Search

Myelomeningocele is the most common congenital neurologic defect, and the only nonlethal disease addressed by fetal surgery. A randomized control trial has established amelioration of the Arnold-Chiari II malformation, reduced ventriculoperitoneal shunt rate, and improvement in distal neurologic function in patients that receive in utero repair. Long-term follow-up of these school-age children demonstrates the persistence of these effects. The use of stem cells in fetal repair is being investigated to further improve distal motor function.

Preoperative short cervical length (CL) remains a major risk factor for preterm birth after laser surgery for twin-twin transfusion syndrome (TTTS), but the optimal intervention to prolong pregnancy remains elusive. The objective of this study was to compare secondary methods for the prevention of preterm birth in twin pregnancies with TTTS undergoing fetoscopic laser photocoagulation (FLP), in the setting of a short cervix at the time of FLP, in five North American Fetal Treatment Network (NAFTNet) centers.This was a secondary analysis of data collected prospectively at five NAFTNet centers, conducted from January 2013 to March 2020. Inclusion criteria were a monochorionic diamniotic twin pregnancy complicated by TTTS, undergoing FLP, with preoperative CL 30 mm. Management options for a short cervix included expectant management, vaginal progesterone, pessary (Arabin, incontinence or Bioteque cup), cervical cerclage or a combination of two or more treatments. Patients were not included if the intervention was initiated solely on the basis of having a twin gestation rather than at the diagnosis of a short cervix. Demographics, ultrasound characteristics, operative data and outcomes were compared. The primary outcome was FLP-to-delivery interval. Propensity-score matching was performed, with each treatment group matched (1:1) to the expectant-management group for CL, in order to estimate the effect of each treatment on the FLP-to-delivery interval.A total of 255 women with a twin pregnancy complicated by TTTS and a short cervix undergoing FLP were included in the study. Of these, 151 (59%) were managed expectantly, 32 (13%) had vaginal progesterone only, 21 (8%) had pessary only, 21 (8%) had cervical cerclage only and 30 (12%) had a combination of treatments. A greater proportion of patients in the combined-treatment group had had a prior preterm birth compared with those in the expectant-management group (33% vs 9%; P = 0.01). Mean preoperative CL was shorter in the pessary, cervical-cerclage and combined-treatment groups (14-16 mm) than in the expectant-management and vaginal-progesterone groups (22 mm for both) (P 0.001). There was no significant difference in FLP-to-delivery interval between the groups, nor in gestational age at delivery or the rate of live birth or neonatal survival. Vaginal progesterone was associated with a decrease in the risk of delivery before 28 weeks' gestation compared with cervical cerclage and combined treatment (P = 0.03). Using propensity-score matching for CL, cervical cerclage was associated with a reduction in FLP-to-delivery interval of 13 days, as compared with expectant management.A large proportion of pregnancies with TTTS and a short maternal cervix undergoing FLP were managed expectantly for a short cervix, establishing a high (62%) risk of delivery before 32 weeks in this condition. No treatment that significantly improved outcome was identified; however, there were significant differences in potential confounders and there were also likely to be unmeasured confounders. Cervical cerclage should not be offered as a secondary prevention for preterm birth in twin pregnancies with TTTS and a short cervix undergoing FLP. A large randomized controlled trial is urgently needed to determine the effects of treatments for the prevention of preterm birth in these pregnancies. © 2021 International Society of Ultrasound in Obstetrics and Gynecology.

Introduction: We sought to characterize the current workforce and training for fetal intervention procedures at fetal centers in North America. Methods: An anonymous electronic survey was developed to query the 34 centers in the North American Fetal Treatment Network regarding the demographics and training of their faculty. Telephone surveys were conducted with directors of known fetal intervention fellowships. Results: More than 50% of maternal-fetal medicine (MFM) faculty at fetal centers were female; more than two-thirds of pediatric surgical faculty were male. Most of the training of faculty was undertaken by visiting more experienced centers or having new faculty work with more experienced ones at the same center. Current fetal intervention fellowships appear to achieve levels of competency for intrauterine transfusions and laser therapy for twin-twin transfusion syndrome. Two-thirds of centers stated that they would be able to offer a position to an MFM who completed a formal fellowship in fetal intervention. Conclusion: A collaborative effort should be undertaken to establish formal fellowships in fetal medicine and intervention.

Objectives Our primary aim was to confirm whether intertwin discordance in ventricular strain and ductus venosus (DV) time intervals predicts twin–twin transfusion syndrome (TTTS). Secondary aims were to create gestational‐age ranges for ventricular strain in uncomplicated monochorionic diamniotic (MCDA) twin pregnancies without selective intrauterine growth restriction (sIUGR) and to characterize the relationship of ventricular strain with gestational age in MCDA twin pregnancies with sIUGR that did not develop TTTS. Methods In the period 2015–2018, we enrolled 150 MCDA twin pregnancies consecutively into this prospective, blinded study of global longitudinal left and right ventricular strain. With the observer blinded to twin pairing and pregnancy outcome, videoclips of the four‐chamber view, which had been recorded during ultrasound surveillance in the usual window for development of TTTS (16–26 completed gestational weeks), underwent offline measurement of strain. Uncomplicated MCDA twin pregnancies, without sIUGR, were used to test the association between strain, gestational age and estimated fetal weight using mixed‐effects multilevel regression. Inter‐rater reliability was tested in 208 strain measurements in 31 fetuses from pregnancies which did not develop TTTS and within‐fetus variation was assessed in 16 such fetuses, in which multiple four‐chamber views were taken on the same day. The effect of sIUGR on strain in otherwise uncomplicated MCDA twin pregnancy was analyzed. MCDA twin pregnancies were defined as ‘pre‐TTTS’ when, having been referred for TTTS evaluation, they did not satisfy Quintero staging criteria, but subsequently developed TTTS requiring laser treatment. MCDA pregnancies which did not develop TTTS comprised the ‘non‐TTTS’ group. Cardiovascular parameters measured in these cases included tissue Doppler parameters and DV early filling time as a percentage of the cardiac cycle (DVeT%). Intertwin strain and DVeT% discordance was compared between non‐TTTS and pre‐TTTS cases, matched for gestational age. Results Paired strain data were available for intertwin comparison in 127/150 MCDA twin pregnancies, comprising 14 pre‐TTTS and 113 non‐TTTS pregnancies, after exclusions. Scans were collected at a median frame rate of 97 (range, 28–220) Hz. Laser therapy was performed at a median gestational age of 20.6 (range, 17.2–26.6) weeks. There were no group differences in right (RV) or left (LV) ventricular strain discordance between 68/113 non‐TTTS and 13/14 pre‐TTTS MCDA twin pregnancies 3.6% was found in eight of 13 pre‐TTTS pregnancies. In non‐TTTS pregnancies, the estimated variability in ventricular strain within each twin during the day was high (RV, 19.7; LV, 12.9). However, within each pair (intertwin variation), variability was low (RV, 5.5; LV, 2.9). Interclass correlation reflecting the proportion of total variability represented by the variability between twin pairs was low (RV, 0.22; LV, 0.18). Both RV (P

OBJECTIVES Fetal surgery for repair of open neural tube defect (ONTD) typically results in decreased need for a ventriculoperitoneal shunt (VPS). Our objectives were to determine the trend in ventricle size (VS) during pregnancy and whether VS and change in VS, as assessed by ultrasound, were predictive of the need for VPS in pregnancy with ONTD. METHODS This was a retrospective analysis of prospectively collected data of consecutive pregnancies with ONTD, evaluated in a single center from January 2012 to May 2018. Two groups were identified: the first consisted of pregnancies that underwent in-utero repair (IUR) and the second those that had postnatal repair (PNR). Penalized B splines were used to determine the trend in VS, across 2-week gestational-age (GA) epochs, between 24 and 36 weeks of gestation. VS at each GA epoch and the change in VS between each GA epoch were compared between the IUR and PNR groups. To determine whether VS at any GA was predictive of VPS, receiver-operating-characteristics (ROC) curves were used and the optimal cut-off at each GA epoch was identified. Univariate analysis and multiple logistic regression were used for further analysis. RESULTS ONTD was diagnosed in 110 fetuses, of whom 69 underwent IUR and 41 had PNR. Fetuses in the IUR group were more likely to have Chiari II malformation (100.0% vs 82.9%; P

Objectives To determine the compliance and effectiveness of fortnightly ultrasound surveillance for detection of twin-twin transfusion syndrome (TTTS) in monochorionic diamniotic (MCDA) twin gestations. Methods This is a retrospective study of ultrasound surveillance of MCDA twins for TTTS. Our surveillance protocol requires fortnightly ultrasounds starting at 16 weeks of gestational age (GA) continuing until delivery. Compliance was assessed by determining the GA of surveillance initiation and time between ultrasounds. GA and Quintero Stage at diagnosis were evaluated to determine whether TTTS was detected prior to advanced disease (Quintero Stage III +) or fetal demise. Results Of 442 women, 264 (59.7%) initiated surveillance after 16 weeks; follow-up ultrasounds were late in 17.4% of cases. TTTS was diagnosed in 43 (9.7%) women at a median GA of 19.7 [17.4, 23.9] weeks. Of 25/43 (58.1%) cases diagnosed during protocol compliance, 12 had advanced disease and two had fetal demise. A similar proportion of diagnoses (n = 18), made while non-compliant, exhibited advanced disease (11/18, 61.1%, P = .40). Thirteen diagnoses occurred during periods of increased ultrasound frequency due to abnormalities (ie, fluid/estimated fetal weight discrepancies or Doppler abnormalities). Conclusions In this population, fortnightly ultrasound compliance was suboptimal. Advanced disease and fetal demise occurred during protocol compliance.

OBJECTIVE Women with placenta accreta spectrum (PAS) having an unplanned delivery may have worse outcome compared with women with a planned delivery. The primary objective of this study was to compare severe maternal morbidity among women with PAS who had a planned scheduled delivery versus an unplanned delivery. Secondary objective was to compare neonatal outcomes. STUDY DESIGN Retrospective cohort study at two tertiary centers (January 2009 to June 2019) of all women who underwent a hysterectomy with a histologic proven PAS. Primary outcome was severe maternal morbidity which defined as any of the following: transfusion of ≥4 RBC units or ureter/bowel injury. Neonatal outcome was a composite neonatal morbidity defined as any of the following: Apgar score's < 5 at 5 minutes, mechanical ventilation, or respiratory distress syndrome. Maternal demographic, clinical, and sonographic characteristics were compared between the two groups (planned vs. unplanned). Descriptive statistics were used as appropriate, and a statistical significance was established if p-value was < 0.05. RESULTS Of 109 women who underwent cesarean hysterectomy for PAS, 41 (37.6%) had an unplanned delivery. There was no significant difference in the number of previous cesarean deliveries or ultrasound findings between the two groups. Women with an unplanned delivery were more likely to bleed during pregnancy than those that had a planned delivery (p = 0.04). Women with unplanned delivery had lower gestational age at delivery (30.3 vs. 33.8 weeks, p = 0.001) had a 75% higher rate of the primary outcome (63 vs. 36%, p = 0.007) and had a higher rate of intensive care unit admissions (39 vs. 17.7%, p = 0.01) compared with women with a planned delivery. The neonatal morbidity did not differ between the two groups. CONCLUSION Since unplanned cesarean hysterectomy among women with PAS occurs in 40% and is associated with significantly higher morbidity, interventions are needed to mitigate the rate of adverse outcomes. KEY POINTS · Only 60% of women with PAS reached planned delivery at 34 weeks.. · PAS unplanned delivery is associated with high morbidity.. · Some women with PAS may need a scheduled earlier delivery..

OBJECTIVEDespite significant improvement in spinal cord function after in utero spina bifida (SB) repair compared with traditional postnatal repair, over half of the children who undergo this procedure do not benefit completely. This lack of benefit has been attributed to closure methods of the defect, with subsequent spinal cord tethering at the repair site. Hence, a regenerative patch or material with antiinflammatory and anti-scarring properties may alleviate comorbidities with improved outcomes. The authors’ primary objective was therefore to compare cryopreserved human umbilical cord (HUC) versus acellular dermal matrix (ADM) patches for regenerative repair of in utero SB lesions in an animal model.METHODSIn vivo studies were conducted in retinoic acid–induced SB defects in fetuses of Sprague-Dawley rats. HUC or ADM patches were sutured over the SB defects at a gestational age of 20 days. Repaired SB defect tissues were harvested after 48–52 hours. Tissue sections were immunofluorescently stained for the presence of neutrophils, macrophages, keratinocytes, meningeal cells, and astrocytes and for any associated apoptosis. In vitro meningeal or keratinocyte cell coculture experiments with the ADM and HUC patches were performed. All experiments were scored quantitatively in a blinded manner.RESULTSNeutrophil counts and apoptotic cells were lower in the HUC-based repair group (n = 8) than in the ADM patch repair group (n = 7). In the HUC patch repair group, keratinocytes were present on the outer surface of the patch, meningeal cells were present on the inner surface of the patch adjacent to the neural placode, and astrocytes were noted to be absent. In the ADM patch repair group, all 3 cell types were present on both surfaces of the patch. In vitro studies showed that human meningeal cells grew preferentially on the mesenchymal side of the HUC patch, whereas keratinocytes showed tropism for the epithelial side, suggesting an inherent HUC-based cell polarity. In contrast, the ADM patch studies showed no polarity and decreased cellular infiltration.CONCLUSIONSThe HUC patch demonstrated reduced acute inflammation and apoptosis together with superior organization in regenerative cellular growth when compared with the ADM patch, and is therefore likely the better patch material for in utero SB defect repair. These properties may make the HUC biomaterial useful as a “meningeal patch” during spinal cord surgeries, thereby potentially reducing tethering and improving on spinal cord function.

Objective Preterm delivery following fetoscopic laser surgery (FLS) of twin–twin transfusion syndrome (TTTS) is associated with severe perinatal morbidity and mortality. The role of steroid hormones in amniotic fluid (AF) after FLS remains unknown. Study Design A prospective cohort study of consecutive case series of FLS for TTTS was performed from April 2012 to February 2017. Cases were divided into early (≤27 weeks) spontaneous preterm delivery (ED) and late delivery (LD; ≥34 weeks) following FLS and compared. AF supernatants were assessed for protein, estradiol, progesterone and cortisol levels (using the ELISA kit), and normalized to total protein levels to adjust for dilution. Results A total of 294 consecutive cases of FLS for TTTS in monochorionic–diamniotic twins were performed during the study period. AF was available in 44 ED patients and 50 LD patients. On logistic regression, ED was associated with higher normalized progesterone levels (odds ratio [OR]: 1.25; 95% confidence interval [CI]: 1.12–1.41), lower normalized cortisol (OR: 0.78; 95% CI: 0.64–0.96), and higher estradiol levels (OR: 1.3; 95% CI: 1.03–1.63). Conclusion Elevated AF normalized progesterone and estradiol, and lower normalized cortisol levels were associated with ED. This novel finding requires further exploration to establish the molecular mechanism operational in pregnancies complicated by TTTS to potentially prevent early preterm birth after fetal surgery.

OBJECTIVES To determine the relationship between intra-amniotic pressure and cervical length (CL) in patients with twin-twin transfusion syndrome (TTTS) undergoing fetoscopic laser photocoagulation (FLP), and to identify pre- or intraoperative factors associated with increased intra-amniotic pressure in this population. METHODS This was a prospective cohort study of patients undergoing FLP for TTTS. Exclusion criteria were triplet or higher-order gestation and prior cervical cerclage, amnioreduction or FLP procedure. CL was assessed using preprocedure transvaginal ultrasound. Intra-amniotic pressure measurements were obtained on initial placement of the trocar into the amniotic cavity, using a direct hydrostatic pressure gauge. The relationship between intra-amniotic pressure and CL was assessed using multivariate linear regression analysis, including relevant preoperative and intraoperative variables. RESULTS In total, 283 pregnancies met the inclusion criteria. Quintero stage of TTTS was I in 33 pregnancies, II in 88, III in 150 and IV in 12. Mean gestational age (GA) at FLP was 20.7 ± 3 weeks. Mean intra-amniotic pressure was 23.1 ± 9 mmHg. On unadjusted linear regression analysis, there was no significant association between intra-amniotic pressure and preoperative CL (P = 0.24) or GA at delivery (P = 0.22). On multivariate analysis, the factors associated significantly with intra-amniotic pressure were: number of prior term deliveries (P = 0.03), recipient maximum vertical pocket (P

OBJECTIVE We evaluated what prenatal genetic counselor's (GCs) practices, attitudes, and barriers are in regards to prenatal microarray since the publication of the American College of Obstetricians and Gynecologists (ACOG) and the Society for Maternal-Fetal Medicine (SMFM) guidelines for microarray use. METHODS This was a survey-based cross-sectional study of English-speaking, board certified or eligible GCs who currently practice prenatal genetic counseling. RESULTS Of 192 respondents, 183 (95%) have incorporated chromosome microarray (CMA) into clinical practice, with 64% believing that the benefits of CMA outweigh the harms and 52% agreeing that CMA should be offered to all women regardless of indication. Those who reported being experts/comfortable in their knowledge of CMA (85%) and familiar with current clinical guidelines (86%) were significantly more likely to offer CMA to patients undergoing invasive testing and patients with fetal anomalies. Patient-specific concerns were the largest reported barrier (51%) when GCs do not offer CMA to patients. CONCLUSION Our study demonstrates GCs follow guidelines for CMA use when specific indications are involved, but further guidelines are needed regarding CMA use for other routine indications where utility of CMA is not clearly understood. On this basis, ACOG and SMFM should continue revising their guidelines as more information comes to light regarding utility of prenatal CMA for all indications, and organizations like the National Society of Genetic Counselors (NSGC) should consider publishing guidelines on prenatal CMA that are specialized to the GCs sphere of practice.

OBJECTIVE: Donor demise after laser surgery for twin-twin transfusion syndrome (TTTS) is well-characterized, but recipient demise is not, nor is neonatal death. This study aims to characterize factors associated with recipient death, donor death, and dual twin death after laser, both before and after birth. METHODS: This is a prospective cohort study of monochorionic twin pairs who underwent laser ablation for TTTS. Risk factors for fetal and neonatal death of both twins were identified using univariable analysis and recursive partitioning, a novel statistical method to quantify contributions of each factor to outcomes. RESULTS: In 413 twin pairs, death of both twins occurred in 9.2% (38/413), donor death in 12.1% (50/413), and recipient death in 2.4% (10/413). Recursive partitioning showed that gestational age at delivery predicts dual twin death (below 23.7 weeks, likely [p

Background: Twin anemia polycythemia sequence is a chronic form of unbalanced fetofetal transfusion through minuscule placental anastomoses in monochorionic twins, leading to anemia in the donor and polycythemia in the recipient. Owing to the low incidence of twin anemia polycythemia sequence, data on diagnosis, management, and outcome are limited. Objective: This study aimed to investigate the diagnosis, management, and outcome in a large international cohort of spontaneous twin anemia polycythemia sequence. Study Design: Data from the international twin anemia polycythemia sequence registry, retrospectively collected between 2014 and 2019, were used for this study. A total of 17 fetal therapy centers contributed to the data collection. The primary outcomes were perinatal mortality and severe neonatal morbidity. Secondary outcomes included a risk factor analysis for perinatal mortality and severe neonatal morbidity. Results: A total of 249 cases of spontaneous twin anemia polycythemia sequence were included in this study, 219 (88%) of which were diagnosed antenatally and 30 (12%) postnatally. Twin anemia polycythemia sequence was diagnosed antenatally at a median gestational age of 23.7 weeks (interquartile range, 9.7–28.8; range, 15.1–35.3). Antenatal management included laser surgery in 39% (86 of 219), expectant management in 23% (51 of 219), delivery in 16% (34 of 219), intrauterine transfusion (with partial exchange transfusion) in 12% (26 of 219), selective feticide in 8% (18 of 219), and termination of pregnancy in 1% (3 of 219) of cases. Perinatal mortality rate was 15% (72 of 493) for the total group, 22% (54 of 243) for donors, and 7% (18 of 242) for recipients (P, SCOPUS: ar.j, info:eu-repo/semantics/published

A consensus definition of selective fetal growth restriction in monochorionic diamniotic twins was recently proposed following a Delphi procedure involving an international panel of experts. The new definition augments the traditional definition with additional sonographic criteria.We sought to determine whether the augmentations of the "Delphi definition" identified additional morbidity and mortality compared with a traditional definition. Furthermore, we sought to determine the benefit of each definition in identifying pathologic growth restriction relative to uncomplicated monochorionic diamniotic twins.This was a retrospective analysis of unselected monochorionic diamniotic twins that underwent fortnightly ultrasound surveillance at a single center between 2011 and 2020. Patients with concomitant twin-to-twin transfusion syndrome, twin anemia polycythemia sequence, or twin reversed arterial perfusion sequence at the time of diagnosis of selective fetal growth restriction were excluded. The diagnosis of selective fetal growth restriction using the Delphi definition required either an estimated fetal weight of3rd percentile or presence of 2 of 4 observations in the smaller twin: (1) estimated fetal weight of10th percentile, (2) estimated fetal weight discordance of25% compared with the larger twin, (3) abdominal circumference of10th percentile, (4) umbilical artery pulsatility index of95th percentile. Diagnosis using the traditional definition required an estimated fetal weight of10th percentile and an estimated fetal weight discordance of25%. To determine the efficacy of the augmentations in the Delphi definition, 3 groups were compared: group I, uncomplicated monochorionic diamniotic twins; group II, twins with selective fetal growth restriction using the traditional definition (and therefore the Delphi definition); and group III, twins with selective fetal growth restriction solely using the Delphi definition. Demographic characteristics, subsequent development of twin-to-twin transfusion syndrome or twin anemia polycythemia sequence, pregnancy outcomes, and neonatal outcomes were compared.There were 325 patients with monochorionic diamniotic twins that met inclusion criteria. Of these, 213 (66%; group I) were uncomplicated, 37 (11%; group II) met the traditional definition for selective fetal growth restriction, and 112 (35%) met the Delphi definition for selective fetal growth restriction with 75 (67%) meeting solely the Delphi definition (group III). Demographic characteristics were similar between groups. Patients in group II delivered earlier than uncomplicated twins (32.1 vs 35.7 weeks of gestation; P.01) and patients in group III (32.1 vs 35.6 weeks of gestation; P.01). Furthermore, they were more likely to have critical umbilical artery Doppler abnormalities (38% vs 4%; P.01) and be delivered for deteriorating fetal status (30% vs 5%; P.01) than those in group III. Overall, survival was lower in group II than groups I and III (89% vs 96% and 100%, respectively; P=.04). Moreover, composite neonatal morbidity and mortality were greater in group II (30%) than either group I (6%; P.01) or group III (9%; P.01). The rates of composite neonatal morbidity and mortality were similar between groups I and III (P=.28).The addition of abdominal circumference and umbilical artery pulsatility index thresholds and isolated estimated fetal weight of3%, as proposed by the Delphi definition, increased the diagnosis of selective fetal growth restriction; however, there was no added benefit in the identification of growth discordant pregnancies at risk of adverse outcomes. Prospective analysis of monochorionic diamniotic twins is required to contextualize these findings.

The aim of this study was to investigate the management and outcome in the post-laser twin anemia polycythemia sequence (TAPS). Data of the international TAPS Registry, collected between 2014 and 2019, were used for this study. The primary outcomes were perinatal mortality and severe neonatal morbidity. Secondary outcomes included a risk factor analysis for perinatal mortality and severe neonatal morbidity. A total of 164 post-laser TAPS pregnancies were included, of which 92% (151/164) were diagnosed antenatally and 8% (13/164) postnatally. The median number of days between laser for TTTS and detection of TAPS was 14 (IQR: 7&ndash, 28, range: 1&ndash, 119). Antenatal management included expectant management in 43% (62/151), intrauterine transfusion with or without partial exchange transfusion in 29% (44/151), repeated laser surgery in 15% (24/151), selective feticide in 7% (11/151), delivery in 6% (9/151), and termination of pregnancy in 1% (1/151). The median gestational age (GA) at birth was 31.7 weeks (IQR: 28.6&ndash, 33.7, range: 19.0&ndash, 41.3). The perinatal mortality rate was 25% (83/327) for the total group, 37% (61/164) for donors, and 14% (22/163) for recipients (p &lt, 0.001). Severe neonatal morbidity was detected in 40% (105/263) of the cohort and was similar for donors (43%, 51/118) and recipients (37%, 54/145), p = 0.568. Independent risk factors for spontaneous perinatal mortality were antenatal TAPS Stage 4 (OR = 3.4, 95%CI 1.4-26.0, p = 0.015), TAPS donor status (OR = 4.2, 95%CI 2.1&ndash, 8.3, p &lt, 0.001), and GA at birth (OR = 0.8, 95%CI 0.7&ndash, 0.9, p = 0.001). Severe neonatal morbidity was significantly associated with GA at birth (OR = 1.5, 95%CI 1.3&ndash, 1.7, p &lt, 0.001). In conclusion, post-laser TAPS most often occurs within one month after laser for TTTS, but may develop up to 17 weeks after initial surgery. Management is mostly expectant, but varies greatly, highlighting the lack of consensus on the optimal treatment and heterogeneity of the condition. Perinatal outcome is poor, particularly due to the high rate of perinatal mortality in donor twins.

OBJECTIVE The optimal outcome after fetoscopic laser surgery (FLS) for twin-twin transfusion syndrome (TTTS) depends on the successful ablation of all placental anastomoses. The objective of this study was to determine the incidence of and risk factors for recurrent TTTS (rTTTS) or twin anemia-polycythemia sequence (TAPS) after FLS, focusing on the impact of cannula diameter. METHODS This was a secondary analysis of data collected prospectively at two centers from 666 consecutive patients undergoing FLS for TTTS. The main outcomes were rTTTS and TAPS following FLS. Variables assessed included gestational age at intervention, stage of disease, recipient maximum vertical pocket, anterior placenta, number of anastomoses ablated, cannula diameter/operative scopes and use of the Solomon technique. Cannula diameter and corresponding scopes used were as follows: 8 Fr and 1.3 mm/0°; 9 Fr and 2.7 mm/0°; 10 Fr and 3 mm/0°; or 12 Fr and 3.3-3.7 mm/30-70°. Cannula diameter was used as a surrogate for scopes during analysis. Multivariate logistic regression analysis was performed to identify risk factors associated with rTTTS or TAPS after FLS; 'center' was considered an independent variable to account for variations in practice. In a nested cohort of pregnancies in which both fetuses survived, placental dye injection was performed in 315 placentae. Multivariate logistic regression analysis was performed to evaluate variables associated with the presence of residual anastomoses. RESULTS rTTTS or TAPS occurred in 61 (9%) cases following FLS (rTTTS in eight (1%) and TAPS in 53 (8%)). Factors associated significantly with the risk of rTTTS/TAPS on multivariate analysis were cannula diameter (when an 8-Fr, 9-Fr, 10-Fr or 12-Fr cannula was used, there was rTTTS/TAPS in 24%, 13%, 2% or 0.8% of cases, respectively (P

OBJECTIVE To compare perinatal outcomes between acute single fetal demise following fetoscopic laser photocoagulation to planned selective reduction (SR) in complicated monochorionic twin pregnancies. METHODS This was a secondary analysis of prospectively collected data in complicated monochorionic twin pregnancies from 2 fetal centers from 2011 to 2016. Group 1 included women undergoing fetoscopic laser photocoagulation for twin-twin transfusion syndrome (TTTS) who experienced a single fetal demise within 24 hours of the procedure. Group 2 consisted of women undergoing planned selective reduction (SR) with TTTS, and Group 3 SR for indications other than TTTS. RESULTS Survival of the remaining co-twin at birth was highest in Group 1 (n = 77 patients; 95%) compared with that in Group 2 (n = 15; 80%) and Group 3 (n = 32; 78%; P = .047). The preterm premature rupture of membrane rate was higher in Group 1 (47%) compared with that in Group 2 (33%) and Group 3 (7%; P

Haptoglobin (Hp) has key immunoregulatory roles that vary with phenotype (Hp1-1, Hp2-1, Hp2-2). Cord blood Hp expression is switched-off in the normal fetus. We hypothesized that in the setting of fetal inflammation placenta becomes inundated with Hp of fetal origin that in turn modulates the output of PGEPlacentas from 40 pregnancies complicated by preterm birth (PTB) (37 weeks), without (n = 15) or with (n = 25) intra-amniotic infection and histological chorioamnionitis (HCA) were scored for intensity of Hp immunostaining. Hp mRNA levels were evaluated by PCR. Cord blood Hp levels, switch-on status and phenotypes were determined by ELISA and Western blotting. Using a villous trophoblast explant system we investigated if Hp can modulate the release of PGEAll cases with HCA had positive Hp immunoreactivity within fetal vascular spaces. Hp staining intensity correlated with cord blood Hp levels and IL-6. Placentas with and without HCA had similar Hp mRNA levels suggesting Hp immunostaining in the fetal spaces is of fetal rather than placental origin. Both Hp1-1 and Hp2-2 up-regulated PGEFetal Hp switch-on status, a marker of antenatal exposure to intra-amniotic infection/inflammation, can be reliably established through evaluation of archived placental specimens. In the setting of infection/inflammation, Hp enhances placental PGE

Fetoscopic laser photocoagulation (FLP) of placental anastomoses is the preferred treatment for twin-to-twin transfusion syndrome (TTTS). Iatrogenic septostomy (IOS) during FLP is associated with increased risk of neonatal morbidity and mortality. We sought to identify risk factors for IOS and quantify the resultant outcomes. This is a secondary analysis of prospectively collected cases of TTTS in monochorionic diamniotic twins following FLP at a single center. Pre-operative ultrasound characteristics and operative technique (i.e., cannula size, total energy used) were compared between cases with vs. without IOS. Pregnancy and neonatal outcomes were also compared. Of 475 patients that had FLP, 33 (7%) were complicated by IOS. There was no association between operative technique and IOS. IOS was more common with later diagnosis, but less likely when selective fetal growth restriction (sFGR) was present. Survival was similar between groups (76% vs. 76% dual survivors, p = 0.95), however, IOS was associated with earlier delivery (29.7 vs. 32.0 wks, p &lt, 0.01) and greater composite neonatal morbidity (25% vs. 8% in both twins, p = 0.02). Risks of IOS at greater gestational ages without sFGR may be related to a larger collapsed intervening membrane area and the resulting increased risk of puncture on entry.

Objective To assess the incidence, clinical course, risk factors and outcomes of preterm prelabor rupture of membranes (PPROM) after fetoscopic laser surgery (FLS) for twin–twin transfusion syndrome (TTTS). Methods This was a prospective observational cohort study of 154 consecutive cases of TTTS. PPROM was defined as rupture of membranes before 34 weeks' gestation. Procedure-to-PPROM and PPROM-to-delivery intervals were determined. Relevant preoperative and intraoperative variables were analyzed by univariate and multivariate logistic regression to determine their impact on PPROM after FLS. Results The incidence of PPROM was 39% (n = 60), occurring at a mean gestational age of 27.2 ± 4.6 weeks. Median procedure-to-PPROM interval was 46 (range, 1–105; interquartile range (IQR), 13–66) days and median PPROM-to-delivery interval was 1 (range, 0–93; IQR, 0–13) day. Mean gestational age at delivery in cases with PPROM was 29.0 ± 4.5 weeks compared with 32.6 ± 3.9 weeks in cases without PPROM (P

Background: Fetoscopic laser photocoagulation (FLP) is the definitive treatment for twin-twin transfusion syndrome (TTTS). Due to variability in geographic proximity to high-volume fetal centers, many patients travel great distances to receive experienced care. We sought to determine whether distance traveled (DT) is associated with gestational age (GA) at delivery and neonatal survival. Methods: A prospective cohort study of patients within the continental United States referred to our center between September 23, 2011 and July 25, 2018 undergoing planned FLP for TTTS (n = 393; GA 20.6 ± 2.5 weeks; stage I: n = 50; stage II: n = 118; stage III: n = 208; stage IV: n = 17) was performed. The great-circle distance to our center was calculated using patients’ home zip codes. DT was stratified into groups containing equal patient numbers and pregnancy outcomes assessed. Results: A total of 393 patients met the inclusion criteria. The threshold distance from our center was n = 181), 250–499 miles (n= 119), and ≥500 miles (n = 93). There was no significant difference between any of the preoperative variables among the three groups, with the exception of race and rural status. Furthermore, there was no significant association between DT and GA at delivery (p = 0.34), time interval from procedure to delivery (p = 0.37), and the number of neonatal survivors (p= 0.21). Preterm premature rupture of membranes (PPROM) at p = 0.04) in the group traveling 250–499 miles. Conclusion: To our knowledge, this is the largest study to show that in TTTS, DT is not associated with GA at delivery, time interval from procedure to delivery, or neonatal survival. Although PPROM at

Background: Prenatal fractional limb volume (FLV) can be used to assess muscle atrophy in fetuses with myelomeningocele. Objective: We hypothesize that FLV in fetal myelomeningocele (fMMC) repair is different from postnatal repair (PNR). Assessing intrauterine muscle development can predict ambulation. Methods: A prospective observational study was performed from July 2012 to April 2016. Demographics, clinical outcomes, and FLV of the fetal thigh were assessed by ultrasound. Ambulation videos were collected from patients over 30 months of age. FLV was compared between the fMMC and PNR groups and between ambulators and non-ambulators. Two-sample t test, ANOVA, Spearman’s rho correlation, and Bland-Altman plots were used for analysis. A p value Results: Fifty-nine patients were included, 24 had fMMC and 35 had PNR. Videos were obtained in 47 cases (73%). There was no difference in baseline demographics between the groups. There was no significant change in the fMMC group between the FLV at initial presentation and the repeat at 34 weeks gestation (54.5 ± 28.2 and 62.2% ± 16.4; p = 0.6). In contrast, the FLV in the PNR decreased between the initial evaluation and the repeat at 34 weeks (54.1 ± 27.7 to 35.8 ± 34.1%; p = 0.04). FLV at 34 weeks gestation was higher in the fMMC group as compared to the PNR group (62.2 ± 16.4 vs. 35.8 ± 34.1%; p = 0.02). There was no difference in FLV between ambulators and non-ambulators either at initial evaluation (p = 0.8) or at 34 weeks gestation (p = 0.6). Conclusion: Lower FLV in the PNR group compared to fMMC may suggest in utero muscle atrophy. No correlation was seen between FLV and subsequent ambulation; however, future larger studies may be needed.

STUDY QUESTION Do management strategies (treatment type and order), including provision of reproductive counseling, differ in patients with non-tubal pregnancies? SUMMARY ANSWER Medical and surgical treatment strategies varied widely for each type of non-tubal pregnancy and reproductive counseling in this patient population is lacking. WHAT IS KNOWN ALREADY Owing to the rarity of non-tubal pregnancies, there is no consensus regarding treatment strategies or protocol. Furthermore, there is limited data on how patients with a non-tubal pregnancy are counseled about future fertility. STUDY DESIGN, SIZE, DURATION This is a descriptive retrospective study. Data were collected from January 2006 to December 2017. A total of 50 patients were included in the study. PARTICIPANTS/MATERIALS, SETTING, METHODS Patients with an ultrasound diagnosis of a non-tubal ectopic pregnancy (e.g. cervical ectopic pregnancy [CEP], Caesarean scar pregnancy [CSP] or interstitial ectopic pregnancy [IEP]) were included. This study was performed at a university-based institution tertiary referral center. Demographic and clinical characteristics, treatment type and order, reproductive counseling and outcomes were collected. Descriptive statistics were used for analyses. MAIN RESULTS AND THE ROLE OF CHANCE Of the 50 patients identified, 13 were CEP (26%), 8 were CSP (16%) and 29 were IEP (58%). Patients with a CSP had a higher parity (median = 3, P = 0.02) and number of prior Caesarean deliveries (mean = 2.1, P LIMITATIONS, REASONS FOR CAUTION This study was performed in a tertiary referral center therefore the management strategy could have been influenced by the prior interventions and patient response. The descriptive retrospective design precluded any assumption of causation. WIDER IMPLICATIONS OF THE FINDINGS The management for non-tubal pregnancies has wide variations. Reproductive counseling in this patient population is lacking. The findings highlight the need for the development of a treatment algorithm and a reproductive counseling protocol for each non-tubal pregnancy to better standardize treatment strategy. STUDY FUNDING/COMPETING INTEREST(S) There was no funding for this study. The authors have no conflict of interest to report.

Preterm birth remains a major complication of fetal laser surgery (FLS) due to twin-to-twin transfusion syndrome (TTTS). Objectives: We tested the hypothesis that neonatal outcomes in fetuses born at >24 weeks are worse in TTTS survivors compared to uncomplicated monochorionic diamniotic (MCDA) twins. Methods: 196 patients with TTTS treated with laser therapy and 91 uncomplicated MCDA gestations were compared. Neonatal outcomes included respiratory distress syndrome (RDS), transient tachypnea of the newborn (TTN), bronchopulmonary dysplasia, intraventricular hemorrhage, necrotizing enterocolitis, and neonatal death. Risk factors assessed were TTTS, maternal age, maternal body mass index, race, premature prolonged rupture of membranes, stage of TTTS, parity, and gestational age (GA) at delivery. Results: GA at delivery was lower in the TTTS group (31.0 ± 4.6 vs. 33.5 ± 2.4 weeks, p < 0.001). RDS and TTN occurred at higher rates in the TTTS than in the uncomplicated MCDA group. After multivariate logistic regression, the only factor significantly associated with the composite adverse outcome was GA at delivery (OR 0.61; 95% CI: 0.58–0.7). Conclusion: TTTS twins treated with FLS are deliver 2.5 weeks earlier than uncomplicated MCDA twins. Respiratory complications were significantly higher in TTTS twins and were mainly the consequence of the early GA at delivery.

Introduction: Optimal uterine relaxation is achieved through higher minimum alveolar concentration (MAC) of inhalational anesthetics, increasing risks to mother and fetus. Our objective was to determine if earlier administration of magnesium sulfate would reduce the requirement of inhalational anesthetics in fetal myelomeningocele repair. Materials and Methods: Prospective observational study of fetal myelomeningocele repair was performed from September 2011 to August 2017. Groups compared were: (1) magnesium sulfate at uterine closure; (2) magnesium sulfate at maternal skin incision. Maternal demographics, anesthetic agents, intraoperative complications, and pregnancy outcomes were reviewed. A Student t test was used for analysis. Results: There were 30 cases in group 1 and 21 cases in group 2. There was no difference in gestational age at intervention (24.92 ± 0.62 vs. 25.22 ± 0.47 weeks, p = 0.07) or at delivery (34.83 ± 2.77 vs. 33.98 ± 3.83 weeks, p = 0.38) between groups 1 and 2, respectively. The maximum MAC of sevoflurane was significantly lower in group 2 (1.84 ± 0.25 vs. 1.05 ± 0.28, p < 0.0001). There was no difference in the average dose of phenylephrine used. Discussion: Magnesium sulfate infusion initiated earlier in open fetal surgery reduces the total anesthetic exposure to the fetus.

We investigated the ability of cryopreserved human amniotic membrane (hAM) scaffold sealed with an underwater adhesive, bio-inspired by marine sandcastle worms to promote healing of iatrogenic fetal membrane defects in a pregnant swine model.Twelve Yucatan miniature pigs underwent laparotomy under general anesthesia at 70 days gestation (term = 114 days). The gestational sacs were assigned to uninstrumented (n = 24) and instrumented with 12 Fr trocar, which was further randomized into four different arms-no hAM patch, (n = 22), hAM patch secured with suture (n = 16), hAM patch with no suture (n = 14), and hAM patch secured with adhesive (n = 9). The animals were euthanized 20 days after the procedure. Gross and histological examination of the entry site was performed for fetal membrane healing.There were no differences in fetal survival, amniotic fluid levels, or dye-leakage from the amniotic cavity between the groups. The fetal membranes spontaneously healed in instrumented sacs without hAM patches. In sacs with hAM patches secured with sutures, the patch was incorporated into the swine fetal membranes. In sacs with hAM patches without sutures, 100% of the patches were displaced from the defect site, whereas in sacs with hAM patches secured with adhesive 55% of the patches remained in place and showed complete healing (p = 0.04).In contrast to humans, swine fetal membranes heal spontaneously after an iatrogenic injury and thus not an adequate model. hAM patches became incorporated into the defect site by cellular ingrowth from the fetal membranes. The bioinspired adhesive adhered the hAM patches within the defect site.

OBJECTIVE: To assess the incidence, timing, and risk factors for death of the donor fetus after fetoscopic laser surgery, we evaluated our cohort of patients who underwent the procedure for twin-twin transfusion syndrome. METHODS: This was a prospective cohort study of 166 consecutive patients with twin-twin transfusion syndrome at a single center. Fetal death was diagnosed by ultrasonography after surgery and before onset of labor. Risk factors for death of the donor twin were identified on univariate analysis and then subjected to multivariate, stepwise, logistic regression analysis. RESULTS: Donor demise occurred in 20 (13%) cases and recipient twin death occurred in four (2.6%). The median procedure to death interval was 4 days (range 1-89 days). Risk factors for donor death were fetal growth discordance greater than 30% (odds ratio [OR] 6.7, 95% confidence interval [CI] 2-23), reverse end-diastolic velocity in the donor umbilical artery (OR 25.0, 95% CI 2-290), a marginal and velamentous cord insertion (OR 4.4, 95% CI 1-19), and an increased number of anastomoses (OR 1.2, 95% CI 1.1-1.5). All four donors with both fetal growth discordance greater than 30% and reverse end-diastolic velocity in the donor umbilical artery resulted in a demise. CONCLUSION: Four risk factors significantly affecting acute and delayed donor demise after fetoscopic laser surgery were identified. The presence of both fetal growth discordance greater than 30% and reverse end-diastolic velocity in the donor umbilical artery was highly predictive of donor demise in our cohort. Knowledge of these risk factors can aid in counseling and assist patients in choosing the most appropriate intervention in the management of twin-twin transfusion syndrome. LEVEL OF EVIDENCE: III.

Background/Aims: Aortic distension waveforms describe the change in diameter or cross-sectional area over the cardiac cycle. We aimed to validate the association of aortic fractional area change (AFAC) with blood pressure (BP) in a fetal lamb model. Methods: Four pregnant ewes underwent open fetal surgery under general anesthesia at 107-120 gestational days. A 4-Fr catheter was introduced into the fetal femoral artery and vein, or the carotid artery and jugular vein. The thoracic aorta was imaged using real-time ultrasound; AFAC was calculated using offline speckle tracking software. Measurements of invasive BP and AFAC were obtained simultaneously and averaged over 10 cardiac cycles. BP was increased by norepinephrine infusion and the association of aortic distensibility with BP was assessed. Results: Baseline measurements were obtained from 4 lambs, and changes in aortic distensibility with increasing BP were recorded from 3 of them. A positive correlation was found between AFAC and systolic BP (r = 0.692, p = 0.001), diastolic BP (r = 0.647, p = 0.004), mean BP (r = 0.692, p = 0.001), and BP amplitude (r = 0.558, p = 0.016) controlled for heart rate. No association was found between BP and maximum or minimum aortic area. Conclusion: AFAC provides a quantifiable measure of aortic distensibility and correlates with systolic BP, diastolic BP, mean BP, and BP amplitude in a fetal lamb model.

Objective To compare outcome of dichorionic–triamniotic (DCTA) triplet gestations with that of a paired cohort of monochorionic–diamniotic (MCDA) twin gestations undergoing laser therapy for treatment of twin-to-twin transfusion syndrome (TTTS). Methods All DCTA triplets treated at four referral centers between specified dates were included. A matched cohort group of MCDA twin gestations affected by TTTS was used for comparison. Results Sixteen sets of DCTA triplets treated with laser photocoagulation of placental anastomoses were compared with 32 matched sets of MCDA twins. All of the ‘singleton’ fetuses in the triplet gestations survived to birth and to the end of the neonatal period. There were no differences in single or double survival or the rate of non-survivors to birth and at age 28 days. A mean 3-week difference was found in the procedure-to-delivery interval in DCTA triplets vs MCDA twins (60 ± 35.8 days vs 83.6 ± 33.2 days; P = 0.029). A mean difference of 3 weeks was also found in the gestational age at delivery in DCTA triplets vs MCDA twins (28.5 ± 3.5 weeks vs 31.9 ± 5 weeks; P = 0.024). A similar post-laser fetal growth pattern in donors and recipients of both groups was noted. Conclusions DCTA triplet gestations with TTTS have a similar rate of post-laser survival but deliver earlier than can be expected for twins treated with laser. These findings have potential implications for the risk of prematurity and long-term outcomes. Copyright © 2014 ISUOG. Published by John Wiley & Sons Ltd.

Preterm birth after fetoscopic laser surgery for twin-twin transfusion continues to be a major challenge despite improved survival rates. The risk factors for the complications have been identified, which include preterm premature rupture of membranes, cervical length shortening, larger cannula diameter, amnioinfusion, complicated procedure, and severe disease. The interventions to prevent preterm birth such as cervical cerclage for short cervix and collagen plug placement have not been successful. Further understanding of the pathophysiology is urgently needed to identify newer methods to prevent preterm birth.

Objective Despite improved perinatal survival following fetoscopic laser ablation (FLA) for twin–twin transfusion syndrome (TTTS), prematurity remains an important contributor to perinatal mortality and morbidity. The objective of the study was to identify risk factors for complicated preterm delivery after FLA. Methods Retrospective cohort study of prospectively collected data on maternal/fetal demographics and pre-operative, operative and postoperative variables of 459 patients treated with FLA in three USA fetal centers. Multivariate linear regression was performed to identify significant risk factors associated with preterm delivery, which were cross-validated using the k-fold method. Multivariate logistic regression was performed to identify risk factors for early compared with late preterm delivery based on median gestational age at delivery of 32 weeks. Results There were significant differences in case selection and outcomes between the centers. After controlling for the center of surgery, multivariate analysis indicated that a lower maternal age at procedure, a history of previous prematurity, shortened cervical length, use of amnioinfusion, a cannula diameter of 12 French (Fr), lack of a collagen plug placement and iatrogenic preterm premature rupture of membranes (iPPROM) were significantly associated with a lower gestational age at delivery. Conclusions Specific fetal/maternal and operative variables are associated with preterm delivery after FLA for the treatment of TTTS. Further studies to modify some of these variables may decrease the perinatal morbidity after laser therapy. Copyright © 2013 ISUOG. Published by John Wiley & Sons Ltd.

Prenatal spina bifida (SB) repair with a regenerative patch may improve neurological outcomes by decreasing inflammatory scarring.This study aims to compare cryopreserved human umbilical cord (HUC) and biocellulose film (BCF) patches sutured over SB lesions for regeneration of native cells and inflammatory response.Sprague-Dawley rats were gavaged with retinoic acid (RA) on embryonic day 10 to induce SB. Hysterotomy was performed on embryonic day 20 and on HUC or BCF patches sutured over the defect. Pups were harvested 30 to 34 h later, and hematoxylin and eosin staining and Trichrome staining assessed basic cellular migration. Immunohistochemistry demonstrated the exact nature of the cellular migration. Patches and surrounding exudates were evaluated with microscopy and cells quantified.Histology showed cellular migration with all HUC patches compared with none with BCF patches. Epithelial cells were noted migrating over the dorsal HUC surface, astrocytes were noted along the HUC surface adjacent to the lesion, and endothelial cells were noted within the HUC. HUC patches showed minimal inflammatory cells. Exudates surrounding the HUC patches had fewer inflammatory cells than exudates around BCF patches.HUC promotes cellular migration of native cells with minimal inflammatory response compared with BCF. HUC may be the superior patch material for prenatal SB repair. © 2017 John WileySons, Ltd.

There is an urgent need is for a regenerative patch for in utero repair of spina bifida in situations in which primary closure is difficult.We present two cases of large myeloschisis defects that underwent in utero spina bifida repair at midgestation with closure of the skin defect by using a cryopreserved human umbilical cord patch. The pregnancies were uncomplicated, and the births occurred at 37 weeks by planned cesarean delivery. The repair sites were intact with no evidence of cerebrospinal fluid leakage, with skin regenerated after delivery over a period of 3-4 weeks. The short-term outcomes after delivery showed reversal of hindbrain herniation, minimal spinal cord tethering, and normal function of the lower extremities.Cryopreserved human umbilical cord may be a suitable patch system for the spina bifida closure.

Objective Preterm delivery after fetoscopic laser surgery (FLS) for twin–twin transfusion syndrome (TTTS) is a major complication. The causative factors leading to preterm delivery continue to be elusive and a better understanding of the risk factors could reduce complications. The objective of this study was to determine the etiology of preterm delivery after FLS for TTTS and its associated risk factors. Methods This was a secondary analysis of a prospective study of 203 patients with TTTS who underwent FLS at a single center between September 2011 and December 2014. Preoperative, operative, postoperative, delivery and neonatal data were reviewed. Preterm delivery was categorized according to etiology into three groups: spontaneous (SPT), indicated (IND) and elective (ELC). Comparisons between groups were performed by ANOVA. Kaplan–Meier survival analysis was performed to compare the procedure-to-delivery interval between groups. To identify risk factors for preterm delivery, logistic regression, with calculation of relative risks (RR), was performed, with P

To estimate the response in lung growth and vascularity after fetal endoscopic tracheal occlusion for severe congenital diaphragmatic hernia in the prediction of neonatal survival.Between January 2006 and December 2010, fetal lung parameters (observed-to-expected lung-to-head ratio; observed-to-expected lung volume; and contralateral lung vascularization index) were evaluated before fetal tracheal occlusion and were evaluated longitudinally every 2 weeks in 72 fetuses with severe isolated congenital diaphragmatic hernia. Thirty-five fetuses underwent fetal endoscopic tracheal occlusion and 37 cases did not.Survival rate was significantly higher in the fetal endoscopic tracheal occlusion group (54.3%) than in the no fetal endoscopic tracheal occlusion group (5.4%, P.01). Fetal endoscopic tracheal occlusion resulted in a significant improvement in fetal lung size and pulmonary vascularity when compared with fetuses that did not go to the fetal intervention (increase of the observed-to-expected lung-to-head ratio, observed-to-expected total lung volume, and contralateral pulmonary vascularization index 56.2% compared with 0.3%, 37.9% compared with 0.1%, and 98.6% compared with 0.0%, respectively; P.01). Receiver operating characteristic curves indicated that the observed-to-expected total fetal lung volume was the single best predictor of neonatal survival before fetal endoscopic tracheal occlusion (cutoff 0.23, area under the curve [AUC] 0.88, relative risk 5.3, 95% confidence interval [CI] 1.4-19.7). However, the contralateral lung vascularization index at 4 weeks after fetal endoscopic tracheal occlusion was more accurate in the prediction of neonatal outcome (cutoff 24.0%, AUC 0.98, relative risk 9.9, 95% CI 1.5-66.9) with the combination of observed-to-expected lung volumes and contralateral lung vascularization index at 4 weeks being the best predictor of outcome (AUC 0.98, relative risk 16.6, 95% CI 2.5-112.3).Fetal endoscopic tracheal occlusion improves survival rate by increasing the lung size and pulmonary vascularity in fetuses with severe congenital diaphragmatic hernia. The pulmonary response after fetal endoscopic tracheal occlusion can be used to predict neonatal survival.

Laser ablation of anastomosis is one method for treatment of twin-twin transfusion syndrome in monochorionic pregnancies. Sonographic mapping of the umbilical cord insertions is an important aspect of the preoperative workup before laser ablation. Proximate umbilical cord insertions can be potential contraindications to laser ablation. This series discusses 6 such cases.

Objective To examine the incidence of preterm premature rupture of membranes (PPROM) in pregnancies affected by twin–twin transfusion syndrome (TTTS) treated with laser photocoagulation where an absorbable gelatin sponge was used as a chorioamnion sealant of the fetoscopic access port. Method A retrospective review was undertaken of consecutive cases undergoing fetoscopic directed laser surgery for TTTS between October 2006 and November 2008 at Texas Children's Fetal Center, in which an absorbable gelatin sponge, used as a chorioamnion ‘plug’, was placed at the conclusion of the intervention as a possible prophylactic measure to prevent PPROM. We excluded cases that had a failure of plug placement and those in which it was not attempted. PPROM was defined as rupture of membranes before 34 weeks' gestation. A comparison was performed between the PPROM group and a no-PPROM group to determine risk factors and outcomes. Results Successful plug placement occurred in 79 of 84 cases (94%) in which it was attempted after laser surgery, with a rate of PPROM of 34% in these patients. PPROM occurred at an average gestational age of 26.5 ± 3.6 weeks, with an average procedure-to-PPROM interval of 5.2 ± 3.5 weeks. There were no statistically significant differences between the PPROM group and the no-PPROM group in maternal demographics or preoperative parameters including: amniotic fluid volumes in the recipient twin's gestational sac, volume of amnioreduction, and location of the placenta. The procedure-to-delivery interval for the total cohort (n = 79) was 9.2 ± 4.7 weeks, without a significant difference between the two groups (P = 0.08). However, after exclusion of one PPROM outlier, the PPROM group had an average procedure-to-delivery time 2 weeks shorter than the group with no PPROM (P = 0.03). The live birth rates were similar in the PPROM and no-PPROM groups, at 77 and 73%, respectively. However, the average recipient's weight in the PPROM group was significantly lower than in the no-PPROM group (1321 ± 493 vs. 1705 ± 576 g; P = 0.02). Conclusion The rate of PPROM and the mean gestational age at delivery in pregnancies in which an absorbable gelatin sponge was used as a sealant of the fetoscopic port following laser photocoagulation for TTTS were comparable to those that have been reported by other laser centers where membrane sealants were not used. A randomized controlled trial should be considered to evaluate the effect of chorioamnion plugging. Copyright © 2010 ISUOG. Published by John Wiley & Sons, Ltd.

Objective To compare a laparoscopy-assisted fetoscopic approach with an ultrasound-directed percutaneous approach for laser photocoagulation of placental anastomoses in cases of twin–twin transfusion syndrome (TTTS) with anterior placentation. Method We performed a retrospective review of all cases that underwent laser ablation of placental anastomoses for TTTS with an anterior placenta at Texas Children's Fetal Center from November 2006 to November 2008. The two cohorts were identified by chart review based on the type of approach: laparoscopy-assisted vs. ultrasound-guided percutaneous uterine entry for fetoscopy. Operative and outcome data were extracted and the groups were compared using statistical methods, taking P < 0.05 as statistically significant. Results In the 100 cases of TTTS studied, 48 had an anterior placenta. Fifteen (31%) of these underwent laparoscopy-assisted fetoscopy (LAF) while a percutaneous approach was used in the remaining 33 (69%) cases. The total procedure time was longer in the LAF group than in the percutaneous group (96.1 ± 25 vs. 67.9 ± 28 min; P < 0.01). There was no difference in the rate of preterm premature rupture of membranes up to 2 weeks and 4 weeks after surgery (7 vs. 15% and 13 vs. 21%, for the LAF group vs. the percutaneous group, respectively; P = 0.7). The gestational ages at delivery were similar: 30.3 ± 4.5 weeks in the LAF group and 29.2 ± 4.6 weeks in the percutaneous group (P = 0.32). The overall survival rate at birth was tending towards better survival in the laparoscopic group than in the percutaneous group (80 vs. 61%, respectively; P = 0.06). The neonatal survival rate was better with the LAF approach than with the percutaneous approach (80 vs. 59%, respectively; P = 0.045). Conclusion Laparoscopy-assisted entry of the uterus is associated with improved neonatal survival for laser photocoagulation in cases of TTTS with a complete anterior placentation. Copyright © 2009 ISUOG. Published by John Wiley & Sons, Ltd.

Objective Twin anemia-polycythemia sequence (TAPS) can occur as a unique disease or as a complication of twin-twin transfusion syndrome (TTTS). Middle cerebral artery (MCA) Doppler studies are not currently part of the routine evaluation of monochorionic twins since they are not used in the Quintero staging system. As such, the true incidence of TAPS is unknown. We aimed to compare the characteristics and outcomes of twin pregnancies with TTTS complicated by spontaneous anemia-polycythemia vs those with TTTS alone. Methods This was a secondary analysis of data collected prospectively from a cohort of 156 consecutive patients undergoing fetoscopic laser surgery for TTTS, between October 2011 and August 2014. TAPS was defined as discordance in the preoperative MCA peak systolic velocity (PSV), with one twin fetus having MCA-PSV ≤ 1.0 multiples of the median (MoM) and the other having MCA-PSV ≥ 1.5 MoM. Maternal demographics as well as preoperative, operative and postoperative variables were analyzed. Results Included in the final analysis were 133 patients with complete records: 11 cases with TTTS with anemia-polycythemia and 122 cases with TTTS alone. There was no difference in maternal body mass index, gestational age (GA) at procedure, rate of preterm prelabor rupture of membranes or GA at delivery between the two groups. Patients with TTTS and anemia-polycythemia were more likely to be older (P = 0.03) and parous (P = 0.04) and had a significantly lower number of placental anastomoses (P = 0.01). The dual live-birth rate was similar for both groups (P = 0.76). Conclusion Cases of TTTS with anemia-polycythemia were more likely to be found in parous and older women and were characterized by fewer vascular anastomoses. TTTS with anemia-polycythemia was not associated with worse perinatal outcome after laser therapy. Copyright © 2015 ISUOG. Published by John Wiley & Sons Ltd.

Traditionally, 2-dimensional ultrasound parameters have been used for the diagnosis of a suspected morbidly adherent placenta previa. More objective techniques have not been well studied yet.The objective of the study was to determine the ability of prenatal 3-dimensional power Doppler analysis of flow and vascular indices to predict the morbidly adherent placenta objectively.A prospective cohort study was performed in women between 28 and 32 gestational weeks with known placenta previa. Patients underwent a two-dimensional gray-scale ultrasound that determined management decisions. 3-Dimensional power Doppler volumes were obtained during the same examination and vascular, flow, and vascular flow indices were calculated after manual tracing of the viewed placenta in the sweep; data were blinded to obstetricians. Morbidly adherent placenta was confirmed by histology. Severe morbidly adherent placenta was defined as increta/percreta on histology, blood loss2000 mL, and2 units of PRBC transfused. Sensitivities, specificities, predictive values, and likelihood ratios were calculated. Student t and χThe following results were found: (1) 50 women were studied: 23 had morbidly adherent placenta, of which 12 (52.2%) were severe morbidly adherent placenta; (2) 2-dimensional parameters diagnosed morbidly adherent placenta with a sensitivity of 82.6% (95% confidence interval, 60.4-94.2), a specificity of 88.9% (95% confidence interval, 69.7-97.1), a positive predictive value of 86.3% (95% confidence interval, 64.0-96.4), a negative predictive value of 85.7% (95% confidence interval, 66.4-95.3), a positive likelihood ratio of 7.4 (95% confidence interval, 2.5-21.9), and a negative likelihood ratio of 0.2 (95% confidence interval, 0.08-0.48); (3) mean values of the vascular index (32.8 ± 7.4) and the vascular flow index (14.2 ± 3.8) were higher in morbidly adherent placenta (P.001); (4) area under the receiver-operating characteristic curve for the vascular and vascular flow indices were 0.99 and 0.97, respectively; (5) the vascular index ≥21 predicted morbidly adherent placenta with a sensitivity and a specificity of 95% (95% confidence interval, 88.2-96.9) and 91%, respectively (95% confidence interval, 87.5-92.4), 92% positive predictive value (95% confidence interval, 85.5-94.3), 90% negative predictive value (95% confidence interval, 79.9-95.3), positive likelihood ratio of 10.55 (95% confidence interval, 7.06-12.75), and negative likelihood ratio of 0.05 (95% confidence interval, 0.03-0.13); and (6) for the severe morbidly adherent placenta, 2-dimensional ultrasound had a sensitivity of 33.3% (95% confidence interval, 11.3-64.6), a specificity of 81.8% (95% confidence interval, 47.8-96.8), a positive predictive value of 66.7% (95% confidence interval, 24.1-94.1), a negative predictive value of 52.9% (95% confidence interval, 28.5-76.1), a positive likelihood ratio of 1.83 (95% confidence interval, 0.41-8.11), and a negative likelihood ratio of 0.81 (95% confidence interval, 0.52-1.26). A vascular index ≥31 predicted the diagnosis of a severe morbidly adherent placenta with a 100% sensitivity (95% confidence interval, 72-100), a 90% specificity (95% confidence interval, 81.7-93.8), an 88% positive predictive value (95% confidence interval, 55.0-91.3), a 100% negative predictive value (95% confidence interval, 90.9-100), a positive likelihood ratio of 10.0 (95% confidence interval, 3.93-16.13), and a negative likelihood ratio of 0 (95% confidence interval, 0-0.34). Intrarater and interrater agreements were 94% (P.001) and 93% (P.001), respectively.The vascular index accurately predicts the morbidly adherent placenta in patients with placenta previa. In addition, 3-dimensional power Doppler vascular and vascular flow indices were more predictive of severe cases of morbidly adherent placenta compared with 2-dimensional ultrasound. This objective technique may limit the variations in diagnosing morbidly adherent placenta because of the subjectivity of 2-dimensional ultrasound interpretations.

Background Chronic hypertension in adults causes arterial lengthening in major arteries, but the effects of early fetal hypertension on the twin-twin transfusion syndrome recipient's vascular architecture remains unknown. Objective We hypothesize that arterial cord redundancy is related to recipient hypertension and subsequent heart failure. Our objectives were to: (1) establish a 3-dimensional color Doppler ultrasound method of measuring umbilical arterial length relative to its corresponding venous segment in the umbilical cord using artery vein angle; (2) compare recipient artery vein angle to gestational age-matched controls; and (3) test the association of artery vein angle with recipient heart failure. Study Design We compared 3 groups prospectively: twin-twin transfusion syndrome pregnancies undergoing fetoscopic laser surgery (preoperatively) and 2 groups of gestational age-matched controls: uncomplicated monochorionic-diamniotic twin pregnancies and healthy singletons. Using a 3-dimensional color-Doppler volume image of 5 cm of cord near the placental insertion, we traced the umbilical artery and vein producing umbilical artery:vein length, (artery vein index) and measured the artery vein angle between umbilical artery and vein. Correlation of artery vein angle to twin-twin transfusion syndrome stage, maximum vertical pocket, umbilical arterial indices, ductus venosus Doppler, and brain natriuretic peptide were performed. We used pulsed-wave and tissue Doppler to measure tissue Doppler velocities and indexed cardiac output and correlated these with artery vein angle. Comparative statistics, including multivariable linear regression, examined the relationship between umbilical arterial Doppler indices and artery vein angle. Results Artery vein angle and artery vein index correlated significantly ( R 2 , 0.86; P P 95th percentile for controls) was measured in 52% recipients. Artery vein angle was higher in twin-twin transfusion syndrome stage 3R vs 1 ( P = .001). Artery vein angle increased with increasing umbilical arterial pulsatility index ( P P = .02) after adjusting for gestational age. Interrater agreements to categorize abnormal artery vein angle values was 95% ( P Conclusion Umbilical arterial lengthening occurs in 52% of recipients and is associated with abnormal Doppler flows, low systolic tissue Doppler velocities, reduced cardiac output, and elevated markers of cardiac failure. This may reflect chronicity and severity of hypertension in the recipient fetus. Further research is needed to explore the mechanisms of elongation and long-term implications.

Preterm premature rupture of membranes remains a major complication after fetoscopic laser surgery (FLS) for twin-twin transfusion syndrome (TTTS). We studied the histologic changes of fetal membranes post-FLS and investigated a possible impact of amniotic fluid (AF) dilution.Fetal membranes of 31 pregnancies that underwent FLS for TTTS were investigated histologically at delivery at different sites: trocar site of recipient sac and at distance, donor sac, and inter-twin membrane.The trocar insertion site on the recipient sac showed no signs of histologic hallmarks of healing. Wide-spread alteration in collagen organization and higher apoptotic index in the amnion of the recipient sac which were absent in donor's and reference membranes. To explain the mechanisms, we analyzed the AF composition of recipient sacs from TTTS pregnancies vs. GA-matched healthy singleton controls and found glucose, protein and lactate dehydrogenase activity were all significantly lower in TTTS sacs consistent with over-dilution of recipient's AF (~2-fold). In-vitro exposure of healthy amniochorion to analogous dilutional stress conditions recapitulated the histologic changes and induced apoptosis and autophagy.Alteration in structural integrity of the recipient's amniochorion, possibly in response to dilution stress, along with ineffective repair mechanisms may explain the increased incidence of preterm birth post-FLS.

Objective Despite fetoscopic laser ablation (FLA) having emerged as an effective treatment for twin–twin transfusion syndrome (TTTS), major postintervention challenges, such as iatrogenic preterm premature rupture of membranes (iPPROM), remain. A chorioamniotic plug (CAP) made of absorbable gelatin sponge has been used to seal the trocar entry site in the chorioamniotic layers to promote healing and prevent iPPROM, yet the results have been equivocal. Our objective was to test the hypothesis that, following FLA for TTTS, iPPROM may be prevented by placement of an absorbable gelatin CAP. Methods A retrospective cohort study was performed on prospectively collected data from 134 consecutive patients who underwent FLA for TTTS. The decision for CAP placement was at the discretion of the physician. Preoperative, operative and postoperative variables were collected and analyzed. The primary outcome was the incidence of iPPROM, and the secondary outcomes were procedure-to-delivery interval and gestational age at delivery. Comparative statistical analysis was performed as appropriate. Results A CAP was placed in 72 (54%) of 134 patients. Factors significantly associated with CAP placement were higher gravidity (P = 0.03), anterior placentation (P = 0.04), general endotracheal intubation (P = 0.02) and a cannula diameter of 12-Fr (P = 0.003). There were no differences between CAP and no-CAP groups in the rate of iPPROM (39% vs 34%, respectively; P = 0.42) or in the procedure-to-delivery interval (65.3 ± 34.7 days vs 58.2 ± 30.8 days, respectively; P = 0.21). The gestational age at delivery was later in the CAP group compared with the no-CAP group (30.7 ± 4.5 weeks vs 28.9 ± 3.9 weeks, respectively; P = 0.02). Conclusions CAP did not reduce the overall risk for iPPROM and did not increase the procedure-to-delivery interval. Further research is needed to identify other methods to prevent iPPROM and prolong pregnancy after laser therapy. Copyright © 2013 ISUOG. Published by John Wiley & Sons Ltd.

The aim of this study was to develop a simple clinical algorithm for prediction of donor and recipient death using 'yes'or 'no' questions through the process of recursive partitioning for patients undergoing laser therapy for twin to twin transfusion syndrome (TTTS). The intent was to identify a subset of patients with very high specificity to whom clinical decisions would be simplified.Secondary analysis of data retrospectively collected from laser procedures was performed for TTTS at NAFTNet centers from 2002 to 2009. Preoperative factors associated with donor and recipient death were identified by recursive partitioning regression analysis. Classification And Regression Trees (CARTs) were developed to refine specificity for prediction of death.There were 466 TTTS patients from eight centers. CARTs were obtained for prediction of donor death. Improved specificity was achieved through recursive partitioning as demonstrated in receiver operator characteristic curves for prediction of death of the donor. There was less than optimal predictive ability for prediction of death in the recipient, as demonstrated by lack of generation of CARTs.Recursive partitioning improves the specificity and refines the prediction of donor fetal and neonatal demise in TTTS treated with laser therapy. This has the potential to improve therapeutic choices and refine counseling regarding outcomes.

The purpose of this study is to evaluate the accuracy of prenatal diagnostic features, particularly congenital cystic adenomatoid malformation volume ratio (CVR), in predicting outcomes for fetuses with lung masses.The records and imaging features of all fetuses referred to the Texas Children's Fetal Center with a fetal lung mass between July 2001 and May 2010 were reviewed retrospectively. Data collected included gestational age (GA) at diagnosis, fetal magnetic resonance imaging findings, CVR, mass size, nature of fetal treatment, surgical findings, pathology, and outcome. Data were analyzed for predicting development of hydrops or the need for fetal therapy using receiver operating characteristic curves.Of 82 fetuses (41 male) evaluated for a lung mass, 53 (65%) were left-sided (1 bilateral), and the mean (SD) GA at diagnosis was 21.5 (4.3) weeks. Seventy-three fetuses underwent fetal magnetic resonance imaging at a mean (SD) GA of 26.1 (4.6) weeks. Thirteen fetuses (16%) had fetal treatment. Four fetuses with hydrops underwent open fetal surgical resection, and 3 survived. Six fetuses with large lung masses and persistent mediastinal compression near term underwent ex-utero intrapartum therapy-to-resection procedures, and 3 fetuses with hydrops underwent serial thoracentesis. Congenital cystic adenomatoid malformation volume ratio correlated strongly with the development of hydrops and the need for fetal therapy with an area under the receiver operating characteristic curve of 0.96 (P.0001) and 0.88 (P.0001), respectively. Of 18 fetuses with a CVR greater than 2.0 compared with 2 (3%) of 60 with a CVR of 2.0 or less, 10 (56%) required fetal intervention (P.0001).Congenital cystic adenomatoid malformation volume ratio correlates strongly with the development of fetal hydrops and the need for fetal intervention. A threshold value of 2.0 yields the most powerful statistical results.

To assess serial changes in the Tei index of the recipient fetus secondary to fetoscopic laser photocoagulation of placental anastomoses (FLP) for twin-twin transfusion syndrome (TTTS) during the peri-operative period.A prospective study of cases that underwent FLP at the Texas Children's Fetal Center was performed. Mean and individual changes in the Tei index measurements were obtained during the pre-operative, intra-operative and the post-operative (12-h and 24-h) periods after surgery and were compared using paired t-test. A p-value of0.05 was considered as significant.A total of 20 patients were included in the study. In both the right and the left side, the mean Tei indices appeared to be increased from baseline during the intra-operative period, with a subsequent decrease to pre-operative values within 24 h of laser surgery. These changes were also noted when individualized patient results were analyzed in a serial fashion.FLP results in a transient increase in the intra-operative myocardial performance index in the recipient fetus. These changes appear to resolve in the post-operative period.

Objective We sought to determine the learning curve (LC) for fetoscopic laser photocoagulation (FLP) as a model for the evaluation of training in surgical procedures. Study Design A retrospective review of consecutive case series of FLP from 2 centers with 3 operators (operator I [O-I], observer trained; operator II [O-II], hands-on trained; and operator III [O-III], clinical fellow) was performed and the LC-cumulative summation (CUSUM) test was plotted. Results The acceptable and unacceptable success rates for at least 1 fetus survival after FLP were set at 82% and 70%, respectively, from a systematic review. A total of 171 consecutive cases were performed by the 3 operators (O-I, 91; O-II, 49; and O-III, 31). From LC-CUSUM test O-I needed 60 procedures, O-II needed 20 procedures, and O-III needed 20 procedures to reach an acceptable performance rate for at least 1 survivor. Conclusion The LC-CUSUM test can be used to accurately assess the LC in a surgical procedure in obstetrics and gynecology. Hands-on trained operators exhibit a shorter LC.

To compare the outcomes in chorioamnion membrane separation with nonchorioamnion membrane separation cases after fetoscopic laser photocoagulation of placental anastomoses for the treatment of twin to twin transfusion syndrome.We performed a retrospective cohort study of twin to twin transfusion syndrome cases that underwent laser ablation at the Texas Children's Fetal Center from October 2007 to March 2009. The cases were identified from our database. We excluded triplet pregnancies. Chorioamnion membrane separation was identified by ultrasound examination performed within 24 hours of intervention. Preterm premature rupture of membranes (PROM) was defined as rupture of the membranes before 34 weeks of gestation.Ninety-seven cases were included in the analysis; 19 (19.6%) had chorioamnion membrane separation. There were no significant differences in preoperative and intraoperative variables, except for a lower recipient maximum vertical pocket (10.3+/-2.1 compared with 12.6+/-3.4; P.01), and higher rate of iatrogenic septostomy (21% compared with 2.5%; P=.01) in the chorioamnion membrane separation group. Postoperatively, the incidence of preterm PROM was 74% in chorioamnion membrane separation group compared with 23% in the nonchorioamnion membrane separation group (relative risk 3.2, 95% confidence interval 1.9-4.4). The procedure-to-delivery interval was reduced by 20 days in the chorioamnion membrane separation group (P.01). The neonatal survival rate was 63.2% in the chorioamnion membrane separation group compared with 84% in the nonchorioamnion membrane separation group (P=.016).A low preoperative recipient maximum vertical pocket is associated with increased risk of chorioamnion membrane separation. Postoperative chorioamnion membrane separation after laser surgery for twin to twin transfusion syndrome is a major risk factor for preterm PROM, early delivery, and an increase in perinatal mortality.II.

Objective To determine practice patterns for evaluation and treatment of Stage I twin-to-twin transfusion syndrome (TTTS) among international physicians. Methods An e-mail cross-sectional survey of members from the IFMSS, NAFTNet and SMFM societies and participants at a Eurofoetus-sponsored TTTS meeting was undertaken between May 2008 and November 2008. Questionnaires consisted of physician demographics and their recommendations for managing Stage I TTTS. Alternative therapies to expectant management were assessed based on the following special circumstances of the patient: residence more than 200 miles from the center, severe symptoms, or a cervical length of ≤ 15 mm. Results Eighty-one surveys were returned, giving a response rate of 84%. Five surveys were excluded as a result of duplication or missing data. Of the remaining 76 surveys, 48 were from North America, 20 were from Europe and eight were from other continents. Expectant management was the predominant recommendation (78%), followed by amnioreduction (11%), laser ablation (11%) and septostomy (1%). Recommendations for amnioreduction were exclusively from North American centers. Laser centers recommended expectant management more frequently than non-laser facilities (89% vs. 59%; P < 0.01). When examples of special patient circumstances were presented, North American centers changed their recommendation from expectant management to amnioreduction more often than did European centers. However, a greater proportion of European centers recommended laser surgery for special patient circumstances. Conclusion Expectant management remains the predominant management of Stage I TTTS. In some patient circumstances, North American centers are more likely to recommend amnioreduction while European centers are more likely to recommend laser therapy. A randomized controlled trial will be necessary to evaluate the most efficacious management strategy for Stage I TTTS. Copyright © 2010 ISUOG. Published by John Wiley & Sons, Ltd.