Treatment of twin-reversed arterial perfusion sequence using high-intensity focused ultrasound.

We describe our experience of high-intensity focused ultrasound (HIFU) for fetal therapy in twin-reversed arterial perfusion (TRAP) sequence. Six pregnant women underwent HIFU therapy, five before 16 weeks and one at 26 weeks. Two types of HIFU system were used: the first-generation system, which comprised a biaxial transducer and continuous exposure pattern, and the second-generation system, which comprised a coaxial transducer and sequential exposure pattern. The first-generation apparatus was used in four cases and the second-generation apparatus was used in two. In three cases, occlusion of the blood vessels mediating flow to the acardiac twin was achieved by HIFU. Two cases experienced intrauterine fetal death despite vessel occlusion. The total survival rate of pump fetuses 2 years after HIFU was 67% and the efficiency rate (the proportion of cases with occlusion or reduced blood flow on ultrasound after HIFU) was 83%. After more than 2 years of follow-up, the surviving infants had no severe clinical complications and no postnatal developmental problems. There was no significant difference in survival rate compared with TRAP cases managed expectantly. Given that complete occlusion of the blood vessels was not achieved in half of the cases, we could not show that HIFU therapy is superior to other treatments. However, HIFU can reduce the cardiac load of the pump fetus and, as it does not require uterine puncture for fetal therapy, there were no fatal complications, such as bleeding, rupture of membranes or infection. Thus, HIFU therapy may represent a less-invasive treatment for TRAP sequence in early pregnancy. Copyright © 2018 ISUOG. Published by John Wiley & Sons Ltd.

First successful case of non-invasive in-utero treatment of twin reversed arterial perfusion sequence by high-intensity focused ultrasound.

High-intensity focused ultrasound (HIFU) has excellent potential as a non-invasive therapeutic tool in various fields of medicine. We present a case of twin reversed arterial perfusion sequence, in which non-invasive blood flow occlusion in the acardiac fetus was successfully achieved by means of HIFU exposure from outside the maternal abdomen. HIFU was applied to blood vessels of the acardiac fetus at the point at which the umbilical cord entered the body in a series of four procedures at 3-day intervals starting at 13 weeks' gestation, and in a final procedure with higher power at 17 weeks. The HIFU intensity was set at approximately 2300 W/cm(2) for the initial series of procedures and at 4600 W/cm(2) for the final procedure, with exposure periods of 10 s. As color Doppler examination revealed absence of blood flow to the acardiac fetus after the second round of HIFU exposure, we concluded that complete occlusion of target vessels had been achieved. Delivery was by Cesarean section at 37 weeks' gestation. A male neonate (the pump fetus) was born weighing 1903 g with Apgar scores of 8 and 9 at 1 and 5 min, respectively. At the time of writing, the baby was healthy and growing normally, with the exception of congenital pseudarthrosis.

High-intensity focused ultrasound treatment for twin reversed arterial perfusion sequence.

Twin reversed arterial perfusion (TRAP) sequence is a serious complication of monochorionic twin pregnancies, in which arterioarterial anastomoses allow blood flow from a 'pump' fetus to an acardiac fetus via reversed flow in the latter's umbilical artery. Several trial treatments for TRAP sequence have been reported, but all of these have been invasive. We present a case of TRAP sequence in which high-intensity focused ultrasound (HIFU) was applied to the umbilical artery of the anomalous twin at 26 weeks as a non-invasive fetal therapy. The HIFU intensity was set at approximately 2300 W/cm(2) with exposure periods of 10 s. Color Doppler ultrasound showed a decrease in blood supply to the anomalous twin, although complete occlusion of the targeted vessel was not achieved. Delivery was by Cesarean section at 29 weeks' gestation and the pump twin survived, without severe clinical complications at 6 months.

PP077. Effect of maternal antioxidant concentrations on expressions of angiogenesis-related genes in the villous trophoblasts from early pregnancy.

INTRODUCTION: The pathogenesis of preeclampsia is associated with oxidative stress of villous trophoblasts in early pregnancy. We have reported that antioxidant supplementations from 12weeks of gestation could prevent mild preeclampsia in pregnant women with oxidative stress. OBJECTIVES: To prove the effect of maternal antioxidant concentrations on expressions of angiogenesis-related genes in the villous trophoblasts from early pregnancy. METHODS: Villous tissue and blood samples were obtained from pregnant women who had artificial abortions between 6 and 11weeks of gestation. RNA was extracted from villous tissue, and gene expression levels of fms-like tyrosine kinase-1 (FLT-1), endoglin (ENG), and placental growth factor (PGF) in villous tissue were measured by reverse transcription polymerase chain reaction. Serum vitamin C and E concentrations were measured by enzyme-linked immunosorbent assay. This study was approved by the Ethics Committee of Showa University Hospital. RESULTS: Negative correlations between maternal serum vitamin C levels and gene expressions of ENG (r=-0.856, p=0.007) and FLT-1 (r=-0.898, p=0.002) in the villous trophoblasts were observed. There was no correlation of maternal serum vitamin E concentration with any other gene expression in the villous trophoblasts. CONCLUSION: It is suggested that maternal oxidative stress may increase the gene expressions of anti-angiogenic factors in villous trophoblasts of early gestation, which may lead to the pathogenesis of preeclampsia.

Location of the placenta or the umbilical cord insertion site in the lowest uterine segment is associated with low maternal blood pressure.

OBJECTIVE: To evaluate whether placental abnormalities and umbilical cord insertion site affect the occurrence of pre-eclampsia and maternal blood pressure. DESIGN: Case-control study. SETTING: Showa University Hospital, Tokyo, Japan. POPULATION: A total of 5722 consecutive women who delivered singleton infants were included in the study. METHODS: The associations of placental abnormalities, the location of the placenta, the location of the cord insertion site, and maternal background with the occurrence of pre-eclampsia and maternal blood pressure at the term of pregnancy were analysed. MAIN OUTCOME MEASURE: Pre-eclampsia and maternal blood pressure at the term of pregnancy. RESULTS: Pre-eclampsia was observed in 236 women (4.1%). Pre-eclampsia was frequently observed in women with placental form abnormalities (odds ratio 3.0) and infarction of the placenta (odds ratio 5.3). Pre-eclampsia was observed in 0 and 4.1% of women with and without placenta praevia, respectively (P = 0.004), and in 0 and 2.5% of women with and without low cord insertion during the first trimester, respectively (P = 0.018). After adjustment for confounding variables, the multivariate regression analyses revealed reductions of 8.4 and 5.0 mmHg in systolic and diastolic blood pressure, respectively, in women with placenta praevia compared with women without placenta praevia, and reductions of 4.3 and 3.1 mmHg in systolic and diastolic blood pressure, respectively, in women with low cord insertion during the first trimester compared with women without low cord insertion. CONCLUSION: Not only placenta praevia but also low cord insertion are associated with low frequencies of pre-eclampsia and low maternal blood pressure.

Detection of umbilical venous constriction by Doppler flow measurement at midgestation.

OBJECTIVES: To investigate whether umbilical venous velocity and venous velocity pulsation are associated with umbilical vein diameter, umbilical ring diameter and umbilical cord coiling index at midgestation. METHODS: Two hundred and eighty pregnant women were enrolled in the study at between 18 and 24 weeks of gestation. The diameter of the umbilical cord and internal diameter of the umbilical vein in a free loop and at the ring, and the umbilical coiling index, were measured using ultrasonography. Umbilical venous velocities were measured by Doppler ultrasonography at the umbilical ring and a free loop of the cord. RESULTS: All variables were successfully measured in 92% of the patients. There were negative correlations between the diameters of the umbilical ring and of the umbilical vein at the ring and the venous velocity at the umbilical ring. The venous velocity at the umbilical ring was significantly higher and the umbilical ring diameter was significantly lower in fetuses with umbilical venous pulsation at the free loop. Significant correlations were observed between the venous velocity and amplitude of pulsation. Venous pulsations at the free loop were frequently observed in fetuses with a hypercoiled cord. CONCLUSION: High venous velocity and increased venous pulsation at the umbilical ring may be associated with umbilical cord constriction.

Prediction of pre-eclampsia by an analysis of placenta-derived cellular mRNA in the blood of pregnant women at 15-20 weeks of gestation.

OBJECTIVE: A panel of cellular mRNA markers was used to predict the occurrence of pre-eclampsia in pregnant women at 15-20 weeks of gestation. DESIGN: Prospective cohort study. SETTING: The Department of Obstetrics and Gynaecology, University of Indonesia, Cipto Mangunkusumo National Hospital, Indonesia. SAMPLE: Peripheral blood samples from asymptomatic pregnant women. METHODS: Among 660 women, 62 developed pre-eclampsia at later gestation (pre-eclampsia group) and each case was matched with five controls. Therefore, the RNA expression levels in the cellular component of maternal blood in 62 women with pre-eclampsia were compared with those in 310 controls. MAIN OUTCOME MEASURES: The cellular RNA expression levels of genes related to angiogenesis and oxidative stress were compared between pre-eclampsia and control groups. A receiver operating characteristic (ROC) curve was used to analyse the sensitivity of each available marker. A logistic regression analysis was performed to calculate the odds for each woman to be classified as a case. RESULTS: The univariate ROC analysis identified soluble vascular endothelial growth factor receptor-1 (Flt-1) and endoglin (ENG) as the markers with the highest sensitivity. The best multivariate model was obtained by combining Flt-1, ENG, placental growth factor (PlGF) and parity. The relative ROC curve yielded a sensitivity of 66% at a 10% 1 - specificity rate with an area under the curve of 0.884 (P < 0.001). CONCLUSION: A panel of cellular mRNA markers in maternal blood can predict the development of pre-eclampsia long before clinical onset.

Predisposing factors for massive hemorrhage during Cesarean section in patients with placenta previa.

OBJECTIVES: To investigate whether maternal history and ultrasound findings can be predictors for massive hemorrhage during Cesarean section in patients with placenta previa and adherence of the placenta. METHODS: We reviewed 127 singleton pregnancies with placenta previa. Maternal history, antenatal ultrasound findings of the placenta, including location, presence of placental lacunae, lack of a clear zone, presence of sponge-like findings of the cervix and presence of a marginal sinus in cases of placenta previa were reviewed retrospectively, and their association with amount of bleeding during Cesarean section was analyzed. RESULTS: Logistic regression analysis revealed that advanced maternal age (odds ratio (OR), 5.4; 95% CI, 1.8-16.4), previous Cesarean section (OR, 20.4; 95% CI, 4.0-105.2) and sponge-like findings in the cervix (OR, 5.6; 95% CI, 1.8-17.0) were associated with massive bleeding (> 2500 mL). Placental adherence occurred in five cases and was more frequent in cases where the placenta was located at the site of the scar of a previous Cesarean section (OR, 123.1; 95% CI, 4.5-3395.2) and where there was lack of a clear zone (OR, 48.0; 95% CI, 3.8-604.7). CONCLUSIONS: Advanced maternal age, previous Cesarean section and presence of sponge-like findings in the cervix are risk factors for massive bleeding during Cesarean section in cases of placenta previa, regardless of whether placental adherence is present. Placental location on the scar of a previous Cesarean section and lack of a clear zone are risk factors for placental adherence. When these findings are identified preoperatively, management should be tailored accordingly.

Application of high-intensity focused ultrasound for umbilical artery occlusion in a rabbit model.

OBJECTIVES: To investigate the application of high-intensity focused ultrasound (HIFU) for fetal umbilical artery blood flow occlusion in a rabbit model. METHODS: A prototype HIFU transducer in combination with an imaging probe with Doppler capability was constructed. Using this transducer, HIFU was applied at 1.4, 2.75 or 5.5 kW/cm(2) through the maternal abdominal skin to the fetal intra-abdominal umbilical arteries of four time-mated Japanese White rabbits (11 fetuses) on gestational day 25. Courses of 5-s HIFU exposure were performed until cessation of umbilical blood flow and cardiac arrest were confirmed by Doppler ultrasonography. Fetal necropsy was performed and exposed lesions were assessed by microscopic histological analysis. RESULTS: The mean diameter of the fetal umbilical artery was 0.6 +/- 0.2 mm and the mean peak systolic velocity of arterial blood flow was 44.7 +/- 18.5 cm/s. When HIFU was applied at 5.5 kW/cm(2), blood flow was completely occluded within 15 courses. HIFU exposure brought about vacuolar degeneration and destruction of elastic fibers in the tunica media of the artery. CONCLUSIONS: HIFU can be used to occlude umbilical artery blood flow in fetal rabbits.

Recombinant human lactoferrin has a potential to suppresses uterine cervical ripening in preterm delivery in animal model.

OBJECTIVE: To investigate the effect of recombinant human lactoferrin (rhLF) on cervical ripening using a rabbit model in which preterm labor was induced by bacterial endotoxin lipopolysaccharide (LPS). STUDY DESIGN: Timed pregnant rabbits (New Zealand White, 3-4 kg, day 14) were randomly assigned to the following treatment groups: Group A, LPS + rhLF (n = 4); Group B, LPS (n = 4); and Group C, control (n = 4). Recombinant human lactoferrin (10 microg) was administrated to pregnant rabbits in Group A and not in Group B. Lipopolysaccharide (100 microg) was given to the rabbits in both groups for 3 days (days14-16). Drugs were administered as a vaginal suppository. On day 18, the rabbits were anesthetized with intramuscular ketamine hydrochloride (20 mg/kg) and diazepam (4 mg/kg). Both cervices of the rabbit uterus, which is bicorpus-bicolli, were taken out. One cervix was placed in 10% formalin solution for a histological study with standard hematoxylin-eosin staining. The other was used for an extension test to assess the grade of ripening. Extension was measured after a 5-mm length of cervical tissue was loaded with 5.8 g. RESULTS: The histological study showed remarkably loose and edematous connective tissue in Group B cervices. Cervical tissues in Group A was not different from those in Group C. Extension lengths were 2.2 +/- 0.2 mm in Group A, 7.0 +/- 2.7 mm in Group B, and 1.7 +/- 0.3 mm in Group C. CONCLUSION: These results suggest that rhLF inhibits cervical maturation induced by LPS in a rabbit model and may have a potential to prevent preterm delivery caused by cervical infection and ripening.

Measurement of mRNA of trophoblast-specific genes in cellular and plasma components of maternal blood.

BACKGROUND: Placental mRNA in maternal plasma is suitable for quantitative analysis regardless of fetal gender and genetic polymorphism status. METHODS: We obtained 155 blood samples from pregnant women to compare human placental lactogen (hPL) and beta-subunit of human chorionic gonadotropin (beta hCG) mRNA and protein levels between the cellular and plasma components of maternal blood. To assess clearance of hPL mRNA expression, we obtained blood samples from nine women immediately before and after delivery by caesarean section. mRNA was extracted from the cellular and plasma components of all samples, and hPL and beta hCG mRNA expression was analysed by reverse transcription-PCR assay. RESULTS: The concentration of beta hCG mRNA in the cellular component positively correlated with the plasma concentration of beta hCG protein and beta hCG mRNA (p = 0.001 for both). The concentration of hPL protein in the plasma correlated with the hPL mRNA concentration of the cellular component (p<0.05). For both hPL and beta hCG, the mRNA concentration of the cellular component was greater than that of the plasma component (22.9-fold higher for hPL and 4.3-fold higher for beta hCG). The half life of hPL mRNA clearance was significantly longer for the cellular fraction (mean half life = 203.8 min, range 150-3465 min) than for the plasma fraction (mean half life = 32.2 min, range 15-385 min) (p = 0.008). CONCLUSION: The present findings indicate that the concentration of hPL and beta hCG mRNA is significantly higher in the cellular component of maternal blood samples than in the plasma component. Cellular mRNA in maternal blood is useful for non-invasive evaluation of placental function.

Cord insertion into the lower third of the uterus in the first trimester is associated with placental and umbilical cord abnormalities.

OBJECTIVES: To assess the feasibility of detecting the cord insertion site during the late first trimester, and to investigate the possible association between perinatal complications and a cord insertion in the lower third of the uterus in the first trimester. METHODS: This was a prospective cohort study in which the positional relationship between the uterus and the cord insertion site was examined using gray-scale transvaginal sonography at 9-11 weeks of gestation. The distance between the internal os and the fundus was divided equally into three parts. Cord insertions located in the upper or middle thirds were defined as normal (controls), and those located in the lower third were defined as cases, regardless of their relationship to the chorion villosum. Third-trimester sonography and pathological examination of the placenta and cord at delivery were performed to check for placental/cord abnormalities. The univariate association between the location of the cord insertion in the first trimester and placental and umbilical cord abnormalities and perinatal complications was assessed. RESULTS: The cord insertion site was identified in 318/340 (93.5%) cases at 9-11 weeks of gestation and it was in the lower third of the uterus in 35 (11.0%) cases. Of these 35, the cord insertion was found at delivery to be low in nine cases (26%) and the placenta was low-lying at delivery in eight (23%). None of the 283 cases with a normal cord insertion in the first trimester was found to have a low-lying placenta at delivery (P < 0.0001). Ten (29%) of the cases with a low cord insertion and four (1.4%) with a normal cord insertion in the first trimester had a velamentous or marginal cord insertion at delivery (P < 0.0001). At delivery, five (14%) of the low cord insertion cases and four (1.4%) of the normal cord insertion cases (P < 0.0001) had some form of placental malformation, including accessory placenta and infarction of the placenta. An emergency Cesarean section was performed in four (11%) and six (2.1%) of the low and normal cord insertion cases, respectively (P = 0.003). CONCLUSION: Developmental abnormalities of the placenta and umbilical cord occur frequently in cases in which the cord insertion is in the lower third of the uterus in the first trimester. We suggest that screening for the cord insertion site at 9-11 weeks of gestation may have clinical significance for predicting abnormalities of the cord and the placenta at delivery.

Uptake of deuterium by dead leaves exposed to deuterated water vapor in a greenhouse at daytime and nighttime.

Dead leaves were exposed to deuterated water vapor (D(2)O) as a substitute of tritiated water (HTO) in a greenhouse at daytime and nighttime to examine uptake and release of tritium by dead leaves because they cover a wide area of the forest floor and are therefore a major target material to be exposed when HTO is atmospherically derived to the forest. The dead cedar needles showed faster uptake and faster release rates during and after the exposure than the fresh ones, and the equilibrium concentration of the dead cedar needles was about two times higher than the fresh ones, indicating a quick response and a high buffering potential of dead leaves. The relation between uptake of D(2)O and number of stoma was examined for dead deciduous leaves; the species with larger number of stoma accumulated more D(2)O at the daytime and nighttime exposures. However, drying of the dead leaves suppressed D(2)O uptake greatly at daytime, suggesting stomata's opening and closing controls the D(2)O uptake of dead leaves.

Velamentous cord insertion into the lower third of the uterus is associated with intrapartum fetal heart rate abnormalities.

OBJECTIVES: To evaluate the accuracy of sonographic identification of the site of umbilical cord insertion (CI) at 18-20 weeks of gestation, to compare the sensitivities for detection of a velamentous cord insertion (VCI) secondary to a CI into the anterior, posterior or fundal wall, and to compare the intrapartum complications secondary to VCI into the upper, middle or lower third of the uterus. METHODS: As part of the routine ultrasound scan at 18-20 weeks' gestation we evaluated abnormal CI (VCI and marginal CI) and the location of the CI in the uterus in 3446 pregnancies. In cases of abnormal CI, the location of the CI was further classified as being in the upper, middle or lower third of the uterus. After delivery, the placenta and the umbilical cord were examined and intrapartum complications were compared with the location of the CI. RESULTS: The values for antenatal detection of VCI were: sensitivity, 25 of 40 (62.5%); positive predictive value, 25 of 25 (100%); and negative predictive value, 3406 of 3421 (99.6%). The sensitivity for cases in which the CI was located on the anterior wall was 12 of 13 (92.3%); when it was located on the posterior wall, the sensitivity was 11 of 22 (50.0%); and when it was fundal the sensitivity was 2 of 5 (40.0%). Variable decelerations were frequently observed with a VCI. In lower VCI cases, non-reassuring fetal heart rate patterns and emergency Cesarean sections occurred with a higher frequency than in cases with upper or middle VCI (P < 0.01). After delivery, the length of the aberrant vessels in cases of VCI by pathologic examination was 3.9 +/- 3.3 cm in the upper third, 4.7 +/- 4.6 cm in the middle third, and 10.6 +/- 6.8 cm in the lower third; thus, the aberrant vessel length was significantly greater when the CI was in the lower third of the uterus (P = 0.024). CONCLUSION: We have demonstrated that VCI with a lower CI site and with longer aberrant vessels is associated with various intrapartum complications. This finding has the potential for improving perinatal outcome.

Cervical inflammatory cytokines and other markers in the cervical mucus of pregnant women with lower genital tract infection.

OBJECTIVE: To evaluate concentrations of interleukin (IL)-6, IL-8, lactoferrin (LF), and alpha defencine (alpha-DF) in the cervical mucus of pregnant women and analyze their relation to cervicitis and bacterial vaginosis (BV). METHODS: Cervical mucus samples were obtained from August 2003 through May 2004 from 157 women who were between the 6th and 36th week of an uncomplicated singleton pregnancy. All women were delivered at term, 69 without BV or cervicitis, 9 with BV, and 79 with cervicitis. RESULTS: Interleukin 8, LF, and mu-DF concentrations were higher in women with cervicitis (0.81 +/- 0.36 pg/mL, 14.8+/-12.3 microg/mL, and 0.60 +/- 0.49 microg/mL) than in women without BV or cervicitis (0.35 +/- 0.34 pg/mL, 8.0 +/- 11.0 microg/mL, and 0.15 +/- 0.12 microg/mL). Interleukin 6 concentration was higher in women with BV (0.26 +/- 0.32 pg/mL) than in women without BV or cervicitis (0.09 +/- 0.15 pg/mL) or in women with cervicitis (0.12 +/- 0.18 pg/mL). CONCLUSIONS: Higher levels of inflammatory cytokines in the cervical mucus of pregnant women may lead to early detection of lower genital tract infection.

Velamentous cord insertion and atypical variable decelerations with no accelerations.

OBJECTIVES: To examine intrapartum fetal heart rate (FHR) patterns in the presence of velamentous cord insertion (VCI). METHODS: The site of cord insertion was determined in 1460 women on antenatal ultrasonographic and postnatal examinations. Each of 24 women with pregnancies complicated with VCI was matched at the onset of labor with 10 women who had a normal pregnancy, and FHR patterns were analyzed for both cases and controls. RESULTS: Compared with controls, cases of variable decelerations with no accelerations (VDNA) during the first and second stages of labor were more frequent in women with VCI (first stage: 25% vs. 5.1%, second stage: 65.0% vs. 21.7%, P < 0.001). The adjusted odds ratio associated with VDNA occurrence in women with VCI was 3.83 (95% confidence interval [CI]: 1.51-9.72, P = 0.005). The mean odds ratio associated with length of aberrant vessels in women with VCI plus VDNA was 1.38 (95% CI: 1.04-1.83, P = 0.026). CONCLUSION: Pregnancies complicated with VCI are associated with a higher rate of VDNA.

Malignant transformation of endometriosis and genetic alterations of K-ras and microsatellite instability.

OBJECTIVES: To clarify the role of specific genetic alterations in the multi-step process of malignant transformation of endometriosis. METHODS: In cases of ovarian endometrioid carcinoma, we separated regions of normal endometriosis, atypical endometriosis and ovarian endometrioid carcinoma by laser microdissection, and examined K-ras mutation and microsatellite instability in each separated tissue sample. RESULTS: We detected K-ras mutation and microsatellite instability in endometrioid carcinoma tissue, but not in normal or atypical endometriosis bordering the cancerous region. CONCLUSIONS: The present findings suggest that K-ras mutation and microsatellite instability are associated with malignant transformation from atypical endometriosis to ovarian endometrioid carcinoma.

Detection of p53 mutations in the plasma DNA of patients with ovarian cancer.

Mutation of p53 is one of the most common genetic abnormalities detected in up to 81% of cases of ovarian cancer. To evaluate the use of plasma DNA analysis as a method for somatic mutation screening, we measured the presence of p53 mutations in DNA isolated from plasma and cancer tissue from patients with ovarian cancer. We analyzed the plasma DNA for the presence of p53 mutations (exons 5-8). Of 27 cases of ovarian cancer, 12 cases (44%) had mutations of p53 in cancer tissue. In two of the 12 cases (16.7%), identical mutations were detected in DNA of their preoperative plasma. In our follow-up of the two patients with p53 mutations in their plasma, mutant DNA was undetectable in their plasma after surgery. In one case, the p53 mutation re-surfaced in their plasma 16 months after surgery, and the patient died 2 months later. We have shown that tumor-derived DNA can be detected in the plasma of some patients with ovarian cancer, particularly in those with more advanced stage.