Yukio Mano

The Loss of Endoglin Promotes the Invasion of Extravillous Trophoblasts

Endoglin is a coreceptor for TGF-β, which is expressed in syncytiotrophoblasts. The soluble form of endoglin (sEng) has been observed to increase in the serum of preeclamptic patients. Several studies have shown that endoglin is involved in cancer invasion. However, the role of endoglin in extravillous trophoblasts (EVT), which have an invasive phenotype, remains unknown. The present study was designed to investigate the expression and role of endoglin in human EVT. We found that endoglin was mainly expressed on cytotrophoblasts within the cell column during the first trimester and its expression decreased in the EVT by immunohistochemistry and immunocytochemistry. The expression of endoglin significantly increased after treatment with TGF-β1 and TGF-β3 in the human EVT cell line, HTR-8/SVneo, as detected by semiquantitative RT-PCR. To investigate the role of endoglin in EVT, the stable knockdown of endoglin was performed by lentiviral short hairpin RNA transfection into the HTR-8/SVneo cells. Although proliferation was not affected, the motility and invasiveness of the HTR-8/SVneo cells significantly increased by the knockdown of endoglin. Both the mRNA expression and secretion of urokinase-type plasminogen activator significantly increased in endoglin knockdown cells. The secretion of sEng was very low in HTR-8/SVneo, and the treatment of endoglin knockdown cells with 10 ng/ml sEng had no effect on their invasiveness. Therefore, the suppression of sEng was not involved in the increased invasiveness of endoglin knockdown cells. These results suggested that EVT increased their invasive function as a result of decreasing expression of transmembrane endoglin.

Maternal molecular hydrogen administration ameliorates rat fetal hippocampal damage caused by in utero ischemia–reperfusion

Molecular hydrogen (H2) scavenges hydroxyl radicals. Recently, H2 has been reported to prevent a variety of diseases associated with oxidative stress in model systems and in humans. Here, we studied the effects of H2 on rat fetal hippocampal damage caused by ischemia and reperfusion (IR) on day 16 of pregnancy with the transient occlusion of the bilateral utero-ovarian arteries. Starting 2 days before the operation, we provided the mothers with hydrogen-saturated water ad libitum until vaginal delivery. We observed a significant increase in the concentration of H2 in the placenta after the oral administration of hydrogen-saturated water to the mothers, with less placental oxidative damage after IR in the presence of H2. Neonatal growth retardation was observed in the IR group, which was alleviated by the H2 administration. We analyzed the neuronal cell damage in the CA1 and CA3 areas of the hippocampus at day 7 after birth by immunohistochemical analysis of the 8-oxo-7,8-dihydro-2׳-deoxyguanosine- and 4-hydroxy-2-nonenal-modified proteins. Both oxidative stress markers were significantly increased in the IR group, which was again ameliorated by the H2 intake. Last, 8-week-old rats were subjected to a Morris water maze test. Maternal H2 administration improved the reference memory of the offspring to the sham level after IR injury during pregnancy. Overall, the present results support the idea that maternal H2 intake helps prevent the hippocampal impairment of offspring induced by IR during pregnancy.

Uterine sutures at prior caesarean section and placenta accreta in subsequent pregnancy: a case–control study

To clarify the effects of uterine myometrial suture techniques at prior caesarean section on the incidence of pathologically diagnosed placenta accreta in placenta praevia with prior caesarean section (PPPC).

Maternal mental disorders and pregnancy outcomes: A clinical study in a Japanese population

Aim:u2002 To assess the maternal and neonatal outcomes of pregnant women with mental disorders in Japan.

Catalytic ferrous iron in amniotic fluid as a predictive marker of human maternal-fetal disorders

Amniotic fluid contains numerous biomolecules derived from fetus and mother, thus providing precious information on pregnancy. Here, we evaluated oxidative stress of human amniotic fluid and measured the concentration of catalytic Fe(II). Amniotic fluid samples were collected with consent from a total of 89 subjects in Nagoya University Hospital, under necessary medical interventions: normal pregnancy at term, normal pregnancy at the 2nd trimester, preterm delivery with maternal disorders but without fetal disorders, congenital diaphragmatic hernia, fetal growth restriction, pregnancy-induced hypertension, gestational diabetes mellitus, Down syndrome and trisomy 18. Catalytic Fe(II) and oxidative stress markers (8-hydroxy-2-deoxyguanosine, 8-OHdG; dityrosine) were determined with RhoNox-1 and specific antibodies, respectively, using plate assays. Levels of 8-OHdG and dityrosine were higher in the 3rd trimester compared with the 2nd trimester in normal subjects, and the abnormal groups generally showed lower levels than the controls, thus suggesting that they represent fetal metabolic activities. In contrast, catalytic Fe(II) was higher in the 2nd trimester than the 3rd trimester in the normal subjects, and overall the abnormal groups showed higher levels than the controls, suggesting that high catalytic Fe(II) at late gestation reflects fetal pathologic alterations. Notably, products of H2O2 and catalytic Fe(II) remained almost constant in amniotic fluid.

Tocilizumab Inhibits Interleukin-6-Mediated Matrix Metalloproteinase-2 and -9 Secretions from Human Amnion Cells in Preterm Premature Rupture of Membranes

Background/Aims: In the present study, we investigated the participation of inflammatory cytokine-induced mediated matrix metalloproteinase (MMP) expressions and inhibition of interleukin (IL)-6-induced MMP secretion in amniotic epithelial cells by tocilizumab. Methods: To investigate the role of MMP expressions, immunohistochemical staining was performed using membranes obtained from 10 patients with preterm premature rupture of membranes (PPROM) and from 10 patients who underwent a nonlabor cesarean section. We also investigated the regulation of MMP expression by inflammatory cytokines in human amnion cells. Results: Immunohistochemical staining showed a significantly higher expression of MMP-2 and -9 in PPROM. Treatment of cultured WISH and primary amniotic epithelial cells with 10–8 or 10–7M IL-6 or tumor necrosis factor (TNF)-α clearly increased the secretion of MMP-2 and -9. Treatment with 10–8M TNF-α or IL-6 significantly increased the invasion of WISH or primary amniotic epithelial cells, respectively, compared with the control. At a low concentration of 1 μg/ml, tocilizumab (anti-human IL-6 receptor monoclonal antibody) inhibited the IL-6-induced MMP secretion. Conclusions: This paper is the 1st report of tocilizumab inhibiting IL-6-induced MMP-2 and MMP-9 secretions from human amnion cells in PPROM.

Maternal molecular hydrogen treatment attenuates lipopolysaccharide-induced rat fetal lung injury

Abstract Maternal inflammation is associated with spontaneous preterm birth and respiratory impairment among premature infants. Recently, molecular hydrogen (H2) has been reported to have a suppressive effect on oxidative stress and inflammation. The aim of this study was to evaluate the effects of H2 on fetal lung injury caused by maternal inflammation. Cell viability and the production of interleukin-6 (IL-6) and reactive oxygen species (ROS) were examined by treatment with lipopolysaccharide (LPS) contained in ordinal or H2-rich medium (HM) using a human lung epithelial cell line, A549. Pregnant Sprague Dawley rats were divided into three groups: Control, LPS, and HW + LPS groups. Rats were injected with phosphate-buffered saline (Control) or LPS intraperitoneally (LPS) on gestational day 19 and provided H2 water (HW) ad libitum for 24 h before LPS injection (HW + LPS). Fetal lung samples were collected on day 20, and the levels of apoptosis, oxidative damage, IL-6, and vascular endothelial growth factor (VEGF) were evaluated using immunohistochemistry. The number of apoptotic cells, and levels of ROS and IL-6 were significantly increased by LPS treatment, and repressed following cultured with HM in A549 cells. In the rat models, the population positive for cleaved caspase-3, 8-hydroxy-2′-deoxyguanosine, IL-6, and VEGF was significantly increased in the LPS group compared with that observed in the Control group and significantly decreased in the HW + LPS group. In this study, LPS administration induced apoptosis and oxidative damage in fetal lung cells that was ameliorated by maternal H2 intake. Antenatal H2 administration may decrease the pulmonary mobility associated with inflammation in premature infants.

Increased expression of sphingosine kinase in the amnion during labor

INTRODUCTIONnSphingosine-1-phosphate (S1P), a bioactive lipid, has been reported to regulate inflammation processes. The onset of labor is thought to be related to inflammation. We therefore hypothesized that S1P might be involved in the onset of labor.nnnMETHODSnThe expression of sphingosine kinase (SPHK)-1, which produces S1P, and S1P lyase (SPL)-1, which irreversibly inactivates S1P, were examined in the fetal membranes. The expression levels were compared between amnions from cases of elective Caesarean deliveries (pre-labor) and those from vaginal deliveries (post-labor). In primary cultured human amnion cells, the expression levels of prostaglandin-endoperoxide synthase (PTGS)-2 were examined in the presence or absence of S1P treatment.nnnRESULTSnSPHK-1 and SPL-1 were both expressed in the amnion. The expression of SPHK-1 in the post-labor amnions increased compared with that in the pre-labor amnions. The expression of PTGS-2, a key regulator of labor, also increased in the post-labor amnion. However, the SPL-1 expression in the pre-labor amnion was not significantly different from that in the post-labor amnion. S1P1-3 and 5, which were coupled with Gi protein, were consistently found in the amnion cells. The treatment with S1P increased the expression of PTGS-2, and this was completely suppressed by a Gi inhibitor in the amnion cells.nnnDISCUSSIONnWe are herein provide the first evidence of increased SPHK-1 expression in post-labor amnions, and that S1P increases the PTGS-2 expression in amnion cells.nnnCONCLUSIONSnOur results suggest that S1P might play a role in the onset of labor via the induction of PTGS-2.

Neuroprotective potential of molecular hydrogen against perinatal brain injury via suppression of activated microglia.

Exposure to inflammation in utero is related to perinatal brain injury, which is itself associated with high rates of long-term morbidity and mortality in children. Novel therapeutic interventions during the perinatal period are required to prevent inflammation, but its pathogenesis is incompletely understood. Activated microglia are known to play a central role in brain injury by producing a variety of pro-inflammatory cytokines and releasing oxidative products. The study is aimed to investigate the preventative potential of molecular hydrogen (H2), which is an antioxidant and anti-inflammatory agent without mutagenicity. Pregnant ICR mice were injected with lipopolysaccharide (LPS) intraperitoneally on embryonic day 17 to create a model of perinatal brain injury caused by prenatal inflammation. In this model, the effect of maternal administration of hydrogen water (HW) on pups was also evaluated. The levels of pro-inflammatory cytokines, oxidative damage and activation of microglia were determined in the fetal brains. H2 reduced the LPS-induced expression of pro-inflammatory cytokines, oxidative damage and microglial activation in the fetal brains. Next, we investigated how H2 contributes to neuroprotection, focusing on microglia, using primary cultured microglia and neurons. H2 prevented LPS- or cytokine-induced generation of reactive oxidative species by microglia and reduced LPS-induced microglial neurotoxicity. Finally, we identified several molecules influenced by H2, involved in the process of activating microglia. These results suggested that H2 holds promise for the prevention of inflammation related to perinatal brain injury.

A case report of placental mesenchymal dysplasia with an increased VEGF-D expression

The pathogenesis of placental mesenchymal dysplasia (PMD) remains unclear. This report presents a case of PMD with a female fetus complicated with intrauterine growth restriction (IUGR). The ultrasound findings were similar to molar pregnancies, but PMD was suspected based on the presence of low β-hCG levels and a normal karyotype. After delivery, pathological examination of the placenta showed dilated villi and thick-walled vessels lacking trophoblast proliferation, which thus led to a diagnosis of PMD. The VEGF-D (Xp22.31) mRNA expression was found to have increased in the abnormal villi. Whether this is an incidental or X-linked gene specific event in, IUGR complicated, PMD pathogenesis warrants further investigation of VEGF-D expression in PMD.