The Role of Membrane Progesterone Receptor Associated Proteins in Gynecological and Reproductive Disorders, And Cancers: An Editor’s Historical Perspective

How can a fetus with half of the antigens from a paternal source not be immunologically rejected? There is evidence that not only does the state of pregnancy fail to preclude invasion of cellular immune cells into uterine tissue, but, in fact, by progesterone (P) blocking the biogenic comma after dopamine, and a lower case t for this: it should read dopamine. This allows a greater infiltration of leukocytes. This invasion seems to be needed to aid in the creation of thinwalled spiral arteries for nutrient exchange between mother and fetus. Related to the speed of the development of these spiral arteries, it is not likely that the main mechanism involves neovascularization, since this is a slow genomic process which would operate by activation of nuclear progesterone receptors (nPRs). Instead, remodeling of the already pre-existing thick-walled uterine arteries by autoimmune mechanisms is more likely. Could the fetal placental unit somehow preclude these cellular immune cells from invading the fetal placental unit? These cells do, in fact infiltrate the fetal placental microenvironment composed of 70% natural killer cells, 20% macrophages, and 10% cytotoxic T-cells. Evidence does exist that one of the main ways of preventing immune rejection of the fetus is by P activating rapid acting membrane (m) PRs to produce immunomodulatory proteins e.g., the progesterone induced blocking factor (PIBF) and the progesterone receptor membrane component-1 protein (PGRMC-1). PIBF, for example, eventually suppresses natural killer cell cytotoxicity by stabilizing perforin granules and granzymes. Understanding these mechanisms has led to a scientifically based treatment regimen to achieve a successful pregnancy.

A unique 34 kDa protein expressed by gamma delta thymic (T) cells during pregnancy suppresses natural killer (NK) cell activity and helps to shift the balance of thymic helper (TH) 1 to TH2 cytokine dominance. The mechanism involved in making this immunomodulaotry protein involves de novo induction of progesterone (P) receptors on gamma delta T cells by the allogeneic stimulus of the fetal semi-allograft and the interaction of the P receptors with a high concentration of P generated at the maternal fetal interface. There is evidence that certain cancer cells may use a similar mechanism to escape NK cell surveillance as evidenced by all 27 different human leukemia cells tested showing a very high level of messenger RNA for this immunomodulatory protein known as the progesterone induced blocking factor (PIBF). Furthermore adding the P receptor antagonist mifepristone down-regulated PIBF expression. PIBF may be synthesized by gamma delta T cells in the tumor microenvironment of solid tumors thus inhibiting NK cell immunosurveillance. Previous studies were performed using an immunocytochemistry technique using a polyclonal antibody because the PIBF protein was not purified. Recombinant DNA technology has allowed the production of a pure protein and thus the development of a monoclonal antibody. The present study was aimed at determining if PIBF is a soluble protein, which if it is could lead to the development of a much more practical and sensitive enzyme linked immunoabsorbent assay (ELISA). Serum samples were blindly assessed for the presence of PIBF using a novel sandwich ELISA we developed with full length (amino acid 1 to 757) recombinant human PIBF, rabbit polyclonal antibody to amino acid 1-300, and affinity purified goat polyclonal antibody to the internal region of PIBF conjugated to horseradish peroxidase. Color was developed using tetramethylbenzidine substrate stopped by 2N H2SO4 and read at 450 lambda. Bovine serum albumin served as negative control. Soluble PIBF was detected in several samples, with highest levels in pregnant patients at over 10 microgram/ml concentration. There were significant inter-individual differences. PIBF, does indeed also exist in a soluble form and can be detected in serum. With further refinement of our ELISA we aim to improve sensitivity and specificity, allowing for the simple measurement of PIBF on a rapid, high throughput basis, making it a more practical test to predict cancers that would benefit from progesterone receptor antagonist therapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 785. doi:10.1158/1538-7445.AM2011-785

We have shown Preeclampsia (PE) to be a progesterone deficient state of pregnancy associated with increased TH1 lymphocytes, cytolytic natural killer (NK) cells, inflammatory cytokine, vasoactive pathways resulting in endothelial function and hypertension. Healthy normal pregnancy (NP) is associated with elevations in progesterone and TH2 lymphocytes, and uterine NK cells favoring immunotolerance toward the fetus. Importantly, NP lymphocytes express progesterone receptors, which stimulate Progesterone Induced Blocking Factor (PIBF) upon binding to progesterone, thus PIBF increases during NP. Therefore we hypothesize that PIBF blockade causes a proinflammatory hypertensive phenotype similar to PE. Rabbit anti-PIBF IgG (0.50 mg/mL) was administered i.p. on gestational day 15 (GD 15) to NP rats, on GD 18 carotid catheters were inserted and on GD 19 mean arterial blood pressure (MAP) and samples were collected. MAP in NP rats (n=7) was 99+ 3 mmHg, which increased to 113+4 mmHg in NP+ anti-PIBF (n=6), p<0.05. Plasma TNF-α was 35+8 pg/mL in NP rats and increased to 84+21 pg/mL in NP+ Anti-PIBF (n=4), p<0.05. Circulating total NK cells were 67+ 11 in NP rats (n=4), which decreased to 36+4 in NP+ Anti-PIBF, while cytolytic NK cells were 0.6 + 0.2 in NP and increased to 3.0+1 in NP+ Anti-PIBF, p<0.05. Circulating nitric oxide was 44+ 11 µM in NP rats (n=5), which decreased to 21+1 µM in NP+ Anti-PIBF (n=6), p<0.05, while, renal cortex preproendothelin-1 increased 15 fold in NP+ Anti-PIBF (n=6) compared to NP rats (n=5). In order to determine if PIBF supplementation improves a PE phenotype during pregnancy, PIBF (2.0 µg/mL) was administered i.p. on GD 15 to the reduced uterine perfusion pressure (RUPP) a rat model of PE and compared to control RUPP and NP rats. On GD19, MAP and samples were collected. MAP in NP rats (n=11) was 100+ 2 mmHg, 105 + 3 in NP+PIBF (n=8), 122+ 1 in RUPP rats (n=10), which improved to 110+2 mmHg in RUPP+PIBF rats (n=11), p<0.05. PIBF lowered circulating and placental cytolytic NK cells in RUPPs from 15+6, 2.4+1% gate to 3+2, 0.4+0.1% in RUPP+PIBF(p<0.05, n=4-8). Moreover PIBF supplementation increased circulating IL-4 and TH2 to 40±8 pg/mL, 2+0.6 % gate in RUPP+PIBF (n=4-8 p<0.05) compared to9 ±2 pg/mL, 0.5+ 0.1 % gate in RUPP. Importantly, vasoactive pathways preproendothelin-1 and nitric oxide were normalized in RUPP+PIBF rats compared to RUPP rats, p<0.05. Collectively, these data demonstrate an important role for PIBF to control the inflammatory milieu thereby normalizing vasoactive pathways and maintaining healthy blood pressures during pregnancy.

Human leukemia cancer cell line studies have demonstrated the presence of a 34 kDa intracytoplasmic splice variant of the parent 90 kDa nuclear protein associated with the centrosome that has significant immunosuppressive activity, especially, but not limited to natural killer (NK) cells. One mechanism of suppressing NK cell cytotoxicity is by stabilizing perforin granules. The 34 kDa intracytoplasmic isoform has been shown to be present only in rapidly growing cells, e.g., embryonic, mesenchymal, trophoblast, and cancer cells. During the luteal phase in ovulating women, and throughout pregnancy, there is a rise in the serum level of this immunomodulatory protein. Thus, it is called the progesterone induced blocking factor (PIBF). Interestingly, the abortafacient, mifepristone, has been shown to suppress the 34 kDa intracytoplasmic concentration of PIBF in cancer cell lines, but it does not lower serum PIBF. Based on the demonstration of significant increase in quality and length of life in controlled animal studies, and anecdotal experience in compassionate use treatment of human patients with a variety of advanced cancer types, the FDA has granted an investigator initiated salvage study of 40 patients with advanced non-small cell lung cancer (NSCLC) who have failed a minimum of two rounds of chemo or immunotherapy to be treated with single agent oral mifepristone (www.clinicaltrials.gov). The first 2 patients are doing extremely well (2 years and 1 year so far) with a high quality life without significant disease progression. Sera PIBF levels were obtained on these patients every 2 months using an experimental ELISA assay. For patient 1, a man with stage IV NSCLC with brain metastasis, his PIBF levels (ng/ml) were all in the range of normal males and women in the follicular phase (27, 38, 53, 59, 52, 46, 39, 45, 44, and 49). For patient 2, who is a female positive for the PD-L1 marker, and who progressed despite 3 rounds of chemotherapy and 1 round of nivolumab, her levels were 10, 30, 12, 21, 23, 50, 64, and 30. Thus, these results suggest that measuring serum PIBF levels in patients with very advanced NSCLC has little value in determining which patients will respond to the progesterone receptor modulator mifepristone, or be able to monitor their response to progesterone receptor modulator therapy. Citation Format: Jerome H. Check, Diane Check, Maya Srivastava, Ann DiAntonio. Sera levels of the immunomodulatory protein the progesterone induced blocking factor (PIBF) are not useful in determining which patients with non-small cell lung cancer will respond to progesterone receptor modulators, e.g., mifepristone [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 586.

Preeclampsia (PE) is characterized by new onset hypertension in association with elevated natural killer (NK) cells and inflammatory cytokines which are likely culprits for decreased fetal weight during PE pregnancies. Progesterone induced blocking factor (PIBF) increases during normal pregnancy and has been shown to decrease inflammation and cytolytic NK cells, both of which are increased during PE. Currently, there is no effective treatment for PE except for early delivery of the fetal placental unit, making PE the leading cause for premature births worldwide. We have previously shown that progesterone supplementation with 17-OHPC improves inflammation, fetal weight and blood pressure in the preclinical RUPP rat model of PE. However the mechanism where by progesterone improves the pathophysiology of PE has never been determined. This study was designed to test the hypothesis that PIBF reduces inflammation while improving hypertension in response to placental ischemia. To test this hypothesis, PIBF (2.0 μg/mL) was administered intraperitoneally on gestation day 15 to RUPP or normal pregnant (NP) rats and on day 18 carotid catheters were inserted and on GD 19 blood pressure and samples were collected. MAP in NP rats (n=9) was 101 + 3 and 110 + 3 in NP+PIBF (n=5), 122 + 2 in RUPP rats (n=7), which improved to 110 + 3 mmHg in RUPP+PIBF (n=9), p&lt;0.05. Pup weight was 2.4 + 0.1 g in NP, 2.5 + 0.1 in NP+PIBF, 1.9 + 0.1 in RUPP and improved to 2.2 + 0.1 in RUPP+PIBF. Neither placental weight nor litter size was affected by PIBF. Fetal reabsorption was lower in RUPP+PIBF compared to RUPP rats. Total placental NK cells were 31 + 9 in NP (n=4), 31 + 5 in NP+PIBF (n=4), 42 + 8 in RUPP rats (n=4) and reduced to 26 + 2 in RUPP+PIBF (n=6). Placental cytolytic NK cells were 0.8 + 0.1 in NP, 0.2 + 0.1 in NP+PIBF, and 1.4 + 0.1 in RUPP rats, which decreased to 0.4+0.1 in RUPP+PIBF. CD4 + T cells were decreased from 21 + 6 in RUPP rats (n=4) to 8 + 2 gate in RUPP+PIBF (n=7). Collectively, our findings demonstrate that PIBF, a produce to progesterone receptor stimulation by progesterone, could be a mechanism to improve fetal growth restriction, inflammation as indicated by CD4+ T cells and cytolytic NK cells while normalizing blood pressure in response to placental ischemia during pregnancy.

There is evidence that treatment with mifepristone a P receptor antagonist will provide a marked palliative effect for many human and murine spontaneous cancers. One proposed mechanism is that it may work by suppressing the expression of a unique immunomodulatory protein that amongst other things stabilizes perforin granules in natural killer (NK) cells and thus negates their cytolytic activity. In humans exposure to P causes a rapid rise in the serum of this protein called the PIBF. The origin for the circulating PIBF is from gamma/delta T cells and is initiated with P interacting with P receptors on gamma/delta T cells. PIBF exists as a 90 kDa nuclear protein occupying a centrosomal position in near proximity to BRCA-1 and is found in all rapidly growing cells including cancer cells. A 34-36 kDa splice variant is found in the cytoplasm of all cancer cells tested. Mifepristone has been found to provide marked palliation despite the absence of evidence of circulating PIBF in some cancers not known to be positive for P receptors. One possible mechanism is suppression of the formation of the 34-36 kDa intracytoplasmic form from the parent 90 kDa form. The objective of the present study was to determine if tumors known to be P receptor positive may demonstrate circulating PIBF. In a blinded manner, serum PIBF was measured in 21 serum samples of women with breast cancer obtained prior to surgery using a research ELISA assay for PIBF. The PIBF levels were then compared according to estrogen (E) and P receptor status. The mean level of PIBF (ng/mL) for those who were ER positive but P receptor negative in 7 cases was 31.0+49.7 with a range of 2.1 to 142.5. For 7 cases that were ER positive and P receptor positive the mean level +SD was 24.8+37.46 and the range 2.4 to 108.32. For ER negative and P receptor positive for 7 cases, the mean PIBF was 24.6+28.3 and the range 6.0 to 86.9. There were no significant differences by ANOVA. It is not clear as yet if P receptor antagonist therapy will provide better palliative effect for P receptor positive vs. P receptor negative tumors, but if so, the mechanism (if it is immunologic) does not seem to be related to suppressing increased levels of circulating PIBF. Thus, testing serum with this new ELISA assay does not seem to be a helpful method to determine which patients may benefit from P receptor antagonist therapy. Some recent data suggest that the P receptor antagonist may suppress intracellular PIBF but not circulative PIBF. Thus, these data do not exclude the possibility that PIBF may play a vital role in escape of cancer cells from immune surveillance but it is the intracytoplasmic form that provides immune protection. Citation Format: Jerome H. Check, Anne Rosenberg, Ann DiAntonio, Hallgeir Rui, Rachael Cohen, Gabrielle DiAntonio. Serum levels of the immunomodulatory protein, the progesterone induced blocking factor (PIBF) are not higher in women with progesterone (P) receptor (R) positive vs. negative breast cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1281. doi:10.1158/1538-7445.AM2015-1281

Recently a sensitive ELISA assay been created to detect serum levels of the immunomodulatory protein, the PIBF. One of the 34 kDa forms of PIBF increases markedly after exposure to progesterone (P) and inhibits natural killer (NK) cytolytic activity by stabilizing perforin granules and causes of shift from thymic helper (TH)1 to TH2 dominance thus diminishing the activity of the cellular immune system. The 34 kDa form is secreted by gamma/delta T cells and is considered a vital step in preventing immune destruction of the fetal semi-allograft. Evaluation of mRNA for PIBF and PIBF protein found that 29 of 29 human leukemia cell lines over-expressed mRNA for PIBF but only 3 of the lines expressed the PIBF protein. Adding P to the culture up-regulated PIBF protein expression whereas adding the P receptor antagonist mifepristone caused down-regulation of PIBF expression. Mifepristone treatment has been demonstrated to show significant palliative effects on a wide variety of murine and human cancers many of which are not known to be P receptor positive. The hypothesized mechanism is that these tumors either secrete PIBF or direct secretion by gamma/delta T cells in the tumor microenvironment and thus inhibit NK cell cytolytic activity. The inhibition is abrogated by inhibiting P receptors and thus a local form of P secretion by the tumor is hypothesized. The objective of this study was to determine if mifepristone will only help inhibit cancer growth in those individuals who have an increased PIBF level present in their serum. An 81 year old woman with chronic lymphocytic leukemia (CLL) started more rapid advancement and became very symptomatic. She refused chemotherapy when she almost had a lethal complication to oral chlorambucil. She requested mifepristone. Her serum PIBF level of 34.9 ng/mL was in the same range as controls without cancer. However within a month of taking 200mg daily of mifepristone orally she showed a dramatic improvement in her CLL with her white blood cell count dropping from 28,000 to 8,000. Her platelet count increased from 40,000 to 240,000 and 2 lung nodules thought to represent possible primary lung cancer (but possibly nodules from her CLL) completely disappeared on repeat CT scan. Also her energy markedly improved and her persistent cough stopped. The serum PIBF did not decrease with mifepristone therapy - 48.3 ng/mL one month later. These data demonstrated that failure to see a marked increase in serum PIBF in a patient with cancer does not predict failure to respond to mifepristone. Though mifepristone has ameliorated murine leukemia in AKR mice this is the first case report of this drug improving human leukemia. The 34-36 kDa PIBF found in cytoplasm of cancer cells may either not be secreted in the serum or may be immunologically different from serum PIBF found after P exposure. Citation Format: Jerome H. Check, Ann DiAntonio, Diane Check, Alex Jaffe, Rachael Cohen, Mojirayo Sarumi, Maya Srivastava. Clinical improvement of symptomatic advancing chronic lymphocytic leukemia following mifepristone therapy despite normal serum levels of the immunomodulatory protein the progesterone induced blocking factor (PIBF). [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 490. doi:10.1158/1538-7445.AM2013-490

The BRCA genes normally provide proteins important in degradating the progesterone (P) receptor. BRCA1 and 2 mutations may make mutated protein, leading to prolongation of P receptor activity which may be involved in the mechanism of how these mutations increase the risk of breast and ovarian cancer. The progesterone induced blocking factor (PIBF) is a protein which suppresses natural killer (NK) cell cytotoxicity, and exposure to P increases both serum levels of PIBF and intracytoplasmic levels in rapidly growing cells (both fetus and cancer cells). The objective of this study was to compare serum PIBF levels in a person with a wild type BRCA2 mutation not known to be associated with malignancy versus the familial type of BRCA2 mutation associated with malignancy. Serum levels of PIBF were obtained in the follicular phase x5 in a woman with a benign wild type heterozygote mutation of BRCA2 (NM000059.3 C.1964 (&amp;gt;(P655R) and in two anovulatory women and a man with familial BRCA2 mutations associated with increased risk of malignancy. The PIBF assay was a research ELISA method using a monoclonal antibody to PIBF. All 5 serum samples of the 21 year old woman with the benign BRCA2 mutation were over 700 ng/mL (levels seen only with high levels of serum P) vs. an average of 55.5 in 2 women with breast cancer at an early age and 1 man positive for the familial BRCA2 mutation. The high levels of serum PIBF seen with the wild type heterozygote mutation show that certain types of BRCA mutations can be associated with increased PIBF levels. Perhaps this BRCA2 wild type mutation is more associated with interfering with degradation of the P receptor found in gamma delta T cells, thus increasing serum PIBF levels. Increased virulence of certain cancers, e.g., breast cancer, has been found with increasing P receptor longevity by either phosphorylation or decreased ubiquitination or sumoylation. Blocking P receptor activity by certain P receptor antagonists, e.g., mifepristone, has been shown to decrease tumor virulence irrespective of the presence or not of P receptors in the tumor. The parent compound for PIBF is a 90 kDa nuclear protein found in the centrosome, but it can be converted to 34-36 kDa immunoprotective intracytoplasmic protein which is similar or identical to the circulating PIBF. Conditions associated with consistent high levels of serum PIBF, e.g., grand multiparous women, are not associated with increased risk of malignancy. Thus an attractive hypothesis (that needs to be proven) is that the BRCA2 mutation (and maybe BRCA1 also) leads to prolongation of the nuclear P receptor leading to increased nuclear PIBF, resulting in increased intracellular PIBF, which, in turn, helps protect the cancer cells especially from NK cell immunosurveillance but potentially other immune factors also. Citation Format: Jerome H. Check, Michael P. Dougherty, Gabrielle DiAntonio, Jamie Vaniver, Marie Duroseau, Maya D. Srivastava. Comparison of serum progesterone levels of the immunomodulatory protein, the progesterone induced blocking factor, in people with BRCA-2 mutations associated with and not associated with a high risk of cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1282. doi:10.1158/1538-7445.AM2015-1282

PIBF is a unique protein that is secreted by gamma/delta T cells and precipitously rises in the sera following progesterone (P) exposure. The 34 kDa protein protects the fetal semi-allograft from immune surveillance by reducing cytoxicity of natural killer (NK) cells by stabilizing perforin granules. The parent 90 kDa form occupies a centrosomal position close to BRCA-1. PIBF can be found in most rapidly dividing cells. One study found an over-expression of the mRNA for the PIBF protein in all 29 leukemia cell lines tested. The P receptor antagonist mifepristone was found to down-regulate PIBF protein. Mifepristone provided significant palliative benefit to a large variety of spontaneous murine and human cancers. P could also be responsible for converting the 90 kDa parent form to a 34-36 kDa split variant in the cytoplasm which may be the immunosuppressive form in cancer cells. Alternatively, some P-like secretion by the tumor could influence gamma/delta T cells in the tumor microenvironment to secrete PIBF and possibly spill over to the sera. The aim of this study was to determine if there is any increase in serum PIBF in women with gynecologic cancer as opposed to controls. Serum was obtained from women about to have surgery for gynecologic problems including malignant and benign disorders. The samples would then be measured for PIBF using a new non-commercial enzyme linked immunoabsorbent assay (ELISA) for PIBF and serum progesterone. The PIBF levels (ng/mL) from lowest to highest in women with various gynecologic cancers (in all cases serum progesterone (P) &amp;lt;2ng/mL) were 10.06, 17.35, 32.59, 35.62, 54,7, and 57.17. The average serum PIBF was 34.6 ng/mL. There were 3 women with benign gynecologic tumors and the serum PIBF was 14.76, 15.7, and 36.64 their average serum PIBF was 22.5 ng/mL). There were 2 women with no tumors having gynecologic surgery and their serum PIBF levels (ng/mL) were 9.56 and 35.27 (with an average of 22.4 ng/mL). At least for gynecologic cancers if PIBF is conferring immune protection it is more likely operating through its intracytoplasmic presence rather than working through sera levels. In women or men exposed to exogenous or endogenous P for just 6 days it is not unusual to see sera PIBF rise to 300 to &amp;gt;800ng/mL. Citation Format: Jerome H. Check, Mojirayo Sarumi, Ann DiAntonio, Krystal Hunter, Gunda Simpkins, Marie Duroseau. A pilot study was initiated to determine if the immunomodulatory protein, the progesterone-induced blocking factor (PIBF), is present in higher quantity in the sera of patients with gynecologic cancer as compared to controls without cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4096. doi:10.1158/1538-7445.AM2014-4096

P33. Decreasing of placental progesterone induced blocking factor expression and spiral artery remodeling disturbance in mice preeclampsia model

31-OR

Preeclampsia (PE), new onset hypertension, is a progesterone deficient state and is associated with an imbalance among CD4 + T lymphocytes, natural killer (NK) cells, and inflammatory cytokines which are likely culprits for decreased fetal weight during PE pregnancies. During normal pregnancy activated lymphocytes express progesterone receptors, which enable progesterone to induce a protein called progesterone induced blocking factor (PIBF). PIBF increases during normal pregnancy and has been shown to decrease inflammation and cytolytic NK cells, both of which are increased during PE. Currently, there is no effective treatment for PE except for early delivery, making PE the leading cause for premature births worldwide. We have previously shown that progesterone supplementation with 17‐OHPC improves inflammation, fetal weight and blood pressure in the preclinical RUPP rat model of PE. However the mechanism where by progesterone improves the pathophysiology of PE has never been determined. This study was designed to test the hypothesis that PIBF reduces inflammation while improving hypertension in response to placental ischemia. To test this hypothesis, PIBF (2.0 μg/mL) was administered intraperitoneally on gestation day 15 to RUPP or normal pregnant (NP) rats and on day 18 carotid catheters were inserted and on GD 19 blood pressure and samples were collected. MAP in NP rats (n=9) was 102± 3 and 110 ± 3 in NP+PIBF (n=5), 124± 3 in RUPP rats (n=4), which improved to 110±3 mmHg in RUPP+PIBF (n=8), p&lt;0.05. Pup weight was 2.4± 0.1 g in NP, 2.5± 0.1 in NP+PIBF, 1.9±0.1 in RUPP and improved to 2.2±0.1 in RUPP+PIBF, p&lt;0.05. Total placental NK cells were 31± 9 in NP (n=4), 31±5 in NP+PIBF (n=4), 42 ±8 in RUPP rats (n=4) and reduced to 26± 2 in RUPP+PIBF (n=6). Placental cytolytic NK cells were 0.8±0.1 in NP, 0.2±0.1 in NP+PIBF, and 1.4±0.1 in RUPP rats, which decreased to 0.4+0.1 in RUPP+PIBF, p&lt;0.05. CD4 + T cells were decreased from 21±6 in RUPP rats (n=4) to 8±2 gate in RUPP+PIBF (n=7). Collectively, our findings demonstrate that PIBF, a produce to progesterone receptor stimulation by progesterone, could be a mechanism to improve fetal growth restriction, inflammation as indicated by CD4+ T cells and cytolytic NK cells while normalizing blood pressure in response to placental ischemia during pregnancy. Support or Funding Information Supported by NIH grants RO1HD067541, HL130456 and AMAG Pharmaceuticals This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .

Based on a conglomeration of basic science research studies a model of hypothetical mechanisms of how the fetal semi-allograft can develop into a live baby was created. One of the tenets of the model is that it is necessary to remodel some of the typically thick-walled uterine arteries found during the proliferative phase into thin-walled spiral arteries to allow nutrient exchange between mother and fetus through an autoimmune mechanism during the luteal phase. The model suggests that the rise in progesterone inhibits dopamine. One function of dopamine is to diminish cellular permeability allowing irritants to infuse into the endometrium thus evoking an inflammatory reaction. The body must suppress these cellular immune cells from attacking the fetus. Progesterone activates membrane progesterone receptors which produces immunomodulatory proteins e.g., the Progesterone Induced Blocking Factor (PIBF), which negates the killing effect of cellular immunity. One hypothesis suggested that PIBF may be secreted by malignant tumors facilitating these tumors with foreign antigens to escape immunosurveillance and thus metastasize. Progesterone receptor antagonists e.g., mifepristone, which suppresses PIBF, have successfully increased length and quality of life in patients with a large variety of end stage treatment resistant cancers. Excessive permeability of various tissues was hypothesized to be related to possible relative deficiency of dopamine. A large variety of medical conditions have been ameliorated significantly by the use of dopaminergic drugs e.g. dextroamphetamine or cabergoline. The model explains why certain conditions may get worse premenstrually e.g., pelvic pain or headaches by the added suppressive effect of progesterone on dopamine. Though possibly further research may modify this model, based on the present hypothesis, a large number of treatment refractory conditions had very successful improvement based on treating with drug releasing dopamine thus correcting tissue permeability disorders.

During the first trimester of pregnancy, nonclassical (CD3-, CD56+, CD16-, perforin [P]bright+) natural killer (NK) cells comprise the major decidual lymphocyte population. These cells, in spite of their high perforin content, exert a low cytolytic activity. Peripheral blood lymphocytes of healthy pregnant women produce progesterone-induced blocking factor (PIBF), which inhibits NK activity. PIBF-producing cells are likely to be present in decidua and might contribute to low decidual NK activity. Decidual cells obtained from elective pregnancy termination were double labeled for CD56 and PIBF. We tested the effect of PIBF on perforin liberation by activated peripheral blood NK cells. Sixty percent of decidual lymphocytes were CD56 + and expressed PIBF at the same time. PIBF-treated and untreated peripheral blood NK cells were incubated with K-562 cells, and perforin content of target conjugated NK cells was detected with immunocytochemistry. PIBF treatment of peripheral blood lymphocytes significantly reduced lysis of K-562 cells. Among target bound lymphocytes in PIBF-treated samples, we found a significantly (P < 0.01) higher rate of P+ cells than in untreated samples. These data suggest that PIBF inhibits cytotoxicity of NK cells via a block of degranulation, and since decidual NK cells are PIBF+, it cannot be ruled out that this effect of PIBF contributes to low decidual NK activity.

Preeclampsia (PE) is characterized by new-onset hypertension in association with elevated natural killer (NK) cells and inflammatory cytokines, which are likely culprits for decreased fetal weight during PE pregnancies. As progesterone increases during normal pregnancy, it stimulates progesterone-induced blocking factor (PIBF). PIBF has been shown to decrease inflammation and cytolytic NK cells, both of which are increased during PE. We hypothesized that PIBF reduces inflammation as a mechanism to improve hypertension in the preclinical reduced uterine perfusion pressure (RUPP) rat model of PE. PIBF (2.0 µg/mL) was administered intraperitoneally on gestational day 15 to either RUPP or normal pregnant (NP) rats. On day 18, carotid catheters were inserted. Mean arterial blood pressure (MAP) and samples were collected on day 19. MAP in NP rats (n = 11) was 100 ± 2 mmHg and 105 ± 3 mmHg in NP+PIBF rats (n = 8) and 122 ± 1 mmHg in RUPP rats (n = 10), which improved to 110 ± 2 mmHg in RUPP+PIBF rats (n = 11), P < 0.05. Pup weight was 2.4 ± 0.1 g in NP, 2.5 ± 0.1 g in NP+PIBF, 1.9 ± 0.1 g in RUPP, and improved to 2.1 ± 0.1 g in RUPP+PIBF rats. Circulating and placental cytolytic NK cells, IL-17, and IL-6 were significantly reduced while IL-4 and T helper (TH) 2 cells were significantly increased in RUPP rats after PIBF administration. Importantly, vasoactive pathways preproendothelin-1, nitric oxide, and soluble fms-Like tyrosine Kinase-1 (sFlt-1) were normalized in RUPP+PIBF rats compared with RUPP rats, P < 0.05. Our findings suggest that PIBF normalized IL-4/TH2 cells, which was associated with improved inflammation, fetal growth restriction, and blood pressure in the RUPP rat model of PE.