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The aim of the study was to compare urine and serum concentrations of PIBF at 24-28 gestational weeks in women with preterm birth, with those of women who delivered at term and to evaluate the impact of PIBF on the outcome of pregnancy. Case-control study was performed in period from 1.6.2010-31.7.2013. Biological samples (urine and serum) were collected from 126 pregnant women. All biological samples were obtained at 24-28 gestation weeks. We measured PIBF concentration and compared women who delivered preterm and those who delivered at term. Thirteen of 126 pregnant women (10.3%) who were included in the study delivered preterm. Among women that actually delivered preterm, median concentrations of PIBF were significantly lower (12.3ng/ml; 101.3ng/ml) than in women who delivered at term (77.0ng/ml; 412.7ng/ml). The serum and urine 24-28 gestational weeks PIBF in those who delivered preterm were generally low from 24 to 37 gestational weeks, while the serum and urine PIBF concentration reached a peak in those delivering between 37-38 gestational weeks, even significantly different from those delivering at 39 to 40 and after 40 gestational weeks. Preterm birth may be predictable at 24-28 gestational week by lower than normal pregnancy PIBF values and measurement of PIBF concentration in biological fluids at that time may be of importance in clinical practice., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

The objective of this study was to analyze the maternal serum concentration of progesterone-induced blocking factor (PIBF) with regard to the prediction and the interval between sampling and the onset of preterm birth. A prospective study was conducted on a sample of 37 women with threatened pre-term birth and 41 healthy pregnant women between the 24th and 28th gestational weeks. Out of 37 patients with threatened preterm birth 11 delivered pre-term and three groups of patients were formed: the preterm delivery group, patients with threatened preterm delivery, and healthy pregnant women. In samples that were taken within 5 days before labor started (6/11, 54.5%), PIBF concentrations were significantly lower than in those obtained more than 5 days before labor (5/11, 45.5%; the mean interval between sampling and the onset of labor was 4.1 ± 1.8 days). Multiple regression analysis of the individual contributions of each observed parameter for preterm delivery demonstrated the significant contribution of a lack of PIBF to preterm birth (p = 0.002). Receiver operating characteristics (ROC) analysis was performed to evaluate the diagnostic accuracy of PIBF for the prediction of preterm birth of women with symptoms of pre-term delivery. The PIBF demonstrated an excellent diagnostic value in the prediction of preterm birth with an area under the ROC curve (AUC) of 0.956 (95% CI = 0.884-0.989; p < 0.0001). Our data suggest that pregnancy termination can be predicted by lower than normal pregnancy PIBF values within 5 days before labor and can contribute to the diagnosis of preterm birth., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

Problem: Progesterone induced blocking factor (PIBF) is required for successful pregnancy. Alternative splicing produces PIBF isoforms with different functions. The full-length (90 kDa) PIBF is involved in cell cycle regulation, whereas smaller secreted forms act as cytokines. We aim to examine the PIBF exon pattern and protein isoform profile in normal and failed murine pregnancies., Method of Study: Pregnant Balb/c mice were killed on gestation days 12-14 or 17-19. Normal and resorbed fetuses, placentae, and uterine tissue were used for RNA and protein analysis with RT-PCR and Western blot, respectively., Results: Late pregnancy and resorption were associated with lower expression of the N-terminal exons, together with significantly reduced production of the full-length protein., Conclusion: Reduced production of the full-length PIBF protein might result in disturbed cell cycle regulation and dysregulated trophoblast invasion, while the absence of PIBF isoforms containing exon 2-4 coded sequences might lead to the loss of local immunosuppression., (© 2013 John Wiley & Sons Ltd.)

Invasiveness is a common feature of trophoblast and tumors; however, while tumor invasion is uncontrolled, trophoblast invasion is strictly regulated. Both trophoblast and tumor cells express high levels of the immunomodulatory progesterone-induced blocking factor (PIBF), therefore, we aimed to test the possibility that PIBF might be involved in invasion. To this aim, we used PIBF-silenced or PIBF-treated trophoblast (HTR8/Svneo, and primary trophoblast) and tumor (HT-1080, A549, HCT116, PC3) cell lines. Silencing of PIBF increased invasiveness as well as MMP-2,-9 secretion of HTR8/SVneo, and decreased those of HT-1080 cells. PIBF induced immediate STAT6 activation in both cell lines. Silencing of IL-4Rα abrogated all the above effects of PIBF, suggesting that invasion-related signaling by PIBF is initiated through the IL-4Rα/PIBF-receptor complex. In HTR-8/SVneo, PIBF induced fast, but transient Akt and ERK phosphorylation, whereas in tumor cells, PIBF triggered sustained Akt, ERK, and late STAT3 activation. The late signaling events might be due to indirect action of PIBF. PIBF induced the expression of EGF and HB-EGF in HT-1080 cells. The STAT3-activating effect of PIBF was reduced in HB-EGF-deficient HT-1080 cells, suggesting that PIBF-induced HB-EGF contributes to late STAT3 activation. PIBF binds to the promoters of IL-6, EGF, and HB-EGF; however, the protein profile of the protein/DNA complex is different in the two cell lines. We conclude that in tumor cells, PIBF induces proteins, which activate invasion signaling, while-based on our previous data-PIBF might control trophoblast invasion by suppressing proinvasive genes.

Problem: The role of progesterone-dependent immunomodulation in the maintenance of normal pregnancy., Methods: In vitro and in vivo data on the effect that progesterone and its mediator progesterone-induced blocking factor (PIBF) exert on the immune functions of pregnant women are reviewed, together with clinical findings., Results: Activated pregnancy lymphocytes express progesterone receptors, which enable progesterone to induce a protein called PIBF. PIBF increases Th2 type cytokine production by signaling via a novel type of IL-4 receptor and activating the Jak/STAT pathway. PIBF inhibits phosholipase A2, thus reduces prostaglandin synthesis. PIBF inhibits perforin release in human decidual lymphocytes and reduces the deleterious effect of high NK activity on murine pregnancy. PIBF production is a characteristic feature of normal human pregnancy, and its concentration is reduced in threatened pregnancies. PIBF mRNA and protein are expressed in a variety of malignant tumors. Inhibition of PIBF synthesis increases survival rates of leukemic mice., Conclusion: Progesterone-induced blocking factor is produced by pregnancy lymphocytes and also by malignant tumors. The PIBF-induced Th2-dominant immune response is favorable during pregnancy but might facilitate tumor growth by suppressing local antitumor immune responses.

Problem: The objective of this study was to compare serum concentrations of progesterone-induced blocking factor (PIBF), anti-inflammatory (IL-10),and pro-inflammatory (IL-6, TNFa, and IFNc) cytokines of women with threatened pre-term delivery, with those of women with normal pregnancy and to evaluate the impact of PIBF on the outcome of pregnancy., Method of Study: A prospective study was conducted on a sample of 30 women with threatened pre-term delivery (study group) and 20 healthy pregnant women (control group) between the 24th and 37th gestational weeks. Serum PIBF, anti-inflammatory (IL-10), and pro-inflammatory (IL-6, TNFa, and IFNc) cytokine concentrations were measured by enzyme-linked immunosorbent assay (ELISA)., Results: Thirteen of 30 patients (43.3%) with symptoms of threatened pre-term delivery, and one of 20 patients (5%) in the control group delivered before the 37th week of gestation. Mean PIBF concentrations in serum samples of patients with threatened pre-term delivery were significantly lower than in those of healthy pregnant women (171.12 +/-162.06 ng/mL versus 272.85 +/- 114.87 ng/mL; P < 0.05).Women with symptoms of threatened pre-term delivery had significantly lower serum levels of IL-10, and higher levels of IL-6 as well as IFNc compared with healthy controls., Conclusion: Our results indicate that measuring PIBF and cytokine concentrations in serum during pregnancy is feasible and may be important for understanding immunological causes of pre-term delivery.

Progesterone-induced blocking factor (PIBF) induces Th2-dominant cytokine production. Western blotting and EMSA revealed phosphorylation as well as nuclear translocation of STAT6 and inhibition of STAT4 phosphorylation in PIBF-treated cells. The silencing of STAT6 by small interfering RNA reduced the cytokine effects. Because the activation of the STAT6 pathway depends on the ligation of IL-4R, we tested the involvement of IL-4R in PIBF-induced STAT6 activation. Although PIBF does not bind to IL-4R, the blocking of the latter with an Ab abolished PIBF-induced STAT6 activation, whereas the blocking of the IL-13R had no effect. PIBF activated suppressor of cytokine signaling-3 and inhibited IL-12-induced suppressor of cytokine signaling-1 activation. The blocking of IL-4R counteracted all the described effects, suggesting that the PIBF receptor interacts with IL-4R alpha-chain, allowing PIBF to activate the STAT6 pathway. PIBF did not phosphorylate Jak3, suggesting that the gamma-chain is not needed for PIBF signaling. Confocal microscopic analysis revealed a colocalization and at 37 degrees C a cocapping of the FITC PIBF-activated PIBF receptor and PE anti-IL-4R-labeled IL-4R. After the digestion of the cells with phosphatidylinositol-specific phospholipase C, the STAT6-activating effect of PIBF was lost, whereas that of IL-4 remained unaltered. These data suggest the existence of a novel type of IL-4R composed of the IL-4R alpha-chain and the GPI-anchored PIBF receptor.

Previous studies from our laboratory showed that the immunomodulatory effects of progesterone are mediated by a 34-kDa protein, named the progesterone-induced blocking factor (PIBF). Lymphocytes of women with threatened abortion fail to produce this factor. Via inducing a Th2 biased cytokine production and blocking of NK activity, PIBF prevents induced pregnancy loss in mice, suggesting that substitution therapy with PIBF could be useful as an alternative treatment of certain forms of recurrent spontaneous abortions. Our study was aimed at mapping the sequence and structure of PIBF coding cDNA and characterizing the encoded protein product. Screening of a human liver cDNA library revealed a 2765-bp clone with a 2271-bp open reading frame. The PIBF1 cDNA encodes a protein of 757 amino acid residues with an 89-kDa predicted molecular mass, which shows no significant amino acid sequence homology with any known protein. PIBF produced via recombinant technique is recognized by the Ab specific for the secreted lymphocyte PIBF Ab, and possesses the biological activities of the secreted lymphocyte PIBF. The full-length PIBF is associated with the nucleus, whereas secretion of shorter forms, such a 34-kDa protein is induced by activation of the cell. The 48-kDa N-terminal part of PIBF is biologically active, and the part of the molecule, responsible for modulating NK activity is encoded by exons 2-4. These data provide an initial step for exploiting the possible diagnostic and therapeutic potential of this immunomodulatory molecule.