transforming-growth-factor-beta and Abortion--Habitual

Regulatory T cells (Tregs) are essential in tolerizing the maternal immune system toward the semi-allogeneic embryo. In this systematic review, we evaluated the association of levels and function of Tregs in peripheral blood and decidua with recurrent miscarriage (RM), defined as two unexplained miscarriages. We included 18 studies. Ten studies showed a significantly decreased level of Tregs in peripheral blood of non-pregnant women with RM, compared to controls (p < 0.05). In pregnant women with RM, levels of Tregs in the peripheral blood were significantly lower compared to control groups (p = 0.0004), as shown in nine studies. Moreover, seven studies described a decrease of Treg levels in the placenta of pregnant women with RM (p < 0.0001) compared to controls. Accordingly, the median of the relative changes (MRC) between cases and controls in the non-pregnant group (peripheral blood), and the two pregnant groups (peripheral blood and decidua) were -0.18 (-0.27-0), -0.26 (-0.35 to -0.17), and -0.52 (0.63--0.31), respectively. In addition to the assessment of Tregs by phenotype, six out of the 18 included studies investigated the functionality of these cells. These studies showed a lower inhibitory effect of Tregs cells on the proliferation of effector T cells of women with RM compared to fertile women. Also, the expression of IL-10 and TGF-beta was diminished. This systematic review shows that Treg levels and their function are significantly decreased in peripheral blood and decidua of pregnant and non-pregnant women with RM. This underlines the hypothesis that Tregs play a role in the pathogenesis of RM.

Topics: Abortion, Habitual; Female; Forkhead Transcription Factors; Humans; Immune Tolerance; Interleukin-10; Placenta; Pregnancy; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Virtually all known cytokines have been demonstrated to be expressed in the placenta and associated fetal and maternal membranes during normal gestation. In addition to playing their traditional roles as modulators of immunological function, cytokines derived from the placenta and extraplacental membranes, together with other locally-derived growth factors, appear to be implicated in various aspects of implantation and placental development. Imbalances in the intrauterine cytokine milieu around the time of implantation and invasion may play a causative role in disorders associated with early pregnancy failure, and are also associated with the abnormal trophoblast development seen in gestational trophoblastic disease. Cytokines thus appear to be an important component of a paracrine/autocrine communication network operating within the feto-maternal interface to ensure the successful establishment of pregnancy.

Topics: Abortion, Habitual; Chemokines; Cytokines; Embryo Implantation; Female; Growth Substances; Humans; Interferons; Maternal-Fetal Exchange; Placenta; Placentation; Pregnancy; Transforming Growth Factor beta; Trophoblastic Neoplasms; Uterine Neoplasms

Recurrent spontaneous abortion (RSA) is a challenging global issue. Although the cause is unknown, increasing evidence suggests that immunological factors play a crucial role in RSA development. Icaritin (ICT), a natural compound derived from Epimedium, has demonstrated diverse biological activities, including anti-inflammatory, anti-cancer, and immunomodulatory effects. In this study, we aimed to investigate the potential therapeutic role of ICT in mitigating RSA in a mouse model. Specifically, we sought to determine whether ICT could modulate the Treg/Th17 cell imbalance via the TGF-β/SMAD signaling pathway and contributed to improved pregnancy outcomes. We conducted experiments on a mouse model with RSA and administered ICT orally. We then examined the effects of ICT on various types of immune cells including Treg and Th17 cells, and assessed the pregnancy outcomes. We also investigated the potential signaling pathway ICT exerted its effects. Our findings revealed that treatment with ICT led to an increase in Treg cells and a decrease in Th17 cells, which restored immune homeostasis and contributed to improved pregnancy outcomes. Furthermore, our data demonstrated that ICT's effects were mediated through the activation of TGF-β/SMAD signaling components. In conclusion, our study suggested that ICT ameliorated RSA by modulating Treg/Th17 cell imbalance via the TGF-β/SMAD signaling pathway. GENERAL SIGNIFICANCE: Our results highlighted the potential of ICT as a novel therapeutic agent for RSA, offering new insights into the underlying mechanisms and opening avenues for future research and clinical translation.

Topics: Abortion, Habitual; Animals; Female; Humans; Mice; Pregnancy; Signal Transduction; T-Lymphocytes, Regulatory; Th17 Cells; Transforming Growth Factor beta

The molecular mechanisms involved in the pathogenesis of recurrent pregnancy loss (RPL) are not completely recognized. The present study aimed to assess the molecules associated with ATP catabolism and hypoxia besides their related miRNAs in patients with RPL. The frequency of Th17 and Treg cells in PBMCs of RPL women and healthy pregnant women were evaluated with Flow cytometry. The expression levels of CD39, CD73, and Hypoxia-inducible factor-alpha (HIF-1α), miR-18a, miR-30a, and miR-206 in PBMCs of two groups were measured with real-time PCR and western blotting. Then, serum levels of IGF-1, TGF-β, and HIF-1α were measured by ELISA. Our results indicated a higher (p = 0.0002) and lower (p < 0.0001) frequency of Th17 and Treg lymphocytes in RPL women, respectively. The expression level of CD39 decreased in PBMCs of RPL women whereas the level of CD73 and HIF-α increased (p = 0.0010, 0.0023, 0.0006 respectively). The results of CD39 and CD37 were also confirmed by protein analysis (p = 0.0047, 0.0364 respectively). Almost, the same results for CD39 and CD73 expression at mRNA and protein levels were observed in isolated Treg cells. Moreover, we found the higher expression of miR-206 and miRNA-30a (p = 0.0038, 0.0123), but the lower expression of miRNA-18a (p = 0.0101) in RPL. The concentration level of IGF-1, and TGF-β reduced (p = 0.0017, 0.0065 respectively) while the level of HIF-α elevated (p = 0.0235) in serum samples of RPL. In conclusion, we observed the dysregulation of molecules that are involved in ATP catabolism and hypoxia, including CD39, CD73, and HIF-1a which is related to miR-18a, miR-30a, and miR-206 change in RPL women. It may be potentially used for RPL prognosis by more comprehensive future studies.

Topics: Abortion, Habitual; Adenosine Triphosphate; Female; Humans; Hypoxia; Insulin-Like Growth Factor I; MicroRNAs; Pregnancy; Transforming Growth Factor beta

Transforming Growth Factor (TGF-β1) is an anti-inflammatory pleiotropic cytokine, crucial for maternal immune tolerance towards semi-allograft. It acts as a mediator in achieving successful implantation and maintenance of pregnancy.. A total of 300 samples; 150 with Recurrent Pregnancy Loss (RPL) and 150 with no pregnancy loss, in their first trimester were evaluated for circulating levels of TGF-β1 using Enzyme-Linked Immunosorbent Assay (ELISA). Further, the Receiver Operating Characteristics (ROC) analysis was performed to assess the potential of TGF-β1 in the risk prediction of RPL and the prognostic importance in the form of favourable and unfavourable outcome in the existing pregnancy.. The results showed significant elevated levels in women without the history of RPL compared to those with the history of RPL (4783.60 ± 522.95 vs. 4252.18 ± 672.26 pg/mL, p < 0.0001).Further evaluation of follow up data of women with the history of RPL, based on favourable (78%) and unfavourable (22%) outcome of the existent pregnancy showed significantly higher TGF-β1 in women with favourable pregnancy outcome in comparison with those who had a foetal loss (4877.12 ± 460.04 vs. 4075.91 ± 616.17 pg/mL, <0.0001). Furthermore, the Receiver Operating Characteristics (ROC) analysis revealed sufficient importance for risk assessment and very good marker to predict unfavourable event (AUC-0.85, SE = 67%, SP = 88%, p < 0.0001).. Certainly TGF-β1 appears to have predictive importance; however additional studies with large sample size are warranted for further validation.

Topics: Abortion, Habitual; Cytokines; Embryo Implantation; Female; Humans; Pregnancy; Prognosis; Transforming Growth Factor beta; Transforming Growth Factor beta1

Thyroid autoimmunity is considered as the most prevalent autoimmune condition in women in fertility age. There are different clinical evidences indicating the association between thyroid autoimmunity and increased risk of RPL. This study aimed to analyze the association of Tregs and Th17 cells development factors and anti-thyroid peroxidase (anti-TPO) antibodies in RPL patients. Healthy controls (n = 36), TPO + controls (n = 25) and TPO + RPL (n = 32) participated in this study. After blood sampling, the frequency of Th17 and Tregs was evaluated using flow cytometry. Real-time PCR and ELISA was used to assess the status of Tregs and Th17 related transcription factors and cytokines in mRNA and protein level, respectively.. TPO + RPL group showed a higher Th17 frequency compared to healthy controls and TPO + controls groups (p = 0.0002 and p = 0.04, respectively). Additionally, mRNA expression levels of RORγT and IL-17 were significantly higher in TPO + RPL compared to healthy controls and TPO + controls groups. In contrast, Foxp3 and TGFβ expression was lower in TPO + RPL. ELISA findings also indicated a significantly higher IL-17 and lower TGFβ secretion in TPO + RPL compared to healthy controls and TPO + controls. Thyroid autoimmunity should intensely be controlled specially in patients with RPL history.

Topics: Abortion, Habitual; Autoantibodies; Female; Humans; Interleukin-17; Peroxidase; Peroxidases; Pregnancy; RNA, Messenger; T-Lymphocytes, Regulatory; Th17 Cells; Transforming Growth Factor beta

VitD3 may contribute to a successful pregnancy through modulation of immune responses. Therefore, VitD3 deficiency may have a role in the immunopathogenesis of unexplained recurrent spontaneous abortion (URSA). However, the mechanisms of immunomodulatory actions of VitD3 in decreasing the risk of recurrent spontaneous abortion have not been understood well.. The purpose of this research was to investigate the influence of 1,25VitD3 on regulatory T cells /Th17 axis, the gene expressions and concentrations of related cytokines including, TGF-β, IL-10, IL-6, IL-23, and IL-17A in peripheral blood mononuclear cells (PBMCs) of healthy women as a control group and women with URSA.. Isolation of PBMCs was performed from peripheral blood of the subjects of the studied groups (20 women with URSA as a case group, and 20 control women). The effects of 1,25VitD3 (50 nM, for 24 hours) on the studied parameters were evaluated and were compared to the positive and negative controls in vitro. Flow cytometry analysis was used to determine the percentages of regulatory T cells and Th17 cells. For gene expression measurement and cytokines assay, Realtime PCR and ELISA were carried out.. The proportion of regulatory T cells was markedly lower, while the proportion of Th17 cells in women with URSA was considerably higher than in the control group (P=0.01, P=0.01). The ratio of the frequency of Tregs to the baseline (1,25VitD3/Untreated) increased, while the ratio of the frequency of Th17 cells to the baseline decreased in women with URSA relative to the controls (P= 0.01, P=0.04). 1,25VitD3 increased IL-10 expressions at both the protein and mRNA levels in PBMCs in women with URSA relative to the control group (P=0.0001, P=0.04). TGF-β levels in the cultured supernatants decreased significantly in the case group in the presence of 1,25Vit- D3 relative to the controls (P=0.03). 1,25VitD3 treatment also significantly decreased gene expressions of IL-6, IL-17A, and IL-23 in PBMCs of women with URSA (P=0.01, P=0.001, P=0.0005), as well as the levels of those cytokines in cell culture supernatants (P=0.03, P=0.02, P=0.01, respectively) in women with URSA relative to the controls.. According to the findings of this research, modulation of immune responses by 1,25VitD3 is accomplished by strengthening Tregs function and inhibiting inflammatory responses of Th17 cells, which may have a positive impact on pregnancy outcome. Thus, as an immunomodulating agent, VitD3 may be effective in reducing the risk of URSA.

Topics: Abortion, Habitual; Cytokines; Female; Humans; Interleukin-10; Interleukin-17; Interleukin-23; Interleukin-6; Leukocytes, Mononuclear; Pregnancy; T-Lymphocytes, Regulatory; Th17 Cells; Transforming Growth Factor beta

Primary cilia regulate cellular signaling and are involved in both sensing and transducing extracellular stimuli. A recent study of patients with recurrent miscarriage (RM) identified mutations affecting DYNC2H1, which were involved in ciliary biogenesis. However, there has been no study concerning primary cilia in the decidua. We compared the number and the length of primary cilia in the decidua of 15 patients with unexplained RM with those of 7 pregnant controls who underwent an artificial termination of pregnancy. Immunohistochemistry was performed using antibodies against primary cilia, extravillous trophoblasts (EVTs), macrophages, uterine Natural Killer (uNK) cells, decidual stromal cells, and the activation of TGF-β and CREB signaling in the decidua of early pregnancy was studied. The density of decidual stromal cells, but not EVTs, macrophages or uNK cells, was found to be significantly higher in the decidua of patients compared to controls. The percentage of ciliated decidual stromal cells was significantly decreased in patients. There was no difference in the primary ciliary length. Regarding TGF-β signaling, p-Smad2 in these cells was diminished significantly in patients, and most of the TGF-β-activated decidual stromal cells of both patients and controls had primary cilia. No difference in the activation of CREB was found. Abnormal primary cilia on decidual stromal cells may be one of the explanatory factors for unknown RM. The inactivation of TGF-β signaling may lead to abnormal ciliogenesis in the decidua.

Topics: Abortion, Habitual; Decidua; Female; Humans; Killer Cells, Natural; Pregnancy; Stromal Cells; Transforming Growth Factor beta; Trophoblasts

The mechanism for traditional Chinese medicine in treating of recurrent spontaneous abortion is not clear. This study aims to explore the mechanism of baotaiyin in the treatment of recurrent abortion by regulating the immune inflammatory axis of interleukin (IL)-23/helper T cell (Th)17.. Spontaneous abortion model mice were randomly divided into a model group, 3 dose (low, medium, and high) groups of baotaiyin, with 10 mice in each group. After 14 days of medication, the levels of IL-17, IL-23, IL-10, and TGF-β in serum were detected with enzyme-linked immunosorbent assay. The proportion of Th17 and regulatory T cells (Treg) cells in spleen lymphocytes was tested with flow cytometry. The expressions of (retinoid-related orphan receptor γt, ROR-γt) and forkhead box P3 (FOXP3) mRNA in decidua tissues was detected with RT-PCR. Embryo absorption rate was counted.. Compared with the model group, the absorption rate of embryo and Th17/Treg cell ratio in baotaiyin medium- and high-dose groups were decreased significantly (all P<0.05); the levels of IL-17 and IL-23 in serum were decreased (both P<0.05), while the levels of TGF-β and IL-10 in baotaiyin medium- and high-dose groups were increased (P<0.05, P<0.01, respectively); the expression of ROR-γt mRNA was decreased and the expression of FOXP3 mRNA was increased (all P<0.01) in decidua tissues of baotaiyin medium- and high-dose groups.. Baotaiyin inhibits the positive feedback cycle of IL-23/Th17 immune inflammatory axis, which regulates Th17/Treg cell balance, mediates the maternal and fetal immune tolerance, and prevents the recurrent abortion.

Topics: Abortion, Habitual; Animals; Female; Humans; Interleukin-10; Interleukin-17; Interleukin-23; Mice; Pregnancy; Transforming Growth Factor beta

Topics: Abortion, Habitual; Biomarkers; Cell Movement; Cell Proliferation; Disease Susceptibility; Female; Gene Expression Regulation; Gene Knockdown Techniques; Humans; Immunohistochemistry; Macrophages; Placenta; Pregnancy; Protein Transport; Transforming Growth Factor beta; Trophoblasts; Ubiquitin Thiolesterase

Pregnancy remains an immune challenge for the uterus that has to adapt to a semi-allogeneic fetus using various regulatory mechanisms. Both HLA-G and regulatory T cells (CD4. We measured the level of both sHLA-G and T. Soluble HLA-G concentrations and T. The results of this study are consistent with previous studies on the role of sHLA-G and T

Topics: Abortion, Habitual; Adult; Case-Control Studies; Cells, Cultured; Female; Forkhead Transcription Factors; HLA-G Antigens; Humans; Immune Tolerance; Interleukin-2 Receptor alpha Subunit; Lymphocyte Activation; Middle Aged; Pregnancy; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Previously, we demonstrated up-regulated activated CD4+ and CD8+ T lymphocytes as well as up-regulated cytotoxic NK cells in the blood of patients with idiopathic recurrent miscarriage. In the present study, we tried to identify deficiencies in counter-regulating immune mechanisms of these patients.. Cytokines were determined in NK cells and in plasma samples of 35 healthy controls, 33 patients with idiopathic recurrent miscarriage, 34 patients with end stage renal disease, 10 transplant patients early and 37 transplant patients late post-transplant using flow-cytometry and luminex. In addition, cytokines were studied in supernatants of cell cultures with peripheral blood mononuclear cells stimulated in-vitro with tumor cell line K562.. Patients with idiopathic recurrent miscarriage exhibited the highest absolute cell counts of circulating TGFß1+ NK, NKT and T lymphocytes and the lowest TGFß1 plasma levels of all study groups (for all p < 0.050). In-vitro, peripheral blood mononuclear cells of patients with idiopathic recurrent miscarriage showed high spontaneous TGFß1 production that could not be further increased by stimulation with K562, indicating increased consumption of TGFß1 by activated cells in the cell culture. Moreover, patients with idiopathic recurrent miscarriage had abnormally high IL4+ as well as abnormally high IFNy+ NK cells (p < 0.010) but similar IL10+ NK cell numbers as female healthy controls and showed the lowest plasma levels of IL10, TGFß3, IL1RA, IL1ß, IL5, IL6, IL8, IL17, TNFα, GM-CSF, TPO and VEGF and the highest plasma levels of G-CSF, FGF-basic, CCL3 and CXCL5 as compared to female HC and female transplant recipients (for all p < 0.050).. Patients with idiopathic recurrent miscarriage show an activated immune system that can hardly be stimulated further and cannot be efficiently down-regulated by up-regulated TGFß1+ and IL4+ NK, NKT and T lymphocytes which are present concomitantly in these patients. The strongly decreased TGFß and IL10 plasma levels indicate deficient down-regulation and reflect a dysbalance of the immune system in patients with idiopathic recurrent miscarriage. These findings may be relevant for explaining the pathogenesis of idiopathic recurrent miscarriage.

Topics: Abortion, Habitual; Biomarkers; Cytokines; Female; Flow Cytometry; Humans; Immunophenotyping; Killer Cells, Natural; Lymphocyte Count; Natural Killer T-Cells; Receptors, Cytokine; T-Lymphocyte Subsets; Transforming Growth Factor beta

Gamma delta (γδ) T cells are known to link innate and adaptive immunity. Decidual γδ T cells are known to provide immunotolerance by producing IL-10 and TGF-β. In recurrent pregnancy loss (RPL) females, the role of peripheral γδ T cells remain unstudied.. To investigate the different phenotypes of γδ T cells in the peripheral blood of women with idiopathic RPL and their possible involvement in RPL condition.. A total of 120 women were recruited for the study. Peripheral blood lymphocytes were isolated and they were stained with appropriate antibodies to determine the phenotype of γδ T cells and major cytokines produced by them in the blood using flow cytometry.. We observed a significant decrease in the proportion of CD3. Increase in IFN-γ and IL-17-producing CD3

Topics: Abortion, Habitual; Adult; Case-Control Studies; Cytokines; Female; Humans; Immunophenotyping; Interferon-gamma; Interleukin-17; Intraepithelial Lymphocytes; Pregnancy; Transforming Growth Factor beta; Young Adult

Immunological mechanisms have been proposed to underlie the pathogenesis of recurrent spontaneous abortion (RSA). Vitamin D has a potent immunomodulatory effect, which may affect pregnancy outcome. The objective of this study was to investigate 25-hydroxyvitamin D [25(OH) D] concentration and vitamin D receptor (VDR) expression in the decidual tissues of RSA patients. Thirty women with RSA (RSA group) and thirty women undergoing elective abortion (control group) were recruited during 2016 from gynecology outpatient clinics. We measured 25(OH) D, interleukin (IL)-17, IL-23, transforming growth factor β (TGF-β), VDR and 1-α-hydroxylase (CYP27B1) in decidual tissues collected during the abortion procedure. In the RSA group, 25(OH) D and TGF-β were significantly decreased while IL-17 and IL-23 were significantly increased compared with the control group. VDR expression was significantly decreased in the RSA group compared with the control group. Logistic regression analysis showed a significant negative correlation between 25(OH) D in decidual tissues and RSA. These results indicated that vitamin D concentrations in the decidua are associated with inflammatory cytokine production, suggesting that vitamin D and VDR may play a role in the etiology of RSA.

Topics: 25-Hydroxyvitamin D3 1-alpha-Hydroxylase; Abortion, Habitual; Adult; Decidua; Female; Humans; Interleukin-17; Interleukin-23; Logistic Models; Pregnancy; Pregnancy Trimester, Third; Receptors, Calcitriol; Risk Factors; Statistics, Nonparametric; Transforming Growth Factor beta; Vitamin D; Vitamin D Deficiency; Young Adult

To investigate the expression levels of adhesion molecule CD49b and negative regulation molecule LAG-3 (CD223) on peripheral blood CD14(+) cells in patients with recurrent spontaneous abortion (RSA).. Peripheral blood samples were collected from 7 normal female individuals and 12 patients with RSA, and then peripheral blood mononuclear cells (PBMCs) and plasma were separated from the peripheral blood via centrifugation. The expression levels of CD49b and LAG-3 on CD14(+) cells in PBMCs were detected by flow cytometry and the levels of interleukin 10 (IL-10) and transforming growth factor β (TGF-β) in plasma were detected by ELISA.. In the RSA patient group, there was no significant difference in the percentage of monocytes compared with that of the normal female group. The numbers of CD14(+) CD49b(+), CD14(+) LAG-3(+) and CD14(+) CD49b(+) LAG-3(+) cells in the RSA patient group were lower than those of the normal female group. The plasma level of TGF-β in the RSA patient group was lower than that of the normal female group. However, there was no significant difference in the plasma level of IL-10 between the two groups.. In RSA patients, the expression levels of CD49b and LAG-3 on CD14(+) monocytes and the plasma level of TGF-β decreased obviously compared with that of the normal females.

Topics: Abortion, Habitual; Adult; Antigens, CD; Cells, Cultured; Enzyme-Linked Immunosorbent Assay; Female; Flow Cytometry; Humans; Integrin alpha2; Interleukin-10; Leukocytes, Mononuclear; Lipopolysaccharide Receptors; Lymphocyte Activation Gene 3 Protein; Transforming Growth Factor beta

To investigate the frequency and function of CD4(+)CD25(+)CD127(dim/-) regulatory T (Treg) cells in decidua of patients with unexplained recurrent spontaneous miscarriage (URSM).. The decidual lymphocytes from patients who experienced URSM and normal pregnant women (controls) were collected by Ficoll density gradient centrifugation. CD4(+)CD25(+)CD127(dim/-) Treg cells were isolated by magnetic cell sorting. The proportion of Treg cells and IL-10, TGF-β in Treg cells were determined by flow cytometry. Inhibitory effects of Treg cells on effecter T cells were detected with or without the presentation of anti-IL-10 antibodies and anti-TGF-β antibodies.. The frequency of CD4(+)CD25(+)CD127(dim/-) Treg cells was decreased in URSM decidua compared to controls (2.09%±0.86% vs. 2.97%±1.19%, p=0.005), and the expression of IL-10 and TGF-β in Treg cells was lower in the URSM group than in the control group (p=0.04 and p=0.01, respectively). Furthermore, the suppressive effect of CD4(+)CD25(+)CD127(dim/-) Treg cells on the proliferation of effector T cells was decreased in URSM decidua (p<0.05). Suppression was mediated predominantly through IL-10 and TGF-β in decidua.. Decreased frequency and immunosuppressive capacity of CD4(+)CD25(+)CD127(dim/-) Treg cells was found in URSM decidua. Treg cells inhibit proliferation of effector T cells mainly via IL-10 and TGF-β in URSM decidua.

Topics: Abortion, Habitual; Adult; Cell Proliferation; Cell Separation; Cells, Cultured; Decidua; Female; Flow Cytometry; Humans; Immune Tolerance; Interleukin-10; Interleukin-7 Receptor alpha Subunit; T-Lymphocyte Subsets; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

CD4(+)CD25(+) T cells (Treg cells) and macrophages play roles in the maintenance of maternal-fetal immunological tolerance. Treg cells suppress the function of macrophages via mechanisms mediated by cell-cell contact and production of soluble factors. The purpose of this study was to investigate regulation of macrophages by Treg cells within decidua from patients with unexplained recurrent miscarriage (RM) and normal control women during early pregnancy. Treg cells and macrophages were isolated from deciduas of unexplained RM (n=15) and control women (n=15) by magnetic cell separation and co-cultured for six days. Regulation of macrophages by Treg cells was assessed in the presence and absence of neutralizing anti-TGFβ antibodies and in transwell experiments. Expression of CD80, CD86, IL10, and IFNγ by macrophages was measured by flow cytometry or ELISA. Macrophage expression of CD80 and CD86 was higher in deciduas of unexplained RM patients compared with controls whereas the expression of IL10 was lower. There was no difference in the expression of IFNγ by macrophages between the two groups. Treg cells inhibited macrophage expression of CD80, CD86 and IFNγ and increased the expression of IL10. The regulatory effects of Treg cells were abrogated in the presence of neutralizing anti-TGFβ antibodies or by transwell culture. The phenotype of macrophages therefore differed in unexplained RM patients compared with normal early pregnant subjects. Macrophage regulation by Treg cells was shown to be mediated by cell-cell contact and TGFβ and this capacity was decreased in unexplained RM patients.

Topics: Abortion, Habitual; Adult; Antibodies, Blocking; Antigens, CD; Cell Communication; Cells, Cultured; Coculture Techniques; Decidua; Female; Gene Expression Regulation; Humans; Interferon-gamma; Interleukin-10; Macrophage Activation; Macrophages; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

We have previously shown a decreased frequency and function of Tregs in women suffering from recurrent spontaneous abortions (RSA). In the current study, we first investigated the expression of FOXP3 after T-cell activation. We observed that expression of FOXP3 in activated PBMCs was already present above baseline before any cell division, indicating that it was induced in cells that were previously negative for this transcription factor. Because RSA women showed a more limited expansion of FOXP3-positive cells, we next assessed the role of IL-2 signaling through STAT5, which is known to be required for generation of inducible Tregs (iTregs). We demonstrated not only that TGF-beta and IL-2 were diminished but also that the IL-2-STAT-5 signaling axis was down regulated in RSA women. Finally, in addition to a limited FOXP3(+) cells expansion in vitro, iTregs from RSA women showed a strikingly lower suppressor activity.

Topics: Abortion, Habitual; Adult; Case-Control Studies; Female; Follicular Phase; Forkhead Transcription Factors; Humans; Immune Tolerance; In Vitro Techniques; Interleukin-2; Interleukin-6; Isoantigens; Kinetics; Lymphocyte Activation; Lymphocyte Culture Test, Mixed; Male; Middle Aged; Pregnancy; Signal Transduction; STAT5 Transcription Factor; T-Lymphocytes, Regulatory; Transforming Growth Factor beta; Young Adult

CD4(+)CD25(+) regulatory T cells (Tregs) are important for the maintenance of immune homeostasis by virtue of their ability to control T-cell proliferation in the peripheral blood (PB). We recently demonstrated that the prevalence of Tregs is decreased, whereas that of Th17 cells is increased, in the PB and decidua samples of patients with unexplained recurrent miscarriage (RM). In this study, we investigated whether the cytokine production of Th17 cells can be suppressed by the Tregs and elucidated the mechanism by which Tregs exert this suppressive effect.. Flow cytometry was used to analyze the surface phenotype and cytokine production of Th17 cells in the PB of women with unexplained RM (n = 17) and healthy women in early stages of pregnancy who underwent elective abortion (n = 20). The suppressive ability of Tregs on Th17 cells was assessed in in vitro co-cultures and transwell experiments. The amount of secreted interleukin-17 (IL-17) in the supernatants was measured by enzyme-linked immunosorbent assay (ELISA). The inhibitory activity of transforming growth factor-β (TGF-β) and IL-10 on IL-17 expression in CD4(+) T cells was assessed using ELISA.. The proportions of IL-17-positive CD4(+) T cells, CC chemokine receptor type 6 (CCR6)-positive CD4(+) T cells and CCR6 expression of IL-17-positive CD4(+) T cells were higher in the PB samples of patients with unexplained RM than in PB of healthy control subjects. In vitro, Tregs could inhibit the expression of IL-17; more Th17 cells were inhibited in the control group than in the unexplained RM group. High-dose TGF-β inhibited the expression of IL-17, whereas IL-10 inhibited IL-17 expression in a dose-dependent manner.. IL-17 expression can be inhibited by Tregs. The suppressive activity of Tregs on Th17 cells was decreased in patients with unexplained RM. The ability of Tregs to suppress cytokine secretion might be effected by a cell-cell contact. TGF-β and IL-10 could inhibit the expression of IL-17.

Topics: Abortion, Habitual; Adult; Cells, Cultured; Female; Humans; Interleukin-10; Interleukin-17; Pregnancy; Receptors, CCR6; T-Lymphocytes, Regulatory; Th17 Cells; Transforming Growth Factor beta

A high proportion of recurrent spontaneous abortions (RSA) remains unexplained. Cytokine genotyping has been investigated. We studied the relationship between unexplained RSA, IL6 (-174 G-->C) and TGFB1 (+869, T-->C; +915, G-->C) gene polymorphisms.. The case-control study composed of 57 south Brazilian women, with unexplained RSA and 74 controls carefully matched was performed. Cytokine genotyping was performed by the polymerase chain reaction-sequence specific primer method, using the 'Cytokine Genotyping Tray'.. The results showed that the genotypic frequencies did not differ from samples for TGFB1 gene. In relation to IL6 gene polymorphism there was a statistical difference in genotypic distribution between samples (P < or = 0.025). The frequency of the C/C genotype was increased in women with RSA in comparison with the frequency observed in controls: 18% versus 4% (P = 0.01).. This result strengthened the importance of IL6 genotypes in the pathogenesis of RSA of unknown cause in the south Brazilian population.

Topics: Abortion, Habitual; Adult; Alleles; Brazil; Case-Control Studies; Female; Gene Frequency; Genotype; Humans; Interleukin-6; Middle Aged; Phenotype; Polymorphism, Genetic; Pregnancy; Transforming Growth Factor beta; Transforming Growth Factor beta1

Transforming growth factor-beta1 (TGF-beta1) is produced by T regulatory lymphocytes (Treg), which play an important role in the physiology of pregnancy. Several polymorphisms of the TGF-beta1 gene (TGFB1) have been reported, some with an important correlation with TGF-beta1 production and disease severity. We performed an association study between TGFB1 polymorphisms and recurrent spontaneous abortion (RSA). We first used a PCR-RFLP method to detect three known TGFB1 cSNPs (coding single nucleotide polymorphisms) among 111 RSA and 110 normal control women from Southern Iran, such as 29T-->C (Leu 10 Pro), 74G-->C (Arg 25 Pro) and 788C-->T (Thr 263Ile), and compared their frequencies between the two groups of subjects. To confirm results of the RFLP study and to identify new SNPs in the RSA women, we then sequenced their DNA samples for seven exons and adjacent intronic regions of TGFB1. Consequently, 10 SNPs were detected; one (-14G-->A) was located in the upstream region of exon 1, three in exons (two in exon 1 and one in exon 5) and six in intronic regions. Two (IVS5+18G-->C and IVS6+910G-->A) of the 10 SNPs were novel. Statistical analysis on the frequency of six most frequent SNPs, including the three cSNPs, as well as on the frequencies of genotypes and 13 haplotypes regarding the 6 SNPs, revealed no significant difference between RSA and control women. Therefore, this study concludes that there is no association between exonic and adjacent intronic polymorphisms of TGFB1 and RSA.

Topics: Abortion, Habitual; Adolescent; Adult; Aged; Exons; Female; Humans; Introns; Middle Aged; Polymorphism, Single Nucleotide; Predictive Value of Tests; Pregnancy; T-Lymphocytes, Regulatory; Transforming Growth Factor beta; Transforming Growth Factor beta1

Recurrent spontaneous abortion (RSA) is regarded as a common pregnancy complication in southern Iran. The exact causes of RSA are not yet known. Transforming growth factor-beta1 (TGF-beta1) is produced by T regulatory lymphocytes (Treg), which play an important role in the physiology of pregnancy. Several polymorphisms of the TGF-beta1 gene have been reported, some with important correlation with disease severity. In this investigation, the polymorphism of the TGF-beta1 gene at promoter region positions -800 (G/A) and -509 (C/T) was studied in 111 RSA and 110 normal female subjects from southern Iran by PCR-RFLP. Results indicated that at position -800 (G/A) polymorphism, 75.7% of RSA cases and 77.3% of normals were homozygote GG. In addition, 23.4% of cases and 22.7% of normal individuals were heterozygote AG. Only one of the patients appeared to be homozygote AA. None of the normal individuals were found to be homozygote AA at this position. In the case of the -509 (C/T) polymorphism, 38.7% of patients and 28.2% of controls were homozygote CC. While 40.6% of cases and 50.9% of normal individuals were heterozygote CT, 20.7% of RSA cases and 20.9% of controls were homozygote TT. The results indicate that there are no statistically significant differences of genotype distribution and allele frequency between RSA cases and controls at both polymorphic sites. In conclusion, the promoter region polymorphisms of TGF-beta1 at positions -800 (G/A) and -509 (C/T) may not be associated with RSA.

Topics: Abortion, Habitual; Adolescent; Adult; Female; Gene Frequency; Humans; Polymorphism, Restriction Fragment Length; Pregnancy; Promoter Regions, Genetic; Transforming Growth Factor beta; Transforming Growth Factor beta1

It has been postulated that a proportion of recurrent miscarriage (RM) might be due to immune causes. The objective was to determine whether cytokine expression in peripheral blood mononuclear cell is altered in patients with a history of RM. We compared the levels of IL-2, IL-4, IL-10, IL-13, TGFbeta1 and IFNgamma in the supernatant of Phytohemagglutinin stimulated mononuclear cells in 21 women with RM at the time of 3rd or higher abortion (group I), 32 women who were at least 3 months past their 3rd or higher abortion (group II) and 32 pregnant women with no history of abortion (group III). Gestational age was matched between groups I and III. Group I had higher level of IL-2 than group III (P=0.001). Group II showed higher level of IL-2 (P=0.001) and IFNgamma (P=0.015) than group III. The production of IL-10 by mononuclear cells of group III was higher than both group I (P=0.002) and group II (P=0.001). There was no difference in the levels of IL-2, IL-10 and IFNgamma between groups I and II. Also, the levels of IL-4, IL-13, and TGFbeta1 were similar among the groups. The data indicate an elevation of Th1 cytokines in women with RM as compared to normal pregnant women, and IL-10 is an important cytokine in the maintenance of pregnancy.

Topics: Abortion, Habitual; Adolescent; Adult; Case-Control Studies; Cell Line; Cytokines; Female; Humans; Interferon-gamma; Interleukin-10; Interleukin-13; Interleukin-2; Interleukin-4; Leukocytes, Mononuclear; Pregnancy; Transforming Growth Factor beta

Topics: Abortion, Habitual; Animals; Apoptosis; Complement System Proteins; Corticotropin-Releasing Hormone; Female; Fertilization in Vitro; Fetus; Histocompatibility Antigens Class I; HLA Antigens; HLA-G Antigens; Humans; Immune System; Immune Tolerance; Infertility, Female; Killer Cells, Natural; Male; Mice; Pre-Eclampsia; Pregnancy; Pregnancy Complications; Reproduction; Semen; T-Lymphocytes; Transforming Growth Factor beta; Trophoblasts; Tryptophan Oxygenase; Uterus

Recurrent spontaneous abortion in DBA/2-mated CBA/J mice has been attributed to the production of Th1 cytokines (tumor necrosis factor [TNF]-alpha and interferon [IFN]-gamma) by asialoGM1+ natural killer (NK) cells and Vgamma1.1delta6.3+ T cells that infiltrate decidua by day 6.5, during the peri-implantation period. Abortions can be prevented by a second population of Vgamma1.1delta6.3 cells, which infiltrate on day 8.5 of gestation, and produce the Th2 cytokine interleukin (IL)-10 and Th3 cytokine transforming growth factor (TGF)-beta2. In low abortion rate immunocompetent mice, most of the TGF-beta2 is derived from gammadelta T cells. However, TGF-beta2-producing cells are present in the decidua of pregnant severe combined immune deficient (SCID) mice, which lack gammadelta T cells.. The cells in day 13.5 decidua of CBA x DBA/2 matings and SCID x SCID matings were identified using flow cytometry and combined surface staining for gammadelta and/or asialoGM1, and intracellular cytokine staining for TNF-alpha, IFN-gamma, and TGF-beta2,3.. TGF-beta2 and TNF-alpha were found in asialoGM1+ NK cells in SCID mouse decidua. In CBA x DBA/2 mated mice, two major and one minor subsets of cytokine-positive cells were identified: -gammadelta-only T cells, double positive asialoGM1+ gammadelta+ (NK-gammadelta T) cells, and a small number of asialoGM1 +gammadelta- NK-only cells. The NK-only and NK-gammadelta T subsets showed a greater Th1/Th2,3 pattern of intracellular staining compared with the gammadelta-only subset. In the CBA x DBA/2 and SCID x SCID systems, Th1/Th2,3 ratios could not predict actual observed abortion rates but did correlate with susceptibility to abortions (if exposed to an additional stimulus such as stress). The known effect of in vivo administration of anti-asialoGM1 antibody on abortion rates within groups of mice exposed to such stresses could also be predicted.. gammadelta+ cells in decidua (e.g. Vgamma1+ cells which can recognize trophoblasts) differ based on the presence or absence of the NK marker-asialo-GM1. NK-gammadelta T cells may be quite important in the Th1 response in early pregnancy that predisposes to abortions in CBA x DBA/2 matings, whereas gammadelta T-only cells appear to be protective. In pregnant SCID mice, the TNF-alpha+/TGF-beta2+ NK population is greatly expanded. An activating stimulus (such as stress or endotoxin) appears to be as important in triggering abortions, as is the Th1/Th2,3 ratio at the feto maternal interface.

Topics: Abortion, Habitual; Animals; Cytokines; Decidua; Female; Interferon-gamma; Interleukin-10; Killer Cells, Natural; Male; Mice; Mice, Inbred C3H; Mice, Inbred CBA; Mice, Inbred DBA; Mice, SCID; Pregnancy; Receptors, Antigen, T-Cell, gamma-delta; Species Specificity; T-Lymphocyte Subsets; T-Lymphocytes, Helper-Inducer; Th1 Cells; Th2 Cells; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

A dichotomous T-helper 1 (Th1) versus T-helper 2 (Th2) cytokine response to trophoblast has been proposed to mediate reproductive failure and success, respectively. Progesterone has immunosuppressive properties. In this study, peripheral blood mononuclear cells from women with and without unexplained recurrent pregnancy loss who had and did not have evidence of embryotoxic, Th1 immunity to trophoblast were cultured with progesterone (10(-5) mol/l) or interleukin (IL)-10 (1500 pg/ml) to determine whether these agents were capable of inhibiting embryotoxic, Th1 immunity to trophoblast. The effects of progesterone on Th2 cytokines and transforming growth factor (TGF)-beta secretion were also assessed. Progesterone was found to specifically block Th1 immunity to trophoblast, as was IL-10. Progesterone also appeared to upregulate TGF-beta secretion in response to trophoblast but had no effect on Th2 cytokine secretion. Our data suggest that assaying Th1 cytokines in supernatants of peripheral blood mononuclear cells cultured with a protein extract from trophoblast may identify individuals more likely to benefit from potentially immunosuppressive doses of progesterone. An appropriately designed clinical trial is needed to determine whether therapies modifying Th1 cytokine secretion in response to trophoblast are useful in the clinical management of recurrent pregnancy loss in women producing these cytokines in response to reproductive antigen stimulation.

Topics: Abortion, Habitual; Animals; Cell Extracts; Cytokines; Dose-Response Relationship, Drug; Embryo, Mammalian; Female; Hormone Antagonists; Humans; Interleukin-10; Leukocytes, Mononuclear; Male; Mice; Mifepristone; Pregnancy; Progesterone; Reference Values; Reproductive History; Th1 Cells; Transforming Growth Factor beta; Trophoblasts

To investigate the significance of transforming growth factor-beta1 (TGF-beta1) in reproduction we have compared plasma levels in normal pregnant women and patients suffering miscarriages. We examined 188 normal pregnant women and 12 pregnant women with miscarriages. Eight women with severe recurrent miscarriages (mean +/- SD of previous number of miscarriages; 10.4 +/- 2.4 times) were also examined before conception; 34 nonpregnant women served as controls. Plasma TGF-beta1 level increased with the gestational week and returned within the normal range 1 month after delivery. The levels among pregnant women with miscarriages (mean +/- SD; 2.44 +/- 0.83 ng/ml) were significantly higher than those of pregnant controls (1.74 +/- 0.95 ng/ml) of matched gestational weeks; levels among nonpregnant women with severe recurrent miscarriages were extremely elevated (4.1 +/- 3.04 ng/ml) compared to the control value (1.34 +/- 0.59 ng/ml). These data suggest that TGF-beta1 may be necessary to maintain pregnancy but also may be a risk factor for recurrent miscarriages.

Topics: Abortion, Habitual; Adult; Female; Humans; Killer Cells, Natural; Pregnancy; Time Factors; Transforming Growth Factor beta; Transforming Growth Factor beta1

Spontaneous recurrent abortion (SRA) has been treated by means of immunization with paternal or third-party white blood cells, yet the immunological basis for SRA and for the role of immunization protocols in pregnancy outcome remains controversial. To elucidate this question, nine women with SRA were immunized with paternal mononuclear cells and studied before and 2 weeks after immunization. Seven women who became pregnant gave birth to live newborns. Secretion of the T helper 1 cytokines IL-2 and interferon-gamma by patients, mononuclear cells decreased, while production of IL-10 increased. The levels of natural killer and lymphokine-activated killer cell-mediated cytotoxicity were markedly decreased. Monocyte functions such as secretion of IL-1 alpha, tumor necrosis factor alpha, IL-6, and cytotoxic activity decreased concurrently with elevations in IL-10 and transforming growth factor beta secretion. Production of IL-12, a pivotal regulatory cytokine, decreased. Furthermore, B7/1 expression on patients' mononuclear cells was downregulated. This resulted in a decrease in monocyte costimulatory activity of purified T cells with soluble anti-CD3, paralleled by a decline in allogeneic proliferative responses. These results suggest that the improved pregnancy success rate in women with SRA following immunization may be partly related to suppression of cell-mediated immunity and monocyte and natural killer cell activity.

Topics: Abortion, Habitual; Adult; Antigen Presentation; B7-1 Antigen; CD3 Complex; Cytotoxicity, Immunologic; Down-Regulation; Female; Humans; Immunity, Cellular; Immunosuppression Therapy; Immunotherapy; Interferon-gamma; Interleukins; Killer Cells, Lymphokine-Activated; Killer Cells, Natural; Leukocytes; Male; Monocytes; Pregnancy; Th1 Cells; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; Vaccination

To determine if patients with unexplained recurrent miscarriage have a deficiency of decidual immunosuppressor cells that produce transforming growth factor beta type 2, as has been found in mice with abortion due to rejection and/or trophoblast failure.. Decidual biopsy specimens were taken as near to the placental attachment site as possible under ultrasound guidance from first trimester legal termination (control) patients with recurrent miscarriage and non-viable pregnancy, and from patients with sporadic missed abortion. The tissue was tested for TGF beta-2+ suppressor cells by in situ hybridization, immunohistochemistry, and analysis of supernatants.. TGF beta-2-related suppressor molecules similar but not identical to those identified in pregnant mice were released by decidual lymphoid cells. Fifty percent of 14 recurrent miscarriage patients showed a lack of suppressor cells and 59% were subnormal in comparison to 20 controls and 5 sporadic miscarriage patients, where 80-85% of the patients had detectable suppressor cells.. Suppressor cell deficiency is compatible with a role for rejection and/or trophoblast failure in some patients with recurrent miscarriage. Presence of suppressor cells in most patients with missed abortion (4/5) is compatible with an alternative cause of fetal death, similar to findings reported in genetic fetal death mice.

Topics: Abortion, Habitual; Cell-Free System; Cells, Cultured; Decidua; Embryo Implantation; Female; Humans; In Situ Hybridization; Pregnancy; Suppressor Factors, Immunologic; T-Lymphocytes, Regulatory; Transforming Growth Factor beta