Effect of Pregnancy Specific ß1-Glycoprotein on the Replicative Potential of Naïve T Cells and Immune Memory T Cells.

We studied the effects of pregnancy-specific ß1-glycoprotein (PSG) on the replicative potential of naïve T cells (CD45RA+) and immune memory T cells (CD45R0+) in vitro by evaluating the expression of the hTERT gene in combination with the proliferative activity of cells. Human PSG was obtained by the author's patented method of immunopurification using a biospecific sorbent with subsequent removal of immunoglobulin contamination on a HiTrap Protein G HP column. We used monocultures of CD45RA+ and CD45R0+ lymphocytes isolated from peripheral blood mononuclear cells of reproductive-age women. It was found that PSG in physiological concentrations inhibited the expression of the hTERT gene mRNA in naïve T cells and immune memory T cells and simultaneously reduced the number of proliferating T cells estimated by the differential gating method. At the same time, PSG reduced CD71 expression only on naïve T cells without affecting this molecule on immune memory T cells. Thus, PSG decreased the replication potential and suppressed the proliferation of T cells and immune memory T cells, which in the context of pregnancy can contribute to the formation of immune tolerance to the semi-allogeneic embryo.

Recombinant Pregnancy-Specific Glycoprotein 1 Has a Protective Role in a Murine Model of Acute Graft-versus-Host Disease.

Acute graft-versus-host disease (aGVHD) is an immune-mediated reaction that can occur after hematopoietic stem cell transplantation in which donor T cells recognize the host antigens as foreign, destroying host tissues. Establishment of a tolerogenic immune environment while preserving the immune response to infectious agents is required for successful bone marrow transplantation. Pregnancy-specific glycoprotein 1 (PSG1), which is secreted by the human placenta into the maternal circulation throughout pregnancy, likely plays a role in maintaining immunotolerance to prevent rejection of the fetus by the maternal immune system. We have previously shown that PSG1 activates the latent form of transforming growth factor ß1 (TGF-ß), a cytokine essential for the differentiation of tolerance-inducing CD4+FoxP3+ regulatory T cells (Tregs). Consistent with this observation, treatment of naïve murine T cells with PSG1 resulted in a significant increase in FoxP3+ cells that was blocked by a TGF-ß receptor I inhibitor. We also show here that PSG1 can increase the availability of active TGF-ß in vivo. As the role of CD4+FoxP3+ cells in the prevention of aGVHD is well established, we tested whether PSG1 has beneficial effects in a murine aGHVD transplantation model. PSG1-treated mice had reduced numbers of tissue-infiltrating inflammatory CD3+ T cells and had increased expression of FoxP3 in T cells compared with vehicle-treated mice. In addition, administration of PSG1 significantly inhibited aGVHD-associated weight loss and mortality. On the other hand, administration of PSG1 was less effective in managing aGVHD in the presence of an alloimmune reaction against a malignancy in a graft-versus-leukemia experimental model. Combined, this data strongly suggests that PSG1 could be a promising treatment option for patients with aGVHD following bone marrow transplantation for a nonmalignant condition, such as an autoimmune disorder or a genetic immunodeficiency.

Pregnancy-specific glycoproteins function as immunomodulators by inducing secretion of IL-10, IL-6 and TGF-beta1 by human monocytes.

PROBLEM: Low levels of pregnancy-specific glycoproteins (PSGs) in maternal serum have been correlated with complications of pregnancy. We investigated the ability of human PSGs to regulate in vitro production of cytokines. METHOD OF STUDY: Human monocytes and murine RAW 264.7 cells were treated with recombinant PSG1, PSG6, PSG11, or a truncated PSG6 consisting of only the N-terminal domain (PSG6N). Cytokine production in response to PSG-treatment was measured by ELISA and/or reverse transcriptase-PCR. RESULTS: All PSGs tested induced secretion of interleukin (IL)-10, IL-6 and transforming growth factor (TGF)-beta1 by both human and murine cells, but not IL-1beta, tumor necrosis factor (TNF)-alpha or IL-12. The N-terminal domain of PSG6 was sufficient for induction of monocyte cytokine secretion. Induction of IL-10 and IL-6 was preceded by an increase in the specific mRNAs. CONCLUSIONS: PSG1, PSG6, PSG6N, and PSG11 induce dose-dependent secretion of anti-inflammatory cytokines by human monocytes. Human and murine PSGs exhibit cross-species activity. Our results are consistent with a role for PSGs in modulation of the innate immune system.

Pregnancy-specific glycoprotein gene expression in recurrent aborters: a potential correlation to interleukin-10 expression.

PROBLEM: The expression of the pregnancy-specific glycoprotein (PSG) genes in the uterine endometrium of women experiencing recurrent first-trimester abortions, and potential correlations to cytokine expression were examined. METHOD OF STUDY: Endometrial RNA, isolated from women with a history of either repetitive first-trimester pregnancy losses or uncomplicated pregnancies, was isolated and analyzed for PSG transcripts by the reverse transcriptase-polymerase chain reaction method. PSG genes showing different patterns of expression were expressed in baculovirus, and the purified proteins examined for their effects on cytokine expression. RESULTS: The expression of PSG11 in the endometria of recurrent aborters was significantly lower than in that of controls (P < 0.01). When tested on monocytes, PSG11 stimulated secretion of interleukin (IL)-10. CONCLUSIONS: The level of expression of the PSG11 gene in the uterine endometrium, during the peri-implantation period, correlates with the risk of pregnancy loss in some women experiencing recurrent spontaneous abortions. The ability of PSG11 to influence the secretion of IL-10 suggests that PSG11 may contribute to the local modulation of the inflammatory T helper-1 response seen in the endometrium of these women.

Effect of pregnancy-specific beta 1-glycoprotein on the development of preimplantation embryo.

The objective of this study is to test the hypothesis that members of the pregnancy-specific beta 1-glycoprotein (PSG) family enhance the growth and maturation of embryos. cDNA encoding two members of the PSG family, namely PSG1 and PSG3, were expressed in Chinese Hamster Ovary (CHO) cells with the expression vector pH beta APr-1-neo. Two-cell stage mouse embryos were co-cultured in a two-chamber system with CHO cells expressing either recombinant PSG1 (rPSG1) or PSG3 (rPSG3) in the presence and absence of neutralizing PSG antibodies. The cleavage and maturation stage of the embryos was assessed at 12-hr intervals. Mouse embryos co-cultured with transfectants expressing rPSG1 showed a significant enhancement of cleavage and maturation rate compared to controls with P < 0.005-0.004. In co-cultures with CHO cells expressing rPSG3, no significant difference from the controls was observed in the early stage of development until late blastocyst formation. At that stage, there was a statistically significant enhancement of development by rPSG3 when compared to controls with P < 0.001. These results suggest that PSG1 and PSG3 exhibit embryotropic activity at different stages of development in the mouse model.

Effect of human pregnancy-specific beta1-glycoprotein on blood cell regeneration after bone marrow transplantation.

Pregnancy-specific beta1-glycoprotein (PSG) is composed of a family of highly homologous proteins initially isolated from human placenta and pregnancy serum. Recent studies showed that PSGs are also present in a number of ectopic sites, including uncultured peripheral blood and bone marrow cells. This report aims at studying the in vivo effect of the PSGs on murine hematopoiesis. The profile of recovery of blood cells after transplantation of viable nucleated bone marrow cells in gamma-irradiated mice with and without the administration of the purified human protein was studied. Five groups of mice were given 0.1 microg human serum albumin, 0.1 microg IL6, 1 microg PSG, 10 microg PSG, and 50 microg PSG, respectively, per mouse per day consecutively for 20 days. The mice were bled once every 2 days, and the platelet and WBC counts were determined using a Nebauer hemacytometer (Hausser Scientific, Buffalo, NY). The recovery of platelet count after bone marrow transplant was much faster in mice receiving 1 microg PSG/day than in animals in any other group. On Day 20 post-transplant, the platelet count of animals in this group reached 178,600 +/- 15,759/microl (mean +/- standard deviation) which was significantly (P < 0.05) higher than that of any other group. On Day 26, the platelet count reached a low normal value of 190,844 +/- 6,380/microl with a range of 185,420-200,500/microl. This value was 3-fold higher than that of the control group (68,600 +/- 15,486/microl in the human serum albumin group). Mice given 1 microg or 10 microg PSG/day also had their WBC count recover significantly faster and achieved a normal value (12,440 +/- 3,680/microl for the 1-microg PSG group, and 12,154 +/- 3,016/microl for the 10-microg PSG group) within the experimental period. On the other hand, the controls, or mice given 50 microg PSG/day did not recover as rapidly and did not achieve a normal WBC count within the experimental period. These results suggest that human placental PSGs enhance platelet and WBC recovery after bone marrow transplant.

[The effect of trophoblast-specific beta 1-glycoprotein and oncoprecipitin A on the growth and morphology of HeLa-M tumor cells]. / Vliianie trofoblast-spetsificheskogo beta 1-glikoproteina i onkopretsipitina A na rost i morfologiiu opukholevykh kletok linii HeLa-M.

Pregnancy-specific beta 1-glycoprotein (PS-1), one of a large family in oncofetal antigens, and oncoprecipitin A, a specific glycoprotein isolated from ascidians, have been shown to reduce cell proliferative activity in HeLa-M cells but almost not to change RNA synthesis. Using scanning electron microscopy, distinct surface transductions in the studied cells were demonstrated. After the incubation of cells with oncoprecipitin A, cell shape alteration and intensive cell spreading were observed. The treatment of tumor cells with PS-1 changed cell shape and essentially reduced cell contacts, as compared to control cells. The changes in cell surface and shape correlated with essential reorganization of actin microfilaments. The use of substances influencing the growth and adhesive characteristics of tumor cells may help in understanding mechanisms of cell transformation.

[Effects of trophoblastic beta 1-glycoprotein (TBG) on the functional activity of different cell lines]. / Vliianie trofoblasticheskogo beta 1-glikoproteina (TBG) na funktsional'nuiu aktivnost' razlichnykh kletochnykh linii.

The effect of TBG on the functional activity of different cell lines, spontaneous and Con A induced proliferation of PBL was studied. If concentration of TBG is higher than 50 mu kg/ml it suppresses the proliferation in many used cell lines, except choriocarcinoma and cancer of uterus. The reliable increasing of spontaneous proliferation of PBL, Jurkat and K-562 cells may be observed if concentration is more lower (0.5-15 mu kg/ml). However proliferation of other cell lines corresponds to control level, and Con A induced proliferation of PBL is inhibited. The effect was more marked at 48, as compared to 24 hours of cell incubation with TBG.

[The hydrophobic properties of beta-glycoprotein from the blood serum of pregnant rats]. / Gidrofobnye svoistva beta-glikoproteina syvorotki krovi beremennykh krys.

During immunoelectrophoresis in the presence of tween-80, triton X-100 and ammonium sulfate blood serum beta-glycoprotein of pregnant rats migrated along with beta-globulins as a main single band; its minor components in zones of alpha- and gamma-globulins were not detected. beta-glycoprotein was completely absorbed by phenyl sepharose in the absence of ligand as well as when the spacer arm for phenyl group was short. When the phenyl group was linked with the template through a long spacer arm, three froms of beta-glycoprotein with different immunoelectrophoretic mobility were detected after absorbtion with phenyl sepharose. Hence, beta-glycoprotein is hydrophobic and is represented by alpha-, beta- and gamma-forms in blood plasma of pregnant rats.

[Effect of trophoblast-specific beta 1-glycoprotein on the ratio of cellular forms in a mast cell population]. / Vliianie trofoblast-spetsificheskogo beta 1-glikoproteina na sootnoshenie kletochnykh form v populiatsii tuchnykh kletok.

The influence of trophoblast-specific beta 1-glycoprotein (TSG) on the degranulation of mast cells and their saturation with heparin was studied. Introduction of the TSG into the population of mast cells of the rat peritoneal fluid practically does not change their degranulation, but lowers the degree of their saturation with heparin. An antibiotic alone increases the saturation of the cells with heparin. The serum of an allergic animal markedly stimulates the degranulation and lowers the degree of saturation of the mast cells with heparin. In an experimental model (antibiotic--the serum of the allergic mast cells) the mast cells transform into very clear (heparin-free) cells and the degree of saturation is at minimum. The TSG introduction into this system stabilizes the population of mast cells and markedly increases the degree of their saturation with heparin. Although the degranulation is rather intensive, it is less expressed, than in the experimental model. This suggests the presence of TSG receptors on the mast cells (targets of allergic reactions). The possibility to use TSG preparations in the therapy of allergic diseases is discussed.

[Comparative study of the immunosuppressive properties of trophoblastic beta 1-glycoprotein and placental alpha 2-microglobulin in vitro]. / Sravnitel'noe izuchenie immunosupressivnykh svoistv trofoblasticheskogo beta 1-glikoproteida i platsentarnogo al'fa 2-mikroglobulina in vitro.

It has been shown that trophoblastic beta 1-glycoprotein (TBG) and placental alpha 2-microglobulin (PAMG-2) in concentrations 60-120 micrograms/ml suppresses both the inductive and proliferative phase of unidirectional mixed lymphocyte reaction in mice, as well as proliferative responses to phytohemagglutinin or pokeweed mitogen. TBG protein was more effective. The proteins were not toxic for lymphocytes.

[Interaction of pregnancy proteins with phytohemagglutinin P and concanavalin A (con A)]. / Vzaimodeistvie belkov beremennosti s fitohemaggliutininom P i konkanavalinom A (kon A).

It has been established that trophoblast-specific beta-glycoprotein and pregnancy-associated alpha 2-glycoprotein specifically react with non-fixed PHA and Con A. Affinity to the former protein is significantly higher than to the latter one. alpha-Fetoprotein has a low affinity to Con A alone. Affinity to this lectin is in an agreement with the content of carbohydrates contained by pregnancy proteins. A considerable part of human serum proteins bind with Con A; receptors for PHA possess only some serum proteins. As the above-mentioned lectins are often used for stimulation of lymphocyte blast transformation, in is recommended that the constituent parts of the culture medium should be preliminarily tested to specify more accurately their affinity to PHA and Con A.

Pregnancy-specific-beta 1-glycoprotein: effect on lymphocyte proliferation in vitro.

The effects of three different preparations of pregnancy-specific-beta 1-glycoprotein (SP1) were investigated in parallel with progesterone, oestradiol and human chorionic gonadotrophin (hCG) on the proliferative response of lymphocytes in the mixed lymphocyte reaction (MLR), and to the mitogens phytohaemagglutinin (PHA) and pokeweed mitogen (PWM). A dose-related inhibition of the MLR was obtained with SP1 at 2.5-10 mg/l. SP1 also inhibited the PHA response at 10 mg/l, but had no effect on the PWM response at these concentrations. Progesterone and oestradiol inhibited all systems at 10-20 mg/l while hCG inhibited all systems at 1500-3000 IU/ml. The observations suggest that SP1 selectively inhibits the proliferative responses of T lymphocytes at the concentrations studied.

[Effect of specific placental proteins--trophoblastic beta 1-glycoprotein chorionic alpha 1-microglobulin--on the proliferation of lymphocytes and malignant fibroblasts in vitro]. / Deistvie spetsificheskikh platsentarnykh belkov--trofoblasticheskogo beta 1-glikoproteida 1 khorionicheskogo al'fa 1-mikroglobulina--na profileratsiiu limfotsitov i zlokachestvennykh fibroblastov in vitro.

A study was made of the action of trophoblastic beta 1-glycoprotein (TBG) and chorionic alpha 1-microglobulin (CAG1) on proliferation of malignant fibroblasts (transplanted L line) and on phytohemagglutinin-stimulated lymphocytes. TBG depressed proliferation of the stimulated lymphocytes and transformed fibroblasts (according to 3H-thymidine incorporation). A dose-response dependence was ascertained. CAG1 did not affect cell division. The inhibitory effect of TBG was seen to be reversed or decreased provided the lymphocyte culture was supplemented with CAG1. The decreased inhibitory effect of TBG in the presence of CAG1 was also noted in the L cell culture. It is likely that in vivo protection of intensely proliferating fetal tissues or tumor from the inhibitors is effected just in this way since placental proteins are synthesized both by embryonic and tumorous cells.