Interleukin-3 induces proliferation but not lymphokine activated killer activity from human and murine mononuclear cells

Recombinant IL-3 (rIL-3) is a potent colony stimulating factor capable of stimulating early hematopoietic pluripotential progenitor cells and of supporting the differentiation of multiple cells. IL-3 has also been shown to have effects on mature, differentiated circulating cells including eosinophils and T cells. We evaluated the role of exogenous rIL-3 in the generation of cells with LAK activity from murine splenocytes and human bone marrow, spleen, unseparated PBMC and purified null cell preparations. rIL-3 was unable to generate lytic activity from any of these populations by itself and appeared to decrease LAK activity in bone marrow cultures containing high dose IL-2, (bone marrow derived cells (n = 3) with LAK activity for fresh tumor, mean lytic units(LU) 94.6 +/- 63.5 vs 32.8 +/- 44.8 for IL-2 and IL-2 plus IL-3 cultures, respectively p2 less than 0.05). Unlike previous reports testing murine cells, IL-3 priming and subsequent culture in IL-2 of human unseparated bone marrow cells or human or murine splenocytes, failed to generate long-term cultures with lytic activity. IL-3 did, however, induce a dose dependent stimulation of bone marrow and null cell preparations (mean null cell stimulation (3H Thymidine incorporation) with IL-3, 436 +/- 168 cpm vs 9802 +/- 9799 cpm, for 0 vs 10(3) units of IL-3, respectively n = 4, p2 less than 0.05). Furthermore, in bone marrow, unseparated PBMC and null cell cultures, the addition of rIL-3 generated characteristic large blastic appearing cells with prominent basophilic granules.(ABSTRACT TRUNCATED AT 250 WORDS)     

Evidence for voltage modulation of IL-2 production in mitogen-stimulated human peripheral blood lymphocytes.

The membrane potential of human PBMC was modulated in culture by isotonic high extracellular K+ (K+e), or the K+ channel blocker, charybdotoxin (ChTX), to determine the effect of depolarization on stimulated proliferation, IL-2 elaboration, and gene expression. In serum-free cultures, ChTX and high [K+]e induced a specific dose-dependent decrease in IL-2 production. ChTX inhibited proliferation of PBMC and purified T cells, decreased IL-2 elaboration 15 h after stimulation by 78.4 +/- 5.3% (n = 5), and decreased IL-2 mRNA steady-state levels by 80% between 8 and 10 h after stimulation. The IC50 for ChTX-inhibition of IL-2 elaboration and IL-2 mRNA were both 1 nM. Similarly, high [K+]e inhibited proliferation with an IC50 of 38.9 +/- 1.1 mM (n = 13), decreased IL-2 elaboration with an IC50 of 21.3 +/- 1.2 mM (n = 6), and decreased IL-2 mRNA steady-state levels with an IC50 of 18 mM. The sensitivities of both IL-2 production and proliferation to depolarization were substantially reduced by calcium, serum, and exogenous rIL-2. From these findings we conclude that membrane potential may contribute to the control of immune responsiveness in vivo.     

Phytohemagglutin-stimulated human T cell: prothymosin alpha as an accessory signal

Thymic peptide factors are known to modulate proliferation of normal human lymphocytes. In this work, we studied the effect of Prothymosin alpha (Pro alpha) on PHA-stimulated PBMC and PBLC. The observed effects of Pro alpha and thymosin alpha 1 (alpha 1) on PBMC were found to depend on the degree of cell stimulation, dose, and preincubation-time. Thymosin beta 4 (beta 4) had no effect on either cell type, regardless of the degree of stimulation, which shows that beta 4 may be used as a control peptide to work in this area. Induction of lymphoproliferation also depended on the presence of macrophages. Addition of monocytes or a cell-free monocyte culture supernatant (not containing IL-2) to the PHA-stimulated PBLC cultures resulted in T cell proliferation. Although IL-1 could not restore the PHA-induced proliferative response of isolated T cells by itself, it would enhance the helper effect of Pro alpha. Moreover, a polyclonal goat anti-human IL-2R (Tac Ag) did block the proliferative response induced by combined rIL-1 and Pro alpha, suggesting that an IL-2-dependent pathway of T cell proliferation was involved.     

Interleukin-2 reverses T cell unresponsiveness to Plasmodium falciparum-antigen in malaria immune subjects

Peripheral blood mononuclear cells (PBMC) from a large proportion of 34 healthy adult native residents in a malaria endemic area showed null or marginal proliferative response (low-responders) to schizont-enriched Plasmodium falciparum malaria antigen (M.Ag) but good response to pokeweed mitogen. In contrast, substantial proliferative response to M.Ag was observed in 8/8 adult temporary residents with a history of one to three acute malaria episodes. Purified CD4+ T cells preferentially responded to M.Ag, however in low-responders CD4+ T cell proliferation was poor. Moreover, no inhibition of CD4+ T cell proliferation was observed when graded numbers of CD8+ T cells were added in culture. The addition of recombinant interleukin 2 (rIL-2) to M.Ag restored the proliferative response of low-responders' PBMC. This response was M.Ag-specific when CD4+ T cells grown in M.Ag plus rIL-2, but not in rIL-2 alone, were tested in secondary cultures.     

Interleukin 2 induces proliferation of normal "resting" human T cells in the absence of other known external stimulation

Interleukin 2(IL-2) is known to stimulate the progression of activated T cells from G1 through the rest of the cell cycle. We have demonstrated that addition of purified recombinant human IL-2 (rIL-2) to fresh normal human peripheral blood mononuclear cells (PBM), which were IL-2 receptor (Tac) negative by FACS analysis, stimulated marked proliferation of the PBM. IL-2-induced proliferation was also observed with umbilical cord blood mononuclear cells. Monocyte depletion of PBM resulted in a marked reduction of rIL-2-induced proliferative response which could be restored by adding back autologous irradiated monocytes but not by interleukin 1. The T cells preincubated with rIL-2 showed a five to six times enhanced autologous mixed-lymphocyte reaction (AMLR) compared to controls. The rIL-2-induced proliferative response of PBM was inhibited in a concentration-dependent fashion by preincubation of PBM with an anti-HLA-DR framework monoclonal antibody. The proliferating cells were shown by two-color flow cytometric analysis to be primarily Leu-1+ and Leu-4+ T cells (both leu-3+ and Leu-2+ subsets); however, 6 to 19% of responding cells had surface markers for B cells or NK cells. The data demonstrate that rIL-2 can induce proliferation of "resting" human T cells. The phenomenon may be related to a monocyte-dependent AMLR which induces IL-2 receptors and IL-2 responsiveness in a subset of T cells.     

Comparative studies on in vitro reactivity of fresh and cryopreserved pig lymphocytes.

The effect of cryopreservation on in vitro reactivity of pig lymphocytes was studied. Peripheral blood mononuclear cells (PBMC) were frozen by controlled-rate freezing and stored in liquid nitrogen (LN2) between 4 and 36 days. Following thawing 74.7 +/- 2.6% of cells were recovered of which 94.5 +/- 0.9% were viable as determined by trypan blue exclusion. Functional parameters measured included the concentration of free intracellular Ca2+ ([Ca2+]i) in resting and mitogen-stimulated PBMC, mitogen and alloantigen-induced blastogenesis, as well as cell-mediated cytotoxicity. Irrespective of storage time and cell donor, [Ca2+]i in frozen-thawed PBMC (67.7 +/- 4.3 nM) was significantly lower (P less than 0.001) when compared to fresh cells (96.2 +/- 4.5 nM). In addition, cryopreserved PBMC only weakly responded with an increase of [Ca2+]i after stimulation by various concentrations of phytohemagglutinin (PHA). Following activation by PHA (2 micrograms/ml) for 4 days fresh lymphocytes (84,047 +/- 5475 cpm) incorporated significantly more (P less than 0.005) [3H]thymidine than frozen PBMC (66,001 +/- 4117 cpm). A similar difference in proliferation rates (P less than 0.05) between fresh (10,046 +/- 1915 cpm) and frozen-thawed PBMC (5852 +/- 1304 cpm) was observed in one-way mixed lymphocyte cultures (MLC), while the spontaneous incorporation of radiolabel was unchanged in frozen stored cells. By using MLC-derived cytotoxic effector cells (E) and [3H]thymidine-labeled concanavalin A blasts as targets (T), cryopreserved PBMC displayed a severe deficiency of cytotoxic effector functions at all tested E:T ratios. These results indicate that pig PBMC are very sensitive to LN2 storage although some immunological functions are more affected by cryopreservation than others.     

Effects of IL-7 and IL-2 on highly enriched CD56+ natural killer cells. A comparative study.

IL-7 has been shown to induce low levels of lymphokine-activated killer cell (LAK) activity in bulk PBMC populations. We report here that immunomagnetically purified CD56+ cells from peripheral blood generated high LAK activity in response to IL-7. The LAK activity induced by IL-7 was comparable to, or slightly lower than, the LAK activity induced by IL-2. When analyzing cells from the same donor, no detectable LAK-generating effect of IL-7 was registered in the PBMC population, in contrast to a substantial effect in the CD56+ population. IL-2 induced 8- to 15-fold higher proliferative activity in CD56+ cells, relative to IL-7. At suboptimal concentrations of IL-2, IL-7 had a synergistic effect on the proliferation. IL-2-neutralizing antibodies did not abrogate the IL-7-induced proliferation or LAK generation. Both IL-7 and IL-2 induced comparable levels of 75-kDa TNFR expression, whereas IL-2R alpha expression was higher in IL-7-stimulated CD56+ cells. Low levels of TNF were produced in response to IL-7 at day 5, as opposed to a 50-fold higher TNF production in response to IL-2. No IL-2 or IL-6 production was detected. Our data indicate that IL-7 has profound and direct effects on CD56+ cells.     

A cytoskeleton-dependent pathway for induction of IL-2 production and a cytoskeleton-independent pathway for IL-2-mediated signal transduction

We have used the synthetic microtubule inhibitor Tubulozole C in order to study the role of the microtubule system in human lymphocyte activation. Microtubule disruption prior to activation with phytohemagglutinin (PHA) resulted in a drastic reduction of IL-2 production. Similarly, using OKT3 or PHA as stimulators, a substantial decrease in proliferation was observed. Although IL-2 receptor analysis performed on the stimulated and antitubular-treated lymphocytes showed a 2-fold decrease in high-affinity and a 100-fold decrease in low-affinity IL-2 receptor expression, a proliferative response to externally added rIL-2 was noticed. This occurred provided the triggering agent was excluded or added in suboptimal concentrations. These results indicate that intact microtubules are necessary for PHA/OKT3-induced proliferation and IL-2 production, but not for IL-2-induced proliferation.     

1,25-Dihydroxyvitamin D3 enhancement of concanavalin-A induced bovine lymphocyte proliferation: requirement of monocytes

The effect of 1,25-(OH)2D3 on Concanavalin-A (ConA) mediated bovine lymphocyte proliferation was dependent on the magnitude of the proliferative response. Lymphocytes from cows with high inherent proliferative responses (134,904 +/- 15,488 cpm/well) were unaffected by 1,25-(OH)2D3; whereas, lymphocytes from cows with low inherent proliferative responses (46,317 +/- 6000 cpm/well) were stimulated 75% (P less than 0.01) by 10(-10)M 1,25-(OH)2D3. At low initial proliferation rates, seen with low density cultures enriched for lymphocytes, the effect of 1,25-(OH)2D3 on proliferation changed from inhibition in the absence of added monocytes to stimulation at 18-23% monocyte addition. These results suggest that 1,25-(OH)2D3 possesses both stimulatory and inhibitory effects on ConA-induced proliferation that are monocyte dependent.     

Interleukin 2 responsiveness of immature T-cell colony-forming cells (T-CFC) from patients with acute T-cell lymphoblastic leukemias

T-cell colony-forming cells (T-CFC) from 13 of 17 patients with T-ALL generated colonies in methylcellulose in the absence of added growth factors or mitogenic stimulation. As previously described, these colonies were composed of immature T cells displaying the same karyotypic abnormalities as fresh leukemic cells. Biochemically purified (bIL-2) and recombinant IL-2 (rIL-2) without any mitogen enhanced colony growth from both unfractionated and blast-enriched cell fractions in patients with a relatively low (less than 50 colonies/5 X 10(4) cells) plating efficiency. However, dose-response experiments revealed that the optimal dose of rIL-2 needed to enhance colony growth varied from patient to patient. Anti-IL-2 (DMS1) and anti-IL-2 receptor (anti-Tac) moAbs inhibited both spontaneous and rIL-2-induced colony formation in a dose-dependent manner. Direct staining of fresh leukemic cells with anti-Tac revealed less than 10% positive cells in all but two patients. However, cell incubation in the absence of growth factors or mitogens for 2-48 hr resulted in an increase of Tac+ cells. These observations indicate that a subset of immature T-CFC from T-ALL patients display functional IL-2-receptors. In addition, our findings strongly suggest that the IL-2/IL-2-R system could be involved in the spontaneous proliferation of some immature T-CFC of T-ALL patients.     

B cell activation during HIV-1 infection. II. Cell-to-cell interactions and cytokine requirement

This study examined the mechanisms underlying the intense activation of HIV-1-specific B cells observed in peripheral blood of HIV-1-infected subjects. Spontaneous in vitro synthesis of anti-HIV-1 antibodies, as well as total Ig production, were dramatically reduced by accessory cell, but not T cell removal. This fall was counteracted by addition of rIL-6, but not other cytokines, to monocyte-depleted cultures; moreover, antisera against IL-6 suppressed spontaneous anti-HIV-1 antibody synthesis in a dose-dependent manner. Although IL-6 apparently sustained HIV-1-specific B cell activation, no increase in serum IL-6 levels was observed; PBMC from seropositive subjects did not produce increased amounts of IL-6 in vitro, compared to seronegative controls, both spontaneously and in the presence of LPS stimulation; finally, no constitutive expression of IL-6 gene could be documented in freshly isolated PBMC. These findings indicate that IL-6 may play a central role in HIV-1-specific B cell activation in seropositive patients, and further stress the importance of this cytokine during HIV-1 infection.     

Identification of IL-6 as a T cell-derived factor that enhances the proliferative response of thymocytes to IL-4 and phorbol myristate acetate

Thymocytes undergo a vigorous proliferative response when stimulated with a combination of IL-4 and PMA. We have found that conA-induced supernatants from a number of Th cell clones could enhance the level of IL-4/PMA-induced proliferation of unseparated thymocytes 0.5- to 2-fold and of peanut agglutinin-positive thymocytes 2- to 10-fold. These supernatants did not contain IL-2 or IFN-gamma, and the enhancing activity could be chromatographically separated from IL-3, -4, -5, and granulocyte/macrophage CSF. The possibility that the thymocyte enhancement factor contained in these supernatants was IL-6 was suggested when murine rIL-6 was found to have similar activity. Further evidence for the identity of these two factors was obtained when an IL-6 assay, based on plasmacytoma growth, was used to test column fractions showing thymocyte enhancement. All fractions active in the thymocyte enhancement assay also had activity in the plasmacytoma growth assay. These observations suggest that the thymocyte-stimulating activity present in the T cell supernatants was due to IL-6.     

Impaired T cell proliferation in acute dengue infection

Decreased proliferative responses to mitogens and recall Ags have been observed in PBMC obtained during several acute human viral infections. To determine whether cell-mediated responses are altered during acute dengue infection, we examined the proliferative responses of PBMC from children enrolled in a prospective study of dengue infections in Thailand. All responses of PBMC during acute illness were compared with the same patients' PBMC obtained at least 6 mo after their infection. Proliferative responses to PHA, anti-CD3, tetanus toxoid, and dengue Ags were decreased significantly in PBMC obtained during the acute infection. The proliferative responses to PHA were restored by the addition of gamma-irradiated autologous convalescent or allogeneic PBMC. Cell contact with the irradiated PBMC was necessary to restore proliferation. Non-T cells from the acute PBMC of dengue patients did not support proliferation of T cells from control donors in response to PHA, but T cells from the PBMC of patients with acute dengue proliferated if accessory cells from a control donor were present. Addition of anti-CD28 Abs restored anti-CD3-induced proliferation of the PBMC of some patients. The percentage of monocytes was reduced in the acute sample of PBMC of the dengue patients. Addition of IL-2 or IL-7, but not IL-4 or IL-12, also restored proliferation of acute PBMC stimulated with anti-CD3. The results demonstrate that both quantitative and qualitative defects in the accessory cell population during acute dengue illness result in a depression of in vitro T cell proliferation.     

Involvement of IL-15 in the pathogenesis of human T lymphotropic virus type I-associated myelopathy/tropical spastic paraparesis: implications for therapy with a monoclonal antibody directed to the IL-2/15R beta receptor.

Human T lymphotropic virus type I (HTLV-I) is the causative agent of an inflammatory neurological disease termed HTLV-I-associated myelopathy/tropical spastic paraparesis (HAM/TSP). An ongoing lymphocyte activation exists in patients with HAM/TSP, which was demonstrated by the spontaneous proliferation of their PBMC ex vivo. It was shown that spontaneous proliferation present in HAM/TSP is due, in part, to an IL-2/IL-2R autocrine loop. However, addition of Abs against IL-2 or IL-2R alpha only partially inhibited the spontaneous proliferation. Since IL-15 is a cytokine with similar functional characteristics to those of IL-2, we reasoned that IL-15 might be an additional growth factor that contributes to the spontaneous proliferation observed in HAM/TSP. In this study, we demonstrated that IL-15 mRNA expression was elevated in PBMC obtained from HAM/TSP patients when compared with those of the normal donors. Furthermore, we showed that the addition of blocking Abs against IL-15 or its receptor inhibited the spontaneous proliferation of HAM/TSP PBMC. Addition of Abs directed toward both IL-15 and IL-2, or their receptors, inhibited the proliferation almost completely. These data suggest the existence of two autocrine loops involving IL-15/IL-15R and IL-2/IL-2R, both contributing to the spontaneous proliferation of HAM/TSP PBMC.     

Functional effects of a monoclonal antibody directed against a distinct epitope on 4F2 molecular complex in human peripheral blood mononuclear cell activation.

The 4F2 antigenic complex is expressed on most human cell lines in culture, on monocytes and activated lymphocytes, but not on resting T and B lymphocytes. Monoclonal antibody (mAb) CB43 recognizes an epitope of the 4F2 heterodimer either located on the light chain or dependent on the conformation of the molecule. The binding of CB43 mAb to peripheral blood mononuclear cells (PBMC) induced a dose-dependent comitogenic effect in the presence of submitogenic concentrations of anti-CD3 mAb. Significant amounts of interleukin (IL)-1 beta but not IL-2 or interferon-gamma were released in the supernatant. Pretreatment of monocytes with CB43 mAb increased the phytohemagglutinin-induced T lymphocyte proliferation. However, CB43 mAb did not exert agonistic effects on activated T lymphocytes. Depletion of CB43+ cells from PBMC decreased the proliferation and generation of cytotoxic effector cells induced by a mannoprotein (MP) derived from Candida albicans cell wall but not by recombinant IL-2. Furthermore, depletion of CB43+ cells from PBMC preactivated with MP or rIL-2 led to a significant decrease in their cytotoxic activity. CB43 mAb did not inhibit the growth of cell lines nor the proliferation of T cells. Thus CB43 mAb identifies a distinct functional epitope on the 4F2 molecular complex and might be useful in further studying the role of this molecule in cellular activation.     

Glutamate modulation of human lymphocyte growth: in vitro studies

Peripheral blood mononuclear cell (PBMC) proliferation induced by phytohemagglutinin, or by anti-CD3 alone or plus anti-CD28 monoclonal antibodies (mAb) was inhibited by glutamate (Glu) in a concentration-dependent manner. This inhibition was not reproduced by selective ionotropic Glu receptor agonists, whereas it was potentiated by l-buthionine-(S,R)-sulfoximine, which depletes glutathione (GSH) stores, and counteracted by 2-mercaptoethanol, a preserver of cell thiols. The inhibitory effects of Glu were related to depletion of intracellular GSH stores, since it decreased GSH levels in a concentration-dependent manner. Furthermore, Glu modulated cytokine secretion by anti-CD3 mAb activated PBMC: it increased IFN-gamma (+44.3+/-8.2%) and IL-10 (+31.6+/-9.7%) secretion, whereas that of IL-2, IL-4, IL-5, and TNF-alpha was not affected. These data suggest that high levels of Glu, which can be reached in damaged tissues, modulate lymphocyte responses to activating stimuli by favouring polarization of the T helper effector response.     

Helper-independent CD8+ cytotoxic T lymphocytes express IL-1 receptors and require IL-1 for secretion of IL-2

The purpose of this study was to examine the role of IL-1 on the activation of CD8+/CD4- class I-restricted helper cell-independent cytolytic T cell (HITc) clones known to produce IL-2 and proliferate in vitro after Ag stimulation with a Friend retrovirus-induced leukemia (FBL). The functional role of IL-1 in Ag-specific proliferation and IL-2 secretion was assessed by stimulating the T cell clones with FBL either in the presence or absence of macrophages (M phi), rIL-1, or rIL-2. Resting cloned HITc cells, purified from residual accessory cells, failed to proliferate in response to FBL alone, but proliferated in response to FBL plus M phi, rIL-1 or rIL-2. Stimulation with FBL alone in the absence of M phi or IL-1 was sufficient for induction of IL-2R expression, and rendered cells responsive to IL-2, but M phi or IL-1 were also required to induce production of IL-2. The activity of IL-1 was further examined by measuring the binding of [125I]rIL-1 alpha, which demonstrated that resting cloned HITc cells expressed IL-1R that increased in number after activation with Ag. This expression of IL-1R and requirement for IL-1 by CD8+ HITc was surprising because previous studies examining T cell populations after mitogen stimulation have not detected IL-1R on the CD8+ population. Therefore, the role of IL-1 in the activation of CD8+ CTL that do not secrete IL-2 after activation was assessed. By contrast to HITc, CD8+ CTL required exogenous IL-2 to proliferate in vitro and did not express IL-1R. These data demonstrate that the subset of CD8+ T cells responsible for IL-2 production express IL-1R and that triggering this receptor with IL-1 after Ag stimulation results in the production of IL-2 and subsequent proliferation.     

Glucocorticoids inhibit cytokine-mediated eosinophil survival

Glucocorticoids characteristically induce eosinopenia in vivo and are effective for treating allergic and other eosinophilic disorders. We studied the effect of glucocorticoids on cytokine-induced survival of human eosinophils in vitro. Eosinophils were purified from normal or mildly atopic volunteers by Percoll density gradient and incubated for 4 days in the presence of cytokine plus steroid. Cell viabilities were determined by staining cells with fluorescein diacetate and propidium iodide. In the absence of glucocorticoids, human rIL-5 enhanced eosinophil survival in a dose-dependent manner, from 22 fM for a minimal effect to 2200 fM for maximal effect. When eosinophils were cultured with a submaximal concentration of rIL-5 (220 fM), dexamethasone, methylprednisolone, and hydrocortisone inhibited eosinophil survival in a dose-dependent manner. Inhibition was time-dependent and required at least 2 days' exposure of eosinophils to dexamethasone. Dexamethasone, methylprednisolone, and hydrocortisone at 1000 nM inhibited survival by 88 +/- 2, 66 +/- 9 and 37 +/- 7%. In contrast, estradiol and testosterone (1000 nM) had no effect on eosinophil survival. When eosinophils were incubated with varying concentrations of human rIL-5 and 1000 nM dexamethasone, survival inhibition was reduced at higher concentrations of human rIL-5, and completely abolished by human rIL-5 23,000 fM. Human recombinant granulocyte-macrophage CSF, human rIL-3, and human rIFN-gamma also enhanced eosinophil survival in a dose-dependent manner and dexamethasone (1000 nM) strongly inhibited cell survival when submaximal concentrations of these cytokines were used. The effects of dexamethasone were reversed by higher concentrations of granulocyte-macrophage CSF (10 U/ml) and IL-3 (3 ng/ml). However, even 1000 U/ml IFN-gamma did not overcome dexamethasone inhibition, indicating a difference between the mechanism of eosinophil survival induced by IFN-gamma and other cytokines. These results suggest that glucocorticoids exert a direct, inhibitory effect on eosinophil survival, which may be important in the treatment of allergic and other eosinophilic disorders. Antagonism of this effect by higher amounts of cytokine may be a mechanism for glucocorticoid resistance.     

The inhibitory effect of human syncytiotrophoblast plasma membrane vesicles on in vitro lymphocyte proliferation is associated with reduced interleukin 2 receptor expression

The mechanisms by which vesicles of syncytiotrophoblast plasma membranes (STPM) prepared from full-term human placentas inhibit lymphocyte proliferation have been investigated. In the presence of STPM, IL-2 secretion and the expression of protein P55 (IL-2R P55) from its receptor were examined in two models of PBMC proliferation: induced by PHA in 3-day-old cultures, and induced by IL-2 in 6-day-old cultures. In the case of PHA stimulation, STPM strongly inhibited IL-2 (but not IL-1) secretion and IL-2R P55 expression at a concentration where lymphocyte proliferation was also blocked. In these conditions, the addition of excess recombinant IL-2 (rIL-2) only partially restored proliferation and IL-2R P55 expression. In addition, STPM inhibited proliferation and IL-2R P55 expression when resting PBMC were stimulated by a high concentration of rIL-2. These results suggest that STPM inhibit lymphocyte proliferation by affecting one or several events occurring in the synthesis and/or expression of IL-2R P55 by a mechanism which is at least partially independent of its inhibitory effect on IL-2 secretion. The significance of these results is discussed in the context of the survival of the fetal allograft.