Arihiro Shiozaki

Proportion of peripheral blood and decidual CD4+ CD25bright regulatory T cells in pre-eclampsia

CD4+ CD25bright regulatory T (Treg) cells have been identified as a principle regulator of tolerance during pregnancy. In the setting of pre‐eclampsia, however, little is known about the dynamics of these cells. In the current study, we determined CD4+ CD25bright Treg cells in the peripheral blood using flow cytometry and forkhead box P3 (FoxP3+) cells at the placental bed using immunohistochemical staining. Peripheral blood mononuclear cells (PBMC) of 38 pre‐eclamptic cases (17 cases Japanese, 21 cases Polish), 40 normal late pregnancy subjects (20 subjects Japanese, 20 subjects Polish), and 21 non‐pregnant healthy controls (10 subjects Japanese, 11 subjects Polish) were included. We found the percentage of CD25bright cells within the CD4+ T cell population in PBMC was reduced significantly in both Japanese and Polish pre‐eclamptic cases than in normal pregnancy subjects (P < 0·001) and non‐pregnant healthy controls (P < 0·001). Also, the percentage of FoxP3+ cells within CD3+ T cells in the placental bed biopsy samples of pre‐eclamptic cases were decreased compared to those in normal pregnancy subjects. These findings suggest that a decreased number of Treg cells was present in pre‐eclampsia, and these changes might break the maternal tolerance to the fetus.

Regulatory T cells and regulatory natural killer (NK) cells play important roles in feto-maternal tolerance

In the early pregnancy decidua, lymphocytes express some activation markers on their surface, suggesting that maternal lymphocytes are activated and recognize the semiallograftic fetus. Therefore, the immunoregulation system must work to prevent fetus rejection. Recent data showed that parts of the immunoregulation system such as CD4+CD25+ regulatory T (Treg) cells, Th3 cells, Tr1 cells, regulatory NK cells, and a tryptophan-catabolizing enzyme, indolamine 2,3 deoxygenase, play very important roles in the maintenance of pregnancy. Not only Treg cells but also regulatory NK cells may inhibit maternal T cell or NK cell fetal attack.

Circulating and decidual Th17 cell levels in healthy pregnancy.

Citation Nakashima A, Ito M, Yoneda S, Shiozaki A, Hidaka T, Saito S. Circulating and decidual Th17 cell levels in healthy pregnancy. Am J Reprod Immunol 2010; 63: 104–109

A role for IL-17 in induction of an inflammation at the fetomaternal interface in preterm labour

Chorioamnionitis (CAM) is a major cause of preterm delivery. Inflammatory cytokines and chemokines play important roles in the pathogenesis of preterm delivery. Interleukin (IL)-17 is a key cytokine which induces inflammation and is critical to host defense. In this study, we examined the role of IL-17 in the pathogenesis of preterm delivery. The levels of cytokines including IL-17, IL-8 and tumor necrosis factor (TNF) alpha were measured by ELISA in amniotic fluid from 154 cases of preterm labor. Flow cytometry and immunohistochemical staining were performed to determine the distribution of IL-17-producing cells. IL-8 secretion was evaluated in primary cultured human amniotic mesenchymal (HAM) cells and human amniotic epithelial (HAE) cells stimulated with IL-17, TNFalpha or IL-1beta. We also studied the signaling pathway of IL-17 and TNFalpha in HAM cells. Levels of inflammatory cytokines in amniotic fluid were higher in preterm delivery cases than in term delivery cases. Furthermore, IL-8, IL-17 and TNFalpha levels were significantly higher in the preterm cases with CAM stage II or III than those without CAM. Flow cytometry and immunohistochemical staining revealed that CD3(+)CD4(+) T cells were the main source of IL-17 in the chorioamniotic membrane. Interestingly, TNFalpha-induced IL-8 secretion was enhanced by IL-17 in a dose-dependent manner in HAM cells. The IKK inhibitor BMS-345541 and mitogen-activated protein kinase (MAPK) inhibitors p38, JNK and p42/44 (ERK1/2 pathway) reduced IL-8 secretion by IL-17-stimulated and TNFalpha-stimulated HAM cells. These results indicate that IL-17, produced by T cells, promotes inflammation at the fetomaternal interface in preterm delivery.

SHORT COMMUNICATION: Circulating and Decidual Th17 Cell Levels in Healthy Pregnancy

Citation Nakashima A, Ito M, Yoneda S, Shiozaki A, Hidaka T, Saito S. Circulating and decidual Th17 cell levels in healthy pregnancy. Am J Reprod Immunol 2010; 63: 104–109

Granulysin produced by uterine natural killer cells induces apoptosis of extravillous trophoblasts in spontaneous abortion.

Immune changes are known to occur in recurrent spontaneous abortion, but it is unclear whether either maternal natural killer (NK) cells or T cells attack fetus-derived trophoblasts. To clarify the immunological causes of spontaneous abortion, we examined the relationship between cytotoxic granule proteins in decidual lymphocytes, such as granulysin, granzyme B, and perforin, and the induction of apoptosis in extravillous trophoblasts (EVTs). The number of granulysin-positive CD56(bright) NK cells increased significantly in the decidua basalis during spontaneous abortion compared with normal pregnancy; however, granzyme B- and perforin-positive cells did not change. Interestingly, the expression of granulysin was also detected in the nuclei of EVTs in spontaneous abortion samples. When IL-2-stimulated CD56(bright) NK cells were cocultured with EVT cells (HTR-8/SV40neo), granulysin was found initially in the cytoplasm and then accumulated in the nuclei of the HTR-8/SV40neo cells. Furthermore, transfected cells expressing a GFP-granulysin fusion protein induced apoptosis in HTR-8/SV40neo cells independently of caspases. Our results suggest that granulysin-positive uterine NK cells attack EVTs; subsequently, the uNK-derived granulysin actively accumulates in the nuclei of EVTs, causing the death of EVTs due to apoptosis. These data support a new apoptosis pathway for trophoblasts via uNK-derived granulysin, suggesting that granulysin is involved in spontaneous abortion.

What is the role of regulatory T cells in the success of implantation and early pregnancy

ProblemThe immune system is well controlled by the balance between immunostimulation and immunoregulation. CD4+CD25+ regulatory T (Treg) cells and an enzyme called indoleamine-2, 3-dioxygenase (IDO) mediate maternal tolerance of the allogeneic fetus. Treg cells, therefore, may prevent early pregnancy loss due to maternal ‘rejection.’MethodsThe latest understanding of tolerance during pregnancy is reviewed.Results and conclusionsRecent data show that CD4+CD25+ Treg cells play essential roles in the induction and maintenance of tolerance, and that they augment the IDO activity in dendritic cells and macrophages. Therefore, CD4+CD25+ Treg cells and IDO enzyme may cooperate in the induction of tolerance during pregnancy. Treg deficiency is associated with very early post-implantation loss and spontaneous abortion in animal models, and low Treg levels are associated with recurrent miscarriages in humans.

Serum granulysin is a marker for Th1 type immunity in pre-eclampsia

Recent studies suggest that pre‐eclampsia is associated with a Th1 predominant state and may be considered a failure of tolerance. Granulysin is a cytotoxic granule protein of natural killer (NK) cells and cytotoxic T lymphocytes (CTLs). Recently, we developed an enzyme‐linked immunosorbent assay (ELISA) system for detecting serum granulysin, and reported that serum granulysin is a useful marker to evaluate the cell‐mediated immunity. In this study, we show that the serum levels of granulysin were significantly elevated in pre‐eclamptic patients compared with those in normal pregnancy subjects. In addition, the serum granulysin levels in pre‐eclamptic patients were well associated with mean blood pressure, percentage of peripheral blood Th1 cells and Th1/Th2 ratios. The present results suggest that the serum granulysin levels would be a useful and novel serum marker to evaluate the Th1/Th2 balance, especially Th1 type immunity in pre‐eclampsia.

Comparison of risk factors for placental abruption and placenta previa: Case-cohort study

Aim:  A case‐cohort study was performed to clarify and compare the risk factors for placental abruption and placenta previa.

Smoking During Pregnancy Increases Risks of Various Obstetric Complications: A Case-Cohort Study of the Japan Perinatal Registry Network Database

Background The adverse effects of maternal smoking on the health of pregnant women have been examined mostly on a disease-by-disease basis. The aims of this study were to evaluate simultaneously the effects of smoking during pregnancy on various obstetric complications, using data from a large medical database, and to investigate the expediency of using a case-cohort design for such an analysis. Methods A case-cohort study was conducted within the Japan Perinatal Registry Network database. Perinatal information on infant deliveries was entered into the database at 125 medical centers in Japan. The base population of the study was 180 855 pregnant women registered in the database from 2001 through 2005. The outcome measures were the incidences of 11 different obstetric complications. Logistic regression models were used to estimate age-adjusted risk ratios (aRRs) and relative excess incidence proportions (REIs). Results The overall prevalence of smoking during pregnancy was 5.8% in the base cohort, and the prevalence was higher among younger women. A comparison of the cases and control cohort showed that smokers during pregnancy had statistically significant higher risks for preterm rupture of the membrane (aRR: 1.67, 95% confidence interval [CI]: 1.43–1.96; REI: 40.2%, 95% CI: 29.9%–49.1%), chorioamnionitis (1.65, 1.36–2.00; 39.4%, 26.4%–50.0%), incompetent cervix (1.63, 1.35–1.96; 38.5%, 25.8%–49.1%), threatened premature delivery (1.38, 1.17–1.64; 27.7%, 14.5%–38.9%), placental abruption (1.37, 1.10–1.72; 27.1%, 8.8%–41.7%), and pregnancy-induced hypertension (1.20, 1.01–1.41; 16.4%, 1.2%–29.3%). Conclusions Maternal smoking was associated with a number of obstetric complications. This highlights the importance of smoking cessation during pregnancy. In addition, case-cohort analysis proved useful in estimating RRs for multiple outcomes in a large database.