Transfer Of Contact Sensitivity Research Articles

Cyclophosphamide-modified murine peritoneal macrophages induce CD4+ T contrasuppressor cells that protect contact sensitivity T effector cells from suppression.

Cyclophosphamide (CY) is one of the most widely used alkylating agents in the treatment of various cancers and some autoimmune diseases. Numerous reports suggest that CY exerts immunoregulatory effects. Animal studies have shown CY affects contact sensitivity (CS) response by depleting CD4+CD25+ T regulatory cells and CD8+ T suppressor (Ts) cells. In a mouse model of CS, we previously showed that in vivo treatment with CY shapes the immunogenic/immunoregulatory balance of peritoneal macrophages. The aim of the current study is to verify if macrophages (Mf) from CY-treated mice are indeed able to induce immunoregulatory cells that could protect from suppression. Adoptive cell transfer of CS was used to examine immunomodulating properties of peritoneal Mf from CY-treated mice. Isolation of peritoneal Mf from animals that were (Mf-CY) or were not (Mf) treated with CY were cultured to identify cytokine repertoire. Further, we assessed spleen cell (SPLC) cytokine production following immunization with trinitrophenyl-conjugated Mf from donors treated (TNP-Mf-CY) or non-treated (TNP-Mf) with CY. In vitro experiments identified that Mf-CY produce more IL-6, TNF-α and TGF-β than naïve Mf. Further, immunization with peritoneal TNP-Mf-CY induces CD4+ T contrasuppressor cells (Tcs) cells that protect CS-effector cells from suppression. Higher IL-17A secretion was observed from TNP-Mf-CY-treated mouse SPLC compared to SPLC from TNP-Mf injected mice suggesting that this cytokine might be important in mediating contrasuppression in this model. Our results show that in vivo treatment with CY influences mouse peritoneal Mf to induce CD4+ Tcs cells that protect CS-effector cells from suppressive signals of Ts cells.

Role of IL-4 in delayed type hypersensitivity.

IL-4 plays a key role in the contact sensitivity skin reaction. This has several implications. First, the view that contact sensitivity (CS) is only mediated by cells with a Th1 profile of cytokine secretion needs modification, in the light of the essential role of IL-4 at the effector stage. Second, the concept of a single cell involved in the systemic transfer of CS is no longer tenable, as it is known that both alpha beta and gamma delta cells are required. Studies with the cell lines (which contain both alpha beta and a few gamma delta cells) suggest that this double requirement may involve the action of IL-4 on gamma delta cells, which bear receptors for IL-4. Finally, the view that T cell lines only transfer CS when injected locally, but not when injected intravenously (systemic transfer), is correct but incomplete, as T cell lines actually give systemic transfer of CS, providing the cell line or the recipient is treated with IL-4.

Three Cell Subsets Are Required for the Transfer of Delayed-Type Hypersensitivity Reaction by Antigen-Specific T Cell Lines

Antigen (trinitrochlorobenzene)-specific T cell lines were obtained by repeated stimulation of lymph node cells from immune mice with antigenin vitro.These T cell lines, consisting of more than 90% CD4+Vβ8.2+and 6 to 9% γδ+T lymphocytes, transfer contact sensitivity (CS) locally when injected at the same site as the challenge antigen, but fail to mediate a systemic passive transfer when injected iv. Injection of T cell lines together with spleen cells from mice immunized 1 day beforehand (1-day cells) allowed a successful, specific systemic transfer of CS. Phenotypic analysis showed that the 1-day immune cell was αβ+, γδ−, sIg−, CD3+, CD4−, CD8−, CD5+, B220 (CD45R)+, Thy 1.2+. The effect of 1-day immune cells occurred through a mechanism involving IL-4, as 1-day immune cells failed to allow systemic transfer of CS by T cell lines in recipient mice treated with mAb to IL-4. These observations strongly indicate that three cell subsets are required for the systemic passive transfer of CS by T cell lines: αβ+CD4+, γδ+, and a third cell subset, which is CD45R+, αβ+, CD3+, but double (CD4, CD8) negative.

Immune or normal gamma delta T cells that assist alpha beta T cells in elicitation of contact sensitivity preferentially use V gamma 5 and V delta 4 variable region gene segments.

In the current study, we confirmed previous findings suggesting that gamma delta T cells were involved in the successful adoptive cell transfer of contact sensitivity (CS) by alpha beta CS-effector T cells. In this study, we used hamster anti-mouse gamma delta-TCR mAb treatment of CS-effector T cells, followed by enrichment and removal of the gamma delta T cells with goat anti-hamster Ig-linked magnetic beads, or by addition of hemolytic rabbit C. This removal of gamma delta T cells abrogated adoptive cell transfers of CS, despite the presence of alpha beta T cells that are known to mediate CS. FACS analysis documented enrichment of gamma delta T cells rising from 1 to 2% of the starting cells, to 60 to 95% of the magnetic bead adherent cells. Adoptive cell transfer of CS was reconstituted by adding back to the alpha beta cells, highly enriched gamma delta cells attached to anti-gamma delta-TCR magnetic beads. Not only were gamma delta-enriched T cells from sensitized mice able to assist immune CS-effector alpha beta T cells, but gamma delta T cells from normal nonimmune mice also had CS-assisting activity, and furthermore, neither were MHC-restricted in this function. Thus, CS-assisting gamma delta T cells were present endogenously in normal mice without prior immunization, and acted without Ag specificity and without MHC restriction, to assist CS-effector alpha beta T cells. Similar studies, with hamster mAbs specific for V gamma and V delta portions of gamma delta-TCR, demonstrated that the gamma delta T cells that assisted the CS-effector alpha beta T cells preferentially expressed V gamma 5 and V delta 4 in their TCR. PCR analysis on extracted mRNA showed that V gamma 5 and V delta 4 gene segments indeed were rearranged and expressed in the sensitized and normal lymph nodes; and one-and two-color FACS analysis of magnetic bead-fractionated cells suggested that V gamma 5 and V delta 4 were expressed on the same T cells. In summary, these results demonstrated that V gamma 5+, V delta 4+, gamma delta T cells were needed to assist alpha beta effector T cells in the adoptive cell transfer of CS.

Gamma delta T cells in normal spleen assist immunized alpha beta T cells in the adoptive cell transfer of contact sensitivity. Effect of Bordetella pertussis, cyclophosphamide, and antibodies to determinants on suppressor cells.

Our prior studies showed that gamma delta T cells were required to assist alpha beta T cells in the successful adoptive cell transfer of contact sensitivity (CS) responsiveness. These TCR-gamma delta+ regulatory T cells in immune spleen and lymph node were CD3+, CD4-, CD8+, nonantigen-specific, and non-MHC-restricted. In the current work, experiments were conducted to determine the mechanisms of how the gamma delta T cells were required to assist the alpha beta T cells in CS. We found that similar regulatory gamma delta T cells were in the spleen of normal mice, but not in the spleen of nude nor SCID mice, suggesting that the regulatory gamma delta T cells were present before immunization and required the thymus for differentiation, and also required rearrangements of gamma delta V gene segments. Treatment of cell transfer recipient mice with Bordetella pertussis (Bp), or with a low dose of cyclophosphamide (50 mg/kg), restored the ability of alpha beta+ gamma delta- T cells to transfer CS. This and other results suggested that Bp caused the CS-assisting gamma delta T cells to leave the lymphoid organs (such as the spleen) and enter the circulation, and only then to be able to assist the TCR-alpha beta+ CS-effector T cells. This effect needed the simultaneous i.v. injection of the CS-effector alpha beta T cells and the CS-assisting gamma delta T cells. The results also suggested that treatment with cyclophosphamide inactivated suppressor T cells in the recipients that acted to inhibit the alpha beta T cell transfer of CS, and thus that the CS-assisting gamma delta T cells acted by protecting the CS-effector alpha beta T cells from this endogenous suppression. This suppression of CS transfers also was eliminated by treatment of recipients with two different mAbs to determinants on suppressor T cells. In conclusion, we have described regulatory TCR-gamma delta+ CS-assisting/protecting T cells that are non-antigen-specific, non-MHC-restricted, CD3+, CD8+ gamma delta T cells that may assist adoptive transferring CS-effector alpha beta T cells by making these effector T cells resistant to suppressor T cells in the normal recipients.

Nonspecific Regulatory Mechanism of Contact Sensitivity: Nonspecific Suppressor Factor Suppresses the Antigen-Presenting Activity of Dendritic Cells to Induce Contact Sensitivity

Nonspecific suppressor factor (NSF), which inhibits the passive transfer of contact sensitivity (CS), is produced from macrophage-like cells induced by oxazolone-conjugated spleen cells. NSF binds selectively to 1a-positive, cyclophosphamide-sensitive, and plastic-adherent non-T cells (named intermediate cells) present in the spleen. NSF-treated intermediate cells acquire the ability to suppress the passive transfer of CS nonspecifically. Because the phenotype of the intermediate cells is similar to that of antigen-presenting cells (APC) of macrophage lineage, we examined the effect of NSF on APC. NSF suppressed the APC activity of draining lymph node cells from mice sensitized with fluorescein isothiocyanate or picryl chloride (FITC-LNC or TNP-LNC) to induce CS. Suppressor molecules which suppress the passive transfer of CS and the APC activity were recovered in the same fraction around 43 kDa by dye-affinity and gel chromatography purification. The results that the suppressor activity of NSF was absorbed with lymph node cells or FITC-LNC, but not 33D1 or anti-1a monoclonal antibody plus complement-treated FITC-LNC indicated that NSF reacts directly with antigen-bearing dendritic cells in the draining lymph node. In contrast with spleen cells, lymph node cells did not have the intermediate function, i.e., NSF-treated LNC could not suppress the passive transfer of CS nor the ability of FITC-LNC to induce CS (APC activity). NSF still blocks the ability of FITC-LNC to induce CS after treatment with cyclophosphamide. Thus, the lymph node and spleen include distinct functional cell populations which express 1a antigen and interact with NSF. APC in the draining lymph node are downregulated directly by NSF and the intermediate cells in the spleen play a modulatory role in the suppressor pathway of NSF.

Immunodeficiency and other clinical immunology: Nedocromil sodium inhibits T-cell function in vitro and in vivo

Nedocromil sodium, a topical antiinflammatory agent recommended as a prophylactic regimen for asthma, is known to inhibit both allergic and nonallergic inflammatory processes in which an essential role for T cells has been implicated. Therefore the direct effects of this drug on several aspects of T-cell activity were analyzed in the present study. By using murine lymphocytes we found that NS at concentrations of 10 −8 to 10 −6 mollL inhibited the mitogen- or antigen-induced proliferative responses of these cells. It is interesting to note that higher concentratrions were ineffective. Preincubation of immune lymph node cells from contact sensitized mice with the drug abrogated their ability to transfer contact sensitivity to naive recipients, an effect that was found to be specific for the treated cells. Nedocromil sodium also interfered with the mitogen-induced interleukin-2 and tumor necrosis factor production by T cells and with their ability to adhere to the bound protein components of the extracellular matrix laminin and fibronectin. All these effects may be attributed to the inhibition of the increase of cytosolic calcium, which accompanies the early phase of T-cell activation and which is an essential step in inducing the aforementioned phenomena.

Down-Regulation of Murine Contact Sensitivity by Hapten-Amino Acid Derivatives

The low responsiveness of contact sensitivity (CS) in sensitized and challenged mice was induced by using hapten-amino acid derivatives. Subcutaneous injection of trinitrophenyl (TNP)-lysine or oxazolone-glycine inhibited ear swelling responses in mice that received rechallenge. The inhibitory effect was long-lasting and antigen-specific. TNP-lysine blocked the transfer of CS in vivo and the binding of immune lymph node cells to antigen in vitro. Thus, the suppression was mediated by inhibition of effector cells, but not associated with the generation of efferent limb-acting suppressor cells. The tool reported herein seems to be quite useful for controlling ongoing murine CS to simple chemicals.

Gamma delta T cells assist alpha beta T cells in adoptive transfer of contact sensitivity.

Cutaneous immune responses to contact sensitizers such as picryl chloride or oxazolone, are classical manifestations of T cell-mediated immunity in vivo. In fact, the first documentation of T cell-mediated immunity was the ability to adoptively transfer contact sensitivity (CS) responses. Although it is now clear that Ag/MHC-restricted alpha beta TCR positive effector T cells are responsible for 24 to 48 h CS responses, other subsets of Thy-1+ cells in mice also participate in the elicitation of CS. Thus, Thy-1+, CD5+, CD3-, B220+, hapten-specific, non-MHC-restricted early-acting cells are required to initiate CS responses by leading to local serotonin release, which allows for extravascular recruitment of the late-acting, alpha beta TCR+, CS effector T cells. This study describes another T cell population that is needed for the adoptive transfer of CS by alpha beta T cells. In vitro treatment of a mixture of CS effector cells with hamster mAb to gamma delta TCR, together with rabbit complement, or by panning on anti-hamster Ig-coated dishes, diminished substantially the subsequent transfer of CS reactivity without affecting either CS-initiating cells, or the later-acting, alpha beta TCR+ CS effector T cells. Immune cells treated with anti-alpha beta TCR mAb, or recovered as adherent cells from petri dishes after anti-gamma delta TCR panning (i.e., gamma delta TCR-enriched cells), reconstituted the ability of anti-gamma delta TCR-treated immune cells (i.e., alpha beta TCR-enriched cells) to transfer 24-h CS responsiveness. The phenotype of the gamma delta T cells that assisted CS effector alpha beta T cells was: CD3+, CD4-, and CD8+. The gamma delta T cells that assisted alpha beta T cells were not Ag-specific since anti-alpha beta-TCR-treated cells (gamma delta T-enriched) from picryl chloride immunized donors aided alpha beta T cells (anti-gamma delta TCR-treated) from oxazolone-immunized donors, and conversely gamma delta T cells from oxazolone-immunized donors aided alpha beta T cells from picryl chloride immunized donors. Furthermore, the CS-regulating gamma delta T cells were not MHC-restricted because gamma delta T cells from H2d or H2b donors could assist alpha beta T cells from H2k donors. It was concluded that a regulatory population of non-Ag specific, non-MHC-restricted gamma delta T cells was needed to assist immune effector, Ag/MHC-specific alpha beta T cells in the adoptive transfer of CS.

Nonspecific regulatory mechanism of contact sensitivity: Nonspecific suppressor factor (NSF)-treated intermediate cells produce a second nonspecific suppressor factor (NSF int)

Nonspecific suppressor factor (NSF), which inhibits the passive transfer of contact sensitivity (CS). is produced spontaneously from macrophage-like suppressor cells induced by intravenous administration of oxazolone (Ox)-conjugated spleen cells. NSF binds selectively to la-positive, cyclophosphamide (CY)-sensitive, and plastic-adherent cells (named intermediate cells) present in the normal spleen. NSF-treated intermediate cells acquire the ability to suppress the passive transfer of CS nonspecifically. In this study, NSF-treated intermediate cells were found to release a second nonspecific suppressor factor (NSF int) during a 2-hr culture, while retaining their suppressor activity. Investigation of the relationship between these two factors showed that both NSF and NSF int were trypsin-sensitive, nondialyzable proteins. However, gel chromatography revealed that NSF was about 43 kDa. while NSF int was about 20 kDa. NSF was released from macrophage-like suppressor cells after RNA-dependent protein synthesis. In contrast, production of NSF int was energy dependent but did not require protein synthesis. Intermediate cells pretreated with lysosomotropic agents, such as ammonium chloride or chloroquine, did not acquire suppressor activity nor release suppressor factors due to NSF treatment. These observations suggest that NSF int is an altered form of NSF released by the intermediate after having undergone some modification; the biochemical mechanism is not known. This study showed that the intermediate cells play an active role in the suppressor cascade of NSF.

Functional equivalence of cryptococcal and haptene-specific T suppressor factor (TsF): II. Monoclonal Anti-cryptococcal TsF Inhibits both Phagocytosis by a Subset of Macrophages and Transfer of Contact Sensitivity

Monoclonal anti-cryptococcal TsF (which inhibits phagocytosis by macrophages) and antipicryl TsF use the same two circuits to block the transfer of contact sensitivity (CS). Both arm macrophages which then release a macrophage suppressor factor (MSF) when exposed to antigen. This MSF depresses the transfer of CS. The evidence suggests that a single molecular species of TsF (MW ca. 70 kDa), which bears an antigen-binding site and I-J determinant, is responsible for MSF production and inhibition of phagocytosis. Anti-cryptococcal TsF also arms the T acceptor cell which then releases nsTsF-1 after triggering with a specific antigen (SCPA). This nsTsF-1, which depresses the transfer of contact sensitivity, was authentic, as shown by its I-J positivity (in contrast to MSF) and its role in the production of nsTsF-2. As anti-picryl TsF also inhibits phagocytosis, it was concluded that anti-cryptococcal TsF, originally detected by the inhibition of phagocytosis, and anti-picryl TsF, originally detected by inhibition of CS, are functionally equivalent.

Detection of immunosuppressive activity in human seminal plasma by an immunobioassay.

Recent evidence has shown that under in-vitro conditions human seminal plasma can interfere directly or indirectly with the function of cells of the immune system. It is however, questionable whether the results generated in vitro can be related directly to in-vivo activity. We have therefore standardized an in-vivo immunobioassay to detect the immunosuppressive property of human seminal plasma using adoptive transfer of contact sensitivity to a specific antigen such as dinitrofluorobenzene. Our results indicate that when sensitized lymphoid cells were incubated in vitro with human seminal plasma, their ability to transfer the delayed hypersensitivity in non-sensitized mice was suppressed or inhibited in comparison with the controls. The percentage suppression varied with different samples but the results indicate clearly that the immunosuppressive properties of human seminal plasma can be demonstrated using an in-vivo immunoassay.

IL-2 Influences the balance between immunity and unresponsiveness in the picryl (TNP) contact sensitivity system by blocking the development or action of an Lyt-2 +, I-J + T suppressor cell

Mice injected with antigen (picrylated spleen cells) intravenously fail to develop contact sensitivity. However, contact sensitivity occurs if these mice are injected with IL-2. This effect of IL-2 was reproduced in vitro by taking spleen cells 2 days after injecting antigen intravenously and culturing them with either 150 u/ml recombinant IL-2 for 2 days or by pulsing with 600–1200 u/ml IL-2 at 4 °C for 1 hr. After 2 days in culture these antigen-exposed cells transfer contact sensitivity to naive recipients in a 24-hr experiment. However, the ability of antigen-exposed cells, pulsed with IL-2, to transfer contact sensitivity is abolished when they are incubated with unpulsed antigen-exposed cells and as few as 1 16 of their number have a significant effect. This phenomenon is specific, as normal cells or cells from mice injected with oxazolonated cells intravenously have no effect. The suppressor cells were Thy-1 +, Lyt-1 −, 2 +, I-J + T cells. It was concluded that IL-2 prevents the development/action of antigen-specific T suppressor cells.

The role of suppression in immunoregulation: in vivo analysis using a monoclonal antibody to T suppressor factors.

We have used a monoclonal antibody (mAb)-specific for murine T suppressor (Ts) cells (mAb 14-12) to study the role of T cells in tolerance and immunoregulation. We demonstrate that mAb 14-12 can block in vivo Ts cell activity in a variety of experimental systems. It prevents the induction of Ts cells induced by i.v. injection of the water-soluble hapten 2,4,6-trinitrobenzene sulfonic acid, and the protein antigen bovine serum albumin. When 14-12 mAb is given prior to the i.v. injection of trinitrophenyl-conjugated spleen cells (TNP-SC) it blocks the induction of Ts cells and sufficiently overcomes suppression so that TNP-SC is able to induce immunity. mAb 14-12 can convert nonresponder mice into responders for the Ir gene-controlled response to the random terpolymer L-glutamic acid60-L-alanine30-L-tyrosine 10 (GAT), and can substitute for cyclophosphamide in overcoming a suppressor barrier in the adoptive transfer of contact sensitivity. Administration of 14-12 mAb just prior to immunization results in the augmentation of contact sensitivity, antibody and plaque-forming cell responses. These results demonstrate the versatility of this reagent for the study of Ts cell activity.

Contrasuppression and tumor rejection.

The growth of a highly progressive MCA-induced tumor 3152-PRO is dependent on the activity of suppressor T cells (Ts). Injection of syngeneic mice with antibodies specific for Ts leads to enhanced tumor transplantation resistance of the 3152-PRO tumor. In addition, injection of recipient mice with highly immunogenic regressor tumors conjugated with trinitrophenyl (TNP) activates a T cell population which also mediates protection to transplantation of TNP-conjugated 3152-PRO tumor cells. One such tumor, 1591-RE, was investigated in detail to determine the phenotype and biologic activity of this T cell population in overcoming Ts cell activity. Induction of transplantation resistance requires the presence of TNP hapten on both the highly regressive immunizing tumor (and not its progressor variant 1591-PRO4), and on the challenge tumor 3152-PRO. The cell population from TNP-1591-RE immunized animals which mediates protection against the transplantation of TNP-3152-PRO is Thy-1+, CD4+, 8-, Lyt1+, I-J+, and Vicia villosa lectin adherent, the identical phenotype to antigen-specific contrasuppressor T cells in the contact sensitivity (CS) response to TNP in vivo. A T cell population of identical phenotype from TNP-1591-RE immunized mice can overcome the effects of antigen-specific Ts cells on PCl-immune cells in the adoptive transfer of CS in vivo. These results suggest that immunoregulatory cells that mediate protection against progressive tumors may be identical in function to antigen-specific contrasuppressor T cells.

Soluble factors in tolerance and contact sensitivity to DNFB in mice: VII. Characterization of a monoclonal, efferent-acting suppressor factor with specificity for DNP/H-2K d

To study further soluble factors which regulate contact sensitivity (CS) 2 2 Abbreviations used: Ars, azobenzenearsonate; CS, contact sensitivity; DNBS, 2,4-dinitrobenzene sulfonate; DNFB, 2,4-dinitrofluorobenzene; DNP-BSA, dinitrophenylated bovine serum albumin; DNP-SC, dinitrophenylated spleen cells; DTH, delayed-type hypersensitivity; EBSS, Earle's balanced salt solution; FCS, fetal calf serum; LNC, lymph node cells; NMIg, normal mouse immunoglobulin; OXA, oxazalone; PBS, phosphate-buffered saline; SSF, soluble suppressor factor; T DH, delayed-type hypersensitivity effector T cells; TNCB, 2,4,6-trinitrochlorobenzene; TNP-BSA, trinitrophenylated bovine serum albumin; Ts, T suppressor cells. to 2,4-dinitrofluorobenzene (DNFB), hapten-primed spleen cells from BALB/c mice were used to make T-cell hybridomas. A hybrid constitutively producing a suppressor factor was identified and cloned (clone 3–10). Incubation of BALB/c DNFB immune lymph node cells (LNC) in the 3–10 supernatant suppressed the ability of the immune cells to transfer CS to DNFB. The passive transfer of CS to oxazalone or to 2,4,6-trinitrochlorobenzene (TNCB) was not suppressed by the 3–10 factor. The hapten specificity of the 3–10 factor further was demonstrated by the ability of DNFB immune LNC but not LNC from unsensitized or from TNCB-sensitized mice to adsorb the factor. The 3–10 factor also was adsorbed by DNFB-immune LNC from mice that were syngeneic with BALB/c mice at the K locus of the MHC (e.g., B10.D2 and D2.GD). Pretreatment of DNFB-immune LNC with monoclonal anti-K d antibody or with anti-DNP antibodies blocked the ability to adsorb the factor. These results indicated that the 3–10 suppressor factor binds to DNP/ H-2K d complexes on immune LNC. Nylon wool-purified T cells (83% Thy-1.2 +) from DNFB-immune LNC were able to adsorb the factor as well as unseparated immune LNC. Furthermore, treatment of immune LNC with anti-Thy-1.2 plus C′ abrogated the ability of the cells to adsorb the factor, indicating that the cellular target of the 3–10 factor is a T cell. In addition, treatment of the immune LNC with an autoantiidiotypic antiserum (CS 231) plus C′, which depletes DNP-specific delayed-type hypersensitivity effector T (T DH) cells, also abrogated the ability of the cells to adsorb the factor. Finally, the suppressor factor was adsorbed and eluted from DNP affinity columns but was not adsorbed by TNP affinity columns. Collectively, these results indicate that although the monoclonal 3–10 suppressor factor has affinity for DNP, focusing of the factor on the T dh cells requires recognition of DNP in the context of the appropriate MHC determinant, K d.

Mechanism of action of a T suppressor factor (TsF) in contact sensitivity: the T cell target for TsF activity in adoptive transfer of immunity is not effector cell.

The passive transfer of contact sensitivity (CS) by immune cells into normal animals requires the interaction of two distinct Ly-1+ T cells, one which is Vicia villosa lectin (VV)-nonadherent, the other which adheres to VV. Functional deletion of either cell type abrogates the adoptive transfer of CS into normal animals, whereas VV-nonadherent cells alone can transfer CS into animals pretreated with cyclophosphamide (Cy). An antigen-specific T suppressor factor, designated TNP-TsF, inhibits the transfer of CS into normal adoptive recipients. TNP-TsF mediates its suppressive activity by inducing an I-J+ subfactor (designated I-J2) from the assay population by the interaction of PC1-F (a TNP-binding subfactor of TNP-TsF) with antigen-primed Ly-2+ T cells. This I-J+ subfactor then complements TNBS-F (an antigen-nonbinding subfactor of TNP-TsF) to form an antigen-nonspecific effector molecule which suppresses DTH responses in an antigen-nonspecific fashion. We report here that TNP-TsF suppresses the adoptive transfer of CS into normal animals but not into animals pretreated with Cy. TNBS-F + I-J2, the effector complex of TNP-TsF, also suppresses the transfer of CS into normal but not Cy-treated animals. When the Ly-1 immune cells were separated into VV-adherent and -nonadherent populations, the TNBS-F + I-J2 suppressor complex suppressed the functional activity of the VV-adherent cell population, but not the VV-nonadherent cells. This suppressive activity correlates with the need for VV-adherent cells in the transfer of CS into normal but not Cy-treated recipients. When an I-J+ molecule (I-J1) from an SRBC-specific TsF was used in place of I-J2 to form a suppressor complex with TNBS-F, this TNBS-F + I-J1 TsF suppressed the transfer of CS into both normal and Cy-treated recipients. This difference in functional suppressive activity correlated with a difference in target cell specificity: TNBS-F + I-J1 suppressed the VV-nonadherent TDTH cell, whereas TNBS-F + I-J2 suppressed the VV-adherent T cell of CS. Immune cells which are transferred under conditions which do not require the VV-adherent cell for transfer are not suppressed by TNBS-F + I-J2 or TNP-TsF, but are suppressed by the TNBS-F + I-J1.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of benzene inhalation on lymphocyte subpopulations and immune response in mice

To clarify the immunotoxicity of benzene, the effects of benzene inhalation on T and B lymphocytes and immune responses in mice were examined, male mice were exposed to 50 or 200 ppm benzene vapor, 6 hr/day for 7 or 14 consecutive days. T and B lymphocytes, in blood and spleen, were detected by the cytotoxicity assay with anti-Thy-1.2 monoclonal antibody and the membrane immunofluorescence test with anti-immunoglobulin antibody, respectively. Humoral immune response to sheep red blood cells was determined by the hemolytic plaque-forming cell assay. Cell-mediated immune response was measured by contact sensitivity (CS) to picryl chloride. The activity of suppressor cells was evaluated in spleen by the suppressive effect on passive transfer of CS. The ratio and absolute number of T and B lymphocytes in blood and spleen were depressed after a 7-day exposure at 50 ppm benzene. The depression of B lymphocytes was dose dependent and more intense than that of T lymphocytes. The ability to form antibodies was suppressed by benzene at all exposure levels, but the CS response was resistant to benzene inhalation and rather enhanced at 200 ppm exposure for 14 days. The activity of suppressor cells could not be detected at this dose level. These data show that benzene inhalation effects on humoral and cell-mediated immune responses are a result of the selective toxicity of benzene to B lymphocytes and suppressor T cells.

Orally induced tolerance generates an efferently acting suppressor T cell and an acceptor T cell that together down-regulate contact sensitivity.

Intragastric administration of the hapten trinitrochlorobenzene (TNCB) suppresses development of contact sensitivity (CS) to attempted epicutaneous sensitization with TNCB. Suppression induced by feeding TNCB is hapten specific and can be transferred to normal animals with lymphoid cells from fed mice. The lymphoid cells in hapten-fed mice that cause suppression of CS have been identified as Thy-1.2-positive cells in spleen and mesenteric nodes. The suppression with Peyer's patch cells from hapten-fed mice appears to be attributable to cells bearing Thy-1.2 antigen (T cell) and to cells with surface Ig (B cell). Feeding TNCB induces an efferent-acting suppressor T cell (Ts eff), as well as an intermediary acceptor T cell (T acc) with which it interacts to block adoptive transfer of CS with immune cells. Ts eff emanating from hapten-fed mice was identified by its specificity for the hapten, insensitivity to pretreatment with cyclophosphamide (CY), ability to produce soluble suppressor factor (SSF), and requirement for T acc to be functional. The presence of T acc in hapten-fed mice, on the other hand, was confirmed by its sensitivity to treatment with CY, interaction with Ts eff or SSF, and the ability to produce nonspecific inhibitor of TDTH cells. Thus, the suppressor T cells that are induced by administering the hapten intragastrically appear to function much like the cells of the suppressor T cell cascade that are induced by giving hapten via parenteral routes.

Polyclonal B Cell Activators Inhibit Contact Sensitivity to Oxazolone in Mice by Potentiating the Production of Anti-Hapten Antibodies that Induce T Suppressor Lymphocytes Acting through the Release of Soluble Factors

Polyclonal B cell activators (PBAs) such as purified protein derivative, lipopolysaccharide, Staphylococcus aureus strain Cowan I (StaCwI), and Pseudomonas aeruginosa inhibit contact sensitivity to oxazolone in mice when given 24 h before sensitization. This suppression, transferable from immunodepressed animals to oxazolone-sensitized recipients with immune serum, has been shown to be due to the early appearance of anti-hapten antibodies. These antibodies elicit T suppressor cells which release soluble factor(s) capable of inhibiting the passive transfer of contact sensitivity.