Immunoactive products of human placenta. I. An immunoregulatory factor obtained from explant cultures of human placenta inhibits CTL generation and cytotoxic effector activity

Supernatants were prepared from short-duration explant cultures of term human placentas obtained after cesarean delivery. These supernatants inhibited murine and human mixed lymphocyte reactions, as well as CTL generation. The effects were reversed by an excess of IL-2-containing medium. Similarly, the material inhibited human natural killer cytotoxicity against K 562 targets. The material was subjected to gel-filtration chromatography on an ACA 44 or Bio-Gel A15m column. The apparent MW of the MLR-CML material was about 60-70 kDa, whereas the NK inhibiting activity was eluted in high-MW components (greater than 200 kDa) as well as in the 50-kDa range. The relevance of this material in local immunoregulation during human pregnancy is discussed.     

Immunoactive products of placenta. III. Suppression of natural killing activity

Placental and trophoblast-enriched cells from iso or allopregnant mice (14 to 17 days of gestation) strongly suppress natural killing by mouse splenocytes. The suppression is not strain restricted and acts at the level of the effector cell. It was not reduced by pretreatment of trophoblast cells with RNA and protein synthesis inhibitors, nor by prostaglandin synthetase inhibitors. However, treatment with trypsin abolished the suppressive activity. Because CTL and ADCC cytotoxicity were also inhibited by trophoblast cells at the effector stage, we postulate that trophoblast cells in vivo are endowed with the ability to inactivate various types of cytotoxic cells in their vicinity that are potentially harmful for the conceptus.     

Hemeoxygenase expression in human placenta and placental bed implies a role in regulation of trophoblast invasion and placental function.

The purpose of this study was to examine the expression of hemeoxygenases HO-1 and HO-2, which are responsible for the production of carbon monoxide (CO), in the human placenta and placental bed and to determine the role of inhibitors of HO on placental perfusion pressure. We hypothesized that HO is expressed within the placenta and that invading cytotrophoblast cells (CTB) express HO isoforms. The expression of HO-1 and HO-2 was studied on placenta and placental bed biopsies, obtained using a transcervical sampling technique, from normal human pregnancies between 8 and 19 wk gestation and at term. In the placenta, HO-2 immunostaining was prominent in syncytiotrophoblast in the first trimester and reduced toward term (P<0.0005). HO-2 endothelial immunostaining was weak in the first trimester, but increased by term (P<0.0005). Within the placental bed, HO-2 was expressed by CTB in cell columns, the cytotrophoblast shell, and cell islands. Both intravascular CTB and interstitial CTB expressed HO-2. HO-1 immunostaining was low in the placenta but intense on the CTB within the placental bed. A striking feature was the absence of HO-1 from the proximal layers of cell columns, with strong expression on the more distal CTB layers of the cell columns. In placental perfusion studies, a significant dose-dependent increase in perfusion pressure was observed in the presence of zinc protoporphyrin, an inhibitor of HO. These results suggest a role for CO in placental function, trophoblast invasion, and spiral artery transformation. Hemeoxygenase expression in human placenta and placental bed implies a role in regulation of trophoblast invasion and placental function.     

Regulation of the immune response to tumor antigen. VI. Differential specificities of suppressor T cells or their products and effector T cells.

Suppressor T cells arising during the development of certain murine methylcholanthrene-induced fibrosarcomas have previously been shown capable of limiting only those effector responses generated against the homologous tumor. Thus, S1509a-induced suppressor T cells inhibit immune reactivity only to the S1509a tumor in S1509a immune mice and have no effect on the rejection of SAI tumors in SAI-immune animals. In contrast to this is the cross-reactivity of effector cells in this system, whereby animals rendered immune to either the S1509a or SAI sarcoma are equally capable of rejecting a challenge of the opposite tumor. The specificity of suppression has been further defined in the present study, which demonstrates that S1509a-induced suppressor cells can inhibit responsiveness only to the S1509a sarcoma, even in the simultaneous presence of both the S1509a and SAI tumors. Furthermore, the suppressor factor that is obtainable from suppressor T cells demonstrates a similar precise specificity in its ability to limit selectively reactivity only against the inducing tumor, regardless of the simultaneous expression of antigens on other tumors recognized by cross-reactive effector cells. These results suggest that the antigenic determinants recognized by effector and suppressor T cells are different, and may provide a model for further dissection of suppressor cell function in vivo.     

Cognate interactions between helper T cells and B cells. IV. Requirements for the expression of effector phase activity by helper T cells.

After activation with anti-CD3, activated Th (THCD3), but not resting Th, fixed with paraformaldehyde induce B cell RNA synthesis when co-cultured with resting B cells. This activity is expressed by Th of both Th1 and Th2 subtypes, as well as a third Th clone that is not classified into either subtype. It is proposed that anti-CD3 activation of Th results in the expression of Th membrane proteins that trigger B cell cycle entry. Kinetic studies reveal that 4 to 8 h of activation with anti-CD3 is sufficient for ThCD3 to express B cell-activating function. However, activation of Th with anti-CD3 for extended periods of time results in reduced Th effector activity. Inhibition of Th RNA synthesis during the anti-CD3 activation period ablates the ability of ThCD3 to induce B cell cycle entry. This indicates that de novo synthesis of proteins is required for ThCD3 to express effector function. The ability of fixed ThCD3 to induce entry of B cell into cycle is not due to an increase in expression of CD3, CD4, LFA-1, ICAM-1, class I MHC or Thy-1. Other forms of Th activation (PMA and A23187, Con A) also induced Th effector function. Furthermore, purified plasma membranes from anti-CD3 activated, but not resting Th, induced resting B cells to enter cycle. The addition of IL-4, but not IL-2, IL-5, or IFN-gamma amplified the DNA synthetic response of B cells stimulated with PM from activated Th. Taken together these data indicate that de novo expression of Th surface proteins on activated Th is required for Th to induce B cell cycle entry into G1 and the addition of IL-4 is required for the heightened progression into S phase.     

Characterization of the cellular basis for the inhibition of cytolytic effector cells by murine placenta

Direct suppression of cytolytic effector cell function by cells of the placenta may represent one mechanism that protects the "fetal allograft" from rejection by maternal transplantation immunity. Collagenase disaggregated murine placental cells block target cell lysis by natural killer, lymphokine-activated killer, and (CTL)-type killer cells. This inhibition is reversible and noncompetitive, similar to a previously described inhibitor of CTL found in spleens of mice undergoing an acute graft vs host (GVH) response. Velocity sedimentation separation of placental cells shows that the inhibitory activity is primarily associated with cells that cosediment with nucleated fetal erythrocytes. When these erythrocytes were lysed, an increased number of non-erythrocytic cells could be separated and under this circumstance, inhibitory activity was seen in association with either small white cells or fetal erythrocytes and with large white cells. There may be several cell populations in murine placenta that can inhibit cytolytic effector cells. The possible relevance of direct placental inhibition of cytolytic effectors to protection of the "fetal allograft" is discussed.     

The brain parenchyma is permissive for full antitumor CTL effector function, even in the absence of CD4 T cells

Effective antitumor immune responses against cerebral malignancies have been demonstrated in several models, but precise cellular function of specific effector cells is poorly understood. We have explored this topic by analyzing the MHC class I-restricted T cell response elicited after implantation of HLA-CW3-transfected P815 mastocytoma cells (P815-CW3) in syngeneic mice. In this model, tumor-specific CTLs use a distinctive repertoire of TCRs that allows ex vivo assessment of the response by immunophenotyping and TCR spectratyping. Thus, for the first time in a brain tumor model, we are able to directly visualize ex vivo CTLs specific for a tumor-expressed Ag. Tumor-specific CTLs are detected in the CNS after intracerebral implantation of P815-CW3, together with other inflammatory cells. Moreover, despite observations in other models suggesting that CTLs infiltrating the brain may be functionally compromised and highly dependent upon CD4 T cells, in this syngeneic P815-CW3 model, intracerebral tumors were efficiently rejected, whether or not CD4 T cells were present. This observation correlated with potent ex vivo cytotoxicity of brain-infiltrating CTLs, specific for the immunodominant epitope CW3170-179 expressed on P815-CW3 tumor cells.     

The physiological interferon response: IV. Production of interferon by the perfused human placenta at term

Human placentas at term, free of bacterial and viral diseases, have been perfused and maintained sterile for up to 13 hr. Several parameters indicate that the organs remained viable and released interferon into the perfusate in a progressive fashion. The amount of interferon was small and the individual variations indicate that there are "poor" and "good" placenta producers. Both interferons -alpha and -beta were produced with a prevalence of the latter type. The partial acid lability and the type heterogeneity suggest that under physiological conditions the placentas produce unusual interferons, the function of which remains speculative.     

Cutting edge: rapid in vivo CTL activity by polyoma virus-specific effector and memory CD8+ T cells

For viruses that establish persistent infection, continuous immunosurveillance by effector-competent antiviral CD8(+) T cells is likely essential for limiting viral replication. Although it is well documented that virus-specific memory CD8(+) T cells synthesize cytokines after short term in vitro stimulation, there is limited evidence that these T cells exhibit cytotoxicity, the dominant antiviral effector function. Here, we show that antiviral CD8(+) T cells in mice acutely infected by polyoma virus, a persistent mouse pathogen, specifically eliminate viral peptide-pulsed donor spleen cells within minutes after adoptive transfer and do so via a perforin-dependent mechanism. Antiviral memory CD8(+) T cells were similarly capable of rapidly mobilizing potent Ag-specific cytotoxic activity in vivo. These findings strongly support the concept that a cytotoxic effector-memory CD8(+) T cell population operates in vivo to control this persistent viral infection.     

Stimulation of proliferation of bovine placental cells by products of activated mononuclear leukocytes

Summary Culture medium conditioned with concanavalin A-stimulated mononuclear leukocytes was tested for its ability to stimulate in vitro proliferation of bovine placental cells. The crude preparation of cytokines caused a dose-dependent increase in [³H]thymidine uptake into cells obtained by trypsinization of fresh bovine placentae and placental cell lines established from cellular outgrowths of long-term bovine placental cultures, but had no effect on growth of 3T3 fibroblasts. Growth of trypsinized placental cells was not enhanced by culture in the presence of interleukin-2, interferon-β₂, interferon-γ, or tumor necrosis factor-α. These results corroborate those of murine studies, suggesting a growth-promoting role for cytokines released into the maternal-fetal interface. Supported in part by a grant from th e Florida Dairy Farmers Milk Checkoff Fund and a grant from CIBA-GEIGY. This is Journal Series R-01079 of the Florida Agricultural Experiment Station.     

Autoimmune effector cells. IV. Induction of experimental allergic encephalomyelitis in Lewis rats without adjuvant

Experimental autoimmune encephalomyelitis (EAE) was induced in Lewis rats without the aid of adjuvant. Radioresistant cells were activated from spleens of nonimmune donor rats by in vitro culture with myelin basic protein (BP). After adoptive transfer, these cells recruited EAE effector cells in normal syngeneic recipients, as demonstrated after transfer to secondary hosts. The induction of EAE without adjuvant may lead to a better understanding of the mechanisms of autoimmune disease induction.     

Interleukin-6 is constitutively produced by human CTL clones and is required to maintain their cytolytic function

Maturation of cytolytic T lymphocytes from nonlytic precursors requires cytokines in addition to IL2. Interleukin-6 is the principal cytokine that cooperates with IL2 in the induction of CTL differentiation from murine and human thymocyte precursors. However, a cytotoxic differentiation factor (CDF) role of IL6 for mature T cells is challenged by data indicating that IL2 alone is sufficient for CTL generation. The aim of this study was to identify a model system in which IL6 acted as a CDF for human peripheral T cells. We noted that IL6 was endogenously produced by CTL clones in the course of their expansion with APC, lectin, and IL2. The majority of several hundred T-cell clones, both CD4+ and CD8+, produced IL6 in response to relatively high doses of IL2. Other experiments that compared the cytolytic function of CTL clones cultured in the presence of IL6 with that of the same clones cultured in the absence of IL6 demonstrated that IL6 contributes to the cytolytic ability of the majority of human CTL clones. Our data suggest that IL6 acts in an autocrine fashion to support CTL differentiation in human T-cell clones.     

Lineage-independent activation of immune system effector function by myeloid Fc receptors.

An emerging theme in immunology finds receptors which initiate cellular effector programs forming multichain complexes in which the ligand recognition elements associate with one or more 'trigger molecules' whose aggregation initiates a signal transduction cascade. The sequence motifs constituting the active sites of these trigger molecules are found in the T cell and B cell antigen receptors, and some Fc receptors, and appear to be central to effector function activation. For example, of the many molecules that mimic or potentiate the action of the T cell antigen receptor (TCR), none have yet been found to initiate effector programs autonomously in cells lacking TCR. We have devised two strategies to study activation mediated by myeloid Fc receptors, which appear not to associate with trigger molecules: the use of primary human cytolytic T cells as surrogate effector cells for genetically delivered receptors, and the use of vaccinia virus vectors to introduce genetically modified receptors into primary human monocytes. Using these approaches, we have found that the cytoplasmic domains of two Fc receptors show comparable function to equivalent domains of the trigger molecule family, but are not homologous to members of that family.     

Features of the initial stage of surface colonization by Eshcherichia coli cells as a function of their mobility and chemotaxis]

Dependence of adhesion and colonization of hydrophilic and hydrophobic surfaces by Escherichia coli strains with different mobility and chemotaxis was studied using E. coli mot+che+, E. coli mot+che-, E. coli mot-che+. Primary adhesion was shown to correlate with mobility of cells and hydrophilic/hydrophobic character of their surface. Secondary adhesion correlated in addition with chemotactic characteristics of bacteria. E. coli populations were shown to vary in electrophoretic mobility and cells capability for adhesion and chemotaxis.