferative responses of long-term MLC cells, cloned cytolytic T cells, and interleukin-2 (IL-2)-dependent T-cell lines. Since these activated T-lymphocyte populations are used to detect IL-2, PMA can interfere with accurate assessment of IL-2 concentration by quantitative bioassay. Further, if greater than 10(-10) M PMA remains as a contaminant in PMA-induced lymphokine preparations, it can mediate lymphokine-like biologic activity, and, therefore, obscure interpretation of experiments involving lymphocyte-lymphokine interactions.

Animals↗

PubMed1983

Analysis of cloned T cell function. II. Differential blockade of various cloned T cell functions by cyclosporine.

Cyclosporine has profound suppressive effects on selected in vitro functions of cloned T lymphocytes. Cyclosporine inhibits the antigen-induced proliferation of the helper T cell clone 12-11. The effective dose required to reduce this response by 50% (ED50) is 28 ng/ml. In contrast, the proliferation of clone 12-11 induced by exogenous growth factors in secondary mixed lymphocyte culture supernatant (2 degrees MLC SN), is relatively insensitive to cyclosporine (ED50 = 4600 ng/ml). Furthermore, cyclosporine abrogates both antigen-induced and mitogen-induced secretion of lymphokines by clone 12-11, indicating that cloned helper T cell function is sensitive to cyclosporine even when interactions between specific alloantigens and their cell surface receptors are bypassed with mitogen. The suppressive effect of cyclosporine is not limited to helper T cell clones. The cytolytic T lymphocyte (CTL) clone 5MD2-2 is also sensitive to cyclosporine. Again, cyclosporine (100 ng/ml) blocks the antigen-driven, but not the exogenous lymphokine-driven, component of clone 5MD2-2 proliferation. This suppression does not result from the occlusion of antigen receptors or from antigen deformation by cyclosporine, because clone 5MD2-2 remains capable of antigen-specific cytolysis in the presence of cyclosporine concentrations that can suppress its proliferation. Finally, the ability of clone 5MD2-2 to remove IL-2 activity from culture media, a function that is significantly enhanced by contact with specific alloantigen, is not influenced by suppressive cyclosporine concentrations.

Animals↗

PubMed1983

Helper cell-independent cytolytic T lymphocytes specific for a minor histocompatibility antigen.

We have isolated several anti-H-2Db-restricted cytolytic T cell clones specific for the minor histocompatibility antigen, H-1, from a long-term B10.129(5M) anti-C57BL/6 mixed leukocyte culture. All proliferate significantly, in the absence of exogenously added lymphokine, and in a specific manner to H-2Db-compatible stimulator cells that carry the appropriate H-1 minor antigen. Moreover, significant proliferation also occurs when T cell-depleted stimulator cells are used as the antigenic stimulus, indicating that the proliferative capacity of these clones is not the result of T cell factors produced by the stimulator cells. After concanavalin A stimulation, IL 2-like lymphokine activity can be detected from the one clone tested, 5M/B6-5. These data demonstrate the existence of antigen-driven, helper T cell-independent cytolytic T cell clones responsive to a single minor histocompatibility antigen. The implications of this finding are discussed.

Animals↗

PubMed1983

Phorbol myristate acetate and in vitro T lymphocyte function. II. Influence of PMA and supernatants from PMA-treated P388D1 cells on the proliferation of cloned T cells.

Neither the culture supernatants from P388D1 cells pulsed with phorbol myristate acetate (PMA), nor PMA itself in concentrations ranging from 10(-6) M to 10(-9) M, are directly mitogenic for murine T lymphocyte clones, yet both markedly augment the antigen-driven proliferation of many, but not all, cloned T lymphocytes. The component of PMA-induced P388D1 supernatant responsible for its co-mitogenic activity is probably PMA, rather than interleukin 1. For responsive clones, the co-mitogenic effect of PMA requires stimulator cells that display the specific allogeneic determinants recognized by the clones. This response persists after T cells are removed from the stimulating population, ruling out induction of mitogenic lymphokines from stimulator T cells by PMA as a primary mechanism for augmentation of clonal proliferation. Both splenocytes and thymocytes cooperate with PMA for enhanced clonal expansion, but heat treatment (45 degrees C, 45 min) of thymocytes destroys their cooperative capacity. PMA can also potentiate the lymphokine-driven proliferation of cloned T cells, indicating that PMA can, under certain conditions, influence T cell clonal expansion by a direct action on T lymphocytes.

Animals↗

PubMed1983

Response against single minor histocompatibility antigens. I. Functional and immunogenetic analysis of cloned cytolytic T cells.

A clonal approach was used to investigate the cellular basis of a T cell response to single minor histocompatibility antigens (miHA). This analysis was performed by functional and immunogenetic characterization of a large number of clones derived from short-term mixed leukocyte culture (MLC) populations generated against the miHA, H-1.3. Forty-nine clones isolated from such MLC were specifically cytolytic for H-1.3-bearing, H-2Db-compatible target cells. Thirty-seven of the 49 cytolytic clones were driven to proliferate when stimulated by spleen cells bearing the H-1.3 alloantigen in the absence of added T cell-derived growth factor(s) (GF). The remaining 12 clones proliferated only when GF was added. A strong positive correlation was observed between antigen-induced proliferation and the production of interleukin 2 (IL 2) activity. A similar correlation was observed when comparing the ability of both antigen and concanavalin A to induce IL 2 activity from the clones. These data suggest that i) antigen-driven or helper T cell-independent cytolytic T cells (HITc) are frequent components of an MLC response to a single miHA, and ii) the ability of HITc to undergo antigen-driven proliferation is related to their ability to produce antigen-induced GF.

Animals↗

PubMed1983

Phorbol ester mediates reversible reduction of cloned T lymphocyte cytolysis.

Prolonged contact with nanomole to micromole concentrations of the tumor promoter phorbol myristate acetate causes a significant reduction in the lytic activity of cloned cytolytic T cells. Diminished lysis is apparent even in the presence of agglutinating lectins. PMA does not exert this effect by minimizing clone viability. In fact, these concentrations of PMA cause significant potentiation of antigen-driven proliferation for many of these same clones. The PMA-mediated loss of cytolysis is reversible. Although contact with antigen does not induce or enhance reexpression of cytolysis, PMA-treated cytolytic T cell clones display normal cytolytic activity after contact with lymphokines.

Animals↗

PubMed1982

Analysis of cloned T cell function. I. Dissection of cloned T cell proliferative responses using cyclosporin A.

CsA interferes in a specific manner with the expansion of T cell clones in that it inhibits the antigen-driven component of the proliferative responses made by cloned helper T cells, cloned conventional cytolytic T cells, and cloned helper-independent cytolytic T cells. Cloned helper T cells and helper-independent cytolytic T cells, which share the ability to proliferate when cultured with specific alloantigen, fail to proliferate when cultured with specific alloantigen, fail to proliferate in response to this stimulus in the presence of CsA (10 to 100 ng/ml). In contrast, the proliferation observed when these cells are cultured with exogenous growth factors (but not alloantigen) is little influenced by as much as 1000 ng/ml CsA. When cloned helper T cells or helper-independent cytotoxic T cells are cultured with alloantigen plus exogenous growth factor, additive or synergistic proliferation occurs. However, CsA (10 to 1000 ng/ml) blocks only the component of proliferation induced by alloantigen, and leaves the lymphokine-driven component intact. CsA has similar effects on the proliferation of cloned conventional cytolytic T cells. Thus, CsA separates cloned T cell proliferation into two components: one driven by contact with alloantigens, the other driven by contact with mitogenic lymphokines.

Animals↗

PubMed1981

A new minor histocompatibility locus linked to H-3.

B10, BALB.B, and A.BY shared an allele at a new locus not shared by B10.LP-H-3b. This locus is linked to the minor histocompatibility gene, H-3, in the 5th linkage group of chromosome 2 of the mouse. The product of the new gene fits the criteria of a minor histocompatibility antigen, since it induced as well as served as a target for cytotoxic lymphocytes (CTL) and its presence accelerated skin graft rejection when compared with that caused by H-3.1. F1 complementation of skin graft rejection and generation of CTL indicated that A.BY and BALB.B do not share H-3a with B10 as previously reported but possess the c allele of their congenic partners A and BALB/c. Thus, B10.LP-H-3b and B10 differ from one another by at least 2 minor histocompatibility loci. The specificity of CTL developed in B10. LP-H-3b and appropriate F1 hybrids indicate that the lytic activity of CTL specific for H-3.1 was restricted to target cells possessing H-2Kb antigens; this contrasted to the H-2Db-restricted activity of CTL specific for the product of the new gene.

Animals↗