T cell synergy in the primary MLR: proliferative kinetics, effector cell generation, and IL 2 production

A primary rat MLR was initiated, and on each of 8 consecutive days during the evolving culture, an aliquot of cells was separated into its constitutive helper/inducer (W3/25+) and suppressor/cytotoxic (OX8+) T cell subsets by a monoclonal antibody, Degalan-bead immunoadsorbent column technique. This allowed a detailed kinetic analysis of T cell proliferation, the generation of effector cells, and the production of IL 2 by each subset relative to net whole culture supernatant IL 2 activity. The primary MLR demonstrates an early period of helper/inducer cell proliferation, IL 2 production and accumulation, followed by a period of suppressor/cytotoxic cell (OX8+) proliferation and IL 2 consumption during which there are distinct waves of allospecific suppressor, followed by cytotoxic activity. If fresh T cells of the helper/inducer or suppressor/cytotoxic phenotype were preseparated and then cultured alone with irradiated allogeneic stimulator cells, proliferation was noted in both subsets despite no demonstrable IL 2 activity in cultures of the suppressor/cytotoxic cells. Finally, a suppressed primary MLR exhibited proliferative inhibition of both T cell subsets.     

Alloantigen-specific suppressor T cells are not inhibited by cyclosporin A, but do require IL 2 for activation

Alloantigen-specific suppressor T cells are activated from normal murine spleen cells in mixed lymphocyte reactions (MLR). These T cells are radioresistant and suppress the activation of cytotoxic T lymphocytes (CTL) in second primary MLR cultures. This report demonstrates that cyclosporin A (CsA) blocks the activation of these suppressor cells at a dose of 1 microgram/ml. However, reconstitution of CsA blocked cultures with IL 2 restores the activation of the suppressor T cells, but fails to significantly restore the activation of CTL in these same cultures. This differential activation requirement was used to establish T cell lines that demonstrate enriched suppressor cell activity but depletion of CTL activity. These findings are discussed in terms of the mechanism of action of CsA in these distinct T cell subsets and the relevance to models of allograft unresponsiveness.     

Soluble suppressor of T cell proliferation in primary in vitro response to murine minor histocompatibility antigens

Normal mouse lymphocytes are not capable of mounting a primary cytotoxic T cell response to Mls encoded, non H-2, allodeterminants, although a strong lymphoproliferative response is observed in primary MLR between Mls incompatible cells. In this study it is reported that in the supernatant of primary cultures between AKR macrophages and CBA/H lymphocytes (H-2 identical, incompatible for Mls and other minor antigens) a suppressor of T cell proliferation in MLR is detected. By contrast, a suppressor is not detected in supernatants from primary cultures between BALB/C macrophages and CBA/H lymphocytes (H-2 incompatible, Mls identical), B10.BR macrophages and CBA/H macrophages and CBA/H lymphocytes (syngeneic) suggesting that the production of the suppressor factor occurs only when an Mls incompatibility exists. The suppressive activity of the Mls incompatible culture supernatant upon MLR between incompatible macrophages and lymphocytes is neither antigen specific nor Mls or H-2 restricted, nor is it due to an irreversible toxic effect on T lymphocytes or macrophages. The inhibition of T cell proliferation could be explained by inhibition of IL 2 production, by blocking its union to T cells or by a combination of both effects. Our findings could help explain previous observations that lymphocytes from mice preimmunized with Mls incompatible cells have a depressed proliferative response as well as depressed cytotoxicity against alloantigens.     

Ia-restricted interaction between normal lymphoid cells and SJL lymphoma (RCS) leading to lymphokine production. I. Enhancement and suppression of antibody formation to TI antigens by supernatants and cells from cocultures of RCS and lymphoid cells

Addition of gamma-irradiated reticulum cell sarcoma (RCS) cells causes suppression of the antibody response to trinitrophenyl (TNP)-keyhole limpet hemocyanin (KLH)-primed syngeneic SJL spleen cells to TNP-polyacrylamide (PAA) in vitro. The response of anti-brain antigen (BAT) + C-treated spleen cells is not suppressed by gamma-RCS, but is suppressed by cells from 48-hr SJL lymph node or thymus + gamma-RCS cultures. Addition of as few as 2.5 x 10(5) cultured (anti-I-A + C treated to remove gamma-RCS) cells causes significant inhibition of the responses of both syngeneic and allogeneic spleen cells. Treatment of gamma-RCS-induced suppressor cells with anti-BAT + C reduces their suppressive activity. In contrast to the cells, supernatants (SN) from (lymph node (LN) + gamma-RCS) cultures greatly enhance, in an antigen-dependent fashion, the responses of untreated or anti-BAT + C-treated Sephadex G10-passed spleen cells to TNP-PAA. TNP-SIII polysaccharide, or TNP-Ficoll, but not as much to TNP-KLH. Addition of SN as late as Day 3 of culture still causes about half as much enhancement as leaving SN in throughout the culture period, but it has no effect if left with the spleen cells for only the first day of culture. SN contains high levels of IL-2 and IFN-gamma; absorption with cells from an IL-2-dependent cytotoxic T-cell line removes the enhancing activity, while treatment with pH 2 to remove the IFN-gamma has no effect. SN from an IL-2-producing T-cell line (LBRM-33) has a similar effect on antibody production to TI antigens as does SN of (LN + gamma-RCS). The results suggest a marked dependency of PFC responses to TI antigen on IL-2 in all strains examined, including SJL, LAF1, DBA/2Ha, and CBA/N, probably through a direct activation of B cells. The findings also suggest that suppressor T cells, induced by gamma-RCS in syngeneic lymphoid cells, absorb the IL-2 needed for responses to TI antigens in vitro.     

The plaque-forming-cell (PFC) response of human blood lymphocytes. II. PFC response by mixtures of allogeneic lymphocytes cultured with formalin-treated staphylococci

Formalin-treated Staphylococci (FSA) induce anti-SRBC PFC in cultures of human lymphocytes. Regulation of the PFC response induced by FSA in cultures containing lymphocytes from two allogeneic donors was studied. The PFC response observed in such cocultures could not be predicted from the responses of lymphocytes from the two donors cultured individually. The PFC response of approximately one half the cocultures was less than expected. The remaining cocultures generated more PFC than expected. The depression or augmentation in the PFC response which was observed in cocultures was reproducible when lymphocytes from the same pair of donors were cocultured. Cocultures containing lymphocytes from identical twins generated the expected PFC response. The data suggest that suppressor or helper activity may be generated during a “two-way” allogeneic mixed lymphocyte reaction (MLR). Much less deviation from the expected PFC response was observed during a “one-way” MLR. Anti-Ia antiserum treatment of either donor's lymphocyte population tended to eliminate the deviation from the expected PFC response in coculture. The data suggest that a feedback loop, involving cells from both donors, may be operating in the “two-way” MLR, which leads to the generation of suppressor or helper activity.     

A partial characterization of suppressor cells in rat fetal liver cells

Rat fetal liver cells (FLC) obtained at 18–20 days gestation suppressed mixed lymphocyte reactions(MLR) of adult lymph node cells. The suppression was not strain specific: both syngeneic and allogeneic FLC were capable of suppressing the MLR. The same suppressor activity was observed with fetal spleen cells but not with fetal thymus cells. Removal of phagocytic cells from FLC failed to inhibit the suppressor activity. The suppressor cells were separated into two different types by BSA density gradient: one is radiosensitive, the other radioresistant. A stronger suppressor activity was observed in radiosensitive cells. The suppressor cells belonged to the fraction agglutinated with peanut agglutinin. The data suggest that the suppressor cells in rat FLC may be a proliferating blastoid-type cell rather than mature lymphocytes or mature macrophages.     

Cell lines derived from human autorosette-forming cells with suppressive activity

Concanavalin-A-stimulated human T lymphocytes from healthy donors and from patients suffering from diverse immune disorders were fractionated into rosette-forming (R) and nonrosette-forming (NR) cells. The separation method is based upon the ability of the lymphocytes to bind autologous erythrocytes and form autorosettes. Long-term cultures of the R and NR subpopulations were established. The activity of the culture supernatants on the T cell proliferation of normal human phytohemagglutinin (PHA)-induced lymphocytes and of a murine, interleukin-2 (IL-2)-dependent cytotoxic T cell line (CTLL) was investigated. Only the R cell line-derived supernatants from almost all patients tested evinced potent suppressor activity, those from healthy donors less so. The suppressive function was demonstrated not to be due to a cytotoxic effect since preincubation of the PHA-induced lymphocytes and CTLL cells with the factor did not diminish their proliferative capacity. Our study indicates the existence of a competitive relationship between the suppressor factor and IL-2. We found that inhibition of the proliferation decreased with the addition of increasing quantities of exogenous IL-2. We also observed that preincubating the CTLL cells with IL-2 prior to exposing them to the suppressive factor precludes inhibition of their proliferation. Phenotypic analysis of the suppressor cell line revealed that they were comprised of a T cell population which included OKT4+ and OKT8+ cells and that 99% of the cells formed autorosettes. Preliminary purification of the suppressive factor was performed by ultrafiltration and maximal suppression was exhibited by the fraction of less than 10,000 daltons. The development of suppressor cell lines from the unique population of autologous rosette-forming cells may be very helpful in studying the immunoregulatory properties of these cells and their suppressor activity.     

IL-4 inhibits IL-2-mediated induction of human lymphokine-activated killer cells, but not the generation of antigen-specific cytotoxic T lymphocytes in mixed leukocyte cultures

Human rIL-4 was studied for its capacity to induce lymphokine-activated killer (LAK) cell activity. In contrast to IL-2, IL-4 was not able to induce LAK cell activity in cell cultures derived from peripheral blood. IL-4 added simultaneously with IL-2 to such cultures suppressed IL-2-induced LAK cell activity measured against Daudi and the melanoma cell line MEWO in a dose-dependent way. IL-4 also inhibited the induction of LAK cell activity in CD2+, CD3-, CD4-, CD8- cells, suggesting that IL-4 acts directly on LAK precursor cells. IL-4 added 24 h after the addition of IL-2 failed to inhibit the generation of LAK cell activity. Cytotoxic activity of various types of NK cell clones was not affected after incubation in IL-4 for 3 days, indicating that IL-4 does not affect the activity of already committed killer cells. No significant differences were observed in the percentages of Tac+, NKH-1+ and CD16+ cells after culturing PBL in IL-2, IL-4 or combinations of IL-2 and IL-4 for 3 days. IL-4 also inhibited the activation of non-specific cytotoxic activity in MLC, as measured against K-562 and MEWO cells. In contrast, the Ag-specific CTL activity against the stimulator cells was augmented by IL-4. Collectively, these data indicate that IL-4 prevents the activation of LAK cell precursors by IL-2, but does not inhibit the generation of Ag-specific CTL.     

In vitro propagation and cloning of murine natural suppressor (NS) cells

During a short period of time after birth or after radiotherapy, the spleens of neonatal and adult TLI-treated mice contain suppressor cells of the mixed leukocyte reaction (MLR) and of graft-vs-host disease. The present report shows that the MLR suppressive activity of spleen cells from TLI-treated adult BALB/c mice can be maintained in long-term tissue culture by using conditioned medium. The suppressor cells can be cloned by limiting dilution, and reproducibly inhibit the [3H]TdR incorporation in the MLR at responder-to-suppressor cell ratios of 50:1. There is no antigen specificity or H-2 haplo-type restriction of the MLR suppression. The suppressor cells do not inhibit [3H]TdR per se, because no inhibition was observed in co-culture experiments with the EL4 tumor line or the IL 2-dependent HT-2 cell line. By using immunofluorescent staining techniques, the surface phenotype of the suppressor cells was found to be similar to that reported previously for cloned NK cells (Thy-1+, Lyt-1-, Lyt-2-, Ig-, Ia-, MAC-1-, asialo-GM1+). However, the suppressor lines showed no natural killer activity when YAC-1 target cells were used. Thus, the suppressor lines have been termed "natural suppressor" cells to indicate surface marker similarities to NK cells, both in vivo and in vitro, but different effector functions.     

Cooperation between mouse T-cell subpopulations in the cell-mediated response to a natural poxvirus pathogen.

Irradiated CBA anti-DBA/2 cells (10⁶ cells/culture) suppressed the production of effector cells in cultures containing 10⁷ unprimed CBA (responder) and 10⁶ irradiated DBA/2 (stimulator) spleen cells per culture. The suppressive element was cellular and suppression was specific for the stimulating antigen. The suppressive activity resided in the cytotoxic cell population in that both suppressive and cytotoxic activities were found in cells of the same size range, predominantly in T-cells, were produced in response to similar doses of stimulator antigen, and were produced with the same time course following establishment of first sensitization cultures. Eventual suppression correlated with the cytotoxic activity introduced into second sensitization cultures by suppressor cells. The short-term cytotoxic activity and suppressor activity were both highly radioresistant. These studies indicate that the suppressor cells formed in an in vitro mixed lymphocyte culture are cytotoxic to stimulator cells.     

Inhibition of human natural killer cell activity by platelet-derived growth factor. II. Membrane binding studies, effects of recombinant IFN-alpha and IL-2, and lack of effect on T cell and antibody-dependent cellular cytotoxicity

We have previously reported that a 2-h pre-treatment with physiologic (nanogram) quantities of the naturally occurring biologic substance, platelet-derived growth factor (PDGF), significantly inhibits human NK cell activity (cytotoxicity) in a dose-dependent manner as measured against the NK-sensitive target K-562, as well as the single-cell binding of human NK cells to K-562 targets (i.e., conjugate formation). In this study, we have completed a more detailed and longer time course evaluation of PDGF-mediated human NK cell inhibition, and demonstrate a more marked inhibition of human NK cells at 8 to 20 h as compared to 2 h. We also show that either rIFN-alpha or IL-2 reverses the PDGF-mediated NK cell inhibition; and furthermore, that PDGF does not inhibit the lymphokine augmented NK activity of recombinant alpha-IFN or IL-2-activated human NK cells. In addition, we have further demonstrated that PDGF does not inhibit cytotoxic T cell activity as generated in the MLR, nor does PDGF significantly inhibit antibody-dependent cellular cytotoxicity. Finally, we have demonstrated that human NK cells have roughly 12,000 high-affinity, surface binding sites for PDGF, as determined by the competitive binding of 125I-labeled purified PDGF and unlabeled PDGF to B73.1+ FACS-sorted lymphoid (NK) cells, and Scatchard analysis of these data. The implications of these findings are discussed.     

Mechanisms of corticosteroid action on lymphocyte subpopulations. IV. Effects of in vitro hydrocortisone on naturally occuring and mitogen-induced suppressor cells in man.

The effects of in vitro hydrocortisone (OHC) on human peripheral blood (PB) suppressor cell function were investigated. Two types of suppressor cells were studied: (i) the naturally occurring PB suppressor cell seen in 10% of normal people whose lymphocytes do not respond to in vitro PWM stimulation with direct anti-SRBC PFC responses, and (ii) Con A-generated suppressor cells. The addition of OHC to PWM-stimulated cultures from nonresponders reconstituted the PFC response in two of three individuals. The addition of OHC to allogenic cocultures of nonresponder and responder lymphocytes completely inhibited the ability of the naturally occurring suppressor cell of the nonresponder cultures to inhibit the PFC responses of normal responders. Preincubating the nonresponder cultures in 10^?5M OHC for 30 min followed by washing did not inhibit suppressor function, whereas readdition of OHC to cocultures did inhibit nonresponder suppressor cell function. The addition of up to 10^?4M OHC to previously generated Con A-activated suppressor cell-fresh cell cocultures in vitro did not prevent or inhibit mitogen-activated suppressor cell function. However, preincubation of PB cells for 6 hr prior to the addition of Con A prevented the generation of suppressor cells and in two of eight experiments generated a population of cells which were in and of themselves mitogenic for autologous fresh PB. Thus, the function of naturally occurring suppressor cells as well as the induction but not the function of Con A-activated suppressor cells is sensitive to pharmacologic levels of OHC. The effect of OHC on naturally occurring suppressor cell function or on the generation of suppressor cells by Con A did not involve cell lysis, but rather was a reversible phenomenon requiring the continued presence of OHC in culture.     

Activation and mechanism of action of suppressor macrophages

Intravenous administration of Corynebacterium parvum to alloimmunized mice activates splenic suppressor macrophages that effectively curtail primary and secondary generation of cytotoxic T lymphocytes (CTLs) in vitro. CTL generation was significantly inhibited in suppressed primary cultures by Day 3, the earliest time point that activity is first detected in control cultures. Suppressor macrophages had to be present during the first 24–48 hr of culture to effectively curtail the generation of CTLs. However, if suppressor macrophages were reactivated by 48-hr in vitro culture and then added to primary sensitizations that had been initiated 48 hr previously, they were capable of significant suppression. Suppressor cells produced a soluble factor that mediated the inhibition of CTL generation. The production or action of this factor could not be counteracted by indomethacin.     

Regulatory mechanisms in cytotoxic T lymphocyte development. II. Dissociation of in vitro cytotoxic T-lymphocyte and suppressor T-cell activities with L-ornithine

L-ornithine was found to differentially affect the induction of allospecific cytotoxic T lymphocytes (CTL) and suppressor T cells (Ts). At a concentration of 10 mM, ornithine inhibited the development of CTL in a mixed-leukocyte culture (MLC). This same population of cells suppressed the generation of CTL when irradiated and cocultured with fresh syngeneic lymphocytes and alloantigen. Suppression was mediated by Lyt-1-2+ cells and was antigen specific. Suppression was abrogated when IL-2 (10 U/ml) was added to the cocultures, but could not be reversed by increasing the antigen dose. Ornithine was not toxic to CTL precursors but rather arrested their development. Cells from MLC plus ornithine developed CTL activity within 2 days of transfer to secondary cultures in the absence of ornithine. Development of CTL effector cells (CTLe) was augmented by but did not require exogenous IL-2. Generation of CTLe from the MLC plus ornithine population was radiation sensitive and could be inhibited by reexposure to ornithine, even in the presence of IL-2. Thus, Lyt-1-2+ T cells allostimulated in vitro in MLC plus ornithine and lacking CTL activity convey radiation-resistant, antigen-specific suppression.     

Effect of cyclosporin A on human lymphocyte responses in vitro. III. CsA inhibits the production of T lymphocyte growth factors in secondary mixed lymphocyte responses but does not inhibit the response of primed lymphocytes to TCGF

The mechanism whereby Cyclosporin A (CsA) inhibits secondary mixed lymphocyte responses was assessed. CsA added to secondary MLR cultures inhibited proliferation and induction of cytolytic lymphocyte activity. This inhibition was found to be associated with the inhibition of T lymphocyte stimulating growth factor(s) (TCGF) production in the supernatants of secondary MLR cultures. As little as 1.0 micrograms/ml of CsA added to secondary MLR cultures resulted in no measurable TCGF activity. In contrast, moderate doses of CsA (1.0, 2.5 micrograms/ml), which completely inhibited the secondary MLR response to alloantigen, did not inhibit the proliferative and CML response of alloantigen-primed lymphocytes to these stimulating growth factors. Even at high doses of CsA (20 micrograms/ml), substantial levels of proliferation (50% of control response) and CML induction (60% of control response) were observed when the primed cells were exposed to secondary MLR supernatants containing TCGF activity. It was concluded that inhibition of secondary mixed lymphocyte responses by CsA may be due in part to the inhibition of TCGF production rather than the inhibition of the effect of TCGF on mature cytotoxic T lymphocytes.     

Development of precytotoxic T cells in cyclosporine-suppressed mixed lymphocyte reactions

Cyclosporine (CsA) blocked the generation of cytolytic activity in a primary MLR of mouse spleen cells. As expected from the known mechanism of action of this drug, it also blocked the accumulation of IL-2 during the MLR. Addition of human rIL-2 did not overcome the inhibition of CTL generation, even when it was added daily to keep its level similar to that produced in a normal MLR. Daily addition was necessary, because the CsA-inhibited MLR consumed IL-2, either by utilization or degradation. The outcome of a 5-day MLR in the presence of CsA (CsA-MLR) depended on whether or not IL-2 was continuously present. In the presence of IL-2, there was no generation of CTL activity, probably because such cultures contained IL-2-dependent suppressive elements described previously. However, when day 5 CsA-MLR cells generated in the absence of IL-2 were washed and recultured with human rIL-2, there was a burst of CTL activity, with a more than 50-fold increase in alloantigen-specific cytotoxicity within 24 to 48 h. This increase is not explainable simply by the proliferation of existing effector CTL. The noncytotoxic cells produced in an MLR in the presence of CsA, and which can be rapidly activated to cytotoxic effector cells by IL-2, are termed "precursor-effector CTL" (peCTL). They could be detected by day 3 of a primary CsA-MLR culture. Their conversion to effector CTL by IL-2 was not inhibited by CsA. Exposure of peCTL to IL-4 also generated CTL activity, to a somewhat lesser degree than IL-2, but the IL-4-induced activation was inhibited by CsA, suggesting that it depended on the induction of another CsA-sensitive lymphokine. The intracellular levels of mRNA encoding the CTL-specific serine esterases CCP1 and CCP2 (granzymes B and C, respectively) increased rapidly during the IL-2-driven conversion of peCTL to effector CTL. This study demonstrates that in the presence of CsA precursors for CTL can accumulate, and that these can be rapidly converted to cytotoxic effector cells by IL-2.     

Suppression of in vitro maintenance and interferon-mediated augmentation of natural killer cell activity by adherent peritoneal cells from normal mice

The ability of adherent peritoneal cells (APC) to inhibit murine natural killer (NK) cell activity was examined. Nylon wool-nonadherent splenic effector cells were incubated overnight with or without different numbers of APC. NK activity was then measured against YAC-1 in a 4-hr 51Cr-release cytotoxicity assay. Proteose peptone-elicited or unstimulated resident APC from normal mice markedly suppressed NK activity of splenic effector cells in the presence or absence of exogenously added interferon. The suppression was dependent on the number of APC added with 10% APC, relative to the number of effector cells, resulting in a greater than 65% inhibition of cytotoxicity. The effector phase of cytotoxicity was not the target of the suppressor cells, because APC did not suppress NK activity when they were present only during the cytotoxicity assay. The addition of APC to alloimmune cytotoxic T cells under similar conditions resulted in no inhibition of cytotoxicity. Both syngeneic and allogeneic APC suppressed NK activity, but several murine macrophage-like cell lines lacked this property. In contrast to APC, incubation of effector cells with adherent spleen cells from normal mice resulted in no inhibition of NK activity. APC from mice injected with C. parvum were less inhibitory for NK activity than normal resident APC. In contrast, C. parvum APC suppressed concanavalin A-induced lymphoproliferation and were directly cytotoxic to tumor target cells in vitro, whereas normal APC lacked these properties. The results indicate that the peritoneum of untreated mice contains suppressor cells that can inhibit the in vitro maintenance and IFN-mediated augmentation of NK activity. In addition, these results indicate a broader spectrum of immune reactivities regulated by APC and suggest that, depending on their level of activation, APC can preferentially inhibit different immune functions.     

Monocyte-derived human macrophages mediate anergy in allogeneic T cells and induce regulatory T cells

Activation of alloreactive T cells by APCs such as dendritic cells (DC) has been implicated as crucial step in transplant rejection. In contrast, it has been proposed that macrophages (Mphi) maintain tolerance toward alloantigens. It was therefore the aim of this study to further analyze the T cell-stimulatory capacity of mature DC and Mphi in vitro using the model of allogeneic MLR. There was a strong proliferative response in T cells cocultured with DC, which was further increased upon restimulation in a secondary MLR. In contrast, T cells did not proliferate in cocultures with Mphi despite costimulation with anti-CD28 and IL-2. Cytokine analysis revealed considerable levels of IL-10 in cocultures of T cells with Mphi, whereas high amounts of IL-2 and IFN-gamma were present in cocultures with DC. There was only minimal T cell proliferation in a secondary MLR when T cells were rescued from primary MLR with Mphi and restimulated with DC of the same donor, or DC of an unrelated donor (third party), whereas a strong primary proliferative response was observed in resting T cells, demonstrating induction of T cell anergy by Mphi. Functional analysis of T cells rescued from cocultures with Mphi demonstrated that anergy was at least partly mediated by IL-10-producing regulatory T cells induced by Mphi. These results demonstrate that Mphi drive the differentiation of regulatory T cells and mediate anergy in allogeneic T cells, supporting the concept that Mphi maintain peripheral tolerance in vivo.     

Immunoregulatory activity of supernatants from short-term cultures of mouse decidual tissue

Supernatants from short-term in-vitro cultures of decidual tissue, obtained from the uteri of pregnant mice from Days 4 to 13 post coitum (Day 1 = day of mating), were assessed for immunoregulatory activity by their addition to a mixed lymphocyte reaction (MLR), an in-vitro analogue of the afferent arm of the immune response. All culture supernatants tested possessed inhibitory activity in the MLR, although the extent of inhibition was affected by seeding density, length of culture, and the day of pregnancy from which decidual tissue was obtained. Inhibitory activity produced by decidual cultures increased from Day 4 to reach a maximum on Day 8, and then declined to Day 11. Two morphologically distinct cell types were present in all decidual cultures; flat dendritic cells, considered to represent decidual cells, and small round cells, but whether immunoregulatory factors are associated with both is uncertain. The results suggest that decidual tissue could fulfil a role in the local partial blockade of the afferent arm of the maternal immune response during pregnancy.     

Inhibition of erythroid progenitor growth is mediated by cytotoxic lymphocytes and not by natural killer cells or IFN-gamma

Alloantigen primed T cells (PTC) were recovered from MLR at day 6 and 12, then added to cultures of erythroid progenitors, erythroid burst-forming units, BFU-E. The PBMC source of BFU-E was prepared either to retain or deplete APC by treatment with appropriate mAb and C. BFU-E grown in cocultures were counted at day 14 and replicate cultures assayed for IFN-gamma production on days 1 to 7. Analysis of MLR cells indicated that large, rapidly cycling cells recovered from MLR at day 6 have significant NK activity, whereas CTL activity is minimal, and production of IFN-gamma requires reexposure to APC. The smaller, noncycling cells recovered from MLR at day 12 have comparable NK activity, also require reexposure to APC for IFN-gamma production, but in addition have significant CTL activity. The addition of day 12 MLR cells to BFU-E cultures results in MHC restricted inhibition of BFU-E growth, suggesting that the CTL activity and not the NK activity contained within this population of cells is responsible for BFU-E inhibition. Studies using enriched population of BFU-E indicated that appropriate APC are needed to trigger both IFN-gamma production and BFU-E inhibition by the PTC. By using various APC-BFU-E combinations it was determined that after reexposure of PTC to appropriate APC, the inhibition of BFU-E was still target-specific indicating a direct effect between the PTC and BFU-E.